STEM Today

Special Edition on Long Duration Spaceflight

STEM TODAY March 2016, No. 6

STEM TODAY March 2016 , No. 6

CONTENTS Analysis and Evaluation of the Functional State of the Cardiovascular System in Cosmonauts Cardiac, arterial and venous adaptation to weightlessness during 6-month MIR spaceflights with and without thigh cuffs (bracelets) Autonomic regulation of circulation and cardiac contractility during a 1 4-month space flight Changes in basal heart rate in spaceflights up to 438 days Main medical results of extended flights on space station Mir in 1 986-1 990 Affect of Microgravity on Cardiac Shape Cardiac and vascular responses to thigh cuffs and respiratory maneuvers Proteomic Profiling of Human Heart Tissue Exposed to Microgravity Effects of Sex and Gender on Adaptation to Space: Cardiovascular Alterations Effect of geomagnetic activity factors on the cardiac rhythm regulation and arterial pressure of cosmonauts

Editorial Editor: Mr. Abhishek Kumar Sinha Advisor: Mr. Martin Cabaniss

STEM Today, March 2016, No.6

Cover Page Mars Global View of Valles Marineris Mosaic of the Valles Marineris hemisphere of Mars projected into point perspective, a view similar to that which one would see from a spacecraft. The distance is 2500 kilometers from the surface of the planet, with the scale being .6km/pixel. The mosaic is composed of 102 Viking Orbiter images of Mars. The center of the scene (lat -8, long 78) shows the entire Valles Marineris canyon system, over 2000 kilometers long and up to 8 kilometers deep, extending form Noctis Labyrinthus, the arcuate system of graben to the west, to the chaotic terrain to the east. Many huge ancient river channels begin from the chaotic terrain from north-central canyons and run north. The three Tharsis volcanoes (dark red spots), each about 25 kilometers high, are visible to the west.

South of Valles Marineris is very ancient terrain covered by many impact craters.

Image Credit: NASA/JPL-Caltech Background NASA Unveils Celestial Fireworks as Official Image for Hubble 25th Anniversary NASA and ESA are celebrating the Hubble Space Telescope’s silver anniversary of 25 years in space by unveiling some of nature’s own fireworks - a giant cluster of about 3,000 stars called Westerlund 2. The cluster resides inside a vibrant stellar breeding ground known as Gum 29, located 20,000 light-years away in the constellation Carina. The comparatively young, 2-million-year-old star cluster contains some of our galaxy’s hottest, brightest, and most massive stars. The largest stars are unleashing a torrent of ultraviolet light and hurricane-force winds that etch away the enveloping hydrogen gas cloud. This creates a fantasy celestial landscape of pillars, ridges, and valleys.

Image Credit: NASA, ESA, the Hubble Heritage Team (STScI/AURA), A. Nota (ESA/STScI), and the Westerlund 2 Science Team Back Cover Global mosaic of Mars.Cerberus region Mosaic of the Cerberus hemisphere of Mars projected into point perspective, a view similar to that which one would see from a spacecraft. The viewer’s distance is 2,000 kilometers above the surface of the planet. The mosaic is composed of 104 Viking Orbiter Images. The images were acquired on February 11, 1980 during orbit 1,323 of Viking Orbiter 1. At that time, it was early northern summer on Mars (aerocentric solar longitude 65 degrees) and the sub-solar declination was 22.6 degrees N. The center of this image is at latitude 12 degrees and longitude 190 degrees. There are thin white clouds dispersed over the northern hemisphere.

Image Credit: NASA.

STEM Today , March 2016

Special Edition on Long Duration Spaceflight Cardiovascular System

At present, there is little evidence suggesting that cardiovascular adaptation to microgravity or space flight increases susceptibility to life threatening arrhythmias in astronauts. From a clinical perspective, according to the "biological model" of sudden cardiac death, both the substrate and the trigger for arrhythmias should be considered to determine whether long-term space flight could lead to an increased risk of sudden death. In this model, structural abnormalities interact with functional alterations, such as exercise, electrolyte disturbances, or neurohumoral modulation, to create an environment in which arrhythmias can be initiated and/or sustained.

In patients with coronary artery disease, the substrate is clear: a myocardial infarction (MI) and/or scar leading to focal areas of slowed conduction, a necessary condition for re-entry. For patients with apparently normal ventricular function, the potential substrate is less certain. In fact, reentry often is not the mechanism of arrhythmia development in these clinical cases: the arrhythmias may be caused by delayed after-depolarizations, and the triggered activity may be mediated via catecholamines. The published report of non-sustained ventricular tachycardia during prolonged space flight supports this hypothesis, in that initiation of tachycardia by a late diastolic premature ventricular contraction (PVC) is more consistent with triggered activity than it is with re-entry.

While there are no definitive data showing that long-duration space flight is associated with cardiac arrhythmias, there are observational data that have been documented over many years that are suggestive of cardiac electrical changes during long flights.

STEM Today, March 2016, No.6

Skylab For example, during Skylab, all 9 American crewmembers exhibited some form of rhythm disturbance.

Most of these rhythm disturbances consisted of single PVCs and were clinically insignificant. However, one crewmember experienced a 5-beat run of ventricular tachycardia during a lower-body negative pressure protocol, and another had periods of "wandering supraventricular pacemaker" during rest and following exercise.

More recently, it has been shown that the corrected QT interval (QTc), a marker of ventricular repolarization, was prolonged slightly in a small number of astronauts after long-duration space flight. In-flight Holter monitoring was not performed during these space flights. Thus, it is not known whether this prolongation was associated with any known arrhythmias. In-flight Holter monitoring was undertaken in the early Space Shuttle era.

Virtually no changes in arrhythmias were documented in flights of 4 to 16 days during either intravehicular or extravehicular operations compared to preflight measurements. Indeed, in these studies, the frequency of arrhythmias may actually have been reduced in flight, though the day-to-day variability of these arrhythmias, which is known to be quite wide, was not quantified. However, aboard the Mir space station, PVCs were detected that were not present before flight and a 14-beat run of ventricular tachycardia was documented.

More recently, several conditions that may predispose crewmembers to arrhythmias have been identified. After long-duration missions QTc intervals are slightly prolonged in crewmembers who did not have prolonged QTc intervals after their short-duration Space Shuttle flights, and several investigators have found decreases in left ventricular mass following space flight.

All of these findings raise the concern that cardiac rhythm disturbances may become an issue during the long in-flight tours of duty planned for ISS and interplanetary missions. The degree to which space flight and its many variables can be considered arrhythmogenic is not clear, but the possibility that serious cardiac rhythm disturbances might occur during space flight is a concern to NASA. Evidence Space Flight There have been no systematic studies of the arrhythmogenic potential of long-duration space flight, and only two studies of short-duration space flight. There have been, however, a number of published reports detailing in-flight arrhythmias.

Page 2

Special Edition on Long Duration Spaceflight One crewmember during Apollo 15 experienced a 22-beat nodal bigeminal rhythm, which was followed by premature atrial beats.This crewmember reported extreme fatigue during the incident, but only when questioned about it by crew surgeons; thus, it was not severe enough to impact the mission. Twenty-one months later the crew member suffered from coronary artery disease and a cardiac infarction without suggestive ECG changes.

STEM Today, March 2016, No.6

Apollo 15 ECG tracings obtained during periods of cardiac arrhythmia. (a) shows the normal heart beat converting to a nodal bigemini rhythm; (b) shows the bigemini rhythm converting to premature auricular contractions. Cardiovascular measurements obtained from Apollo crewmembers during their evaluations, heart rate was the most easily measured and yielded the most accurate and predictable values.

Table contains heart - rate data on individual crewmembers during three conditions of orthostatic stress evaluations: (1) resting supine control, (2) the highest level of LBNP [-SO mm Hg (-67 x 102 N/m2 )], and (3) passive standing. Resting supine heart rate is elevated significantly in 13 of 24 crewmen (54 percent) at the first postflight evaluation; the group response is elevated at the two-percent level of confidence.

A trend toward preflight values is subsequently evident. By the third postflight evaluation, only three of fifteen individuals (20 percent) show significant elevations in resting supine heart rate, and the group mean value is not statistically different from the preflight group mean heart rate (n = 15, paired).

Following the same comparisons, the application of -50 mm Hg (-67 x 102 N/m2 ) LBNP produced significantly elevated heart rates in 14 of 17 Apollo crewmen (82 percent) at the first postflight evaluation, with a group elevation significant at the O.1-percent level.

-40 mm Hg (-53 x 102 N/m2 ) LBNP and was not tested at -50 mm Hg (-67 x 102 N/m2 ) LBNP on recovery day. Five other crewmembers (the Apollo 8 CMP, the Apollo 8 LMP, the Apollo 9 LMP, the Apollo 16 CMP, and the Apollo 16 LMP) developed presyncopal symptoms at some point before protocol completion during their immediate postflight.

50 mm Hg (-67 x 102 N/m2 ) stress; the Apollo 15 Commander experienced similar symptoms during his second postflight evaluation. Although more crewmembers, immediately postflight, demonstrated a larger heart rate increment over preflight values during LBNP stress than during the resting control period, statistically significant group differences disappeared by the third postflight evaluation. Passive vertical standing results indicated a similar increase in heart rate immediately postflight, with eight of nine crewmembers (89 percent) having heart rates above their 95 - percent preflight envelope, and the group mean value being elevated at the 0.1 percent level.

Page 3

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

Fundamental differences in the Apollo spacecraft, in its operational environment, and in program goals were expected to produce physiological responses that differed from those seen after the Gemini flights. The two-gas (oxygen and nitrogen) atmosphere and the capability to move about in the spacecraft led to speculation that returning Apollo crewmen might show little or no change in orthostatic tolerance.

On the other hand, there was some concern regarding the ability of the cardiovascular system to withstand acceleration stresses associated with lunar descent and ascent. Headward acceleration (+Gz) was imposed during the Lunar Module descent after three to four days of weightlessness, and a near one-g (+Gz) force was produced by the ascent profile after a day or more of 1/6-g exposure. Also, the results of postflight tests were expected to show important differences in cardiovascular responsiveness between crewmen who walked on the moon and those who remained in weightless flight. These speculations and many other unanswered questions emphasized the need to gain as much understanding as possible about the cardiovascular system and its adaptation, first to zero g and, later, to one g.

Page 4

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

For several years before the first manned Apollo flight, investigators had studied the effects on the cardiovascular system of the application of lower body negative pressure (LBNP). Lower body negative pressure involves the application of reduced pressure usually to that portion of the body below the level of the iliac crests. Evaluations of its use as a simulator of orthostatic stress and as a preventer of cardiovascular deconditioning bad been made. Lower body negative pressure, at levels ranging from 40 to 60 mm Hg (-53 x 102 to -80 x 102 N/m2 ) ) as determined by individual tolerance, produced changes in heart rate and blood pressure similar to those resulting from upright tilting. Clearly, the cardiovascular responses initially induced by either stress procedure depended primarily on displacement of blood, chiefly from central blood volume reservoirs, to the lower extremities.

Although qualitatively alike, differences in the magnitude of cardiovascular compensatory responses induced by LBNP have been reported. Stevens (1966) and Stevens and Lamb (1965) found a greater increase in heart rate during upright tilting than during LBNP adjusted to produce the same cardiac output reduction (-19 percent).

Later, Musgrave and co-workers (1969; 1971) reported that even though LBNP at 40 mm Hg (-53 x 102 N/m2 ) ) and the upright posture displaced essentially equal volumes of blood to the lower extremities, negative pressure levels of -SO mmHg (-67 x 102 N/m2 ) ) were required to produce equivalent elevations of heart rate. Both groups of investigators attributed the smaller heart rate response during LBNP to the absence of stimulation of carotid and other baroceptors by gravity-induced hydrostatic pressure and flow changes. Further, the absence of

Page 5

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

hydrostatic pressure gradients along the lower extremities during LBNP caused displaced blood to be distributed differently than during tilt.

In the Skylab missions, several instances of ventricular PVCs, supraventricular PVCs, and nodal arrhythmia were recorded. The arrhythmias occurred during effort tests, extravehicular activities (EVAs), lower body negative pressure sessions, and throughout the entire mission. These included two consecutive PVCs in one astronaut during exercise and an episode of atrioventricular dissociation preceded by sinus bradycardia in two astronauts.

Skylab Crew Health Crew Surgeons’ Reports During the flight phase of the Skylab missions, the Crew Surgeons relied to a great extent on the daily private medical conference with the crews over an air-to-ground loop from the NASA Mission Control Center to monitor crew health. For continuous clinical evaluation of the crew, the Crew Surgeon had access to medical parameters derived from the experiment data and was also dependent on the following monitored areas for clinically related information: • Radiological health • Skylab environmental data, incuding toxicological evaluation • Medical data obtained from the Operational Bioinstrumentation System during the scheduled extravehicular activities Postflight, the Crew Surgeon coordinated all the medical activities relating directly to the crew. He was the medical team leader on the recovery ship and had prime responsibility for the continuous clinical care of the crew especially during the medical experiments, and later at Johnson Space Center.

Skylab 2 Medical examinations performed on the three crewmen at specified intervals beginning 40 days preflight did not reveal any major change in any crewmember’s health status. They remained in good health throughout the

Page 6

Special Edition on Long Duration Spaceflight preflight phase, except for the Pilot who developed a 24-hour illness resembling a viral gastroenteritis about 1 month before flight, just coincident with the initiation of the Flight Crew Health Stabilization Program.

In-flight, on mission day 1, the Commander developed a left serous otitis media, which required the extended use of an oral decongestant as well as a topical nasal decongestant. On mission days 3 through 7, the Commander also used a topical steroid cream to relieve the symptoms of a probable mild contact dermatitis of his right arm. Complying with a preflight decision, the Scientist Pilot took one scopolamine/dextroamphetamine sulfate capsule just after insertion, and the medication was not repeated. Prior to extravehicular activity, the Scientist Pilot and the Pilot utilized a topical nasal decongestant prophylactically; the Pilot also took a systemic decongestant.

Skylab

STEM Today, March 2016, No.6

No significant arrhythmias developed in-flight. Early terminations of the Lower Body Negative Pressure experiment by the Scientist Pilot and Pilot were sporadic, and in this mission the maximum level of exposure to lower body negative pressure was reduced following early termination of the Lower Body Negative Pressure test. The Commander and Pilot took hypnotic medication of choice on the night of mission day 27 to help accommodate a change to their work/rest schedule for entry and splashdown.

Entry itself on the 29th day was nominal. Postsplash (on the water) the heart rates were: Commander, 84;Scientist Pilot, 84; and Pilot, 76 beats per minute.

Aboard the ship on recovery day vertigo, postural instability (especially with eyes closed), reflex hyperactivity, and paresthesias of the lower extremities were prominent findings. The Scientist Pilot developed seasickness while still in the Command Module and the most prominent symptoms cleared in 4 to 6 hours. Scaling of the skin of the hands was noted on the Commander and the Scientist Pilot. The Pilot experienced a vagal response (decreasing heart rate, pale and sweaty appearance) in the recovery period of the Metabolic Activity experiment , which lasted just a few minutes. Muscle and joint soreness, generally confined to the lower back and lower extremities, were first noted on the first day post recovery. During the ongoing postflight period of surveillance, no significant medical problems developed as an apparent result of the long duration in weightless space flight.

No drugs were taken except for vitamins.

Skylab 3 Preflight, no infectious diseases or other medical problems were experienced by the crew during the 30-day preflight period, the last 21 days of which included the Flight Crew Health Stabilization Program. Launch and orbital insertion were nominal. Shortly after orbital insertion, the Pilot began to experience nausea; this was aggravated by head movement. One hour after insertion, the Pilot took an antimotion sickness capsule, scopolamine/ dextroamphetamine sulfate, with good relief. The crew entered the Orbital Workshop 9 hours and 45 minutes after lift-off. Following strenuous work to activate the Orbital Workshop, the Pilot vomited once.

During the second mission day, the Commander and Scientist Pilot also experienced some motion sickness during continued Orbital Workshop activation; they took scopolamine/dextroamphetamine sulfate, as required, for alleviation of symptoms. This indisposition caused a loss of work time during the first 3 days of flight. Two additional days elapsed before all symptoms had dissipated. Since medical experiments were not run until mission day 5, subjective voice reports by the crew were the only means of health assessment during this time.

On mission day 5, after the first medical experiments were conducted, objective clinical data were available to aid in evaluating the crew’s health. In general, the crewmembers remained in excellent health except for a few minor clinical problems and rare sporadic early terminations of the Lower Body Negative Pressure experiment by the Commander and the Scientist Pilot.

The Pilot reported a painless sty on the left upper eyelid on mission day 29, which responded to an ophthalmic antibiotic ointment and cleared by mission day 32. On mission day 33, the Commander reported the beginning of a boil under his right arm. Instructions from the ground to the Commander were to avoid using stick-type deodorant, and the wearing of garments which fitted tightly under the arms. No medications were recommended and the condition cleared in about 48 hours. A recurrence of the boil in approximately the same area on mission day 50 again lasted only 48 hours, and did not require any medication. The crew maintained high levels of daily exercise during the mission. Extravehicular activities were successfully completed on mission days 10, 28, and 57 without medical problems.

Page 7

Special Edition on Long Duration Spaceflight

The crew slept 6 hours on the night prior to entry and were awake approximately 15 hours prior to splashdown on mission day 60. The Scientist Pilot took an antimotion sickness capsule approximately 40 minutes prior to the entry burn, while the Commander and Pilot took their antimotion sickness medication approximately 5 to 10 minutes after the burn. Prior to the burn, all three crewmen inflated their orthostatic counter measure garments. The entry was nominal. At about 20 to 30 minutes after splashdown while still in the Command Module, the Scientist Pilot checked the pulse rate of each crewman and obtained the following values: Commander, 88; Scientist Pilot, 70; and Pilot 62 beats per minute. Pulse checks by the Crew Surgeon immediately after the Command Module was aboard the recovery ship were similar. Blood pressures were within acceptable ranges for these crewmen. All three crewmen egressed the Command Module on their own power.

Skylab

STEM Today, March 2016, No.6

Postflight the cardiovascular deconditioning observed was carefully documented, but no clinically serious events occurred. As in Skylab 2, vertigo, postural instability, hyperreflexia, dry skin, and slight fissuring of the hands were noted. On recovery, a previous back strain suffered by the Commander recurred from a situation combining "lifting" and loss of balance. On recovery day the Commander developed presyncope during the stand test. The Pilot had a vagal response, also associated with presyncope, during the recovery phase of the Metabolic Activity experiment M171 . The overall rate of recovery postflight was more rapid than that observed in the first manned Skylab mission.

Skylab 4 In Skylab 4, the Flight Crew Health Stabilization Program lasted 27 days due to a 6 - day slip in the launch for evaluating and correcting potential launch vehicle problems. The crew underwent preflight evaluations, which were augmented by several new experiments, such as echocardiography and pulmonary function evaluation.

Several items noted in the medical history and clinical examinations and requiring attention for the upcoming flight were: a history of low back pain (lumbo-sacral strain) experienced by the Commander in the preflight period, and the concern as to whether there would be recurrence of this pain on his return to Earth; some recurring variable left ear drum injection and lability of blood pressure noted during the preflight period in the Scientist Pilot; and the history of recurrent nasal congestion and a tendency toward lability of blood pressure in the Pilot.

Skylab Cardiovascular review of these men showed no evidence of nor tendencies toward arrhythmias. These findings were well documented in order to permit evaluation of any in-flight changes. The crew remained in good health throughout the preflight period. This crew also had no formal scheduled in - flight medical examinations. Data from experiments and "as necessary" medical evaluations continued to provide the necessary information for monitoring of health status. A heart rate and blood pressure stress evaluation for clinical reasons would be obtained on any individual at least every 4 days, if for some reason the experiments Lower Body Negative Pressure experiment M092, Vectorcardiogram experiment M093 , and Metabolic Activity experiment M171 could not be run. This longest mission happily was characterized by the absence of any major illness or injury. However, it is important to point out that in this mission there were numerous symptomatic events that required variable amounts of medication .

For all Skylab 4 crewmen, the initial medication was the prescribed antimotion sickness drugs; the Scientist Pilot did not experience motion sickness and the Commander had minimal malaise for 3 days. The Pilot had significant nausea with vomiting for 1 day and then malaise for 2 more days. The second major recurrent use of medication was lip balm and skin cream to prevent drying of the lips and skin, respectively. The sleep medications were utilized intermittently throughout the mission by all the crewmen. Decongestants (topical and systemic) were used during the mission. These were used both prophylactically during the extravehicular activities and for specific symptomatic relief of the feeling of fullness in the head, nose, and ears. The Scientist Pilot

Page 8

Special Edition on Long Duration Spaceflight utilized aspirin twice for transient headaches on mission days 17 and 67. On mission days 75 through 79, he utilized wet packs to help resolve a minimal papular rash on the left neck and ear area. The Pilot had a rash in the upper mid-back area, which was treated as a fungal infection, and which did resolve after about a week and a half.

STEM Today, March 2016, No.6

The observed in-flight problems were not related to preflight problems except remotely; one could state that the Pilot’s prior history might have indicated the greater susceptibility to upper respiratory congestion. In following the crew, the Daily Health Status Summary sheet was a comprehensive guide. It was updated for this particular mission and was kept by the person in aeromedical monitoring position working in direct support of the Mission Operation Control Room Surgeon. Data for this summary were prepared from the Evening Status Report which gave sleep, medication, exercise, and experiments M071 (Mineral Balance, ch. 18), M073 (Bioassay of Body Fluids, ch. 23), and M172 (Body Mass Measurement, ch. 19) data, from the dump tapes, and from the private medical conference. The latter permitted subjective and objective crew observations about their responses to the stressor tests [Lower Body Negative Pressure and Metabolic Activity ] as well as the general status of living in zero-g. Vectorcardiographic data became especially valuable as the Pilot began demonstrating vectorcardiographic parameters differing significantly from preflight. None of these deviations from preflight "norms" were considered clinically abnormal. In summary, there were neither clinically significant cardiac arrhythmias nor vectorcardiographic changes in-flight.

Instrumental in maintaining crew health was maintenance of a proper environment. It should be stressed, there were no significant problems in maintaining the limits of environmental conditions of total pressurre, oxygen, and carbon dioxide. Other parameters, such as temperature and relative humidity, were more variable.

These parameters were influenced by the orbital inclination and Sun angle of the Skylab complex and the performance of the supplementary thermal protection devices; additionally, potential off-gassing from the heated spacecraft was satisfactorily circumvented. Personal cleanliness was fairly well maintained by use of the shower or by sponge baths but proved to be time consuming.

The increased quantity and quality of exercise available to the crew were important in maintaining the crew health of Skylab 4. For each successive mission the exercise time had been increased from one-half hour, to 1 hour, to 1 and one-half hours per day, respectively. In Skylab 4 the bicycle ergometer, the Mark I (an isokinetic force generating pulley), the Mark II (springs), and Mark III (the standard Apollo exercise device), the treadmill, and isometric exercises were available to counteract the effect of the zero-g environment; the crew had the highest overall average of quantifiable work output from their exercise.

The maintenance of nutrition was satisfactory; the Skylab 4 crew ate at essentially preflight caloric levels and were quite satisfied with the taste of the food. The high density food bars, utilized to extend provisions when the Skylab 4 mission was extended to 84 days, were tolerated well by the crew although they left a subjective sense of hunger. As in Skylab 3, vitamin supplementation was maintained. The weight losses for the Skylab 4 crewmen were less than those for the crewmen of the other two missions.

The work/rest cycle was a key problem in this last mission. During the early phase of this mission the crew was scheduled at a pace comparable to the pace attained by Skylab 3 crewmen in the latter part of their mission. New experiments, stowage confusion, onboard equipment malfunctions, and the sheer length of the mission were all contributing factors to produce psychological stresses which were slowly resolved over the first half of the mission. As the end of the mission approached, two late single-block shifts of sleep time were made, as the preferred mode, to adjust the crew to the circadian shift required. Crew comments postflight indicated this was a suitable and effective approach to the time shift required. Earlier piecemeal shifting in Skylab 2 and Skylab 3 was not subjectively as effective. In preparation for entry, scopolamine/dextro-amphetamine sulfate was prescribed for all three crewmen at approximately 2 hours prior to intended splashdown. The crew inflated their counter measure garments prior to burn and reinflated them to compensate for the increasing internal pressure as the Command Module was pressurized during descent.

As in Skylab 3, the splashdown was initially in stable-2 (heat shield up), and changed to stable-1 (heat shield down) within a nominal time frame.Initial "on water" pulse rates were: Commander, 70; Scientist Pilot, 80; and Pilot, 80 beats per minute. Blood pressure and pulse readings taken inside the spacecraft were acceptable and the crew egressed and walked essentially unassisted.

The triad of vertigo, postural instability and reflex hyperactivity was again noted postflight. This time it was the Commander who experienced a vagal response with presyncope at the end of forced expiration in pulmonary function testing. Petechiae were noted in the lower legs of all three crewmembers late on recovery day, and during the day afterwards. Muscle and joint soreness during exercise developed postflight, but only to a

Page 9

Special Edition on Long Duration Spaceflight minimal degree. The postflight period was free of any illnesses or injuries. Postflight physiological readaptation, as measured by the experiments revealed the crew to be in as good or better status than the crews of the two earlier missions.

STEM Today, March 2016, No.6

In addition, an isolated incident of a non-sustained 14-beat ventricular tachycardia (Figure 2), with a maximum heart rate of 215 beats per minute, was recorded using in-flight Holter monitoring abroad the Mir.Although not part of a systematic scientific study,this case provides additional evidence of arrhythmias during long-duration space flight.

Analysis and Evaluation of the Functional State of the Cardiovascular System in Cosmonauts The state of the cardiovascular system of cosmonauts during space missions has been studied since the beginning of manned spaceflights. Systematic studies of the effect of microgravity on the human cardiovascular system began during the missions on board the Salyut-6 orbital station (OS) and continued on board Salyut-7; the greatest amount of information came from research aboard the Mir OS . During the 15 years of operation of the Mir OS, 35 Russian cosmonauts, 7 astronauts from the United States, 1 astronaut from the European Space Agency (ESA) (a German citizen), and 1 French astronaut participated in 28 main missions.

Eighteen cosmonauts participated in more than one mission. Medical monitoring of the cosmonauts’ health was very important for the safety of long-term spaceflights. The factual data accumulated in the course of medical examinations made it possible to determine the general patterns of changes in the functional parameters of the cardiac activity and the circulation system at rest and under functional loads during 73- to 438-day spaceflights.

According to current views, the cardiac index (CI) is one of the main functional parameters of the cardiovascular system reflecting its regulatory mechanisms and reserve capacity . Quantitative estimates of hemodynamic parameters characteristic of the three types of blood circulation, hypokinetic, eukinetic, and hyperkinetic, at about the same arterial blood pressure (ABP) were developed and suggested for practical use. The results of examination of cosmonauts on the Mir OS were analyzed in this respect.

It was found that, during spaceflight, the ratio between blood circulation types in resting subjects changed compared to that on the earth and that this change depended on various factors, including the conditions of the examination. Authors summarized and analyzed the results of the GE tests in the same cosmonauts taking into account the types of blood circulation. The crews of the first eight main missions (MM-1 to MM-8) performed

Page 10

Special Edition on Long Duration Spaceflight a two-step test (GE-2) according to the following scheme: a 125-W exercise for 5 min, a rest for 1 min, and a 175-W exercise for 3 min, with the total load amounting to 1150 W. Beginning from the ninth main mission (MM-9 to MM-28), a modified test was used. The load was made close to submaximal (75% of the maximal power), and the GE test consisted of three steps (GE-3), with the load increasing from step to step (125-, 150-, and 175-W exercises for 3 min each).

STEM Today, March 2016, No.6

Results The distributions of the participants of MM-1 to MM-8 and MM-9 to MM-28 with respect to blood circulation type at rest before the spaceflight were different. In MM-1 to MM-8, 28, 56, and 16% of cosmonauts had hypokinetic (CI = 2.66 ± 0.0525 l/min/m2 ), eukinetic(CI =3.59± 0.0625 l/min/m2 ), and hyperkinetic (CI = 4.73 ± 0.1512 l/min/m2 ) circulation types, respectively. In MM-9 to MM-28, a smaller proportion of cosmonauts (9%) had the hypokinetic type of circulation (CI = 2.71 ± 0.1060 l/min/m2 ), while the frequencies of the eukinetic and hyperkinetic types of circulation (with CI values of 3.64 ± 0.049 and 4.78 ± 0.1512 l/min/m2 , respectively) were about the same (47 and 43%, respectively).

Thus, in general, the eukinetic circulation type (as measured at rest before the exercise) was prevalent in all crews of the Mir OS before a flight. The CI values for each circulation type in both groups of cosmonauts (those performing GE-2 and GE-3) were almost equal and agreed with data reported by other authors. Possibly, some of the differences between the MM-1- MM-8 and MM-9-MM-28 groups in the frequencies of circulation types before the flight were determined by individual genetic factors and differences in age. This possibility was indicated earlier by other researchers and was confirmed by the results of dynamic follow-up monitoring for several years of all cosmonauts that participated in missions on the Mir OS. In cosmonauts that participated in more than one mission, the CI values before the first and subsequent spaceflights were about the same (3.56 ± 0.0740 and 3.60± 0.1096 l/min/m2 , respectively).

Page 11

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

The HR, SV, and CO before the spaceflight were minimum in cosmonauts with the hypokinetic circulation type and maximum in those with the hyperkinetic type in both groups (GE-2 and GE-3) (table; Fig. 1); in cosmonauts with the eukinetic circulation type, these values were intermediate. The differences between the groups with respect to these parameters were statistically significant in most cases. The sBPs of cosmonauts with all types of circulation varied insignificantly (table; Fig. 2).

The distribution of the cosmonauts with respect to circulation type at rest during the flight was different. In cosmonauts participating in MM-1 to MM-8, the frequency of the hypokinetic type decreased to 9%, that of the eukinetic type remained the same (57%), and that of the hyperkinetic type increased to 34%. The cosmonauts participating in MM-9 to MM-28 exhibited a different distribution with respect to circulation type during the flight, as was the case with this distribution before the flight. The frequency of the hypokinetic circulation type remained almost unchanged (10%), the frequency of the eukinetic type somewhat decreased (to 40%), and that of the hyperkinetic type increased (to 50%). Thus, in general, the frequency of the hypokinetic circulation type decreased and that of the hyperkinetic type increased during the spaceflight. This agrees with the results of analysis of all medical studies during the flight.

However, the eukinetic circulation type was prevalent in the participants of MM-1 to MM-8 both before and during the spaceflight, whereas, in cosmonauts participating in MM-9 to MM-28, the hyperkinetic type was the most frequent. Most changes during the flight in the parameters studied as compared to their values before the flight were small and had no definite trends. The ratio between hemodynamic parameters for different types of blood circulation was preserved only for CO in cosmonauts participating in MM-1 to MM-8 and for SV and CO in those participating in MM-9 to MM-28. In these cases, the parameters remained minimum for the hypokinetic type of blood circulation and maximum for the hyperkinetic type. Apparently, this reflected different mechanisms of adaptation of the cardiovascular system to microgravity.

Page 12

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

Before the flight, the GE was accompanied by blood circulation changes adequate to this load, namely, a significant increase in HR, ABP, and CO in cosmonauts with the hypo- and eukinetic types of circulation (table; Figs. 1, 2). However, quantitative changes in each parameter were specific for different circulation types. Changes in SV also depended on the type of blood circulation. The maximum increase in HR during the GE-2 test was 150 and 111% in cosmonauts with the hypokinetic and eukinetic types of circulation, respectively. The absolute HRs were almost equal in these groups (125 and 116 beats per minute, respectively) (Fig. 1). The same was true for the changes in CO. Its relative increase was less in cosmonauts with the hypokinetic than with the eukinetic type of circulation (130 and 97%, respectively). However, the CO after the load was significantly higher in subjects with the eukinetic type of circulation (13.21 l/min). The increase in SV was virtually the same (17 and 18%, respectively) (table).

Changes in CO are known to be determined by changes in both of its components, SV and HR. In response to a functional load, the CO increases to ensure a sufficient ABP. This increase is determined, depending on the individual functional capacity of the cardiovascular system, by an increase in HR and, to a varying extent, in SV. The responses of the cardiovascular system were classified according to the pattern of the relationship between these parameters. According to this classification, the responses of the cosmonauts with the hypoand eukinetic types of circulation participating in MM-1 to MM-8 were normotonic (with a slight trend to hypotonic due to a relatively small increase in SV).

In subjects with the hyperkinetic type of circulation, the increase in CO was entirely determined by the increase in HR; this response was classified as hypotonic. Therefore, in terms of the compensatory capacity of the cardiovascular system, the response of the subjects with the hyperkinetic type of circulation even to a small load (1150 W) was less favorable than the responses of subjects with other circulation types. FsBP increased to the same extent (by 28-31%) in subjects with all types of blood circulation and was about the same (149-154 mm Hg) at the first minute of the recovery period in them.

The three-step load increased HR in cosmonauts with the hypokinetic and eukinetic types of circulation (by 147 and 136%, respectively). The relative increase in CO in subjects with the hypokinetic type of circulation was larger than in subjects with the eukinetic type (193 and 129%, respectively) and was accounted for by a considerable (41%) increase in SV. In contrast to the other types of blood circulation, the response to the exercise was purely normotonic in the case of the hypokinetic type. In cosmonauts with the eukinetic type of blood circulation, the relative increase in SV did not exceed 16%. In the case of the hyperkinetic type, SV tended to increase (by 9%) during the GE-3 test, as was the case with the GE-2 test. The increase in CO was mainly accounted for by the increase in HR. The ABP increase by about the same value and reached about the same level (193-203 mm Hg) in cosmonauts with all types of blood circulation (Fig. 2).

Comparison of the relative changes and absolute values of HR, ABP, and CO demonstrated that they were substantially higher in the case of the three-step load (GE-3) in cosmonauts with all types of blood circulation.

This agrees with the results of the comparative analysis of the cardiovascular responses to GE-2 and GE-3 , which demonstrated that the three-step load strained the compensatory mechanisms of the cardiovascular system to a higher degree because of the greater (by 17%) total load. The observed characteristic features of the patterns of changes in HR, SV, and CO in response to exercise using a bicycle ergometer before the flight indicated that, even then, cosmonauts with different types of blood circulation exhibited different types of cardiovascular responses to the functional load. This response was the least favorable in the case of the hyperkinetic type of blood circulation.

When analyzing the results of GE tests during the flight, we found that the most characteristic feature of the cardiovascular responses in all cases was an almost complete absence of increase in SV, or sometimes even a decrease in it, at the first minute after the end of the exercise (table). These characteristic features were noted earlier during examination cosmonauts on board the Salyut-6 OS; in cosmonauts participating in Salyut-7 and

Page 13

Special Edition on Long Duration Spaceflight Mir missions, they were regular and statistically significant . Changes in SV during physical exercise are known to be accounted for by two factors: the blood inflow and the contractile function of the myocardium. Ultrasound examination demonstrated that the myocardial contractile function did not decrease during the spaceflight.

Therefore, authors may assume that the main reason for decreased SV reaction is insufficient venous return of blood to the chambers of the heart after the exercise and a resultant decrease in the final diastolic volume of the left ventricle because of the retention of a part of the blood in the blood vessels of the lower extremities. This suggestion seems reasonable if one takes into account that, in microgravity, the decreased transmural pressure may result in the formation of zones of free extensibility in crural veins, the vascular tone is changed, and blood is retained in the arterioles of leg muscles that have recently ceased functioning.

For all of the three types of blood circulation, authors observed the following significant (p < 0.001) differences in hemodynamic parameters after GE-3 during the flight compared to the values measured after GE-3 before the flight: a reduced relative increase in HR without changes in its absolute value and reductions of both the relative increase in and the absolute values of ABP and CO.

STEM Today, March 2016, No.6

Thus, the results of this study demonstrated that spaceflights caused changes in the frequencies of blood circulation types under conditions of physiological rest. In general, the frequency of the hypokinetic circulation type decreased and that of the hyperkinetic type decreased. The eukinetic type of blood circulation was the most frequent both before and during the flight. The response of the cardiovascular system to graded exercise on a bicycle ergometer before the flight was less favorable in cosmonauts with the hyperkinetic type of blood circulation. They exhibited a hypotonic response, i.e., an increase in CO that was entirely accounted for by an increase in HR due to an enhanced chronotropic function of the myocardium, with the SV remaining almost unchanged.

In microgravity, the mechanism of the formation of CO changed in all cosmonauts irrespective of the type of blood circulation: the contribution of SV was virtually absent, and the role of HR was dominant. This was especially evident in cosmonauts with the hyperkinetic circulation type upon performance of three-step graded exercise. Taking into account that microgravity alters the entire hemodynamic pattern of the human body, we may assume that these conditions are the main factor determining the decrease in the contribution of SV to the maintenance of the CO value necessary for performing physical exercise during a spaceflight. In estimating the tolerance of cosmonauts to physical load in microgravity, correct interpretation of the dynamics and absolute values of HR, ABP, and CO is of crucial importance.

Cardiac, arterial and venous adaptation to weightlessness during 6-month MIR spaceflights with and without thigh cuffs (bracelets) The objectives of this investigation were to study the effects of thigh cuffs (bracelets) on cardiovascular adaptation and deconditioning in 0g. The cardiovascular parameters of six cosmonauts were measured by echocardiography, Doppler, and plethysmography, during three 6-month MIR spaceflights. Measurements were made at rest during preflight (-30days), inflight (1, 3-4, and 5-5.5months) without cuffs (morning) and after 5h with cuffs, and during postflight (+3 and +7days).

Lower-body negative pressure (LBNP) measurements were performed 1 day after each resting session. Inflight values of left ventricle end-diastolic volume and stroke volume measured without the thigh cuffs (-8 to -24% and -10 to -16%, respectively, both P<0.05) were lower than corresponding preflight values. The jugular and femoral vein cross-sectional areas (A jv and A f v , respectively) were enlarged (A jv : by 23-30%, P<0.001; A f v : by 33-70% P<0.01). The renal and femoral vascular resistances (R ra and R f a , respectively) decreased (R ra : by -15 to -16%, P<0.01; R f a : by -5 to -11%, P<0.01). Inflight, the thigh cuffs reduced the A jv (by -12 to -20%, P<0.02), but enlarged the A f v (A f v : by 9-20%, P<0.02) and increased the vascular resistance (R ra : by 8-13%, P<0.05; R f a : by 10-16%, P<0.01) compared to corresponding inflight, without-cuffs values. During LBNP (-45mmHg, where 1mmHg=133.3N/m2 ), R f a and the ratio between cerebral and femoral blood flow (Q ca /Q f a ) increased less inflight and postflight (+25% for R f a and +30% for Q ca /Q f a ) than during preflight (60% for R f a and 75% for Q ca /Q f a , P<0.01). This reduced vasoconstrictive response and less efficient flow redistribution toward the brain was associated with orthostatic intolerance during postflight stand tests in all of the cosmonauts. The calf circumference increased less inflight and postflight (6% P<0.05) than preflight (9% P<0.05). The vascular response to LBNP remained similarly altered throughout the flight. The thigh cuffs compensated partially for the cardiovascular changes induced by exposure to 0g, but did not interfere with 0g deconditioning.

Page 14

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

Autonomic regulation of circulation and cardiac contractility during a 14-month space flight The space flight of physician cosmonaut V.V. Polyakov, the longest to date (438 days), has yielded new data about human adaptation to long-term weightlessness. Autonomic regulation of circulation and cardiac contractility were evaluated in three experiments entitled Pulstrans, Night, and Holter. In the Pulstrans experiment electrocardiographic (ECG), ballistocardiographic (BCG), seismocardiographic (SCG), and some other parameters were recorded. In the Night experiment, only the ballistocardiogram was recorded, but a special feature of this experiment is that the BCG records were obtained with a contactless method. This method has several advantages, the most important of which are the possibility of studying slow-wave variations in physiologic parameters (ultradian rhythms) on the basis of recordings made under standard conditions over a prolonged period.

The Holter experiment (24-hour electrocardiographic monitoring) used a portable cardiorecorder (Spacelab, USA). The obtained electrocardiographic data were used to analyze heart rate variability. In the first 6 months of the 14-month flight, the dynamics of cardiovascular parameters in V.V. Polyakov was virtually the same as in the other cosmonauts.

Results Heart-rate variability and cardiac contractility in the Pulstrans experiment Figures 3 and 4 show temporal variations of the basic cardiologic parameters measured in the Pulstrans experiment in the course of the 14-month space flight. As can be seen in Fig. 3A, systolic and diastolic pressures during the flight were lower by 10-15 mm Hg than before it. The decreases were most marked on days 147-189 and 257 of the flight. The heart rate rose somewhat at first but reached the preflight level on days 200 and 340 of flight. The heart rate was at its maximum on day 250 of flight when it exceeded the preflight level by 10 beats/min. The SCG amplitude had increased by day 55 of flight, was markedly increased on day 250-257 (Table l), and was highest near the end of the flight, on days 340 and 410. Of note is the drastic increase of the SCG amplitude in the first few days after landing when it exceeded about 10-fold the preflight level. The

Page 15

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

BCG amplitude showed less marked increases and had three maxima - on days 189 ,257 and 410 of flight. Of much interest seems to be the considerable (nearly 1.5-fold) increase in the SCG/BCG ratio at the end of flight , although a much larger (nearly 20-fold) increase in this ratio was noted in the first few days after landing.

Page 16

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

Heart-rate variability values (Table 1 and Fig. 3B, 3C) demonstrated phasic changes. The SD showed a tendency toward a reduction during the first 6 months, a marked increase in the next two months of flight (days 220-257)

Page 17

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

and a reduction near the end of straying for 14-months in weightlessness. Worthy of note are also big changes in the ratio of spectral components ( PNS and SNS) of the heart rate variability during months 7 and 8 of the flight. The power of respiratory waves fell sharply (indicating reduced activity of the parasympathetic component of regulation), whereas the power of LF waves with periods of 20-50 seconds showed a similarly large increase (indicating heightened activity of the sympathetic component). The power of MF spectral component showed a decrease during first 5 months of flight with minima at day 147, but it increase till preflight level at days 250-257 and than fall sharply, especialy in postflight period. The relative power of MF had other dynamics ( Fig.3B). It minima is observed at days 250-257.

Diurnal variations in heart-rate variability values The data on alterations in mean 24-h values of heart rate variability parameters are indicative of at least two important features of circulation regulation during the 14-month flight (fig.4): 1. The heart rate significantly decreased in the course of the flight, whereas the SD (standard deviation) increased; 2. The absolute power of the heart-rateâ&#x20AC;&#x2122;s variability high frequency (HF) component decreased, whereas that of its MF component increased.

Mean 24-h values of the heart rate and SD were closest to their background (terrestrial) values on day 165 of flight, but spectral components differed considerably from their background values during that period. Of interest is separate consideration of mean diurnal and nocturnal data. Of particular interest in this respect is the graphic representation of "morning-evening-night" relationships in the form of a so-called phase plane (Fig.5). Here, heart rate values are plotted on one axis while values of the absolute power of MF component are plotted on the other. It can be seen that the "regulation area" (i.e. the area of the triangles formed by the

Page 18

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

"morning-evening" and "evening-night" vectors) during the preflight period is approximately the same as on day 165 of flight. The "regulation area" is greatly reduced and is characterized by a considerable elongation of the "evening-night" vector early during the flight (day 12) and at its end ( day 391 ). Moreover, the direction of the "morning-evening" vector changed by the end of flight. These data show that the circadian organization of the regulation of circulation changes in the course of a prolonged flight, which may be due to the inclusion of the hypothalamic-pituitary level in regulation processes.

Page 19

Special Edition on Long Duration Spaceflight

Ultradian rhythms during the 14-month flight Ultradian rhythms reflect the activity of regulatory mechanisms, by which appropriate interactions among various systems of the human body are assured . One of the best-known ultradian rhythms is the 90-minute rhythm reflecting rest-activity cycles during wakefulness and sleep (non-REM sleep and REM sleep). A sine qua non for successfully addressing the problem of ultradian periodicities and their role in the adaptation of the body to unusual conditions, including prolonged weightlessness, is nocturnal recording of physiologic parameters. The relevant data obtained during Polyakov’s 14 month flight are presented in Fig. 6. It is the dynamics of factor K. Fig. 6 compares the mean values of factor K obtained in the Night experiments of V.V. Polyakov and of the four cosmonauts who were on a 6-month flight. It can be seem that factor K increased during the first 3 months of flight in all five cosmonauts, and that its values in the 6th-7th months of flight were close in all of them to those recorded preflight. A second increase in this factor was observed for V.V. Polyakov in the 8th-10th months of flight.

STEM Today, March 2016, No.6

Particular attention was paid to analyzing waves in the hour range. The presence of distinct 90-minute cycles and their clearly defined structures reflect sleep of good quality. The 90-minute periodicities were evaluated by cross-spectral analysis. The baseline data used were the means of heart rate, respiratory rate, motor activity, BCG amplitude, and heart rate/respiratory rate ratio. The periodicities of each parameter pair that correlated better than others are shown in Table 2. The basic crosspectral period increased in the 2nd-3th months of flight and then returned to a normal value of 102 minutes in the 5th-6th months before showing a second increase to 227-240 minutes in the 9th-10th months.

In the first 6 months of the 14 month flight, the dynamics of cardiovascular parameters in V.V.Polyakov was virtually the same as in the other cosmonauts. Thereafter, however, the following differences were noted for Polyakov: • A tendency of the heart rate to decrease, particularly in night time, to values below those recorded early during the flight; • Alterations in the amplitude and duration of superslow oscillations of physiologic parameters in the 7th8th months, with a fall in the total power of the oscillations and a rise of their relative power in the interval of 3.5-35 minutes; • The tendency of SCG and BCG amplitudes to substantially increase, particularly in the 8th-9th and 14 th months of flight; • A considerable rise in daily average values of the absolute power of the heart rate’s variability MF component accompanied by a fall in its HF component.

The first 5-6 months of flight were, therefore, sufftcient only to stabilize physiologic functions, in particular functions of the circulatory system, at levels close to terrestrial ones. Stronger adaptational processes appear to develop later.

Mention should be made of at least three important aspects after the first 6 months of Polyakov’s sojourn in space: • Activation of a new, additional adaptive mechanism in the 8th-9th months of flight, as is evidenced by alterations in the periodicity and power of superslow wave oscillations (ultradian rhythms) reflecting the activity of the subcortical cardiovascular centers and of the higher levels of autonomic regulation; • Growth of cardiac contractility (both the SCG and BCG) accompanied by a decrease in heart rate during the last few months of flight; and • A considerable increase in the daily average values of absolute power of heart rate’s variability MF component, which reflects the activity of the vasomotor center.

The above-mentioned increases, toward the end of the flight, of vasomotor wave amplitudes in the MF range well agree with the growth of cardiac contractility (SCG and BCG ampkitude). If an increase in vascular tone

Page 20

Special Edition on Long Duration Spaceflight in the upper part of the body does not result in elevated arterial pressure but occurs when the arterial pressure is somewhat lowered, this may be interpreted as a consequence of augmented filling with blood of the vascular bed because of an increase either in the stroke volume or in the volume blood flow rate. Since the circulating blood volume is known to be reduced and the stroke volume not to increase during a prolonged space slight, the only way in which the volume blood flow rate can be increased may be a rise in the rate of blood ejection into large vessels, which will require additional energy expenditure. That such a compensatory mechanism operates is suggested by the increased amplitudes of seismoand ballistocardiograms in the second half of the 14 month flight. On the other hand, we can see the activation of the vasomotor center which is responsible for maintaining adequate vascular tone. This appears to be a second mechanism by which arterial pressure is sustained. This mechanism is associated with elevated activity of subcortical sympathetic centers which appear to be in turn activated by higher levels of regulation.

Changes in basal heart rate in spaceflights up to 438 days The long-term acclimation of heart rate to microgravity was studied in a cosmonaut who stayed onboard the MIR space station for 438 d. This was the longest mission in the history of manned space exploration. The results are evaluated in the context of findings from three other cosmonauts who lived onboard MIR for a shorter time.

STEM Today, March 2016, No.6

For the record mission sleep polygraphies were obtained prior to mission on the ground, between the 3rd and the 30th d in space, after 6 mo in space, and toward the end of mission. From each of the sleep polygraphies beat-to-beat intervals of cardiac rhythms were determined and analyzed as the time series of the average beat-to-beat interval.

RESULTS A lengthening of the average beat-to-beat interval by 176 ms was found during the record flight compared with measurements on the ground. This increase in the average beat-to-beat interval corresponds to a reduction of heart rate by about 20%. The lengthening of the average beat-to-beat interval was more pronounced for nonREM sleep than for REM sleep. During the first month, a lengthening by 82 ms was observed. Measurements after 6 mo showed a further lengthening by 94 ms, and at the end of the mission no further change in average beat-to-beat interval was observed.

Testing the response of heart rate to microgravity across distinct and stationary behavioral states appears to be appropriate to investigate the cardiovascular system. The long-term acclimation of heart rate is possibly due to an increased dominance of the parasympathetic control of cardiac rhythms in space.

Main medical results of extended flights on space station Mir in 1986-1990 During 1986-1990, seven prime crews (PC) carried out missions on the Mir space station and an 8th prime crew has started its space work which will be finished in 1991. A total of 18 cosmonauts (including the 8th prime crew) have participated in extended missions on the Mir space station, one cosmonaut being launched twice.

The primary medical goals for these extended space missions have been maintenance of good health status and performance of space crews and making medical investigations.

CARDIOVASCULAR SYSTEM The most regular inflight changes have occurred in the latter part of electrocardiographic tracings (12 standard leads). These demonstrate a decrease of T-wave amplitude in most leads in all cosmonauts. Analysis of hemodynamic parameters has shown a tendency for an increase in mean heart rates (HR) and a lack of change in stroke volume (SV), cardiac output (CO) and actual specific peripheral resistance (SPRa). Ultrasonic examinations of the physician cosmonaut made during months 7-8 inflight did not reveal changes in left ventricular volumes, SV or ejection fraction (EF). Arterial pressure (AP) measurements showed that diastolic pressure (DBP) had a tendency to decrease (P < 0.18) and pulse pressure (PP) to increase (P < 0.07).

Abdominal ultrasonic examinations carried out by V. S. Bednenko et al. both inflight (7 cosmonauts examinedâ&#x20AC;&#x201C;4 at the end of 5-6 months, 3 at the end of 8-9 months) and postflight (16 cosmonauts) demonstrated: a moderate increase in the size of the liver, spleen, kidneys, pancreas, blood filling of the lungs, cross-sectional area of the large ventral vessels, and a clearer vascular pattern of liver; decreased acoustic density of the pancreas; signs of lateral renal pelvic enlargement with a decrease in renal parenchyma (decreased parenchymal-pelvic system area ratios). These changes are considered to be echographic signs of venous engorgement. An increased gall bladder area and dilatation of the common bile duct also point to the development of bile congestion in the biliary tree. In contrast to the preflight period graded physical exercise tests on a bicycle ergometer during flight (at a work load of 125 W and 175 W for 5 and 3 min, respectively with a l min interval) resulted in insignificant rises of HR, a decrease of SV and CO (by 14.5 and 15.1%) decline in D BP by 6.8% and a PP increase of 13.5% (P < 0.01). These reactions point to the nature of adaptive inflight processes uncovered by exposure to graded

Page 21

Special Edition on Long Duration Spaceflight exercise when findings are compared to the preflight period.

Postflight hemodynamic studies during graded step-wise exercise conducted by A. F. Zhernakov et al. demonstrated a more distinct increase in absolute values of HR, systolic AP and diastolic AP (in most cases); and smaller increment of SV and shortening of the left ventricular ejection period vs its corrected value for changes in HR. No HR and ECG changes of an ischemic type were found. Severity of hemodynamic changes did not depend on flight duration. When compared to pretest hemodynamic parameters, lower body negative pressure (LBNP) tests (at -25, -35, and -45 mmHg for 1, 3, and 5 min respectively) applied in flight led to decreases of SV (5.2%) and CO (8%) as well as an increase in SPRa (15.7%). Blood pressure did not change significantly.

Inflight relative and absolute increases in SPRa were considered to be reactions to prevent pronounced changes in SV, CO and BP. It should be noted that cardiovascular responses to postflight postural tests had no correlation with flight duration.

STEM Today, March 2016, No.6

AFFECT OF MICROGRAVITY ON CARDIAC SHAPE There is little data on how ventricular shape and function changes when exposed to microgravity. The sphericity index is a measure of ventricular shape. As the heart becomes more spherical its ability to perform efficiently decreases. We analyzed pre- and in-flight data of cardiovascular shape in spaceflight in comparison with finite element modeling of the impact of gravity on cardiac shape.

Twelve astronauts with vertical preflight (1g) and in-flight (microgravity) echocardiograms available were identified. Pre-flight echos were obtained approximately 3-4 months prior to launch using a Philips IE-33 (Andover, MA) echo machine. Microgravity echocardiograms were obtained at least two weeks after exposure to a microgravity environment. Astronauts received basic training on operating the echo machine, and live remote guidance from the study team was employed to direct astronauts aboard the International Space Station in obtaining optimal images. Prior to July 2011, studies (6 subjects) were performed on a ATL HDI5000 (Bothel, WA) while those afterwards were obtained using a GE Vivid Q (Milwaukee, WI). SI was calculated. A finite element model was constructed using realistic fiber architecture and orthotropic tissue properties, and subjected to gravitational force from 0 to 1g with SI similarly calculated. Statistical analysis for significance was performed employing the two-tailed paired t-test.

Results Studies from a total of 12 astronauts were available for analysis. The mean sphericity index from the vertical preflight (1g) echocardiograms was 2.01 which was significantly reduced to 1.82 in the microgravity environment (p = 0.0008). This 9.4% reduction in SI compared favorably to the 7.9% reduction predicted by the aforementioned finite element model.

The human heart becomes more spherical as it is exposed to a microgravity environment, a change predicted by mathematical modeling. The impact of this change on cardiac function in space will be addressed as part of the on-going study. Ultimately, authors hope to identify effective countermeasures that astronauts can employ to prevent the untoward cardiovascular effects of long-duration space flight.

Cardiac and vascular responses to thigh cuffs and respiratory maneuvers The transition to microgravity eliminates the hydrostatic gradients in the vascular system. The resulting fluid redistribution commonly manifests as facial edema, engorgement of the external neck veins, nasal congestion, and headache. This experiment examined the responses to modified Valsalva and Mueller maneuvers measured by cardiac and vascular ultrasound (ECHO) in a baseline steady state and under the influence of thigh occlusion cuffs available as a countermeasure device (Braslet cuffs).

AN IMMEDIATE EFFECT of the transition to microgravity during early spaceflight is a loss of the hydrostatic gradient in the cardiovascular system, resulting in a cephalad venous and lymphatic fluid shift . The loss of lower extremity volume is on the order of 1-2l , i.e., larger than on transition to supine or even head-down-tilt (HDT) posture on Earth . Not withstanding the massive fluid shift, invasive central venous pressure (CVP) measurements in spaceflight have shown it to decrease to 2 mmHg with no clinically significant long-term changes in cardiac output . Increased jugular venous distension in spaceflight is seen consistently and persists throughout the mission , but it is no longer a reliable index of right atrial pressure or right ventricular preload.

The effect of fluid redistribution in astronauts is commonly observed as visible facial edema ("puffiness") and engorgement of the external neck veins, with a concomitant decrease in leg size. Subjectively, crewmembers report "fullness in the head," nasal congestion, and diminished sense of taste and smell. Research projects involving echocardiography have been conducted on almost every space vehicle since 1980. Occlusive inflatable cuffs and the Braslet have been investigated in an attempt to reverse the fluid shifts seen during HDT and

Page 22

Special Edition on Long Duration Spaceflight microgravity . Investigations have documented the peripheral vascular effects; however, ultrasound technology at the time was not adequate to confirm the effect on cardiac preload . The International Space Station (ISS) ultrasound system operated with real-time remote guidance has heightened both the scope and the fidelity of imaging studies in space ; furthermore, the ISS crew represents an unprecedented cohort of long-duration microgravity test subjects for prospective investigations.

STEM Today, March 2016, No.6

The Braslet-M thigh cuff countermeasure device is intended to ameliorate the symptoms associated with the microgravity induced cephalad volume shifts in the early hours and days of microgravity exposure by impeding the venous outflow and creating commensurate fluid sequestration in the lower extremities. The cuffs are custom built for each crewmember before the mission and consist of segments of elastic and nonelastic materials to conform to the shape of the upper thigh (Fig. 1). Each device contains an adjustment belt that can be tightened to achieve the desired degree of compression selected in preflight calibration, which consists of a special negative 30◦ tilt-table procedure based on subject feedback and cranial impedance rheography to determine the appropriate compression of the extremity.

RESULTS Table 1 summarizes results from 15 experiment sessions conducted on nine subjects. Out of the 17 sessions scheduled, 15 successfully produced a useful set of data. Some anomalies included the following.

One session not performed due to ultrasound hardware failure.

One session in which cardiac image data were not captured properly due to a procedural error.

One session in which no ECG tracing was obtained with vascular images.

One session in which partial data were lost due to hardware failure.

One session not performed due to scheduling constraints.

One session in which respiratory maneuvers were not properly synchronized, causing difficulties with data analysis.

Phase 1 analysis: effect of Braslet on cardiovascular parameters After correcting for multiple testing (see Statistical Methods), the Braslet cuff was identified as producing a statistically significant effect on the mean response during at least one type of maneuver (including baseline) in 10 of the 27 parameters measured. In particular, significant decreases with application of the cuff were observed in cardiac output, LV stroke volume, left lateral E’, mitral A and E wave velocity, and right isovolumic relaxation time (IVRT) during baseline; left lateral A’ and E’ during the modified Valsalva maneuver; and left lateral E’, right IVRT, and right Tei index (49a) during the modified Mueller maneuver. Significant increases were observed in mitral deceleration time (baseline) and in the femoral vein area (baseline) and Valsalva (∗ in Table 1). Echocardiographic normal parameters were studied on the ISS by Hamilton et al. and, using their data, the magnitude and direction of the changes reported in Table 1 can be approximated for these subjects.

Page 23

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

Phase 2 analysis: comparison of Valsalva, Mueller, and baseline - maneuvers with and without Braslet Again after correction for multiple testing, we observed statistical evidence that the effect of the Braslet cuff (on vs. off) was significantly different in seven parameters between at least two of the maneuvers (including baseline; last column of Table 1). More specifically, parameters showing evidence of this differential change were cardiac output, heart rate, IJV area, LV diastolic volume, left lateral Sâ&#x20AC;&#x2122;, mitral deceleration time, and right lateral Eâ&#x20AC;&#x2122;.

Page 24

Special Edition on Long Duration Spaceflight Braslet Release After completing all the imaging components with maneuvers in Braslet-off and Braslet-on states, the cardiac probe was positioned to get a continuous four-chamber view. The operator crewmember rapidly released the Velcro straps on both Braslet cuffs, and at least 10 cardiac cycles were recorded. Unfortunately, the four-chamber view (n = 6) was difficult to maintain during the Braslet release, resulting in deviation of the imaging plane from its original position; therefore, it was replaced with the more stable TD view (n = 9) in later sessions.

STEM Today, March 2016, No.6

Herault et al. reported that despite extensive training of Mir cosmonauts before flight, the quality of the ultrasound data obtained in-flight may be inadequate. To assure precise execution of the study protocol and quality of the data of our study, both the ultrasound video and the cabin view video were streamed in real time to the NASA Telescience Center, enabling continuous control of the experiment and verbal guidance of the crew by investigators. Each image and each cine-loop used in the analysis were acquired by the astronauts only when the ground-based expert considered the image adequate and commanded acquisition. Thus an established system of balanced expertise distribution enabled data acquisition with quality that was acceptable for real-time assessment and thorough retrospective analysis. TD was used for the first time in spaceflight during this investigation. As an excellent means to assess cardiac performance , TD method is of particular importance for long-duration spaceflight (Fig. 4). TD spectra were taken from the LV lateral wall, septum, and RV free wall.

Furthermore, TD spectra are easier to obtain and less vulnerable to motion artifacts than two-dimensional echocardiographic views. In subjects, LV lateral Eâ&#x20AC;&#x2122; decreased by >20% with the application of Braslet regardless of the maneuver, and LV Lateral Aâ&#x20AC;&#x2122; decreased only when Braslet was applied and a Valsalva maneuver was performed. RV isovolumic relaxation time decreased by >20% with the Braslet applied and by >30% when a Mueller was simultaneously performed. This indicates that preload was reduced with Braslet and that at these reduced end-diastolic volumes the reduced intrathoracic pressure from Mueller made the tricuspid valve open sooner. Mitral pulsedwave Doppler velocity of the LV inflow is a load-dependent parameter and decreases in response to reduced preload . By decreasing preload with Braslet, both Valsalva and Mueller seemed to have caused a modest reduction in LV preload as demonstrated by smaller velocities or extended relaxation slopes.

Impaired LV filling is seen with Valsalva secondary to increased thoracic pressure while Mueller produces a similar result as a consequence of increased RV afterload from the subatmospheric intrathoracic pressure and the small end-diastolic cavitary and pericardial volumes created by the Braslet (Fig. 5).

Femoral vein area increased by 89% with the application of the Braslet. This change was less pronounced (51%) when a Valsalva maneuver was compared pre/post-Braslet, since without Braslet, Valsalva already increased the femoral vein area. This observation provides evidence that the Braslet, when worn and calibrated correctly, still allows for thoracic maneuvers to have a demonstrable effect on lower extremity venous filling. The pressure from occlusion cuffs compresses the superficial veins more than the deep circulation. Although Herault et al. reported the IJV area decrease in changes in IJV area only when Braslet was combined with thoracic maneuvers.

Cardiac output significantly decreased by 19% and stroke volume decreased by 12% when the Braslet was worn, and the heart rate compensation changed significantly in the increasing direction during a Valsalva maneuver. These findings are consistent with Pourcelot and Pottier and colleagues who reported a decrease in

Page 25

Special Edition on Long Duration Spaceflight stroke volume (≈20%) and cardiac output (≈20%) using echocardiography with thigh compression during a short-duration French-Soviet Salyut flight in 1982. On flight day 4, they used pneumatic thigh cuffs at 40 and 60 mmHg, which is similar to the constraining stress of the Braslet measured by Hamilton et al. (unpublished results) using balloon pressure transducers . Diridollou and Maillet reported that the Braslet used in their HDT study provided the equivalent of 30 mmHg "counterpressure" over the area of its application on the upper thigh.

Lindgren et al. observed a 3% increase in leg volume with 12◦ HDT with thigh cuffs inflated to 50 mmHg for 15 min; however, these subjects would have been considered hypervolemic compared with microgravity.

STEM Today, March 2016, No.6

Herault et al. measured the effects of wearing the Braslet for 5 h on six cosmonauts during a 6-mo stay on the Mir space station and reported stroke volume and cardiac output reduction of 15 and 14%, respectively, after 1 mo of spaceflight. It is interesting to note that these changes became minuscule after 3 mo of spaceflight, and the Braslet actually increased stoke volume and cardiac output after 5 mo of spaceflight. They also report that femoral vein area increased by ≈20% with the Braslet applied at 1 and 3 mo, but 5 mo into the flight this increase was only 9%. Authors did not observe this trend on any subject. Muscle atrophy was more prevalent in pre-ISS missions, altering the fit and, therefore, the efficacy of the Braslet ; the ISS countermeasures system is far more effective at preventing muscle atrophy and exercise deconditioning. Hamilton et al. performed a prospective echocardiographic study in six ISS crewmembers 152, 116, 149, 34, 190, and 41 days after launch and found no clinically significant differences between cardiovascular parameters acquired on-orbit compared with pre- and postmission data.

Authors therefore believe that mission length up to 6 mo does not have a significant effect on the outcome of this study. The myocardial performance index [Tei index (50)] is calculated using diastolic and systolic time intervals derived from TD spectra as a combined measure of myocardial performance. The RV Tei index exceeded the normal terrestrial value of <0.3 in all but one subject. Diridollou and Maillet used high-frequency ultrasound to demonstrate the increase in forehead skin thickness (2.8%) and decrease in tibial skin thickness (3.5%) when subjects are placed in 6◦ HDT. When Braslet was applied during 7 days of HDT, the forehead thickness decreased 0.6% from baseline compared with the control group, which increased by 6.4%.

Arbeille et al. measured IJV distension and facial edema in a HDT study. Compared with supine values they found that facial skin thickness increased by 5% after 7 days of HDT but, after wearing the Braslet for 8 h, the skin thickness was reduced by 5%. This agrees with the subjective comments by Herault et al. , which document that all astronauts who wore the Braslet during their study in space claim to have had a "sensation of comfort."

Matsnev et al. reported that when Braslet was employed on Soyuz-38 the cosmonauts reported a reduction in space adaptation syndrome symptoms (dizziness, congestion, and headaches).

Kirsch et al. measured CVP after 22 h of microgravity during the Spacelab 1 mission and found it to be less than the preflight supine levels in two subjects. They repeated this during the Spacelab D1 missions and found that the CVP again fell to levels below preflight supine 20-40 min after liftoff. This is consistent with the findings

by Buckey et al., which found that invasive CVP fell to 2.5 cmH2 O immediately at the transition to microgravity on the Space Life Sciences 1 mission. Foldager et al. reported that CVP decreased to 6.5-2.0 mmHg after 3 h of microgravity exposure during the Spacelab D2 (n =4) and Space Life Sciences 2 (n = 2) missions.

Although Braslet did not induce a profound change in IJV area despite the obvious reduced cardiac preload, thoracic maneuvers seemed to have a profound additive effect when the Braslet was applied. This implies that the IJV is close to the pressure required to maintain its unstressed volume (i.e., to saturate its filling capacity) and that a Mueller maneuver will decrease its cross-sectional area. The Mueller - Valsalva difference is instructive since Valsalva caused a small increase in IJV area, implying the vein was close to being maximally distended despite the reduced cardiac preload caused by the Braslet. Nonetheless, the IJV area decreases significantly when a Mueller maneuver is performed (Fig. 6). This finding is consistent with the IJV, which takes very little CVP to distend it maximally and has a low enough CVP to be manipulated with limited thoracic pressures when Braslet is applied. This can be replicated at the bedside on a healthy patient by observing the change in fullness of the jugular venous pulse when raising or lowering the patient’s neck by only 1 in. Therefore, a Mueller maneuver with Braslet applied in microgravity seems to decrease the CVP acutely to <2 mmHg. This is consistent with the maintenance of RV preload under normal terrestrial conditions where RV transmural pressures are ≈1.5 mmHg in humans .

The Braslet device acutely reduces the effective circulating volume by sequestering fluid in the lower extremities, as directly observed by vascular ultrasound and supported by the reduced preload indexes measured by echocardiography. Vascular ultrasound confirmed reduced distention of the jugular venous system and increased sensitivity of the jugular vein area to thoracic maneuvers. These findings combined with subjective comments from crewmembers suggest that the relative cranial venous insufficiency caused by microgravity is partially

Page 26

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

alleviated by the Braslet. The hazards secondary to wearing the occlusive Braslet cuffs in microgravity for an extended time (>1 h) are unknown, although in some studies the device was worn for longer than 6 h.

Validation of On-Orbit Methodology for the Assessment of Cardiac Function and Changes in the Circulating Volume Using Ultrasound and Braslet-M Occlusion Cuffs Braslet device acutely reduced the effective circulating volume by sequestering fluid in the lower extremities, as directly observed by vascular ultrasound and supported by the reduced preload indexes measured by echocardiography. Vascular ultrasound confirmed reduced distention of the jugular venous system and increased sensitivity of the jugular vein area to thoracic maneuvers. These findings combined with subjective comments from crewmembers, suggest that the relative cranial venous insufficiency caused by microgravity is partially alleviated by the Braslet device. These respiratory maneuvers were shown to cause statistically significant hemodynamic effects. A statistically significant reduction in jugular venous (IJV) filling was also observed during Braslet application, which coupled with negative airway pressure to consistently collapse the IJV. Remotely guided ultrasound provided an effective and objective means of measuring the physiological effects of the Braslet and produced data quality that was superior to previous investigations in space. Fourteen of eighty-one conditions (27 parameters measured at baseline, Valsalva, and Mueller maneuver) were significantly different when the Braslet-M cuff was applied. Seven of twenty-seven parameters were found to respond differently to respiratory maneuvers depending on the presence or absence of thigh compression. The Valsalva and Mueller maneuvers appeared to enhance the ability at the microgravity bedside to determine volume status.

Impaired cerebrovascular autoregulation and reduced CO2 reactivity after long duration spaceflight THE MICROGRAVITY environment, which causes cephalic fluid shifts with increased arterial pressure at the level of the brain relative to normal daily life on Earth , might cause alterations in cerebrovascular structure and function. The impact of microgravity on human cerebrovascular function has primarily been examined during and after short-duration spaceflights with only a few measurements of cerebral blood flow velocity (CBFV) during or after long-duration flights. During spaceflight, only modest changes in CBFV have been reported with small increases in cerebrovascular resistance after months in space that were speculated to reflect increased sympathetic vasoconstriction . Postflight CBFV in the supine posture was unchanged or slightly elevated from preflight.

Postflight measurements of dynamic cerebrovascular autoregulatory indexes, reflecting vascular smooth muscle responses to changes in arterial blood pressure, were slightly enhanced in one group of astronauts but were reduced in astronauts with reduced postflight orthostatic tolerance. Results from Earth-based analogs of spaceflight using head down-bed rest (HDBR) have revealed increases, decreases, or no changes in supine CBFV. After HDBR, some studies have shown a greater reduction in CBFV with lower body negative pressure (LBNP) or assuming an upright posture , suggesting an impairment in the ability to regulate cerebral blood flow when faced with an orthostatic stress. However, other research has found no difference in cerebrovascular responses to tilt or LBNP or with rapid deflation of leg cuffs.

Page 27

Special Edition on Long Duration Spaceflight Animal models of spaceflight using hind limb suspension have provided convincing evidence of cerebrovascular structural and functional changes.

Cerebral arteries from suspended rats have demonstrated vascular smooth muscle hypertrophy with a smaller luminal cross-sectional area and increased basal myogenic tone . Further work has also shown that these changes are associated with reduced cerebral blood flow and greater vasoconstrictor responses possibly acting through nitric oxidedependent or renin-angiotensin system-dependent mechanisms.

STEM Today, March 2016, No.6

To date, there have been no investigations of the cerebrovascular response to carbon dioxide (CO2 ) after spaceflight. Cerebrovascular CO2 reactivity reflects endothelial function through activation of the nitric oxide system that was impaired in animals by hind limb suspension . In addition, cerebrovascular CO2 reactivity might also be altered as a consequence of elevated partial pressure of CO2 in the ambient air on the International Space Station (ISS) that could chronically influence arterial CO2 (PACO2 ) and cerebral acid-base balance.

Cerebral hemodynamic data from long-duration spaceflight were collected as a part of the Cardiovascular and Cerebrovascular control on return from the International Space Station (CCISS) project. Methods and procedures were reviewed and approved by the Office of Research Ethics at the University of Waterloo and the Committee for the Protection of Human Subjects at Johnson Space Center. Each volunteer signed an approved consent form after receiving full verbal and written details of the experiment. The experiment conformed to the guidelines in the Declaration of Helsinki.

Seven astronauts (1 woman) with an average age of 48 ± 4 years participated. Preflight data were collected 36 ± 22 days before launch. Postflight data were collected from astronauts returning via the space shuttle within 3 to 4 h of landing (R + 0; n = 4) and, for astronauts returning via the Russian Soyuz spacecraft, the morning after landing (R + 1; n = 2) or two mornings after landing (R + 2; n = 1) for an astronaut whose return was delayed by weather. For R + 0 testing, the astronauts landed on the shuttle in a supine posture and remained in that posture for transportation to the research facility and until the experiment was completed. Experiments on days R + 1 and R + 2 were conducted first thing in the morning. The astronauts did not assume upright posture on test day before being transported to the laboratory in a supine posture in an attempt to minimize the effects of re-adaptation to Earth’s gravitational environment. The astronauts spent an average of 147 ± 49 days in space ranging from 58 days to 199 days with all but one spending greater than 100 days on orbit.

RESULTS There were no differences in resting CBFV, CVRi, or PI from pre- to post-spaceflight during the LBNP phase of the study (Fig. 1). In response to LBNP, CBFV (Fig. 1A) and PI (Fig. 1C) were reduced at -20 mmHg with no changes in CVRi (Fig. 1B). BPM CA was not different (P = 0.142) from pre- to postflight at rest (92.9 ± 15.0 and 99.9 ± 8.4 mmHg) or at -20 mmHg of LBNP (92.2 ± 13.4 and 97.4 ± 8.8 mmHg). There were also no differences in PETCO2 (P = 0.737) at rest from preflight (42.3 ± 2.5 mmHg) to postflight (42.7 ± 1.2 mmHg) or at -20 mmHg LBNP from preflight (42.4 ± 3.0 mmHg) and postflight (41.8 ± 1.9 mmHg). The BPM CA and cerebrovascular responses to the two-breath test with intermittent increases of inspired CO2 are displayed for a typical subject in Fig. 2. The transient elevation in PCO2 reduced CVRi and resulted in increases in CBFV, whereas BPM CA maintained spontaneous variations without obvious effect of PCO2 .

Page 28

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

Group mean responses from the ARMA analyses are displayed as the time course of change to a unit input in BPM CA and PCO2 in Fig. 3 and individual step responses in Fig. 4. When BPM CA was considered as the input signal the averaged gain value for BPMCA CBFV was not different pre- to postflight (-0.225 ± 0.453 vs. 0.069 ± 0.119 cm s−1 mmHg−1 BPM CA , P = 0.13; Figs. 3A and 4A), but the BPM CA CVRi gain was reduced 17% after spaceflight (0.035 ± 0.007 vs. 0.029 ± 0.005 units/mmHg BPM CA , P = 0.047; Figs. 3C and 4C). The indicators of CO2 reactivity from the responses of CBFV and CVRi revealed a strong trend for reduced CBFV response from pre- to postflight (2.113 ± 1.228 vs. 1.300 ± 0.595 cm s−1 mmHg−1 PCO2 , P = 0.056; Figs. 3B and 4B) and a reduction in the CVRi response (-0.038 ± 0.018 vs. -0.027 ± 0.012 units/mmHg PCO2 , P = 0.017; Figs. 3D and 4D).

The major findings of this study showed that long-duration spaceflight was associated with reductions in the indexes of cerebrovascular dynamic autoregulation and CO2 reactivity with no differences seen in resting CBFV or responses to low levels of LBNP. These findings, following long-duration spaceflight on the ISS, were consistent with hypotheses that suggested cerebrovascular consequences from chronic elevation in cerebral blood pressure and chronic exposure to elevated atmospheric PCO2 . The results contrast with improved dynamic autoregulation after the short-duration Neurolab spaceflight mission but are similar to findings in astronauts who presented with orthostatic intolerance after spaceflight.

Cerebrovascular indicators at rest and during LBNP Authors anticipated greater reduction in CBFV during LBNP after long-duration spaceflight, indicative of impaired cerebral blood flow regulation, but both at rest and during LBNP the CBFV, PI, and CVRi were unchanged from preflight. Resting CBFV is expected to reflect the metabolic demands of the brain as well as the constant influence of PaCO2 . This research finding of no change in supine CBFV after spaceflight was consistent with other observations after short- and long-duration flights . The unchanged estimate of PaCO2 taken from the end-tidal PCO2 made it unlikely that alterations in CO2 had an effect on resting cerebral blood flow.

Page 29

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

The potential effects of long-duration spaceflight on cerebrovascular structure and function have been inferred from HDBR studies with humans and from animal models of hind limb suspension . The human HDBR studies reported variable results with increases, decreases, and no change in CBFV. Animal models have revealed reduced flow accompanied by significant structural changes with increased thickness of vessel walls and reduced internal dimensions . The animal models also suggested an involvement of renin-angiotensin system might in structural modifications with hind limb suspension . An upregulation after 4 wk, but not after 7 days, of proteins involved in the synthesis of angiotensinogen and of angiotensin II type 1 receptors was observed in cerebral and carotid arteries, and these changes were associated with increased arterial wall thickness . Based on these results from animal models, it is possible that human cerebral arteries might also undergo structural change with prolonged exposure to microgravity. At the moment, it is not possible to rule out the possibility that, although CBFV was unchanged, cerebral blood flow might have been reduced as a function of a smaller MCA. This can only be tested in future studies by quantitative measurements of cerebral blood flow or measures of MCA dimensions.

In the current study, CBFV was reduced during -20 mmHg of LBNP, but there was no difference in this response from pre- to postflight. These results contrast with observations after the short-duration Neurolab mission where there was a smaller reduction in CBFV during LBNP as well as during upright tilt compared with preflight. The LBNP results after long-duration spaceflight were consistent with observations from cosmonauts after 6 mo in space; however, at greater levels of LBNP, cosmonauts showed a trend toward greater reductions in CBFV after flight . In the current study, it is therefore possible that -20 mmHg LBNP was not sufficient to challenge cerebral blood flow regulation, since these same astronauts only showed small changes in postflight baroreflex function in the seated posture ; therefore, it is still possible that changes in CBFV might have occurred with greater levels of LBNP or in assuming an upright posture. Reduced PI during LBNP might have reflected a relative reduction in cerebrovascular resistance under the conditions of constant PETCO2 in the current study , but there were no apparent pre- to postflight differences in this indicator of static cerebrovascular autoregulation.

Dynamic cerebrovascular autoregulation Dynamic cerebrovascular autoregulation, in contrast with static autoregulation , is an index of the rapid responses of the cerebrovascular system to acute changes in BPM CA . Because an inverse relationship exists between the dynamic autoregulatory index and PCO2 , it was important that resting PCO2 was not different from pre- to postflight and further that the simultaneous effects of BPM CA and PCO2 were accounted for by the ARMA model. The 17% reduction in gain for CVRi relative to the change in BPM CA provided an index of impaired dynamic cerebrovascular autoregulation after spaceflight. Similarly, there was a trend toward a reduced

Page 30

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

CBFV response to a change in BPM CA (P = 0.128). Reduced cerebrovascular dynamic autoregulation after long-duration spaceflight was consistent with some but not all findings after HDBR and with astronauts who had orthostatic intolerance after short-duration spaceflights . Altered postflight cerebrovascular control might reflect the structural and functional changes observed in the animal models of spaceflight .

Cerebrovascular reactivity to CO2 No previous investigation has examined cerebrovascular CO2 reactivity after spaceflight. Animal studies showing reductions in nitric oxide- dependent dilation of cerebral arteries after hind limb suspension suggested that the CO2 response might be altered due to the link between CO2 -induced dilation of human cerebral arteries and nitric oxide- dependent mechanisms . CO2 reactivity was investigated in the current study by the two-breath method , which accounts for the spontaneous variability in PCO2 that can complicate interpretation of cerebrovascular responses . The reduced gain for the CBFV and CVRi responses computed with respect to the input of PCO2 support the hypothesis that the nitric oxide- dependent dilatory mechanisms of the cerebrovascular system might be impaired by long-duration spaceflight. The environment of ISS had an average inspired PCO2 of 3.34 mmHg during the experiments described in this study. This value is over 10-fold greater than ambient conditions on Earth. Data from a clinical population suggest that chronic exposure to hypercapnia leads to blunting of cerebrovascular CO2 reactivity ; however, it is unclear if chronic exposure to increased atmospheric PCO2 , as seen on ISS, produces reductions in cerebrovascular CO2 reactivity in a healthy population. Slightly elevated inspired CO2 might have had a chronic effect on PaCO2 and arterial acid-base, which could in turn affect cerebrovascular resistance. To date, no arterial blood samples have been taken on ISS to determine whether changes in PaCO2 have occurred.

Individual variability and potential implications The responses of dynamic autoregulation and CO2 reactivity, and the pre- to postflight differences, varied between astronauts. Finding a range of preflight baseline CBFV and CVRi responses to the input of BPM CA (Fig. 4, A and C) is similar to previous observations . Likewise, differences between subjects in CO2 reactivity (Fig. 4, B and D) are expected and might be related to differences in nitric oxide-dependent dilation, but this was not directly tested. Of interest in the current study is the magnitude of change following spaceflight. One individual (solid circle, Fig. 4) had the largest change from pre- to postflight in both CBFV and CVRi responses to BPM CA along with one of the larger changes in response to PCO2 . The other astronauts had relatively consistent changes. The functional consequences of these changes cannot be identified from the current study; however, a recent study noted that reductions in dynamic cerebrovascular autoregulation were associated with orthostatic intolerance. No tilt tolerance tests were performed by the astronauts in the current study.

Page 31

Special Edition on Long Duration Spaceflight

Implications of individual variability in the changes in cerebrovascular dynamic autoregulation and CO2 reactivity beyond orthostatic tolerance are not known. However, NASA has recently identified idiopathic intracranial hypertension as a risk factor for postflight visual problems . The current data showing a range of changes in postflight cerebrovascular responses might provide impetus to investigate the potential for transcranial Doppler measurements to yield information on risk for these visual problems.

STEM Today, March 2016, No.6

Measurements of jugular, portal, femoral, and calf vein cross-sectional area for the assessment of venous blood redistribution with long duration spaceflight One of the major cardiovascular disturbances induced by exposure to microgravity is the passive fluid shift towards the thorax and the head. Such a shift in fluid results in a transfer of fluid to thoracic and cephalic tissues (facial edema) in addition to increasing cardiac chamber volume which triggers neuro-hormonal mechanisms resulting in reduced plasma volume and decreased cardiac mass. However, the increase in jugular vein diameter (spaceflight and bed rest), femoral vein diameter (with spaceflight but not bed rest) , and in portal vein diameter (bed rest) induced by this fluid shift do not trigger any compensatory phenomena allowing for pooling of fluid and potentially resulting in negative physiological responses. A previous 1 week bed rest study reported an increase in jugular vein diameter and signs of liquid stagnation during eye fundus examinations (increased papilla area and ocular vein diameter) without an increase in eye pressure.

Recently, astronauts have reported visual deficiencies during long duration spaceflight which may be related to an increase in fluid similar to that observed with bed rest, or possibly due to an increase in pressure around the ocular globe . The objective of the current study was to investigate, during 6 months of spaceflight on the International Space Station (ISS), the jugular vein (JV), portal vein (PV), femoral vein (FV), tibial vein (TibV), and gastrocnemius vein (Gast V) properties to quantify venous blood flow redistribution during the early and late phases of exposure to microgravity.

It was hypothesized that during spaceflight: (a) jugular vein volume would increase and remain increased for the duration of the flight due to the cephalic fluid shift, (b) portal vein cross-section would increase during the flight as there is no direct communication between the splanchnic vascular output (portal vein) and the systemic venous network (inferior vena cava) indicating pooling of blood flow in the splanchnic region, (c) femoral vein would also increase despite the venous blood volume gravity centre being moved toward the upper part of the body, (d) although changes in vein cross-sectional area or volume will be of different amplitude, all astronauts will show the same directional change in vein size regardless of inflight physical activity and nutrition.

Ten astronauts (7 male, 3 female, age: 47 ± 5 years, mass 69 ± 12 kg, height: 1.72 ± 8m) participated in the Vessel Imaging study. The protocol of this study was approved by the NASA and ESA ethical committees under the reference IRB pro 0340 with all procedures and protocols being conducted in accordance with the Helsinki declaration. Each astronaut was informed about the content and schedule of the experiment, signed an informed consent form before participation, and was aware of his or her right to withdraw from the study at any time without prejudice. During the ISS spaceflights astronauts had calibrated meals (meals designed for each individual astronaut by nutritional specialists) but food and liquid intake were not strictly controlled.

Astronauts also performed daily exercise (treadmill approximately 3 h per day) but this was not standardized and differed between astronauts.

Results Ninety percent of the video files downlinked from the ISS were of sufficient quality to be processed allowing for measurement. Due to missing data points (equipment failure and delayed replacement), two astronauts were excluded from the inflight analysis (n = 8). With spaceflight, PV volume (Fig. 2a) and JV volume (Fig. 2b) were both significantly increased with respect to pre-flight values both early and late in the spaceflight (PV: +36 and + 45 %; JV +178 and +225 %). The magnitudes of the increase in PV volume and JV volume were not equivalent resulting in a significant increase in the JV/PV ratio (JV volume to PV volume ratio–Fig. 2c, p < 0.05) both early and late in the spaceflight (JV/PV: +102 and +120 %). Similar to the PV and JV results, FV area (Fig. 3a) was also found to be increased early (+124 %) and late (+169 %) during the spaceflight. In contrast, TibV (Fig. 3b) and Gast V (Fig. 3c) were found to be decreased with spaceflight (TibV: -46 and -52 %; Gast V -68 and -55 %). All values measured returned to pre-flight levels on R + 4 with the exception of TibV which was still reduced (-19 %). With the transition from a supine to a seated posture, both the TibV area and Gast V area were seen to increase. There was no difference in this change post-spaceflight compared to the pre-flight response (Table 1).

Page 32

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

Increase in jugular, portal, femoral vein size The current study demonstrated significant changes in venous dimensions with long duration spaceflight which

Page 33

Special Edition on Long Duration Spaceflight may be indicative of venous blood pooling.

These results support the hypotheses of the study in that all astronauts showed the same directional changes in JV volume, PV volume, and FV crosssectional area indicating that the observed venous changes are the result of exposure to the spaceflight environment and are consistent across the astronaut population. As diet and exercise were not strictly controlled during the flight, and astronauts reported both increased and decreased physical activity levels and salt intake, it would appear that the observed responses occur independently of physical activity or nutritional status.

STEM Today, March 2016, No.6

Additionally, the differences in the magnitude of venous changes in different vascular beds may indicate significant blood pooling and fluid shifts with spaceflight which may have additional physiological consequences requiring further study. Despite a possible reduction in circulating blood volume of approximately 10 % and carotid and femoral arterial flows remaining stable as already reported during long term flights , the jugular and femoral veins in the current study were found to be significantly increased in all astronauts during spaceflight.

This increase in size of the jugular and femoral veins could indicate a passive stowage of blood at the cephalic and pelvic areas, respectively, and not an increase in flow rate or circulating blood volume. In contrast to the femoral and jugular veins, the increase in portal vein cross-sectional area and volume may indicate that more blood was stowed in the portal venous network, and that possibly, more blood was traveling through the splanchnic area. A similar increase in portal vein area was previously observed during bed rest and corresponded both to an increase in blood volume inside the portal vein network (increased crosssectional area) and in flow volume (ml/min) crossing the vessel (increased blood velocity and cross-sectional area). The increase in portal vein cross-sectional area with bed rest was observed despite a significant reduction in plasma volume and cardiac volume , and a decrease in mesenteric arterial vascular resistance. These observations support the idea that the increase in portal vein cross-sectional area might be indicative of increased splanchnic blood volume and flow; however, in the present study, we did not have access to flow velocity and thus can only comment on flow volume (ml) and not flow volume change (ml/min) in the splanchnic area.

Finally, the increased venous dimensions observed in this study suggest the presence of higher amounts of blood in the cephalic, splanchnic, and pelvic regions during the flight which confirm the 3 first hypotheses.

Additionally, despite the high amplitude variability of these changes among the astronaut group, each show the same directional change in vein size which confirm the 4th hypothesis.

Potential consequence of the cephalic, splanchnic, and pelvic, regional venous blood pooling These networks are not designed to store large amounts of fluid over long periods of time which may have an impact on organ structure and function. The hydrostatic pressure against cephalic organs (eye, brain, thyroid, superficial muscle skin tissue) may increase and provoke a higher filtration towards these tissues as evidenced by the presence of a facial skin edema. A similar phenomenon may also be present inflight at the eye fundus level (increase papilla and vein) and may contribute to the visual impairment reported by several astronauts inflight.

Additionally, the blood stagnation inside the jugular vein may contribute to increased intracranial pressure , potentially resulting in impaired cerebrovascular reactivity . Several studies have reported an increase in circulating insulin in astronauts during spaceflight and an increase in glycemia and insulin resistance during bed rest . These changes may be in part due to altered abdominal organ function from splanchnic venous pooling.

However, this has yet to be investigated. Similarly, the increase in femoral vein cross-sectional area confirms the presence of venous blood stagnation in the pelvic region but to date, there is no evidence to suggest a potential impact on pelvic organ structure or function.

The jugular vein volume to portal vein volume ratio (JV/PV) was calculated with the objective to quantify the proportion of venous blood volume stowed at the cephalic level compared to the splanchnic one. This index of venous blood volume (blood pooling) redistribution was significantly increased with spaceflight by approximately 102 and 120 % which indicated that the increase in venous blood volume was higher at the jugular

Page 34

Special Edition on Long Duration Spaceflight level than at the splanchnic one. Previous reports have indicated that only a subset of astronauts experience visual problems with long duration spaceflight . This may be related to individual differences in venous blood pooling as not all astronauts in the current study showed the same magnitude increase in the JV/PV ratio with spaceflight. Future studies will look to determine if the magnitude of cephalic venous blood pooling relates to the incidence and degree of visual symptoms observed during long duration spaceflight.

STEM Today, March 2016, No.6

Reduction of calf vein in relation with microgravity induced fluid shift In contrast to the jugular, portal, and femoral veins, the calf veins were found to be significantly reduced for the duration of the spaceflight. During spaceflight, changes in body posture do not result in distension of the leg veins that would be normally seen on Earth. However, despite the veins of the calf being significantly smaller during spaceflight, the distension response to a change in body position (supine to sit) upon return to Earth was not altered. This is in contrast to results from long duration bed rest which reported greater distensibility of the veins during lower body negative pressure (LBNP) or stand tests . It is possible that the exercise countermeasures used by astronauts on the ISS may have contributed to the maintenance of the response of leg veins to posture change.

Additionally, it should be noted that supine to sit transitions may not be equivalent to stand test or LBNP. Therefore, it is unclear if the lower leg vein response would be consistent if a greater orthostatic stress was applied. Although the inflight countermeasures may have contributed to maintain leg vein responses, it did not appear to attenuate the venous blood pooling observed in the current study. Other physical countermeasures have also been suggested to reduce the liquid stagnation at the cephalic level. The thigh cuffs called "Braselets" apply a pressure of approximately 30 mmHg at the upper part of the thigh which traps blood and other fluids in the superficial leg veins and tissues, and consequently, reduces the jugular vein area inflight, restoring it to pre-flight or pre-bed rest levels . The use of thigh cuffs inflight also reduced the discomfort at the head level.

Unfortunately, during the time the thigh cuffs were applied (approx. 5-8 h on daytime), the femoral veins remained markedly enlarged which may serve to negatively influence the mechanical properties of these veins.

Lower body negative pressure has also been suggested as a countermeasure to reduce the jugular vein size and the amplitude of the fluid shift towards the head. However, during spaceflight, LBNP could not be applied for extended periods of time. Therefore, presently, no effective countermeasures have been identified to reduce the accumulation of venous blood in various vascular territories.

The results of the echographic examinations showed increased jugular vein and portal vein volume, increased femoral vein area, and reduced calf vein area. These observations suggest that significant venous blood pooling is evident with spaceflight that persists throughout the duration of the flight. The consequences of this stagnation of fluid still remain unclear and future work is required to determine these consequences and develop effective countermeasure for use in long duration spaceflight.

Proteomic Profiling of Human Heart Tissue Exposed to Microgravity Microgravity is a stress experienced during space travel and has been linked to changes in cardiac structure and function in astronauts and cosmonauts. In order to define the mechanisms by which microgravity alters cardiac structure and function, author performed a proteomic investigation of the effects of simulated microgravity on the cardiomyocyte proteome. Using a combination of label-free relative quantitation and dynamic stable isotope labeling by amino acids in cell culture (dynamic SILAC), author compared both the protein expression changes and protein turnover rates between two experimental groups: simulated microgravity and normal gravity.

Label-free data revealed that microgravity markedly alters protein expression over time, specifically altering the levels of proteins involved in muscle contraction and structure, translation, metabolism, protein folding and transport, and calcium handling. Dynamic SILAC data demonstrated that protein turnover is diminished in response to simulated microgravity. This observation in combination with the decline in translational proteins suggests that protein translation may mediate the decline in contractile and structural proteins observed in the microgravity group. To validate this finding, author examined three independent measures of protein translation and three surrogates of cell damage and protein degradation. Relative to normal gravity, protein translation was decreased in simulated microgravity, while cell damage and protein degradation was not statistically significant between groups, leading to an overall conclusion that decreased protein translation may mediate microgravityinduced changes to cardiac structure and function.

In order to define the mechanisms by which microgravity alters cardiac structure and function, author performed a proteomic investigation of the effects of simulated microgravity on the cardiomyocyte proteome.

Using a combination of label-free relative quantitation and dynamic stable isotope labeling by amino acids in cell culture (dynamic SILAC), author compared both the protein expression changes and protein turnover rates

Page 35

Special Edition on Long Duration Spaceflight between two experimental groups: simulated microgravity and normal gravity (1x gravity) over time. From this studies the following general concepts were obtained: • Protein expression is altered in cardiomyocyte following simulated microgravity. Author measured proteinlevel expression for 848 proteins (6,174 peptides) across all timepoints and gravity conditions. As expected very few proteins are differentially expressed (p<0.05) at 12 hours, and the number grows over time, until nearly 100 proteins are differentially expressed. Proteins were classified into three groups: 1) unchanged over time under either condition, 2) equally changed over time under either condition, and 3) differentially changed over time between microgravity and 1x gravity. The significantly altered proteins were then grouped by biological function; seven functional groups were formed: Muscle contraction/structure (26%); Translation (23%); Metabolism (21%); Protein transport and folding (12%); Calcium handling (5%); and Other.

STEM Today, March 2016, No.6

• Cardiomyocytes under simulated microgravity have diminished protein turnover. Using the ratio of the heavy-isotope label to the total of heavy and light peptide in the dynamic SILAC labeling component of the experiment, data strongly suggest that amino acid incorporation and thus protein turnover is diminished in the cardiomyocyte subjected to simulated microgravity. No statistical difference exists between measured turnover at 12 hours, however p<1e-6 at both 48 hr and 120 hr. This data is highly suggestive that protein turnover is drastically decreased under the microgravity condition, and since access to the translational machinery to precursor amino acid did not seem to be different, the difference points to transcriptional or translational control.

• Protein translation may be slowed in cardiomyocytes under simulated microgravity. Following the independent quantitative and label-incorporation analyses, two features of the data were connected. The protein expression data revealed that many proteins were downregulated following 120 h of microgravity, specifically, a number of translational proteins were diminished. The decrease in translational proteins suggests that protein translation is slowed, which could be mediating the decline in expression of the other significantly reduced proteins. This revelation was strengthened by the markedly reduced protein turnover observed in the label-incorporation data. Because decreased translation is a characteristic that has been observed in microgravity-induced skeletal muscle atrophy, author decided to perform independent experiments that could validate this potential phenomenon: azidohomoalanine (AHA) incorporation, luciferase activity using adenoviral technology, and total protein-to-total RNA ratio.

AHA is a methionine analogue that is incorporated into an individual peptide in a similar fashion as arg or lys. However, AHA was presented to the groups only at 120 h time point and only for 2 h; this design was merely to obtain a snapshot of protein synthesis at the time when the largest separation in protein turnover was observed. AHA incorporation is robust in 0 h controls but is dramatically reduced after 120 h in microgravity relative to 1x gravity. Adenoviral technology was used to deliver the luciferase gene to each group. This technology allows the gene to be transcribed and translated into protein using the cell’s inherent machinery. For this experiment, the Adenoviral-Luciferase was delivered to all cells 3 h prior to group separation. The adenovirus was removed upon group separation, and the luciferase protein activity was measured at 120 h. Luciferase activity is a direct correlate to luciferase protein content, since no posttranslational modifications are necessary to initiate luciferase enzyme function. Light detection revealed that luciferase protein was significantly diminished 7.5 fold in microgravity compared to 1x gravity. Cell pellets were collected from each group at 120 h, and total protein and total RNA were assessed in order to provide a global examination of protein synthesis relative to the amount of RNA. Data revealed that the total protein-to-total RNA ratio was significantly attenuated 4 fold in microgravity relative to 1x gravity.

In all, these independent measures support the SILAC label-incorporation data, indicating that protein turnover in the cardiomyocyte is diminished following 120 h of exposure to simulated microgravity.

• Indicators of cell damage and protein degradation are not significantly affected in response to simulated microgravity. Author next examined indicators of cell damage, death, and protein degradation in attempt to determine if microgravity was inducing damage to the cardiomyocyte, thereby altering the proteome, or enhancing protein degradation, thereby slowing protein turnover. Lactate dehydrogenase is a soluble cytosolic enzyme, and its release is an indication of cell membrane permeability or damage. The media was examined from each group at 0, 1, 2, 3, 4, 8, 12, 24, 48, 96, and 120 h. No statistically significant difference was observed between microgravity and 1x gravity at any time period.

Caspase-3 is a classical end-effector protease involved in apoptosis. An increase in caspase-3 is a surrogate for enhanced cell death. Author surveyed caspase-3 activity at 0, 12, 48, and 120 h, and did not observe a statistically significant difference between microgravity and 1x gravity at any time point. Finally,

Page 36

Special Edition on Long Duration Spaceflight we assessed global ubiquitination of protein lysates at 0 h and from each group at 120 h. Again, no difference was observed between microgravity and 1x gravity. Together, these data indicate that cardiomyocytes under microgravity are not markedly damaged in response to simulated microgravity. Moreover, because ubiquitination of proteins is a way proteins are tagged for degradation, these data suggest that microgravity is not enhancing protein degradation, which also indicates that protein degradation may not be playing a critical role in the diminished protein turnover induced in microgravity.

STEM Today, March 2016, No.6

Long-Term Simulated Microgravity Causes Cardiac RyR2 Phosphorylation and Arrhythmias in Mice

Exposure to microgravity during space flight causes various changes in the human cardiovascular system, including a cephalic fluid shift, changes in cardiac systolic volume, and, over time, a loss of left ventricle mass. It

Page 37

Special Edition on Long Duration Spaceflight has been suggested that microgravity leads to a reduction in cardiac output and stroke volume due to cardiac remodeling, triggered by a reduction in circulating blood volume. In addition, documented observations in crew members over several years suggest that long-term exposure to microgravity can alter the electrical properties of the heart, increasing the propensity towards cardiac arrhythmias.

One factor thought to contribute to enhanced arrhythmogenic risk is the cardiac sympathovagal imbalance observed in response to microgravity. However, at present, it remains unclear whether microgravity is a direct cause of arrhythmias and cardiac dysfunction, or whether it is caused indirectly by previously asymptomatic cardiovascular disease.

STEM Today, March 2016, No.6

It is well established that abnormal intracellular Ca handling can contribute to both contractile dysfunction in heart failure (HF) and arrhythmogenesis. Diastolic release of Ca from the sarcoplasmic reticulum (SR) via the type-2 ryanodine receptor (RyR2) can activate the Na/Ca-exchanger, causing delayed afterdepolarizations (DADs). DADs can trigger premature action potentials that can initiate arrhythmias through local reentry. In addition, the leak of Ca from the SR (via RyR2) can interfere with SR reloading, associated with weakened cardiac contractility. One major reason for enhanced RyR2 activity, or "leaky" RyR2 channels, in heart failure is increased phosphorylation by Ca/calmodulindependent protein kinase II (CaMKII). This enzyme becomes activated in diseased hearts, whereas inhibition of CaMKII can improve cardiac contractility and prevent cardiac arrhythmias. Specifically, CaMKII phosphorylates residue S2814 on RyR2 in normal and diseased hearts, while genetic inhibition of S2814 phosphorylation in mice subjected to pressure-overload ameliorates cardiac failure and ventricular arrhythmias. At this time, however, it is unknown whether this signaling pathway also contributes to the pathogenesis of HF and arrhythmias associated with microgravity-induced cardiac remodeling.

Rrsults Validation of the mouse hindlimb unloading (HU) model of simulated microgravity To assess the functional effects of microgravity on cardiac physiology, we subjected C57Bl/6 mice to experimental hindlimb unloading (n=15). Wire rings were implanted between the 6th and 7th intervertebral space of the tail, allowing for suspension at a 30- degree angle (of the HU group) (Fig. 1A). Control mice (n=19) also receive the rings but were not suspended. The design of the suspension assembly enabled mice to move around the cage freely, with unrestricted access to food and water (Fig. 1B,C). To validate the model, the body weight of each animal was monitored on a weekly basis, prior to and following HU (or sham). The body-mass of HU mice prior to suspension (26.1 ± 1.5g) was not significantly different from sham (26.4 ± 1.5g; P=0.65). After 7 days of HU, mice displayed a minor loss in body weight (-5.4%). However, there were no statistically significant changes in body weight following 14, 21, or 28 days of HU, compared to day 7. Furthermore, no significant changes in body weight were evident between mice subjected to HU for 28 days (25.0 ± 1.0g) and sham mice (26.4 ± 0.8g; P=0.21), consistent with previous studies.

Dual-energy X-ray absorptiometry (DXA) was also performed to analyze body composition in these HU and sham mice, prior to and at day 14 and 28 following HU (Fig. 2A). Analysis of whole body images revealed no significant changes in the amounts of lean muscle tissue or bone mineral content (BMC) in mice following 14 or 28 days of HU compared to their baseline values or sham mice (data not shown). In contrast, analysis of the femur region revealed a progressive decline in lean muscle mass (Fig. 2B) and BMC (Fig. 2C), starting at 14 days and continuing up to 28 days after HU. Following completion of the experiments, hindlimb muscles were dissected to confirm the extent of muscle mass loss. The soleus muscle weight was reduced by 54% in HU mice (5.9 ±1.7 mg) compared with sham mice (12.8 ± 3.6 mg; P<0.05). The gastrocnemius weight was also reduced, by 20%, in HU mice (117.5 ± 16.2 mg) compared with sham mice (147.5 ± 14.6 mg; P<0.05). Thus, simulated microgravity by means of HU caused changes in body composition including bone demineralization and muscle atrophy of the lower extremities.

Development of cardiac dysfunction following hindlimb unloading To evaluate the functional consequences of simulated microgravity on cardiac function, transthoracic echocardiography was performed prior to, 28, and 56 days after HU. At baseline (1 day before HU), left ventricular ejection fraction (LV-EF) and cardiac dimensions were similar in sham and HU mice (Fig. 3). However, after 28 days of HU, LVEF was significantly decreased in HU mice (57.5 ± 1.4) compared to baseline (64.9 ± 1.3; P<0.001) and sham (66.7 ± 0.7; P<0.001) (Fig. 3A). After 56 days of HU in a separate group of mice, the LV-EF was decreased further (47.8 ± 1.1) compared with baseline (64.9 ± 1.3; P<0.001) or sham (62.1 ± 1.9; P<0.001). In contrast, there were no significant changes in sham mice over time (P=0.64). The end-systolic LV internal diameter (LVIDs) increased in the HU mice starting at 28 days and further progressed up to 56 days after HU (Fig. 3B). Specifically, the LVIDs was significantly increased in HU mice at 56 days of HU (3.2 ± 0.1 mm) compared with HU mice at baseline (2.5 ± 0.1 mm; P<0.001), and compared to sham mice at 56 days of HU (2.6 ± 0.1 mm; P<0.001). Furthermore, the end-diastolic LV internal diameter (LVIDd) was also signifi-

Page 38

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

cantly enhanced after 56 days of HU (P<0.05; Fig. 3C). Together, these data show that HU leads to progressive cardiac remodeling associated with cardiac dilatation and mild systolic heart failure

Page 39

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

Increased susceptibility to ventricular arrhythmias following hindlimb unloading To determine whether simulated microgravity alters the electrical properties of the heart, authors first performed surface electrocardiograms (ECGs) on anesthetized mice using the MouseMonitor (Indus Instruments, Houston, TX) heating pad with embedded ECG leads to reduce noise (Fig. 4A,B). Analysis of ECG intervals revealed no significant differences in RR interval and PR intervals (Fig. 4C,D). There was a mild trend towards prolongation of the QTc interval (QT interval corrected for RR) in experimental mice (n=10) at 28 days after HU (40.9 Âą 1.0 ms) compared to sham (n=14) controls (38.3 Âą 1.3 ms; P=0.14) (Fig. 4E), a possible occurrence consistent with the possible presence of a potentially arrhythmogenic substrate.

Because spontaneous arrhythmias were not observed during the brief ECG recording periods (<5 min), basal surface ECG recordings were evaluated for the presence of spontaneous premature ventricular contractions (PVCs) during a 30-min period. Whereas none of the sham controls exhibited any PVCs (0 of 14 mice), 20%

Page 40

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

(2 of 10) of the HU mice showed PVCs (P<0.05). Next, authors performed programmed electrical stimulation (PES) using an intracardiac pacing catheter. Ventricular pacing was performed at basic cycle lengths of both 90 and 70 ms (Fig. 5A, first 2 arrows below tracings depict 90 ms cycle length), followed by a premature beat (last arrow below tracings). PES provoked runs of non-sustained ventricular tachycardia (VT) in 50% of mice at 28 (n=10) or 56 days (n=4) of HU, respectively, compared to sham mice (16%; P=0.08) combined from both time points (n=19) (Fig. 5B). To gain more insight into the potential differences in arrhythmogenesis among the groups, we expressed the induction of VT as a percentage of total induction attempts (Fig. 5C). The percent of pacing protocols leading to VT induction was significantly greater in mice at 28 (30%) and 56 days (36%) after HU, compared to sham mice (13%; P<0.05) from both groups (Fig. 5C). Together, these data suggest that simulated microgravity increases the susceptibility of VT.

Altered sarcoplasmic reticulum Ca handling following hindlimb unloading To determine the mechanisms underlying enhanced arrhythmogenesis and contractile dysfunction, authors studied SR Ca handling in ventricular myocytes isolated from mice at 28 days of HU (n=3) or sham (n=2) (Fig. 6).

After a steady-state pacing train at 3-Hz, spontaneous SR Ca release events (SCaREs) were observed in myocytes from HU (n=11 cells) but not sham (n=7 cells) mice (Fig. 6A, B). Quantification of these SCaREs revealed more frequent, significant events in HU mice (2.25 [interquartile range, 0.0 to 5.3 events/min]) compared with sham mice (0 events/min; P<0.05; Fig. 6C). Next, tetracaine (TTC) was applied to measure the diastolic Ca leak from the SR, as described. Myocytes from HU mice exhibited enhanced SR Ca leak (16.3 [interquartile range, 10.0 to 28.2%]) compared with sham mice (9.7 [interquartile range, 8.9 to 10.8%]; P<0.05) (Fig. 6D). Finally, application of caffeine caused the release of Ca stored in the SR, resulting in a large Ca transient from which the SR Ca load was calculated. There was a trend toward reduced SR Ca content in myocytes from HU mice,

Page 41

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

consistent with enhanced SR Ca leak (P=0.09; Fig. 6E). These cellular studies show that simulated microgravity causes profound alterations in intracellular Ca handling in ventricular myocytes, consistent with depressed contractility and enhanced arrhythmogenesis.

Hindlimb unloading causes enhanced CaMKII activation and phosphorylation of Ca handling proteins RyR2 and PLN To gain more insight into the signaling pathways possibly involved in the enhancement of SR Ca leak, authors determined whether CaMKII was activated in the hearts of mice subjected to 28-days of HU. Western blotting of ventricular lysates revealed unaltered levels of total CaMKII levels, but increased levels of CaMKII autophosphorylation at T287 (Fig. 7A). Thus, the degree of CaMKII auto-phosphorylation was increased by about 40%, indicative of increased CaMKII activity in the heart (Fig. 7D). To assess the potential effects on key Ca handling proteins, the CaMKII phosphorylation levels of RyR2 (Fig. 7B) and PLN (Fig. 7C) were determined.

Normalized to total protein levels, CaMKII phosphorylation of RyR2 at S2814 and PLN at T17 were both significantly elevated in hearts from HU mice compared with sham (Fig. 7D). In contrast, PKA phosphorylation of RyR2 at S2808 (Fig. 7B,D) and PLN at S16 (not shown) were unaltered in HU mice. Together, these data show that simulated microgravity activates CaMKII and promotes CaMKII-dependent phosphorylation of Ca-handling proteins RyR2 and PLN, which might be responsible for enhanced SR Ca leak.

Page 42

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

Cardiac arrhythmias in astronauts during space flight have been relatively benign, but arrhythmias do occur quite commonly. A number of reports have been published with detailed information regarding in-flight arrhythmias. With only limited medical assistance and supplies available on spacecrafts and the international space station, concerns exists about the potential detrimental consequences of cardiac rhythm disorders. Cardiac arrhythmias could jeopardize mission objectives and possibly the lives of crewmembers. The risks of fatal arrhythmias could be even more profound in case of potential future flights to Mars, which could last over one year. In addition, the recent expansion of commercial space flights exposes participants during short-term orbital flights to developing serious arrhythmias, in particular since these individuals are typically older and less healthy than career astronauts. However, the mechanisms that contribute to arrhythmogenesis during longduration spaceflight have not been elucidated. Several factors might contribute to arrhythmogenesis during spaceflight. Once exposed to microgravity, the human cardiovascular system undergoes profound changes, including alterations in autonomic regulation that may adversely influence cardiac repolarization and precipitate rhythm disturbances, In addition, long duration spaceflight has been shown to prolong cardiac conduction and repolarization. In astronauts involved in the Skylab missions, several instances of arrhythmias including PVCs premature atrial contractions and re-entrant tachycardias have been recorded. These events occurred throughout the entire mission and in particular during effort tests, extravehicular activities, and lower body negative pressure sessions. In addition, an isolated incident of a non-sustained 14-beat ventricular tachycardia was recorded in a crewmember, using in-flight Holter monitoring, aboard the Mir.

Page 43

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

Although not part of a systematic scientific study, these specific cases suggest that exercise and/or sympathetic activation might promote or exacerbate cardiac arrhythmias during long-duration space flight. A recent study suggests that exposure to microgravity increases the predisposition to cardiac arrhythmias during an acute stressor as a result of altered function of gap junctions, in particular connexin 43 (Cx43). In this study, electrocardiographic data were obtained over a short period of HU using an implantable telemetry device following administration of a sympathetic stressor, isoproterenol, in combination with animal restraint.

The findings of this study suggest that activation of simulated microgravity in mice also leads to activation of CaMKII, an enzyme also activated by sympathetic activation. Activated CaMKII was associated with enhanced phosphorylation of S2814 on RyR2 and T17 on PLN (Fig. 7). The absolute increase in CaMKII autophosphorylation levels and RyR2-S2814 phosphorylation were statistically significant but relatively modest (about 30%) in HU mice (Fig. 7). However, prior studies have demonstrated that such increases can functionally alter RyR2 open probability and may contribute to arrhythmogenic Ca release events. Moreover, recent studies have demonstrated that diastolic Ca leakage via CaMKII phosphorylated RyR2 is a major contributor to proarrhythmic events and cardiac failure resulting from dilated cardiomyopathy (DCM). Indeed, our findings revealed enhanced SR Ca leak in ventricular myocytes isolated from mice subjected to hindlimb unloading (Fig. 6). Diastolic SR Ca release can lead to delayed afterdepolarizations, PVCs, an enhanced susceptibility to arrhythmias, and heart failure. Interestingly, Jennings et al. reported that treatment with a calcium channel blocker suppressed reoccurring PVCs in a spaceflight participant. However, it remains very difficult to obtain mechanistic insights from such observational studies in human subjects. Other ion channels can also contribute to triggered activity promoted by enhanced CaMKII activity, in particular activation of potassium and sodium

Page 44

Special Edition on Long Duration Spaceflight channels. Current experimental animal studies suggest that abnormal intracellular Ca release due to CaMKII activation might contribute to arrhythmogenesis, and the eventual decline of cardiac performance due to longterm exposure to simulated microgravity.

STEM Today, March 2016, No.6

Effects of Sex and Gender on Adaptation to Space: Cardiovascular Alterations Cardiovascular (CV) disease is the leading cause of death in women, with women developing CV disease about a decade later than their male peers. There are major differences in risk factors (e.g., smoking and diabetes have a more adverse impact on women). Clinically, women have greater CV morbidity and mortality, in part because they do not consistently receive optimal preventive strategies, diagnostic procedures, and treatments, although gender gaps continue to narrow. The sex and gender differences of effects of radiation exposure during spaceflight on cardiovascular risk factors and cardiovascular disease require ascertainment; the physiologic "aging" characteristics of spaceflight should be examined for their effect on both cardiovascular risk factors and cardiovascular disease.

Unique to women and their cardiovascular disease are hormonal concerns. Oral contraceptives designed to suppress menstruation and prevent pregnancy may increase blood pressure. There is no evidence that they increase risk of myocardial infarction, but they do increase the risk of venous thromboembolism. Differences with newer oral contraceptive formulations are unexplored (these newer agents likely will be used in future spaceflights). Menopausal hormone therapy, a new consideration as female astronauts age, has potential effects on autonomic blood pressure, volume status, and orthostatic tolerance, all of which must be investigated at baseline, during and after spaceflight. Little ground-based data are available regarding the effects of hormonal levels/oral contraception as it relates to space flight. Some studies state that no oral contraceptives were used, while other studies do not identify use or lack of use of oral contraceptives. Therefore, the lack of coherent study design among experiments to look at sex differences as they may relate to space flight presents a challenge. Not all studies of hormonal levels as they relate to space flight synchronized menstrual cycles. Exercise effects in women in simulated space studies show that exercise can improve orthostatic tolerance, preserve cardiac volume, and increase cardiac mass. Gender comparisons will be important as well as the gender effects of preconditioning prior to spaceflight.

During the past decade, little new research has been conducted in space to address cardiovascular differences related to sex and gender. Orthostatic tolerance is an important variable, with an increased prevalence of orthostatic intolerance in female compared with male astronauts, although few studies have targeted women (and even fewer studies involve minority women). Women have a greater loss of plasma volume than men following spaceflight, which mandates gender-based evaluation of countermeasures. There are known gender differences in response to cardiovascular stress, with women characteristically responding with an increase in heart rate and men with an increase in vascular resistance. It is uncertain whether this explains the male advantage for reentry orthostasis, but delineation of mechanisms should provide the rationale for countermeasures.

It is not known whether spaceflight increases the risk of arrhythmias, nor have sex or gender differences been studied. Baseline gender differences in supraventricular and ventricular arrhythmias and baseline gender differences in the electrocardiogram are likely relevant.

Cardiovascular alterations after simulated microgravity exposure, such as 6â&#x2014;Ś head-down bed rest (HDBR), appear to be similar between sexes. For example, it was found that men and women had similar cardiovascular responses to lower body negative pressure (LBNP) after short-duration HDBR compared with a control condition. Cardiac atrophy occurs in women similar to men following sedentary 60 days of HDBR. A similar reduction in blood volume of about 9% in men and women has been reported after 7 days of HDBR. However, Fortney et al. found less reduction of plasma volume in women than men ( - 10% versus - 15%) after 13 days of HDBR. Conversely, Vernikos et al. observed a higher decrease in plasma volume after 3 days of HDBR in women compared with men.

These inconsistent findings may be attributable to differences in the duration of HDBR or the time of inclusion in relation to the menstrual cycle in women, as well as the inherent noise in the measurement techniques. The lower orthostatic tolerance in women than men after HDBR cannot be modified by midodrine. Different from men, cardiovascular responses to exogenous nitric oxide (sublingual nitroglycerin) are not altered by HDBR in women. Prolonged bed rest may cause impairment of endotheliumdependent function at the microcirculation level in women.

The largest study to examine the responses of women to HDBR was the Womenâ&#x20AC;&#x2122;s International Space Simulation for Exploration (WISE)-2005 project that examined the effects of 60 days of HDBR in three groups of 8 women (control, exercise and nutrition). When the data were analyzed with respect to those who could com-

Page 45

Special Edition on Long Duration Spaceflight plete a 10-minute tilt after HDBR compared to those who could not, less arterial vasoconstriction and greater change in vein cross-sectional area were identified as key factors that might have contributed to poorer orthostatic tolerance. Finding some variability in orthostatic tolerance within a group is expected, as large differences between individuals have been identified in men and women, with a strong genetic component determined in studies with identical twins. In this latter study, the reduction in orthostatic tolerance after 28 days of HDBR was less with an LBNP-exercise routine similar to the WISE study (13%) than in control subjects (34%), and no differences were reported between men and women.

STEM Today, March 2016, No.6

Long-duration spaceflight may result in eccentric cardiac atrophy and impair cardiac compliance, leading to a prominent reduction in upright stroke volume and orthostatic tolerance in astronauts upon returning to the Earth. Female astronauts are more susceptible to orthostatic intolerance after spaceflight than male astronauts.

It has been proposed that low vascular resistance responses, a strong dependence on volume status, and/or a smaller stroke volume secondary to a smaller and less compliant left ventricle may be the underlying mechanisms.

After spaceflight, a greater reduction in plasma volume was reported in female astronauts. The change in lower extremity vein capacitance resulting from a loss of external fluid forces in the dehydrated extracellular compartment was proposed to be another potential mechanism associated with reentry orthostasis. This condition appears accentuated in women due to their inherent lower center of gravity and proportionately larger mass in the lower extremities. In both male and female astronauts, systemic peak oxygen uptake was well maintained during 9 to 14 days of spaceflight but was significantly reduced immediately on return to Earth, most likely because of reduced intravascular blood volume, stroke volume, and cardiac output. Up to now, there is no information available regarding sex differences in the degree of aerobic deconditioning after spaceflight in humans.

However, in response to simulated microgravity exposure, the relative changes in aerobic capacity are similar between sexes despite the marked differences in absolute values.

Effect of geomagnetic activity factors on the cardiac rhythm regulation and arterial pressure of cosmonauts Investigations of the effect of geomagnetic activity factors on the cardiac rhythm regulation and arterial pressure of cosmonauts during the expeditions onboard the Soyuz spacecraft, and the MIR and ISS orbital space stations was carried out for various durations of flight in weightlessness and, under control. Groups of cosmonauts were inspected under flight conditions outside the geomagnetic disturbances and in ground preflight conditions, during disturbances without them.

Based on the data of medical control of cosmonautsâ&#x20AC;&#x2122; cardiac rhythm changes at the 32nd orbit of the flight were analyzed for all crews of the Soyuz transporting vehicles (TV) for the period of 1986-1995. This orbit was the last before Soyuz TV docking with the orbital MIR station, and it was the orbit, on which cosmonautsâ&#x20AC;&#x2122; organism functions have already been relatively normalized after their activation at previous stages of flight. In total, 49 records were selected for analysis (for 49 crew members including the members of both main and visitation crews). A group of 8 cosmonauts was selected, who were in flight during all days of geomagnetic disturbances for the analyzed period. In addition, two control groups were selected: 9 cosmonauts whose flight has fallen on the time 3- 7 days apart from magnetic storms, and the second group included the remaining 32 cosmonauts .

Each record of a dynamic series of cardiointervals represented a text file of duration values of cardiointer- vals in milliseconds. The series length varied from 150- 200 values (2-3 min) up to 1000 values (15 min).

When analyzing the files the discrete-sliding method was used, where the samples with standard duration of 128 s were analyzed with a step of 10 s. After this all calculated indicators were averaged. 40 various indicators were calculated by the techniques described above, a part of which is shown in Table 2 with estimation of the confidence of distinctions by the Student criterion.

The obtained data testify that during the magnetic storm the following effects took place simultaneously: reduction of heart rate (HR) and displacement of the vegetative balance to the side of sympathetic link of regulation (decrease of HF %, CV, MxDMn, pNN50, and RMSSD, as well as increase of stress index SI and sympathetic link activity VLFs). The increase of indicators SNCA and LFs/HFs testifies that there were specific changes of vascular regulation. This is confirmed by growing LFt indicator. The mentioned changes can be interpreted physiologically as activation of a vasomotor (vascular) center and slowing of the time of reception and processing of information in it.

Some authors note that the most prominent deviations of physiological functions come in 24-48 hours after a magnetic storm and become apparent, most frequently, as increasing arterial pressure and arising vegetovascular distony. In this study authors have also observed more significant changes of cardiac rhythm variability indicators on the 1st-2nd day after the magnetic storm. These changes manifested themselves in essential

Page 46

Special Edition on Long Duration Spaceflight growth of the values of indicators 1C, VLF %,VLFs. LF decreases to a lower degree, but HF drops considerably. Attention is drawn to the fact of considerable increase of the indicator of activity of regulatory systems (IARS) and significant growth of the number of arrhythmias (Narr).

STEM Today, March 2016, No.6

Physiological interpretation of the effects, revealed during magnetic storms and, especially, in the next 24-48 hours, indicated that such effects took place for heart (appearance of arrhythmias) and vascular system (lengthening of vascular tone regulation time and a functional stress of the vasomotor center).

Investigations onboard the MIR Station The results of daily continuous monitoring of the Holter electrocardiogram were studied for two crew members of the MIR EI-22 expedition on the 176-179 days of flight (after 6 months, on February, 9-12, 1997). As evidenced by the experience of numerous past expeditions of duration up to 6 months, the functional state of cosmonauts in this period of flight was characterized by occurrence of the stage of relatively steady adaptation. The age of cosmonauts no. 1 and no. 2 was 44 and 37 years, respectively. Based on the Holter monitoring results, the time database was generated, which included the files of initial values of all cardiointervals for the day (70-100 thousand numerical values, approximately). For statistical processing, the data on 5-min intervals were used, which then were averaged over all days, over the periods of a day (day, evening, night), and over hour intervals. Figure 1 presents the diurnal dynamics of indicators of cardiac rhythm variability of cosmonauts during geomagnetic storms and under quiet conditions at the 6-th month of flight.

Attention should be paid to the changes of values of basic indicators in the control group (as compared to the one-month flight data). This is, first of all, a higher heartbeat rate and lowered cardiac rhythm variability, as well as redistribution of frequency components of the spectrum to the side of prevalence of slow second-order waves and increase of IC. All this indicates to activation and stress of regulatory mechanisms of a cardiovascular system after half-year stay in weightlessness conditions.

Against this background, under the magnetic storm effect the HR becomes confidently lower, and the cardiac rhythm variability (pNN50, RMSSD) grows. In this case, the relative and absolute power of vasomotor waves (LF %, LFs) confidently grows, and the slowwave components (VLFs) become stronger. The LFt indicator is confidently lower, which means the absence of the vascular center overstress. However, in general, the indicated changes suggest increased activity of regulatory systems (IARS). After the magnetic storm the increased total power of spectrum is conserved in all ranges, and the number of arrhythmias (NArr) considerably grows.

On the whole, it is seen from the presented results, that after six-month stay under weightlessness conditions the cardiac rhythm regulation system of cosmonauts under geomagnetically quiet conditions is already in the state of functional stress because of the long flight itself. The magnetic storm effect, as a new stressor, disturbs the vegetative balance, established to this time, and causes short-term overstress of regulatory mechanisms. In this case, the parasympathetic link of regulation strengthens.

Page 47

STEM Today, March 2016, No.6

Special Edition on Long Duration Spaceflight

Page 48

Special Edition on Long Duration Spaceflight

STEM Today, March 2016, No.6

Investigations of Cosmonauts’ Vascular Tone under Ground Conditions and during Space Flight according to the Data of 24-hour Monitoring of Arterial Pressure and Heartbeat Rate For checking the statistical confidence of the results obtained above, the 24-hour monitoring of arterial pressure (AP) and heartbeat rate (HR) were analyzed for a rather large group of cosmonauts: 24 ground preflight records and 12 records for the 4-6th months of flight were studied. Indicators were measured each 15 min during 24 h before and during the space flight. For statistical estimation of revealed effects the Statistica6 package (Student’s t-criterion) was used (Table 3).

The analysis of cosmonauts’ records distribution over the experimental groups has shown that various records of a single person fall into some groups. Preflight daily average values of arterial pressure varied from 106/75 to 122/85 mm Hg, and HR ranged from 55 to 79 beats per minute. The AP daily average values during stay in weightlessness conditions varied from 108/64 up to 126/83 mm Hg and the HR values from 57 to 76 beats per minute. For smoothing individual distinctions, the primary data were normalized for each person separately, so that the individual average values of indicators be equal to 0, and the individual standard deviations be equal to 1.

It is seen from Table 3 that the response of the cardiovascular system to the effect of geomagnetic disturbances exists both before and during the flight. In this case, the response under ground conditions and in weightlessness conditions is opposite. This can be due to the following causes. As a result of adaptation to weightlessness conditions, the cardiovascular system of a man passes to a new level of functioning.

Under ground conditions the response of the cardiovascular system to a geomagnetic disturbance is mainly characterized by changing vascular tone, namely, by its increase. This becomes apparent in reduction of the arterial pressure and heartbeat rate. Under weightlessness conditions at the 6th month of flight the state of cosmonauts is characterized by lower values of pulse and pressure as compared to the ground data on the background of stress of regulatory systems. Their increase under an effect of magnetic storms (Fig. 2) reflects strengthening of a nonspecific component of the adaptive stress-response.

There are essential shifts in the variability of cosmonauts’ cardiac rhythm during magnetic storms and after their termination, which reflects changes in the autonomous regulation of heart activity.

At the initial stage of a long-term space flight (at the second day of flight), under the magnetic storm effect, a general nonspecific stress-response of adaptive syndrome type was observed (increase of pulse rate, decrease of power of the spectrum of respiratory waves, etc.). The most prominent deviations of physiological functions appeared in 24-48 h after the magnetic storm. Immediately in the period of magnetic storm effect the specific response is observed, associated with regulation of a vascular tone (meteo-responses). At the instants of reconstruction of regulatory processes the heart work instability can arise with increasing number of arrhythmic contractions.

At the end of a half-year stay in space under the magnetic storm effect, destabilization of the functional stress state that arose to this time because of additional stressor effect of a magnetic storm is observed. The revealed effects are of principal significance for the concept of helio-geomagnetic factors as the time sensors of living organisms.

Page 49

STEM Today, March 2016, No.6

R EFERENCES R

Musgrave, F. Story; Zechman, Fred W.; and Mains, Richard C.: Changes in Total Leg Volume During Lower Body Negative Pressure. Aerospace Med., vol. 40, no. 7, Judy 1969, pp. 602-606.

R

Stevens, Paul M.: Cardiovascular Dynamics During Orthostasis and the influence of Intravascular Instrumentation. Am. J. Cardiol., vol. 17, Feb. 1966, pp.211.218.

R

Stevens, Paul M.; and Lamb, Lawrence E.: Effects of Lower Body Negative Pressure on the Cardiovascular System.

Am. J. Cardiol., vol. 16, Oct. 1965, pp. 506-515.

R

Herault S, Fomina G, Alferova I, Kotovskaya A, Poliakov V, Arbeille P. Cardiac, arterial and venous adaptation to weightlessness during 6-month MIR spaceflights with and without thigh cuffs (bracelets). Eur J Appl Physiol.

2000 Mar;81(5):384-90.

R

Alferova IV, Turchaninova VF, Golubchikova ZA, Liamin VR, Analysis and evaluation of the functional state of cardiovascular system in cosmonauts during long-term space flights, Fiziol Cheloveka, 2003 Nov - Dec;29(6):511.

R

Baevsky RM, Moser M, Nikulina GA, Polyakov VV, Funtova II, Chernikova AG.Autonomic regulation of circulation and cardiac contractility during a 14-month space flight. Acta Astronaut. 1998 Jan-Apr;42(1-8):159-73.

R

Biomedical Results of Apollo, NASA-SP-368 .

R

Biomedical Results of Skylab ,NASA-SP-377.

R

Grigoriev AI, Bugrov SA, Bogomolov VV, Egorov AD, Polyakov VV, Tarasov IK, Shulzhenko EB. Main medical results of extended flights on space station Mir in 1986-1990. Acta Astronaut. 1993 Aug;29(8):581-5.

R

Gundel A, Drescher J, Spatenko YA, Polyakov VV. Changes in basal heart rate in spaceflights up to 438 days.

Aviat Space Environ Med. 2002 Jan;73(1):17-21.

R

Chris May, Allen Borowski, David Martin, Zoran Popovic, Kazuaki Negishi, Jagir R. Hussan, Patrick Gladding, Peter Hunter, Ilana Iskovitz, Mohammed Kassemi, Michael Bungo, Benjamin Levine, James Thomas, AFFECT OF MICROGRAVITY ON CARDIAC SHAPE: COMPARISON OF PRE- AND IN-FLIGHT DATA TO MATHEMATICAL MODELING, Journal of the American College of Cardiology, Volume 63, Issue 12, Supplement, 1 April 2014, Page A1096, ISSN 0735-1097.

R

Hamilton DR, Sargsyan AE, Garcia K, Ebert DJ, Whitson PA, Feiveson AH, Alferova IV, Dulchavsky SA, Matveev VP, Bogomolov VV, Duncan JM. Cardiac and vascular responses to thigh cuffs and respiratory maneuvers on crewmembers of the International Space Station. J Appl Physiol (1985). 2012 Feb;112(3):454-62.

R

Zuj KA, Arbeille P, Shoemaker JK, Blaber AP, Greaves DK, Xu D, Hughson RL. Impaired cerebrovascular autoregulation and reduced CO2 reactivity after long duration spaceflight. Am J Physiol Heart Circ Physiol. 2012 Jun 15;302(12): H2592-8.

R

Arbeille P, Provost R, Zuj K, Vincent N. Measurements of jugular, portal, femoral, and calf vein cross-sectional area for the assessment of venous blood redistribution with long duration spaceflight (Vessel Imaging Experiment). Eur J Appl Physiol. 2015 Oct;115(10):2099-106.

STEM Today, March 2016, No.6

R

Respress JL, Gershovich PM, Wang T, Reynolds JO, Skapura DG, Sutton JP, Miyake CY, Wehrens XH. Long-term simulated microgravity causes cardiac RyR2 phosphorylation and arrhythmias in mice. Int J Cardiol. 2014 Oct 20;176(3):994-1000.

R

Belevych AE, Radwanski PB, Carnes CA, Györke S. ’Ryanopathy’: causes and manifestations of RyR2 dysfunction in heart failure. Cardiovasc Res. 2013 May 1;98(2):240-7.

R

Dobrev D, Wehrens XH. Role of RyR2 phosphorylation in heart failure and arrhythmias: Controversies around ryanodine receptor phosphorylation in cardiac disease. Circ Res. 2014 Apr 11;114(8):1311-9 .

R

Xu D, Shoemaker JK, Blaber AP, Arbeille P, Fraser K, Hughson RL. Reduced heart rate variability during sleep in long-duration spaceflight. Am J Physiol Regul Integr Comp Physiol. 2013 Jul 15;305(2):R164-70.

R

Platts SH, Bairey Merz CN, Barr Y, Fu Q, Gulati M, Hughson R, Levine BD, Mehran R, Stachenfeld N, Wenger NK. Effects of sex and gender on adaptation to space: cardiovascular alterations. J Womens Health (Larchmt).

2014 Nov;23(11):950-5.

R

Harvey PA, Leinwand LA. The cell biology of disease: cellular mechanisms of cardiomyopathy. J Cell Biol. 2011 Aug 8;194(3):355-65.

R

Breus, T. K. , Baevskii, R. M. , Funtova, I. I. , Nikulina, G. A. and Alexeev, E. V. , Chernikova, A. G. ,Effect of geomagnetic field disturbances on the adaptive stress reaction of cosmonauts , ,Cosmic Research , August 2008, Volume 46, Issue 4, pp 367-372.

R

Xu D, Shoemaker JK, Blaber AP, Arbeille P, Fraser K, Hughson RL. Reduced heart rate variability during sleep in long-duration spaceflight. Am J Physiol Regul Integr Comp Physiol. 2013 Jul 15;305(2):R164-70.

R

Moffitt JA, Henry MK, Welliver KC, Jepson AJ, Garnett ER. Hindlimb unloading results in increased predisposition to cardiac arrhythmias and alters left ventricular connexin 43 expression. Am J Physiol Regul Integr Comp Physiol. 2013 Mar 1;304(5):R362-73.

R

Nakashima T, Ohkusa T, Okamoto Y, Yoshida M, Lee JK, Mizukami Y, Yano M. Rapid electrical stimulation causes alterations in cardiac intercellular junction proteins of cardiomyocytes. Am J Physiol Heart Circ Physiol. 2014 May;306(9):H1324-33.

R

Harvey PA, Leinwand LA. The cell biology of disease: cellular mechanisms of cardiomyopathy. J Cell Biol. 2011 Aug 8;194(3):355-65.

R

Baker JE, Moulder JE, Hopewell JW. Radiation as a risk factor for cardiovascular disease. Antioxid Redox Signal.

2011 Oct 1;15(7):1945-56.

R

Arbeille P, Provost R, Vincent N, Aubert A. Adaptation of the main peripheral artery and vein to long term confinement (Mars 500). PLoS One. 2014 Jan 27;9(1):e83063.

R

S.M.C. Lee , M.B. Stenger , S.M. Smith , S.R. Zwart,S.S. Laurie, R.J. Ploutz-Snyder,S.H. Platts , Defining the Relationship Between Biomarkers of Oxidative and Inflammatory Stress and the Risk for Atherosclerosis in Astronauts During and After Long-duration Spaceflight (Cardio_Ox) ,NASA.

R

Zhang LF, Cheng JH, Liu X, Wang S, Liu Y, Lu HB, Ma J. Cardiovascular changes of conscious rats after simulated microgravity with and without daily -Gx gravitation. J Appl Physiol (1985). 2008 Oct;105(4):1134-45.

S

T

Science

Technology

E Engineering

Today

M Mathematics