Changes in organ perfusion and weight ratios in post-simulated microgravity recovery

Head-down tilt models have been used as ground-based simulations of microgravity. Our previous animal research has demonstrated that there are significant changes in fluid distribution within 2 h after placement in a 45 degrees head-down tilt (45HDT) position and these changes in fluid distribution were still present after 14 days of 45HDT. Consequently, we investigated changes in fluid distribution during recovery from 16 days of 45HDT. Changes in radioactive tracer distribution and organ/body weight ratio were examined in rats randomly assigned to a 45HDT or prone control group. The 45HDT rats were suspended for 16 days and then allowed to recover at the prone position 0, 77, 101, or 125 h post-suspension. Animals were injected with technetium-labeled diethylenetriamine pentaacetate (99mTcDTPA, MW=492 amu, physical half-life of 6.02 h) and then killed 30 min post-injection. Lungs, heart, liver, spleen, kidneys, and brain were harvested, weighed, and measured for radioactive counts. Statistical analyses included two-way analysis of variance (ANOVA) that compared 45HDT versus controls at the four experimental time points. The organ weight divided by the body weight ratio for the brain, heart, kidneys and liver in the 45HDT rats was significantly different than the control rats, regardless of time (treatment). There was no difference between the different time points (time). The average 99mTcDTPA count divided by the organ weight ratio values for the heart, liver, and spleen were significantly higher in the 45HDT group than the control group. The average counts for the heart and spleen were significantly higher at 77, 101, and 125 h than at time zero. We conclude that the major organs have different recovery patterns after 45HDT for 16 days in the rat.     

Early renal changes in 45 degrees HDT rats

Background: Both microgravity and simulated microgravity models, such as the 45HDT (45 degrees head-down tilt), cause a redistribution of body fluids indicating a possible adaptive process to the microgravity stressor. Understanding the physiological processes that occur in microgravity is a first step to developing countermeasures to stop its harmful effects, i.e., (edema, motion sickness) during long-term space flights. Hypothesis: Because of the kidneys' functional role in the regulation of fluid volume in the body, it plays a key role in the body's adaptation to microgravity. Methods: Rats were injected intramuscularly with a radioactive tracer and then lightly anesthetized in order to facilitate their placement in the 45HDT position. They were then placed in the 45HDT position using a specially designed ramp (45HDT group) or prone position (control group) for an experimental time period of 1 h. During this period, the 99mTc-DTPA (technetium-labeled diethylenepentaacetate, MW=492 amu, physical half-life of 6.02 h) radioactive tracer clearance rate was determined by measuring gamma counts per minute. The kidneys were then fixed and sectioned for electron microscopy. A point counting method was used to quantitate intracellular spaces of the kidney proximal tubules. Results: 45HDT animals show a significantly (p=0.0001) increased area in the interstitial space of the proximal tubules. Conclusions: There are significant changes in the kidneys during a 1 h exposure to a simulated microgravity environment that consist primarily of anatomical alterations in the kidney proximal tubules. The kidneys also appear to respond differently to the initial periods of head-down tilt.     

Effect on the cardiac function of repeated LBNP during a 1-month head down tilt

Cardiovascular assessment by ultrasound methods was performed during two long duration (1 month) Head Down Tilt (HDT) on 6 healthy volunteers. On a first 1 month HDT session, 3 of the 6 subjects (A, B, C) had daily several lower body negative pressure tests (LBNP), whereas the 3 subjects remaining (D, E, F) rested without LBNP. On a second 1 month HDT session subjects D, E, and F had daily LBNP tests and the A, B and C subjects did not. The cardiac function was assessed by Echocardiography, (B mode, TM mode). On all the "6 non LBNP" subjects the left ventricule diastolic volume (LVDV), the stroke volume (SV) and the cardiac output (CO) increase (+10%, -15%) after HDT then decrease and remain inferior (-5%, -5%) or equal to the basal value during the HDT. Immediately after the end of the HDT the heart rate (HR) increase (+10%, +30%) whereas the cardiac parameters decrease weakly (-5%, -10%) and normalize after 3 days of recovery. On the "6 LBNP" subjects the LVDV, SV and CO increase (+10%, 15%) after 1 h HDT as in the previous group then decrease but remain superior (+5%, +15%) or equal to the basal value. After the HDT session, the HR is markedly increased (+20%, +40%) the LVDV and SV decrease (-15%, -20%) whereas the CO increases or decreases depending on the amplitude of the HR variations. These parameters do not completely normalize after 3 day's recovery. Repeated LBNP sessions have a significant effect on the cardiovascular function as it maintains all cardiac parameters above the basal value. The LBNP manoeuvre can be considered as an efficient countermeasure to prevent cardiac disadaptation induced by HDT position and probably microgravity.     

Effects of 12 days exposure to simulated microgravity on central circulatory hemodynamics in the rhesus monkey

Central circulatory hemodynamic responses were measured before and during the initial 9 days of a 12-day 10 degrees head-down tilt (HDT) in 4 flight-sized juvenile rhesus monkeys who were surgically instrumented with a variety of intrathoracic catheters and blood flow sensors to assess the effects of simulated microgravity on central circulatory hemodynamics. Each subject underwent measurements of aortic and left ventricular pressures, and aortic flow before and during HDT as well as during a passive head-up postural test before and after HDT. Heart rate, stroke volume, cardiac output, and left ventricular end-diastolic pressure were measured, and dP/dt and left ventricular elastance was calculated from hemodynamic measurements. The postural test consisted of 5 min of supine baseline control followed by 5 minutes of 90 degrees upright tilt (HUT). Heart rate, stroke volume, cardiac output, and left ventricular end-diastolic pressure showed no consistent alterations during HDT. Left ventricular elastance was reduced in all animals throughout HDT, indicating that cardiac compliance was increased. HDT did not consistently alter left ventricular +dP/dt, indicating no change in cardiac contractility. Heart rate during the post-HDT HUT postural test was elevated compared to pre-HDT while post-HDT cardiac output was decreased by 52% as a result of a 54% reduction in stroke volume throughout HUT. Results from this study using an instrumented rhesus monkey suggest that exposure to microgravity may increase ventricular compliance without alternating cardiac contractility. Our project supported the notion that an invasively-instrumented animal model should be viable for use in spaceflight cardiovascular experiments to assess potential changes in myocardial function and cardiac compliance.     

Increased urinary excretion rates of serotonin and metabolites during bedrest

Astronauts are often on a voluntarily reduced energy intake during space missions, possibly caused by a metabolic or emotional stress response with involvement of the central serotonergic system (SES). We investigated 24 h urinary excretion (24 h-E) of serotonin (5-HT) and 5-hydroxyindol acidic acid as indicators of the SES in healthy males under two different normocaloric conditions: normal physical activity (NPA) and -6 degree head-down-tilt (HDT). HDT or NPA were randomly arranged with a recovery period of 6 months in between. 24 h-E of hormones varied widely among individuals. Values were higher in HDT compared to NPA. Assuming that the 24 h-E values are, beside being indicators for alterations in the number and metabolism of platelets. Also indicators of central SES, HDT condition seems to activate central SES in a higher degree compared to NPA. Therefore, changes in central SES might be involved in the mechanisms associated with space flight or microgravity, including possible maladaptations such as voluntary undernutrition.     

Immobilization induces a very rapid increase in osteoclast activity

We studied in a randomized, strictly controlled cross-over design, the effects of 6 days 6 degrees head-down tilt bed rest (HDT) in eight male healthy subjects in our metabolic ward. The study consisted of two periods (phases) of 11 days each in order to allow for the test subjects being their own controls. Both study phases were identical with respect to environmental conditions, study protocol and diet. Two days before arriving in the metabolic ward the subjects started with a diet. The diet was continued in the metabolic ward. The metabolic ward period (1l days) was divided into three parts: 4 ambulatory days, 6 days either HDT or control and 1 recovery day. Continuous urine collection started on the first day in the metabolic ward to analyze calcium excretion and bone resorption markers. On the 2nd ambulatory day in the metabolic ward and on the 5th day in HDT or control blood was drawn to analyze serum calcium, parathyroid hormone, and bone formation markers. Urinary calcium excretion was, as early as the first day in immobilization, increased (p<0.01). CTX- and NTX-excretion stayed unchanged in the first 24 h in HDT compared to the control. But already on the 2nd day of immobilization, both bone resorption markers significantly increased. We conclude from these results--pronounced rise of bone resorption markers--that already 24 h of immobilization induce a significant rise in osteoclast activity in healthy subjects. Thus, it appears possible to use short-term bed rest studies as a first step for the development of countermeasures to immobilization.     

Efficacy of periodic centrifugation of primates during 4-week head-down tilt.

Creation of artificial force of gravity (AFG) to counteract the negative consequences of microgravity in manned space missions of extended duration is one of the high-priority problems of space biology and medicine. However, there are a number of especial effects of AFG (namely, structural changes in muscles and bones, and some other system) which need implantation of electrodes and sensors and are possible only with animals. That is why it is of particular interest to make studies with monkeys whose reactions to changed gravity bear much resemblance with human. The purpose of the investigation was development of a protocol of periodic gravity loads as a counter-measure against the hypokinetic syndrome in Macaca mulatta. Two series of experiments were performed. In the series, animals were split into two groups of 6 species each who were motor restrained with the head end tilted downward at 5 degrees (HDT) for 28 days. Monkeys of group-2 were periodically subjected to centrifugation (HDT+G). During the first series of experiments rotation was conducted in the +Gz direction at g-loads from 1.2 to 1.6 units for 30-40 minutes 4-5 times a week. In the second series, g-load was equal to 1.2 units and the animals were rotated 30 min. 2-3 time a week. The criterion of Y-training protocol efficacy was a test +Gz run at 3 units for 30 s. during which functioning of the cardiovascular systems and its controls was evaluated. The test run was performed prior to and after HDT. Following HDT the animals of group HDT+G were more resistant to the test than their counterparts who had not been trained on the centrifuge. Data of the investigation imply that following HDT and HDT+G alike reduced the amount of total bodily fluids (by approximately 5%), the intracellular component (approximately 4%), and plasma volume (by 6-7%). Yet, there are radical differences between the groups in the levels of reduction in extracellular fluids (by 11% and 6.5%, respectively, P<0.05) and the interstitial component (by 11.5% and 6.5, respectively, P<0.05). Prophylactic centrifugation during HDT was also positive to the muscular blood flow in lower extremities.     

Human thermohomeostasis onboard "Mir" and in simulated microgravity studies.

Significant changes of thermogomeostatic parameters was obtained by thermotopometric method using the techniques simulate of microgravity effects: bed rest, pressurized isolation, suit immersion (SI). However, each of ground models made rectal temperature (T) trend downward. The autothermometric study (24 and 12 sessions, 2-13th and 6-174th flight days) was carried out onboard "Mir" by two flight engineers who had preliminary tested at SI (1-2 days). Studies of German investigators onboard "Mir" confirmed: rectal T must be higher in space flight as compared to the normal environment (n=4). Comparative studies suggest that microgravity is a key factor for the human body surface T raise and abolishment of the external/internal T-gradient. T-homeostasis was not really changing during missions and could be regarded as acute effect of microgravity. After delineation of changes in body surface T--by Carnot's thermodynamic law--rectal T raise should have been anticipated. Facts pointing to the excess entropy of human body must not be passed over.     

Choroidal responses in microgravity. (SLS-1, SLS-2 and hindlimb-suspension experiments)

Fluid and electrolyte shifts occuring during human spaceflight have been reported and investigated at the level of blood, cardio-vascular and renal responses. Very few data were available concerning the cerebral fluid and electrolyte adaptation to microgravity, even in animal models. It is the reason why we developed several studies focused on the effects of spaceflight (SLS-1 and SLS-2 programs, carried on NASA STS 40 and 56 missions, which were 9- and 14-day flights, respectively), on structural and functional features of choroid plexuses, organs which secrete 70–90 % of cerebrospinal fluid (CSF) and which are involved in brain homeostasis. Rats flown aboard space shuttles were sacrificed either in space (SLS-2 experiment, on flight day 13) or 4–8 hours after landing (SLS-1 and SLS-2 experiments). Quantitative autoradiography performed by microdensitometry and image analysis, showed that lateral and third ventricle choroid plexuses from rats flown for SLS-1 experiment demonstrated an increased number (about x 2) of binding sites to natriuretic peptides (which are known to be involved in mechanisms regulating CSF production). Using electron microscopy and immunocytochemistry, we studied the cellular response of choroid plexuses, which produce cerebrospinal fluid (CSF) in brain lateral, third and fourth ventricles. We demonstrated that spaceflight (SLS-2 experiment, inflight samples) induces changes in the choroidal cell structure (apical microvilli, kinocilia organization, vesicle accumulation) and protein distribution or expression (carbonic anhydrase II, water channels,…). These observations suggested a loss of choroidal cell polarity and a decrease in CSF secretion. Hindlimb-suspended rats displayed similar choroidal changes. All together, these results support the hypothesis of a modified CSF production in rats during long-term (9, 13 or 14 days) adaptations to microgravity.     

Dynamic change of ERPs related to selective attention to signals from left and right visual field during head-down tilt

To study further the effect of head-down tilt (HDT) on slow positive potential in the event-related potentials (ERPs), the temporal and spatial features of visual ERPs changes during 2 hour HDT (-10 degrees) were compared with that during HUT (+20 degrees) in 15 normal subjects. The stimuli were consisted of two color LED flashes appeared randomly in left or right visual field (LVF or RVF) with same probability. The subjects were asked to make switch response to target signals (T) differentially: switching to left for T in LVF and to right for T in RVF, ignoring non-target signals(N). Five sets of tests were made during HUT and HDT. ERPs were obtained from 9 locations on scalp. The mean value of the ERPs in the period from 0.32-0.55 s was taken as the amplitude of slow positive potential(P400). The main results were as follows. 1) The mean amplitude of P400 decreased during HDT which was more significant at the 2nd, 3rd and 5th set of tests; 2) spatially, the reduction of mean P400 amplitude during HDT was more significant for signals from RVF and was more significant at posterior and central brain regions than that on frontal locations. As that the positive potential probably reflects the active inhibition activity in the brain during attention process, these data provide further evidence showing that the higher brain function was affected by the simulated weightlessness and that this effect was not only transient but also with interesting spatial characteristics.     

The effect of head-down tilt on brain potentials related to visual attention

To study the possible effect of simulated weightlessness on brain function state, the brain event-related potentials (ERPs) in a simple visual selective response task were compared between HDT and HUT in 9 normal subjects. The results were: The Target(T) and non-Target(NT) flash signals both induced significant slow positive potentials which were supposed to related to the attention state; the amplitude of the positive potentials in frontal regions decreased significantly especially for NT-ERPs during HDT comprared with that during HUT. The data reported provide new evidence indicating that more attention should be paid on the brain function study during space flight.     

Impaired T-wave amplitude adaptation to heart-rate induced by cardiac deconditioning after 5-days of head-down bed-rest

The study of QT/RR relationship is important for the clinical evaluation of possible risk of acquired or congenital ventricular tachyarrhythmias. In the hypothesis that microgravity exposure could induce changes in the repolarization mechanisms, our aim was to test if a short 5-days strict 6° head-down bed-rest (HDBR) could induce alterations in the QT/RR relationship and spatial repolarization heterogeneity. Twenty-two healthy men (mean age 31±6) were enrolled as part of the European Space Agency HDBR studies. High fidelity (1000 Hz) 24 h Holter ECG (12-leads, Mortara Instrument) was acquired before (PRE), the last day of HDBR (HDT5), and four days after its conclusion (POST). The night period (23:00–06:30) was selected for analysis. X, Y, Z leads were derived and the vectorcardiogram computed. Selective beat averaging was used to obtain averages of P–QRS–T complexes preceded by the same RR (10 ms bin amplitude, in the range 900–1200 ms). For each averaged waveform (i.e., one for each bin), T-wave maximum amplitude (Tmax), T-wave area (Tarea), RTapex, RTend, ventricular gradient (VG) magnitude and spatial QRS-T angle were computed. Non-parametric Friedman test was applied. Compared to PRE, at HDT5 both RTapex and RTend resulted shortened (−4%), with a decrease in T-wave amplitude (−8%) and area (−13%). VG was diminished by 10%, and QRS-T angle increased by 14°. At POST, QT duration and area parameters, as well as QRS-T angle were restored while Tmax resulted larger than PRE (+5%) and VG was still decreased by 3%. Also, a marked loss in strength of the linear regression with RR was found at HDT5 in Tmax and Tarea, that could represent a new dynamic marker of increased risk for life-threatening arrhythmias. Despite the short-term HDBR, ventricular repolarization during the night period was affected. This should be taken into account in astronauts for risk assessment during space flight.     

Electrolytes in femur and plasma of rats during prolonged restriction of motor activity.

Hypokinesia (decreased motor activity) induces insignificant bone mineral changes. The aim of this study was to measure mineralization, density, and also electrolyte content in the femur of rats during prolonged hypokinesia (HK). Studies were done on 144 male Wistar rats (370-390 g) during 15 days period of pre-HK and 90 days period of HK. Rats were equally divided into two groups: hypokinetic rats (HKR) and vivarium control (VCR). The HKR group of rats was kept in small individual cages. Femur mineralization density, ash mineral content, calcium (Ca) and phosphate (P) content, and plasma Ca and P concentration were measured. In the HKR group body weight, femur mineralization, density, ash mineral content, Ca and P concentration decreased significantly (p < or = 0.01) while plasma Ca and P concentration increased significantly (p < or = 0.01) when compared with the VCR group. The measured parameters did not change significantly in the VCR group when compared with the baseline control values. It was concluded that prolonged HK induces a significant reduction in electrolyte concentration accompanied by decreased mineralization, density, and ash mineral content of the femur of rats.     

The human cardiovascular system during space flight

Purpose of the work is to analyze and to summarize the data of investigations into human hemodynamics performed over 20 years aboard orbital stations Salyut-7 and Mir with participation of 26 cosmonauts on space flights (SF) from 8 to 438 days in duration. The ultrasonic techniques and occlusive plethysmography demonstrated dynamics of changes in the cardiovascular system during SF of various durations. The parameters of general hemodynamics, the pumping function of the heart and arterial circulation in the brain remained stable in all the space flights; however, there were alterations in peripheral circulation associated with blood redistribution and hypovolemie in microgravity. The anti-gravity distribution of the vascular tone decayed gradually as unneeded. The most considerable changes were observed in leg vessels, equally in arteries (decrease in resistance) and veins (increase in maximum capacity). The lower body negative pressure test (LBNP) revealed deterioration of the gravity-dependent reactions that changed for the worse as SF duration extended. The cardiovascular deconditioning showed itself as loss of descent acceleration tolerance and orthostatic instability in the postflight period.     

Central and regional hemodynamics in prolonged space flights.

This paper presents the results of measuring central and regional (head, forearm, calf) hemodynamics at rest and during provocative tests by the method of tetrapolar rheography in the course of Salyut-6-Soyuz and Salyut-7-Soyuz missions. The measurements were carried out during short-term (19 man-flights of 7 days in duration) and long-term (21 man-flights of 65-237 days in duration) manned missions. At rest, stroke volume (SV) and cardiac output (CO) as well as heart rate (HR) decreased insignificantly (in short-term flights) or remained essentially unchanged (in long-term flights). In prolonged flights CO increased significantly in response to exercise tests due to an increase in HR and the lack of changes in SV. After exercise tests SV and CO decreased as compared to the preflight level. During lower body negative pressure (LBNP) tests HR and CO were slightly higher than preflight. Changes in regional hemodynamics included a distinct decrease of pulse blood filling (PBF) of the calf, a reduction of the tone of large vessels of the calf and small vessels of the forearm. Head examination (in the region of the internal carotid artery) showed a decrease of PBF of the left hemisphere (during flight months 2-8) and a distinct decline of the tone of small vessels, mainly, in the right hemisphere. During LBNP tests the tone of pre- and postcapillary vessels of the brain returned to normal while PBF of the right and left hemisphere vessels declined. It has been shown that regional circulation variations depend on the area examined and are induced by a rearrangement of total hemodynamics of the human body in microgravity. This paper reviews the data concerning changes in central and regional circulation of men in space flights of different duration.     

Severe disruption of the cytoskeleton and immunologically relevant surface molecules in a human macrophageal cell line in microgravity—Results of an in vitro experiment on board of the Shenzhou-8 space mission

During spaceflight the immune system is one of the most affected systems of the human body. During the SIMBOX (Science in Microgravity Box) mission on Shenzhou-8, we investigated microgravity-associated long-term alterations in macrophageal cells, the most important effector cells of the immune system. We analyzed the effect of long-term microgravity on the cytoskeleton and immunologically relevant surface molecules. Human U937 cells were differentiated into a macrophageal phenotype and exposed to microgravity or 1g on a reference centrifuge on-orbit for 5 days. After on-orbit fixation, the samples were analyzed with immunocytochemical staining and confocal microscopy after landing. The unmanned Shenzhou-8 spacecraft was launched on board a Long March 2F (CZ-2F) rocket from the Jiuquan Satellite Launch Center (JSLC) and landed after a 17-day-mission. We found a severely disturbed actin cytoskeleton, disorganized tubulin and distinctly reduced expression of CD18, CD36 and MHC-II after the 5 days in microgravity. The disturbed cytoskeleton, the loss of surface receptors for bacteria recognition, the activation of T lymphocytes, the loss of an important scavenger receptor and of antigen-presenting molecules could represent a dysfunctional macrophage phenotype. This phenotype in microgravity would be not capable of migrating or recognizing and attacking pathogens, and it would no longer activate the specific immune system, which could be investigated in functional assays. Obviously, the results have to be interpreted with caution as the model system has some limitations and due to numerous technical and biological restrictions (e.g. 23 °C and no CO₂ supply during in-flight incubation). All parameter were carefully pre-tested on ground. Therefore, the experiment could be adapted to the experimental conditions available on Shenzhou-8.     

Effect of a venotonic agent on the main arteries and veins during a 5 day HDT.

Objective: To evaluate the cardiac, arterial and venous effect of a venotonic drug (Cirkan "Ck") administrated orally daily to 6 subjects in HDT position during 5 days. These subjects underwent a second 5d HDT without Ck treatment one month later. Pre and post HDT the subjects were submitted to a stand test. Method: The cardiovascular parameters were measured by echography and Doppler. Results and discussion: The Cirkan treatment contributed to reduce the increase in cerebral resistance, and to maintain the lower limb resistance at a higher level than on controls. It reduces the vein section at the extremities (Jugular, femoral) and in the central vein system connected to the right heart (sub hepatics). On the other hand it increases the portal vein section which means that the blood stagnation at the splanchnic level is increased. Despite these arterial and venous significant modifications the clinical and ECG and Blood pressure response to the Stand test was similar in both groups.     

Executive function on the 16-day of bed rest in young healthy men

Microgravity due to prolonged bed rest may cause changes in cerebral circulation, which is related to brain function. We evaluate the effect of simulated microgravity due to a 6° head-down tilt bed rest experiment on executive function among 12 healthy young men. Four kinds of psychoneurological tests—the table tapping test, the trail making test, the pointing test and losing at rock–paper–scissors—were performed on the baseline and on day 16 of the experiment. There was no significant difference in the results between the baseline and day 16 on all tests, which indicated that executive function was not impaired by the 16-day 6° head-down tilting bed rest. However, we cannot conclude that microgravity did not affect executive function because of the possible contribution of the following factors: (1) the timing of tests, (2) the learning effect, or (3) changes in psychophysiology that were too small to affect higher brain function.