An international collaboration studying the physiological and anatomical cerebral effects of carbon dioxide during head-down tilt bed rest: the SPACECOT study
Abstract
Exposure to the microgravity environment results in various adaptive and maladaptive physiological changes in the human body, with notable ophthalmic abnormalities developing during 6-mo missions on the International Space Station (ISS). These findings have led to the hypothesis that the loss of gravity induces a cephalad fluid shift, decreased cerebral venous outflow, and increased intracranial pressure, which may be further exacerbated by increased ambient carbon dioxide (CO2) levels on the ISS. Here we describe the SPACECOT study (studying the physiological and anatomical cerebral effects of CO2 during head-down tilt), a randomized, double-blind crossover design study with two conditions: 29 h of 12° head-down tilt (HDT) with ambient air and 29 h of 12° HDT with 0.5% CO2. The internationally collaborative SPACECOT study utilized an innovative approach to study the effects of headward fluid shifting induced by 12° HDT and increased ambient CO2 as well as their interaction with a focus on cerebral and ocular anatomy and physiology. Here we provide an in-depth overview of this new approach including the subjects, study design, and implementation, as well as the standardization plan for nutritional intake, environmental parameters, and bed rest procedures.
NEW & NOTEWORTHY A new approach for investigating the combined effects of cephalad fluid shifting and increased ambient carbon dioxide (CO2) is presented. This may be useful for studying the neuroophthalmic and cerebral effects of spaceflight where cephalad fluid shifts occur in an elevated CO2 environment.
exposure to microgravity results in multiple physiological and anatomical changes, some of which may alter cerebral and ocular structure and function. Currently, more than one-half of astronauts present with ophthalmological changes during 6-mo missions on the International Space Station (ISS). The National Aeronautics and Space Administration (NASA) has identified this as a high-priority risk to human health in space and named it spaceflight-induced intracranial hypertension/vision alterations, also referred to as the visual impairment and intracranial pressure (VIIP) syndrome (1) or microgravity ocular syndrome (8). The findings include swelling of the optic nerve head, increased optic nerve sheath diameter, choroidal folds, cotton wool spots, flattening of the posterior eye globe, and hyperopic shifts up to 3 diopters (5, 7). Although the etiology of these ocular changes is currently unknown, there are similarities to patients with idiopathic intracranial hypertension. Several astronauts with papilledema also had mildly elevated intracranial pressure (ICP) measured postflight through lumbar puncture (7). These findings have prompted the hypothesis that ICP is increased during spaceflight secondary to a microgravity-induced headward fluid shift and decreased cerebral venous outflow due to the loss of physiological hydrostatic pressure gradients in space.
Because the ISS is a closed environment, ambient carbon dioxide (CO2) levels average 0.5%, i.e., approximately 10 times higher than terrestrial levels (0.04%) (6). However, with no natural convection currents in microgravity, large fluctuations in atmospheric CO2 occur over hours and days, and pockets of high CO2 concentrations may form in areas of low ventilation (14). As a potent arterial vasodilator, CO2 can evoke manifold physiological responses. Notably, hypercapnia increases cerebral blood flow (9) and may lead to acutely increased intracranial volume and thus elevated ICP. Therefore, exposure to increased ambient CO2 may exacerbate the hypothesized microgravity-induced increase in ICP during spaceflight (6). For decades, head-down tilt (HDT) bed rest has been used as a ground-based microgravity analog for physiological research (10a, 20). However, HDT bed rest studies performed in the 6° HDT position with ambient air do not reproduce most of the cerebral or ophthalmic findings seen in astronauts (17, 18). Conversely, short-duration exposure to steeper HDT angles have demonstrated decreased cerebral venous outflow (9) and increased ICP (4, 11), consistent with hypothesized contributing factors to microgravity-induced ocular changes. Here, we present a novel ground-based approach to study two of the major hypothesized contributing factors to ocular changes in space: headward fluid shifting and elevated ambient CO2, as well as their possible interaction. Because this study was the first to implement this new approach to HDT bed rest, vital signs, blood parameters, and standardization techniques are presented as standard measures.
Overall, our aim was to perform a comprehensive evaluation of the cerebral, ocular, and cardiopulmonary physiological effects of HDT with and without elevated ambient CO2. As a secondary aim, we evaluated the effects of short-duration (3 h) exposure to 3% CO2 as a simulation of CO2 accumulation that may occur on the ISS. The SPACECOT Study (studying the physiological and anatomical cerebral effects of CO2 during head-down tilt) was an international collaboration of more than 10 institutions and, to our knowledge, the first study to investigate the combined effects of HDT and increased ambient CO2 for >24 h. This approach was accompanied by a suite of both gold-standard and innovative noninvasive monitoring technologies to provide a comprehensive understanding of the HDT + CO2 challenge on whole body physiology. In this overview paper, we focus on the framework of our new approach and present baseline subject characteristics and general results. Specific results related to the various procedures and physiological systems will be presented in subsequent papers.
MATERIALS AND METHODS
The SPACECOT study was performed at the : envihab facility at the German Aerospace Center in Cologne, Germany. The :envihab is a state-of-the-art environmental medicine research facility that allows for full environmental and atmospheric conditioning of the bed rest facility, which consists of 12 bedrooms, a common area, bed rest compatible showers, bathrooms, a metabolic kitchen, laboratories, and a control room. The study procedure was approved by the local ethical commission of the regional medical association (Ärztekammer Nordrhein) as well as the Baylor College of Medicine Institutional Review Board and was registered at ClinicalTrials.gov (identifier number: NCT02493985). A test run of the study schedule and procedures was conducted in May 2015, and the full study was performed in June to July of 2015. All of the subjects provided written informed consent at the German Aerospace Center in Cologne, Germany.
Subjects.
The SPACECOT study included six healthy male subjects (means ± SD: age, 41 ± 5 yr; weight, 82 ± 7 kg; height, 177 ± 4 cm). Baseline subject demographics and anthropometrics are shown in Table 1. Body surface area (BSA) was calculated from the classical equation of Du Bois and Du Bois (3) as follows: BSA = 0.007184 × W0.425 × H0.725, where W = weight of subject in kilograms and H = height of subject in centimeters. Subject inclusion criteria consisted of healthy male subjects, ages 30–55 yr, body mass index of 20–26 kg/m2, nonsmoking for at least 6 mo, no current prescription medication use, and with an exercise capacity as well as cardiovascular and mental health that approximately matches the astronaut corps. Prior to inclusion in the study, all subjects underwent a comprehensive medical screening. Subject exclusion criteria were as follows: V̇o2max outside of the range of 30–60 ml·kg−1·min−1, history of migraines, history of ophthalmological conditions (including glaucoma, retinopathy, and cataracts), history of chronic back pain, increased risk of thrombosis, history of kidney stones or kidney disorders, and any neurological, cardiovascular, or psychiatric conditions. Subjects also underwent psychological evaluations, including the big five inventory and the Freiburger Persönlichkeitsinventar. The recruitment process also included 1 h in the 12° HDT position, including 10 min in a magnetic resonance imaging (MRI) scanner while in the HDT position with a face mask, to test eligibility with these conditions, as well as ultrasound screening of ophthalmic arteries to ensure suitability for noninvasive ICP monitoring (Vittamed, Kaunas, Lithuania).
| Subject | Age, yr | Weight, kg | Height, cm | BMI, kg/m2 | BSA, m2 | Thorax Vol, cm3 | Abdomen Vol, cm3 | Thigh Vol, cm3 | Calf Vol, cm3 | Total Vol, cm3 |
|---|---|---|---|---|---|---|---|---|---|---|
| 1 | 47 | 82.5 | 174 | 27.2 | 1.97 | 18,911 | 9,937 | 2,707 | 1,579 | 27,483 |
| 2 | 43 | 91.4 | 183 | 27.4 | 2.13 | 21,846 | 12,999 | 7,422 | 2,259 | 41,208 |
| 3 | 39 | 68.8 | 175 | 22.6 | 1.83 | 12,863 | 5,856 | 3,648 | 1,423 | 23,006 |
| 4 | 33 | 80.1 | 178 | 25.4 | 1.98 | 13,331 | 6,555 | 5,021 | 1,771 | 26,915 |
| 5 | 39 | 85.0 | 180 | 26.2 | 2.05 | 16,550 | 7,698 | 3,793 | 1,926 | 27,987 |
| 6 | 43 | 84.5 | 173 | 28.2 | 1.99 | 20,477 | 10,493 | 3,778 | 1,850 | 31,733 |
| Means ± SD | 41 ±5 | 82 ±7 | 177 ± 4 | 26.2 ± 2 | 1.99 ± 0.1 | 17,330 ± 3,724 | 8,923 ± 2,706 | 4,395 ± 1,656 | 1,801 ± 290 | 29,722 ± 6,276 |
Study design.
The SPACECOT study implemented a randomized, double-blind crossover design with two conditions: 26.5 h of 12° HDT with ambient atmosphere and 26.5 h of 12° HDT with 0.5% CO2 atmosphere (Fig. 1). At the end of each condition (while still in the 12° HDT position), subjects were also exposed to a 3% CO2 atmosphere for 2.5 h through a mask and tank system, to determine the effects of short-term exposure to high CO2 concentrations, similar to what astronauts may experience in areas with poor ventilation on the ISS, leading to a full 29 h of HDT bed rest. The two HDT conditions were preceded by a baseline data collection day that exhibited identical study procedures. The only exception was that, before the first baseline condition, subjects came in 1 day prior for a familiarization session with the facility, staff, techniques, and equipment involved in the study. All baseline data were collected in the seated, upright position. Exceptions include baseline flow-mediated dilation ultrasound and MRI-derived measures, which were collected in the supine position, and several other parameters took baseline measures in both the upright and supine positions. Between each HDT condition, there was a washout period of 1 week to avoid carryover effects. The six subjects were tested in two groups of three, with each group experiencing a different atmospheric conditioning order. Notably, both the investigators and the subjects were blinded to the atmospheric conditions during the two conditions and were unmasked after data analyses were completed. In addition, : envihab technicians and engineers who controlled and monitored atmospheric conditions offsite were not scientifically involved in the study and operated independently from study investigators.
Fig. 1.The SPACECOT study was a randomized, double-blind crossover design study with two conditions: 29 h of 12° head-down tilt (HDT) with ambient air and 29 h of 12° HDT with 0.5% CO2. Before each condition, subjects underwent 1 day of baseline data collection (BDC) in the seated, upright position.
All study procedures were in accordance with the standardization plan issued by the European Space Agency and/or the International Academy of Astronautics Study Group (15, 16). During the full 29 h of bed rest, all subjects strictly adhered to the 12° HDT position including during all daily activities such as eating, showering, and going to the bathroom. Subjects were required to keep a daily log of critical incidents and were not allowed a pillow of any kind during the HDT phases of the study, as this would cause deviations from the full body HDT position. Further, subjects were allowed to lie on their side for short durations; however, one shoulder remained in contact with the mattress at all times (sitting or standing not permitted), and the 12° HDT angle was maintained. Each subject was accommodated in a single-person room, and the daily schedule consisted of 6:30 AM wake up and 10:30 PM lights out. Light intensity (100 lux) and spectrum, enriched in green (~540 nm) wavelengths to entrain the circadian system, were standardized and periodically verified with an external spectral irradiance meter in the subject bedrooms and measurement rooms. To track their activity during the nighttime as an indicator of sleep quality, the subjects wore an Actiwatch activity monitor (Actigraph, Pensacola, FL) on the wrist of the nondominant arm.
Nutrition.
All subjects received a strictly controlled diet, for the entire duration of the study, tailored to individual resting metabolic rates (RMRs; Table 2). The RMR was estimated based on body weight and age from equations provided in the Human Energy Requirements Report by the United States Food and Agriculture Organization/World Health Organization/United Nations University Expert Consultation (21). Then, for each individual, the daily energy intake during stationary phases was set to 1.5 × RMR (corresponding to light intensity daily physical activities), whereas during HDT the daily energy content was set to 1.3 × RMR (corresponding to the energy need of a sedentary adult). The carbohydrate content of the diet was set to 50–55% and dietary fat intake was set to 30–35%, in line with international bed rest standards (15). The selected food items corresponded to a standard German diet. Fluid intake was regulated and set to 42 ml·kg body wt−1·day−1 and, apart from the fluid portion contained in the meals, was provided in the form of water and diluted apple juice. Intake of caffeine, alcohol, and chocolate was prohibited. All meals were planned and supervised by registered dieticians and prepared in the : envihab metabolic kitchen. Nutrient content was calculated using PRODI software (Kluthe Prodi 6.3 expert; Nutri-science, Germany).
| BDC | 12° HDT (1st Day) | 12° HDT (2nd Day) | |
|---|---|---|---|
| Energy, kcal/day | 2,649 ± 133 | 2,287 ± 114 | 2,656 ± 143 |
| Protein, g/day | 98 ± 9 | 97 ± 8 | 97 ± 9 |
| Protein, g·kg body wt−1·day−1 | 1.21 ± 0.00 | 1.20 ± 0.01 | 1.20 ± 0.01 |
| Protein, %TEE/day | 15 ± 1 | 17 ± 1 | 15 ± 1 |
| Fat, g/day | 95 ± 5 | 83 ± 4 | 95 ± 5 |
| Fat, %TEE/day | 33 ± 0 | 34 ± 0 | 33 ± 0 |
| Carbohydrates, g/day | 333 ± 13 | 274 ± 12 | 336 ± 15 |
| Carbohydrates, %TEE/day | 52 ± 1 | 49 ± 0 | 52 ± 1 |
| Fiber, g/day | 33 ± 1 | 31 ± 1 | 33 ± 2 |
| Fluid, ml/day | 3,393 ± 305 | 3,407 ± 304 | 3,405 ± 301 |
| Fluid, ml·kg body wt−1·day−1 | 42.0 ± 0.0 | 42.0 ± 0.0 | 42.0 ± 0.0 |
| Calcium, mg/day | 1,068 ± 26 | 1,152 ± 43 | 1,133 ± 63 |
| Chloride, mg/day | 6,311 ± 472 | 6,478 ± 506 | 6,598 ± 587 |
| Chloride, mmol·kg body wt−1·day−1 | 2.20 ± 0.04 | 2.25 ± 0.06 | 2.29 ± 0.00 |
| Potassium, mg/day | 4,434 ± 194 | 3,972 ± 206 | 4,153 ± 198 |
| Sodium, mg/day | 4,077 ± 323 | 4,152 ± 329 | 4,168 ± 368 |
| Sodium, mmol·kg body wt−1·day−1 | 2.20 ± 0.03 | 2.23 ± 0.06 | 2.24 ± 0.00 |
| Magnesium, mg/day | 463 ± 19 | 447 ± 15 | 421 ± 26 |
| Phosphorus, mg/day | 1,778 ± 106 | 1,641 ± 44 | 1,606 ± 104 |
| Iron, mg/day | 13.3 ± 0.6 | 13.8 ± 0.9 | 14.5 ± 1.0 |
| Fluoride, μg/day | 2,893 ± 88 | 1,597 ± 143 | 1,318 ± 61 |
| Iodide, μg/day | 273 ± 19 | 119 ± 11 | 189 ± 12 |
| Zinc, mg/day | 11.9 ± 0.6 | 11.3 ± 0.4 | 15.8 ± 1.2 |
| Copper, μg/day | 2,059 ± 81 | 1,821 ± 83 | 2,022 ± 105 |
| Biotin, μg/day | 44.4 ± 1.5 | 82.2 ± 2.4 | 51.4 ± 2.3 |
| Folic acid, μg/day | 790 ± 20 | 605 ± 19 | 365 ± 12 |
| Niacin equivalent, μg/day | 35,229 ± 2,943 | 24,229 ± 1,552 | 34,294 ± 2,672 |
| Pantothenic acid, mg/day | 7.5 ± 0.5 | 6.0 ± 0.4 | 5.2 ± 0.3 |
| Retinol equivalent, μg/day | 1,902 ± 160 | 2,020 ± 234 | 812 ± 15 |
| Vit A retinol, mg/day | 0.6 ± 0.0 | 0.3 ± 0.0 | 0.5 ± 0.0 |
| Vit B1, mg/day | 1.4 ± 0.1 | 2.1 ± 0.2 | 1.7 ± 0.2 |
| Vit B12, μg/day | 6.6 ± 0.8 | 5.0 ± 0.4 | 5.5 ± 0.5 |
| Vit B2, mg/day | 1.8 ± 0.1 | 1.8 ± 0.1 | 1.5 ± 0.1 |
| Vit B6, mg/day | 2.7 ± 0.2 | 2.7 ± 0.3 | 2.8 ± 0.2 |
| Vit C, mg/day | 324 ± 22 | 408 ± 78 | 98 ± 6 |
| Vit D, IU/day | 1,202 ± 19 | 1,105 ± 3 | 1,065 ± 2 |
| Vit E, mg/day | 19.9 ± 0.6 | 13.2 ± 0.8 | 13.0 ± 0.6 |
| Vit K, μg/day | 384 ± 36 | 128 ± 90 | 108 ± 8 |
Measurements.
As part of the standardized bed rest protocol, general health indicators including three blood pressure and heart rate readings (Intellivue MMS X2; Philips, Best, The Netherlands) were obtained every morning in the fasted state immediately following the scheduled wake-up at 6:30 AM. In addition, body mass was also assessed daily in bed with a bed scale (DVM 5703; Sartorius, Goettingen, Germany) following the first urine void of the day. Subjects were weighed in standardized dedicated clothing, the weight of which was subtracted from the total weight for accurate subject weight. Body temperature was also measured in the ear three times, with the mean used for statistical analysis. Urine output was collected and pooled every 24 h for assessment. Furthermore, venous blood samples were taken at baseline (morning of HDT day, before entry into 12° HDT position) and after 4.5 h HDT, 24 h HDT, and 28.5 h HDT + 3% CO2. Various parameters were measured including white blood cells (WBC), lymphocytes (LYM), monocytes (MON), neutrophils (NEU), eosinophils (EOS), basophils (BAS), red blood cells (RBC), hemoglobin (HGB), hematocrit (HCT), mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), red blood cell distribution width (RDW), platelet count (PLT), and mean platelet volume (MPV). The aforementioned measured parameters are presented as standard health indicators, along with core environmental data.
To fully understand the physiological effects of HDT with and without increased ambient CO2, a comprehensive set of cerebral, ocular, and cardiopulmonary measurements were performed on a carefully timed and adhered-to schedule. This ensured that measurements with particular devices or assays were taken at the same time of day across conditions for each subject as well as within 3 h of the other subjects within the campaign group. Deviations from the planned schedule were minimal (no more than 15 min), allowing for cross-comparison and multimodality investigation. All data acquisition systems were time-synchronized to a master clock which was displayed prominently, and the resulting individual data sets were merged into a master data table in Stata v14. Results from the various physiological and anatomical measurements will be published in a series of discrete papers.
Statistical analysis.
Statistical analysis was carried out in IBM SPSS Statistics Version 20 (IBM, Armonk, NY). ANOVA and linear mixed effect models were constructed with time and atmosphere as main effects allowing for a time-atmosphere interaction, and subject as a random effect. When a main effect was found to be significant, Bonferroni post hoc analyses were implemented (adjusted for multiple comparisons by dividing α by the number of comparisons) to determine differences between time points and/or atmospheric conditions. All data are presented as means ± SD. The level for statistical significance was set to α = 0.05, and β was set to 0.2.
RESULTS
Environmental conditions.
Environmental parameters in the bedrooms, measurement rooms, and common spaces of the : envihab bed rest facility were recorded every minute, 24 h/day (Fig. 2). On days with ambient atmosphere (baseline data collection days and 12° HDT with ambient air), the mean CO2 level was 0.04 ± 0.01%, whereas on days with increased ambient CO2 (intervention day 12° HDT with 0.5% CO2), the mean CO2 level was 0.48 ± 0.02%.
Fig. 2.Atmospheric CO2 percentage in the various rooms within the : envihab bed rest facility as detected by wall-mounted CO2 sensors.
General health indicators.
General health indicator data are presented in Table 3. Twenty-four-hour pooled urine volume had a significant main effect of time (P = 0.005) and was found to increase from 2,533.8 ± 324.7 ml at baseline data collection to 3,038.5 ± 506.1 ml during 12° HDT with ambient air (P = 0.04) and from 2,671.8 ± 704.3 ml at baseline data collection to 3,185.2 ± 325.8 ml during 12° HDT + 0.5% CO2 (P = 0.03). However, there was no significant main effect of atmosphere (P = 0.4). No significant main effects of time or atmosphere were found for mean arterial pressure (P = 0.97, P = 0.6), systolic blood pressure (P = 0.5, P = 0.6), diastolic blood pressure (P = 0.5, P = 0.8), or heart rate (P = 0.2, P = 0.4). Body temperature, however, was found to have a main effect of atmosphere (P = 0.02) but not time (P = 0.6). Body weight showed a significant main effect of time (P = 0.001) but not atmosphere (P = 0.97), with a slight decrease in mean body weight after 19.5 h HDT (P = 0.02).
| Condition | Baseline | 19.5 h HDT |
|---|---|---|
| Body Weight, kg | ||
| HDT + 0.5% CO2 | 82.03 ± 7.71 | 81.33 ± 7.36* |
| HDT + amb | 82.1 ± 7.93 | 81.42 ± 7.56 |
| Body Temperature, °C | ||
| HDT + 0.5% CO2 | 35.25 ± 0.93 | 35.27 ± 0.98 |
| HDT + amb | 34.88 ± 0.86 | 34.88 ± 0.86 |
| Mean Arterial BP, mmHg | ||
| HDT + 0.5% CO2 | 86.4 ± 4.77 | 85.32 ± 5.04 |
| HDT + amb | 85.88 ± 6.66 | 86.58 ± 5.93 |
| Systolic BP, mmHg | ||
| HDT + 0.5% CO2 | 113.33 ± 3.2 | 113.85 ± 5.86 |
| HDT + amb | 112.88 ± 8.86 | 117.05 ± 5.74 |
| Diastolic BP, mmHg | ||
| HDT + 0.5% CO2 | 72.93 ± 6.72 | 71.05 ± 5.95 |
| HDT + amb | 72.38 ± 6.94 | 71.32 ± 7.72 |
| Heart Rate, beats/min | ||
| HDT + 0.5% CO2 | 60.5 ± 6.82 | 61.22 ± 9.92 |
| HDT + amb | 62.88 ± 8.51 | 59.05 ± 8.85 |
Blood parameters.
As shown in Table 4, significant main effects of time were found for WBC (P < 0.001), LYM (P < 0.001), MON (P = 0.003), NEU (P < 0.001), EOS (P < 0.001), HGB (P = 0.001), HCT (P < 0.001), RBC (P = 0.004), MCH (P = 0.003), and RDW (P = 0.035). Significant main effects of atmosphere were found for BAS (P = 0.036), MCV (P = 0.045), PLT (P = 0.002), and MPV (P < 0.001). No main effects were found for MCHC.
| BDC (Morning of HDT) | 4.5 h HDT | 24 h HDT | 28.5 h HDT + 3% CO2 | |
|---|---|---|---|---|
| White Blood Cells, × 103/µl | ||||
| HDT + 0.5% CO2 | 5.67 ± 1.37 | 6.67 ± 1.63 | 7 ± 1.79* | 8.17 ± 2.32‡ |
| HDT + amb | 6 ± 1.26 | 7.17 ± 2.23 | 7.17 ± 1.47 | 7.83 ± 2.14‡ |
| Lymphocytes, % | ||||
| HDT + 0.5% CO2 | 42.17 ± 4.79 | 35.17 ± 5.15‡ | 33 ± 4.29‡ | 30.17 ± 5.64‡ |
| HDT + amb | 42.83 ± 3.43 | 36 ± 5.9‡ | 32.33 ± 5.16‡ | 31.17 ± 6.65‡ |
| Monocytes, % | ||||
| HDT + 0.5% CO2 | 10.67 ± 1.63 | 10.5 ± 1.05 | 11.33 ± 1.86 | 11.67 ± 1.37 |
| HDT + amb | 11.17 ± 1.33 | 10 ± 1.79 | 10.83 ± 0.98 | 10.83 ± 1.17 |
| Neutrophils, % | ||||
| HDT + 0.5% CO2 | 43.67 ± 5.13 | 51.67 ± 5.32‡ | 53.5 ± 4.76‡ | 55.83 ± 6.82‡ |
| HDT + amb | 42.83 ± 4.12 | 51.5 ± 5.96‡ | 54.33 ± 5.24‡ | 55.33 ± 6.22‡ |
| Basophils, % | ||||
| HDT + 0.5% CO2 | 0 ± 0 | 0 ± 0 | 0.17 ± 0.41 | 0.33 ± 0.52 |
| HDT + amb | 0.5 ± 0.55 | 0.17 ± 0.41 | 0.17 ± 0.41 | 0.17 ± 0.41 |
| Eosinophils, % | ||||
| HDT + 0.5% CO2 | 2.83 ± 0.98 | 2.17 ± 0.75 | 2 ± 0.89 | 1.83 ± 0.75* |
| HDT + amb | 3.33 ± 0.82 | 2.33 ± 0.52* | 2.17 ± 0.75† | 2 ± 0.89‡ |
| Hemoglobin, g/dl | ||||
| HDT + 0.5% CO2 | 15 ± 0.89 | 15.17 ± 1.17 | 15.5 ± 0.84 | 15.5 ± 0.84 |
| HDT + amb | 14.83 ± 1.17 | 15.33 ± 1.03 | 15.5 ± 1.05 | 15.83 ± 1.17† |
| Hematocrit, % | ||||
| HDT + 0.5% CO2 | 43 ± 2.76 | 43.17 ± 3.31 | 44.5 ± 3.02 | 45 ± 2.19 |
| HDT + amb | 42.67 ± 3.39 | 43.5 ± 3.08 | 44.83 ± 2.99* | 45.5 ± 2.35† |
| Red Blood Cells, × 106/µl | ||||
| HDT + 0.5% CO2 | 5 ± 0 | 5 ± 0 | 5.33 ± 0.52 | 5.33 ± 0.52 |
| HDT + amb | 5 ± 0 | 5 ± 0 | 5.17 ± 0.41 | 5.33 ± 0.52 |
| Mean Corpuscular Volume, fl | ||||
| HDT + 0.5% CO2 | 85.67 ± 4.5 | 85.33 ± 4.18 | 85.67 ± 4.5 | 85.67 ± 3.93 |
| HDT + amb | 85.67 ± 4.5 | 86 ± 4.52 | 86 ± 4.52 | 85.83 ± 4.22 |
| Mean Corpuscular Hemoglobin, pg | ||||
| HDT + 0.5% CO2 | 29.33 ± 1.63 | 29.83 ± 1.47 | 30 ± 1.41† | 29.83 ± 1.47 |
| HDT + amb | 29.67 ± 1.37 | 30 ± 1.41 | 30 ± 1.41 | 29.83 ± 1.47 |
| Mean Corpuscular Hemoglobin Concentration, g/dl | ||||
| HDT + 0.5% CO2 | 34.5 ± 0.55 | 34.67 ± 0.52 | 34.67 ± 0.52 | 34.83 ± 0.41 |
| HDT + amb | 34.67 ± 0.52 | 35 ± 0.63 | 34.67 ± 0.52 | 34.83 ± 0.41 |
| Red Blood Cell Distribution Width, % | ||||
| HDT + 0.5% CO2 | 12.67 ± 0.52 | 12.5 ± 0.84 | 12.5 ± 0.84 | 12.33 ± 0.82 |
| HDT + amb | 12.67 ± 1.03 | 12.5 ± 1.05 | 12.5 ± 1.05 | 12.33 ± 0.82 |
| Platelets, × 103/µl | ||||
| HDT + 0.5% CO2 | 224.33 ± 41.25 | 231.33 ± 42.87 | 228.17 ± 42.93 | 233.5 ± 44.23 |
| HDT + amb | 221.67 ± 35.64 | 245 ± 24.9 | 245.17 ± 22.12 | 252.67 ± 32.89 |
| Mean Platelet Volume, fl | ||||
| HDT + 0.5% CO2 | 8.83 ± 1.17 | 8.5 ± 1.05 | 8.33 ± 1.03 | 8.17 ± 1.17 |
| HDT + amb | 8.17 ± 0.75 | 7.83 ± 0.98 | 8 ± 0.89 | 8 ± 0.89 |
DISCUSSION
The SPACECOT study demonstrates the feasibility of an innovative approach to studying the effects of headward fluid shifting and elevated ambient CO2 on cerebral and ocular anatomy and physiology. The implemented approach utilized a steeper HDT angle (12°) than what is typically used (6°), both with and without increased ambient CO2, to decipher the potential role of atmospheric conditioning during HDT bed rest. In addition, the study implemented a closely controlled environment and strict adherence to standardization plans and the HDT position. A full comprehensive examination of cerebral, ocular, and cardiopulmonary physiological parameters was implemented with a combination of gold-standard and novel noninvasive technologies, allowing for a full body system physiological analysis. While it is unclear whether HDT bed rest is a valid model for the ocular changes that occur during spaceflight, our approach aimed to investigate the cerebral and ocular effects of two major hypothesized contributing factors to the VIIP syndrome: headward fluid shifting and exposure to chronically elevated ambient CO2 concentrations.
The :envihab facility was able to provide full environmental control and continuous monitoring of atmospheric conditions, light intensity and spectrum parameters, temperature, and humidity. Atmospheric condition during HDT had little or no additional effect on daily blood pressure, heart rate, urine output, body weight, or blood count parameters. However, HDT affected many of the parameters, regardless of atmospheric condition, notably inducing increased urine output, WBC, NEU, HCT, and HGB. Diuresis is a known response to return the perceived HDT-induced increases in central blood volume close to baseline values (2). Increased HCT and HGB are also likely due to the shift in central blood volume. Notably, WBC count increased with HDT and was further aggravated by the short duration exposure to 3% CO2. The percentage share of LYM and granulocytes was shifted to NEU in HDT, and when translated into absolute counts, there is little change in LYM count while innate immune cells are increased. The observed increase in polymorphonuclear cells in our study is consistent with an acute stress response. Typically, physiological stress leads to demargination of NEU from the blood vessel wall into the lumen of the blood, where they enter the systemic circulation and are thus measurable. This is consistent with a previous HDT study in which eight volunteers underwent 6° HDT for 42 days and polymorphonuclear cells increased during HDT, whereas the T-lymphocytes and monocytes did not change (12).
In contrast to the slight elevation of WBC count, the vital signs including blood pressure did not significantly change from baseline to the late HDT time point (Table 3) in either the HDT alone or HDT with 0.5% CO2 condition. This is consistent with findings from previous bed rest studies (10, 13), and there may be several explanations for this. First, the subjects may have adapted to the stress of the environment and, therefore, the blood pressure remained unchanged from baseline. Second, a decrease in plasma volume related to HDT-induced diuresis may have occurred, which would lower the central venous pressure and preload, thus reducing cardiac output given a stable heart rate.
Nutritional intake was highly standardized and regulated in the presented study and is important to take into consideration when interpreting results of other physiological systems, because diet may affect multiple organ system function including cardiovascular and cerebrovascular parameters. Notably, sodium content was set to ~4 g/day in this study, standard for bed rest studies, and is an important factor to monitor closely, because sodium intake could have effects on blood pressure and fluid retention. Whether sodium intake has an effect on cerebral and ocular changes in spaceflight is unknown; however, if vascular fluid content and thus volume increases, this may also have effects on the cerebrovascular system. Given that ICP is related to cerebral venous outflow and central venous pressure, it is possible that a high sodium diet may contribute to a higher circulating venous volume and affect ICP.
The unique platform of the SPACECOT study with a steeper degree of HDT as well as atmospheric conditioning is considered to be more challenging for subjects than standard HDT bed rest studies, and, therefore, new procedures were implemented to ensure both subject comfortability and successful completion of the study. First, we selected several subjects with prior bed rest experience who were mentally prepared for the bed rest experience. Furthermore, we extended subject recruitment to include brief HDT exposure for both mental and physical familiarization with the demands of bed rest at a steeper HDT angle, as well as an MRI session to exclude unknown anxiety disorders or claustrophobia. In addition, a full day of familiarization was added before the first day of baseline data collection, to introduce the subjects to all of the study measurement techniques and the busy schedule. This is thought to have alleviated some of the stress associated with the complex procedures of the study, allowing subjects to be more relaxed and comfortable during actual baseline measurements the following day. Furthermore, pillows are normally provided to subjects in bed rest studies; however, the use of a pillow places the head and eyes slightly above the intended degree of full body tilt. Thus, pillows were not allowed in the SPACECOT study, an important consideration for future bed rest studies focused on cerebral and ocular physiology. In addition, bed rest studies often involve procedures that require subjects to move into the supine position (e.g., MRI scans and exercise regimes). In the present study, we made every effort to ensure the 12° HDT position was maintained during the entire bed rest period. This included having dedicated subject monitors assigned to every subject to ensure subject safety and comfort as well as maintain head, neck, and body position at 12° HDT during bed rest, transfers to other beds, and meal times. In addition, custom-built wedges at 12° HDT were implemented into the MRI scanner to maintain the HDT position. Furthermore, a mask and tank system was used when subjects were required to leave the bed rest facility for all conditions (ambient and increased CO2) to retain the blinded aspect of the study and atmospheric conditioning when in the MRI scanner. It is important to note that subjects were unlikely to be in a fully relaxed state due to the stressful environment of the busy study. However, this is thought to be similar to the stressful environment that astronauts may endure during spaceflight.
With the implementation of a steeper HDT position to investigate the effects of cephalad fluid shifting, we determined inherent limitations and hereby provide recommendations for successful future implementation of this approach. First, the 12° HDT position can lead to subjects slowly slipping down the bed. We therefore lined the mattress with a nonskid pad to prevent slippage. In addition, lower back pain was a common complaint, and countermeasures against this should be in place, including heating pads and daily physical therapy massages applied to the lower back. Given the relatively short duration of the exposure to 12° HDT, we cannot extrapolate as to whether the back pain would have resolved over time; however, back pain also frequently occurs with a milder degree of HDT and usually resolves spontaneously after adaptation. Finally, eating was found to be more difficult at this steeper HDT angle, and therefore dedicated subject monitors also assisted subjects with eating, and the menu was adapted to provide more finger food such as sandwiches. Adverse events of the study included one episode of emesis, and one subject was unable to urinate in the 12° HDT position during the first campaign and had to have a urinary catheter inserted. In the subsequent campaign, this subject was briefly moved to the 0° position for urination. Data were examined to determine whether measurements obtained from this one subject were outside of one standard deviation of the group mean, and this was determined not to be the case. With this one exception, in general, the subjects were able to urinate and defecate at the 12° HDT position without difficulty.
There are important differences between real microgravity and HDT bed rest that must be taken into account when interpreting results, most notably, the presence of gravitational vectors in the latter. Unlike in microgravity where all hydrostatic gradients are abolished, a small foot-to-head vector is created during HDT to create a headward fluid shift similar to what may be experienced in microgravity. In addition, a Gx (chest to back) gravitational vector is present in HDT bed rest, which may result in different cardiovascular and pulmonary physiological responses compared to microgravity. Furthermore, this study aimed to investigate the immediate effects of headward fluid shifting in the presence of elevated ambient CO2 to understand potential mechanistic underlying factors that, if persistent, could lead to ophthalmic changes associated with the VIIP syndrome. However, because ophthalmic changes in astronauts appear after months of microgravity exposure, we cannot assume that observed changes can be directly applied to longer term HDT bed rest or microgravity.
To our knowledge, this is the first study to implement a steeper HDT position with increased ambient CO2 for more than 24 h of bed rest in healthy human subjects as an approach to investigate two hypothesized contributing factors to ocular changes in astronauts: headward fluid shifting as well as exposure to chronically elevated ambient CO2. Further work is needed to determine whether a longer duration of bed rest within this model is well tolerated and leads to physiological and anatomical changes similar to those associated with spaceflight.
GRANTS
This study was supported by the National Space Biomedical Research Institute through NASA NCC 9–58, the Baylor College of Medicine Center for Space Medicine, and the German Aerospace Center Institute of Aerospace Medicine.
DISCLOSURES
No conflicts of interest, financial or otherwise, are declared by the authors.
AUTHOR CONTRIBUTIONS
K.M.-G., E.M., D.D., G.S., J.I.S., C.V.R., P.F.-M., U.L., J.R., and E.M.B. conceived and designed research; K.M.-G., E.M., D.D., G.S., J.I.S., C.V.R., P.F.-M., U.L., J.R., and E.M.B. performed experiments; K.M.-G., E.M., P.F.-M., and E.M.B. analyzed data; K.M.-G., E.M., U.L., and E.M.B. interpreted results of experiments; K.M.-G. and J.R. prepared figures; K.M.-G. drafted manuscript; K.M.-G., E.M., D.D., G.S., J.I.S., C.V.R., P.F.-M., U.L., J.R., and E.M.B. edited and revised manuscript; K.M.-G., E.M., D.D., G.S., J.I.S., C.V.R., P.F.-M., U.L., J.R., and E.M.B. approved final version of manuscript.
ACKNOWLEDGMENTS
We acknowledge the SPACECOT investigators group (in alphabetical order): Mathias Basner, Christine Becker, Eusebia Calvillo, Jonathan Clark, Rahul Damani, Wolfgang Doering, Christian Dohmen, Peter Gauger, Darius Gerlach, Olga Hand, Khader Hasan, Elfriede Huth, Bernd Johannes, Larry Kramer, Gabriele Kraus, Uwe Mittag, Klaus Muller, Jad Nasrini, Ben Niederberger, Dirk Poddig, Matthias Putzke, Martina Sagner, Haleh Sangi-Haghpeykar, Irmtrud Schrage-Knoll, Wolfram Sies, Claudia Stern, Henning Stetefeld, Brian Stevens, Annette von Waechter, Tobias Weber, and Martin Wittkowski. We also thank the subjects for their time and steadfast dedication. We appreciate the technical support and advice provided by Mark Shusterman, Mitch Levinson, Rolandas Zaeklis, Adi Tsalach, Avihai Ron, and Tracy Johnson. Finally, this study would not have been possible without the encouragement and support of Rupert Gerzer, Julie Do, and Jeff Sutton.
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