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4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 0
|
After a 16-year hiatus, Russia has resumed its program of biomedical research in space, with the successful 30-day flight of the Bion-M 1 biosatellite (April 19–May 19, 2013). The principal species for biomedical research in this project was the mouse. This paper presents an overview of the scientific goals, the experimental design and the mouse training/selection program. The aim of mice experiments in the Bion-M 1 project was to elucidate cellular and molecular mechanisms, underlying the adaptation of key physiological systems to long-term exposure in microgravity. The studies with mice combined in vivo measurements, both in flight and post-flight (including continuous blood pressure measurement), with extensive in vitro studies carried out shortly after return of the mice and in the end of recovery study. Male C57/BL6 mice group housed in space habitats were flown aboard the Bion-M 1 biosatellite, or remained on ground in the control experiment that replicated environmental and housing conditions in the spacecraft. Vivarium control groups were used to account for housing effects and possible seasonal differences. Mice training included the co-adaptation in housing groups and mice adaptation to paste food diet. The measures taken to co-adapt aggressive male mice in housing groups and the peculiarities of
|
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4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 1
|
“space” paste food are described. The training program for mice designated for in vivo studies was broader and included behavioral/functional test battery and continuous behavioral measurements in the home-cage. The results of the preliminary tests were used for the selection of homogenous groups. After the flight, mice were in good condition for biomedical studies and displayed signs of pronounced disadaptation to Earth's gravity. The outcomes of the training program for the mice welfare are discussed. We conclude that our training program was effective and that male mice can be successfully employed in space biomedical research.
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 2
|
After a 16-year hiatus, Russia resumed in 2013 its program of biomedical research in space, with the successful 30-day flight of the Bion-M 1 biosatellite (April 19–May 19, 2013), a specially designed automated spacecraft dedicated to life-science experiments. “M” in the mission's name stands for “modernized”; the epithet was equally applicable to the spacecraft and the research program. The principal animal species for physiological studies in this mission was the mouse ( Mus musculus ). Unlike more recent space experiments that used female mice, males were flown in the Bion-M 1 mission. The challenging task of supporting mice in space for this unmanned, automated, 30-day-long mission, was made even more so by the requirement to house the males in groups. Russian biomedical research in space traditionally has employed dogs, rats, monkeys, and more recently Mongolian gerbils. The flight of Laika in 1957 was one of the early dog experiments and became world famous for demonstrating that a living organism can withstand rocket launch and weightlessness, thus paving the way for the first human spaceflight. Laika's success also promoted biomedical research with other non-human animals in space that culminated with the Bion biosatellites program. A total of 212 rats and 12
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 3
|
monkeys were launched on 11 satellites and exposed in microgravity for 5.0–22.5 days between 1973 and 1997. Animal experiments on the Bion missions have contributed comprehensive data on adaptive responses of sensorimotor, cardiovascular, muscle, bone and other systems to spaceflight conditions and the mechanisms underlying these adaptations [1] , [2] . The use of mice for space experiments offers numerous advantages. Probably the most apparent one is their small size and thus the possibility of utilizing more animals per flight, thus increasing scientific output and the cost-efficiency ratio. Comparisons of data obtained with mice, with those obtained from larger species or humans can also reveal how factors affecting adaptation to spaceflight conditions depend on the size of the organism. The mouse has become the most prevalent “mammalian model” in biomedical research, with a fully described genome and an established role in genetically engineered mutants. While mice are preferred mammalian models for molecular biology studies, their small size is a debated limitation rather than an advantage for physiological studies. Miniaturization of scientific hardware has reduced some of the disadvantages of the species small size. Finally, the use of genetically controlled mice offers a means to reduce inter-individual variability and obtain potentially
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 4
|
more consistent results. Despite the advantages of the mouse as a model organism for space research, their use was rather limited (apart from a number of experiments with mice during early space exploration in the 1950's and 1960's, which were aimed primarily at testing if living organisms can survive the launch or a brief exposure in microgravity) [3] . Flight experiments with mice were performed aboard STS-90 (“NeuroLab”), STS-108, STS-129, STS-131 (“Mouse Immunology I”), STS-133 (“Mouse Immunology II”), and STS-135 with exposure times ranging from 12 to 16 days. Research programs of these flights were largely focused on studies of muscle, bone/tendon/cartilage, nervous, and cardiovascular systems, and innate and acquired immune responses. Experiments were performed with groups of 30 or fewer female C57BL/6J mice, which were dissected typically shortly after return [4] – [9] . The Mice Drawer System (MDS) experiment of the Italian Space Agency is by far the longest spaceflight of mice to date [10] . In this mission, 6 mice were exposed for 91 days aboard the International Space Station. The advantages offered by the possibility of genetic manipulations with mice were utilized in this experiment; three mice were transgenic with pleiotrophin overexpression (C57BLJ10/ PTN) and three
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 5
|
mice were their wild-type counterparts. The MDS habitats required periodic replenishment and servicing by by astronauts. Sadly, half of the mice died during the course of this mission due to various estimated reasons. In the present paper we aim to present a brief overview of the Bion-M 1 mission scientific goals and experimental design. Of particular interest we will focus on the program of mouse training and selection for the experiments, and some outcomes of the Bion-M 1 mission. The scientific program of the Bion-M 1 project was aimed at obtaining data on mechanisms of adaptation of muscle, bone, cardiovascular, sensorimotor and nervous systems to prolonged exposure in microgravity and during post-flight recovery. To this aim, functional measurements in vivo were combined with complementary morphological, biochemical, cellular and molecular studies performed in vitro . To our knowledge, Bion-M 1 is the first study with mice to follow the dynamics of re-adaptation to normal gravity or to assess post-flight in vivo functions. The in vivo measurements in-flight included video recording of mice behavior and continuous recording of arterial pressure during all stages of the flight. After the flight, testing of animal behavior, memory and learning, vestibular function, physical activity, strength and
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 6
|
fitness, as well as continuous registration of cardiovascular parameters were performed. The set of in vivo measurements was designed to provide data on the effects of microgravity on the physiological systems of interest, and to follow the time course of re-adaptation to Earth's gravity after the flight. To reduce the possibility of inadvertently stressing the mice with numerous post-flight tests and thus obscuring the effects of spaceflight, the decision was made to monitor mice in their home cage using an automated data collection system (TSE PhenoMaster) and implantable telemetry, while reducing the number of tests [11] . The program had to be performed with a limited number of animals; thus the use of a repeated measures design. Mice were tested before and after the flight (or its simulation on-ground) [12] . The in vitro studies were focused on metabolism and the structure and function of tissues from microgravity-exposed mice. More specifically, the individual studies were selected to maximize the scientific yield and targeted the impact of microgravity and other factors specific to spaceflight on biochemical, morphological and, primarily, cellular and molecular mechanisms involved in the adaptation of main physiological systems, such as the central nervous system, sensory systems, skeletal muscle,
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 7
|
supporting tissues, including bone and connective tissues, cardiovascular, respiratory, digestive, neuroendocrine and immune systems. The following organs and tissues were harvested: brain structures, vestibular organs, retina and crystalline lens, spinal cord, muscles of hind limbs and back, vertebral column, fore and hind paw bones, articular and cartilage structures, ligaments, bone marrow, cardiac muscle, arteries from brain and hind legs, lung, intestines, liver, kidney, spleen, thymus, skin, salivary glands, testis and blood. These samples were analyzed using morphological (light and electron microscopy, histochemistry and immunohistochemistry), biochemical and a variety of molecular-biology techniques (enzyme-linked immune assays, high-pressure liquid chromatography, western-blot, real-time PCR, whole-genome gene expression). Functional properties of skeletal muscle, cardiac muscle and resistive arteries samples were evaluated using myography and atomic force microscopy. Mice for in vitro studies were euthanized by cervical dislocation; because of the diversity and number of studies, this method was chosen as the common procedure to reduce variability and one that provided the least interference with the objectives of the in vitro studies.
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 8
|
The scientific program of the Bion-M 1 project was aimed at obtaining data on mechanisms of adaptation of muscle, bone, cardiovascular, sensorimotor and nervous systems to prolonged exposure in microgravity and during post-flight recovery. To this aim, functional measurements in vivo were combined with complementary morphological, biochemical, cellular and molecular studies performed in vitro . To our knowledge, Bion-M 1 is the first study with mice to follow the dynamics of re-adaptation to normal gravity or to assess post-flight in vivo functions. The in vivo measurements in-flight included video recording of mice behavior and continuous recording of arterial pressure during all stages of the flight. After the flight, testing of animal behavior, memory and learning, vestibular function, physical activity, strength and fitness, as well as continuous registration of cardiovascular parameters were performed. The set of in vivo measurements was designed to provide data on the effects of microgravity on the physiological systems of interest, and to follow the time course of re-adaptation to Earth's gravity after the flight. To reduce the possibility of inadvertently stressing the mice with numerous post-flight tests and thus obscuring the effects of spaceflight, the decision was made to monitor mice in their home cage using
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 9
|
an automated data collection system (TSE PhenoMaster) and implantable telemetry, while reducing the number of tests [11] . The program had to be performed with a limited number of animals; thus the use of a repeated measures design. Mice were tested before and after the flight (or its simulation on-ground) [12] . The in vitro studies were focused on metabolism and the structure and function of tissues from microgravity-exposed mice. More specifically, the individual studies were selected to maximize the scientific yield and targeted the impact of microgravity and other factors specific to spaceflight on biochemical, morphological and, primarily, cellular and molecular mechanisms involved in the adaptation of main physiological systems, such as the central nervous system, sensory systems, skeletal muscle, supporting tissues, including bone and connective tissues, cardiovascular, respiratory, digestive, neuroendocrine and immune systems. The following organs and tissues were harvested: brain structures, vestibular organs, retina and crystalline lens, spinal cord, muscles of hind limbs and back, vertebral column, fore and hind paw bones, articular and cartilage structures, ligaments, bone marrow, cardiac muscle, arteries from brain and hind legs, lung, intestines, liver, kidney, spleen, thymus, skin, salivary glands, testis and blood. These samples were analyzed using morphological (light and
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 10
|
electron microscopy, histochemistry and immunohistochemistry), biochemical and a variety of molecular-biology techniques (enzyme-linked immune assays, high-pressure liquid chromatography, western-blot, real-time PCR, whole-genome gene expression). Functional properties of skeletal muscle, cardiac muscle and resistive arteries samples were evaluated using myography and atomic force microscopy. Mice for in vitro studies were euthanized by cervical dislocation; because of the diversity and number of studies, this method was chosen as the common procedure to reduce variability and one that provided the least interference with the objectives of the in vitro studies.
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 11
|
The study was approved by IACUC of MSU Institute of Mitoengineering (Protocol № 35, 1 November, 2012) and of Biomedical Ethics Commission of IBMP (protocol № 319, 4 April, 2013) and conducted in compliance with the European Convention for the Protection of Vertebrate Animals used for Experimental and Other Scientific Purposes [13] . Experiments were performed with C57BL/6N male mice. The choice of strain was based on the requirements of the tissue harvesting program participants and the fact that C57/BL6 is one of the most widely used strains. The selection of male mice was based on the same requirement. The advantages offered by using males are their substantially larger size and the absence of sex steroid cycling compared to females; the vast majority of animal research is conducted using males, so their use in this study made the results more readily comparable to the data in the published literature. Nevertheless, the risks associated with group-housing mature males were recognized and greatly affected the preparatory procedures. It was decided that the optimal age of mice at launch was 4–5 months, when the males were still young but fully mature. The growth rate in mice is known to slow down by this
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 12
|
age, and obesity, which C57/BL6 mice are prone to exhibit, is not yet considered a factor. Space environment affects microorganisms by increasing their virulence; major shifts in immune status in space are well documented [5] , [8] . It is reasonable to think that even pseudo pathogenic species may increase morbidity during prolonged exposure in space [14] , [15] . These considerations underscored the necessity for the use of specific pathogen-free mice in the experiments, a side benefit of this approach being the reduced variability of data obtained in SPF-animals [16] . Male C57BL/6N mice (n = 300) weighing 22–25 g were purchased from the Animal Breeding Facility - Branch of Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry. Mice were specific pathogen free. The main site for mouse training was the animal facility of Moscow State University's Institute of Mitoengineering. At the time of launch and the start of the related control experiments, the mice were 19–20 weeks old. Mice were individually identified using RFID chips (El1000, Felixcan, Spain) and earmarks. Routine manipulations of the mice consisted of daily handling, examination, and weighing. The Bion-M 1 biosatellite, launched on April 19, 2013 from cosmodrome Baikonur, and the descent module landed
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 13
|
on May 19, 2013 in the vicinity of Orenburg, successfully fulfilling the plan for an unmanned, 30-day-long orbital spaceflight. Housing and climate parameters were replicated in the subsequent ground control (GC) experiment (July 26 to August 26, 2013). A total of 4 experimental groups were used for the flight and ground control experiments (n = 45 per group). An additional fifth group included the backup mice for the main flight group (n = 45). Mice of the space flight group (SF) were exposed to microgravity for 30 days. Concurrent with the SF mice, another group of 45 mice remained in the animal facility (SFV). The ground control (GC) experiment was conducted, after the landing, in the refurbished BOS flight habitats. The habitats were installed in a climatic chamber that replicated the temperature, humidity, gas composition and other flight-specific climate parameters. The corresponding vivarium control (GCV) mice were housed in the animal facility. The separate and concurrent GCV groups were used to account for possible seasonal differences between SF and GC mice. Each of the groups (SF, GC, SFV, GCV and backup SF mice) included mice designated for in vivo studies and recovery (n = 10) and mice for dissection and
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 14
|
in vitro measurements (n = 35). Each in vivo study subgroup, in its turn, consisted of 5 mice implanted with telemetry probes to monitor blood pressure and 5 intact animals. Mice were handled and trained before the flight and ground control experiments. Basically, training consisted of shaping the groups of three mice each for social housing and adaptation to paste diet. The training of mice designated for in vivo studies was more comprehensive. It started with implanting the telemetry probes and, following recovery, a set of preliminary behavioral and functional tests ( Figure 1 , Table 1 and 2 ). Mice of SF, SFV and backup SF groups were co-adapted in housing groups of 3 mice each, SF mice were adapted to paste food diet, and mice of in vivo subgroups passed through preliminary tests. After transportation to and adaptation at Baikonur, SF mice were flown aboard the Bion-M 1 satellite for 30 days. After landing mice were examined and transported to Moscow, where animals of the in vitro subgroup were dissected, while recovery dynamics was followed in the in-vivo subgroup before dissection 7 days after landing. Ground control experiment replicated the principal stages of the spaceflight experiment. The BOS
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 15
|
habitats were designed to house up to three mice per group [17] . Stable cohorts of three mice were maintained throughout the training program and consequent experiments, with the exception of brief periods of post-surgery recovery (5–6 days) and behavioral tests (7 days), when they were housed individually. Individually ventilated cages (GM 500, Techniplast, floor area 500 cm 2 ) were used. Techniplast mice shelters (red plastic triangles, 15Lx11Wx8H cm), paper tubes and paper tissues for nesting material were provided to enrich the environment. Cage cleaning was done once weekly, and water was refreshed twice weekly. All materials were sterilized prior to use by autoclave. Techniques aimed to prevent animal contamination were thoroughly followed. The hardware for mice housing in-flight was an adapted version of the habitats used for rats in the original Bion program. It was developed by Biofizpribor, according to the IBMP requirements and was tested by IBMP in 2011. Each block consisted of five individual habitats (BOS, Block Obespecheniya Soderzhaniya, Russian for - literally - Unit for the Provision of Housing). The living compartment of each habitat was an acrylic cylinder 98±2 mm in diameter and 200±10 mm in length with a total volume of approximately 1.7
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 16
|
l ( Figure 2 ). A stainless steel feeder (9 ml) was mounted at one end (the “top”) of the cylinder. The feeder had a flap, used in the refill procedure. The food distribution system (except for the feeders) was common to all five habitats in the block. Paste “space” food was pumped from the container and distributed between individual habitats of the assembly 6 times daily (every 4 hours at 00:00, 04:00, 08:00, 12:00, 16:00 and 20:00), a total of 54 g/day per 3 mice. Food distribution was adjusted through manipulations of pressure developed in the container and in the feeders to follow this pattern prior to experiments. Bulbs for day-phase (08:00–20:00, 45±5 Lux) and night-phase (20:00–08:00, 5±2 Lux) lighting, an air supply fan (0.3 m/s) and an air distribution system were positioned on one side of the cylinder. A video camera was installed between the light bulbs; video was recorded 6 times daily (simultaneously with feeding, each session lasted 2 hours) at 8 frames per second. The opposite side of the cylinder had the grid for waste removal. The waste removal system was individual for each habitat. The door at the “bottom” of the cylinder had a glass
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 17
|
window. In order to minimize impact of variability arising from possible differences in mice housing conditions, experimental subgroups were formed from mice that were housed in different habitats. Air composition and climate parameters in the satellite were maintained by the centralized life support system and remained in the designed range throughout the flight. During the initial stage of training, mice were provided with standard pelleted food (Assortiment Agro, Russia) and deionized water given ad libitum . Two weeks prior to launch, and the start of the GC experiment, the animals were adapted to the flight paste food diet. Both the paste and standard diets (pelleted food and water) were first presented together; after 2–3 days the water bottle was removed, followed in another 2–3 days by the removal of the pelleted food. Vivarium control (SFV and GCV) groups were fed standard chow and water throughout the experiments. The “space” paste food diet was developed at IBMP by T. Gurieva and E. Mednikova. It was produced based on the standard chow, with water and casein as gelling agent added. Consumption of standard pelleted chow and water (in SFV and GCV) or paste diet (in SF and GC) was monitored daily for
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 18
|
ten days in ten randomly chosen cages per group. Data for the cage was averaged across the days and expressed as g per 10 g bodyweight. A separate preliminary experiment in metabolic chambers was conducted to quantify urination in mice fed standard chow and paste food. Four groups of mice, of 3 mice each, were housed in metabolic chambers (Techniplast, Italy) and fed pelleted food (grinded) or paste food. Body weight, food and water consumption, urine volume and feces weight were monitored daily (3 days for each diet). Arterial pressure was monitored in sub-groups of mice designated for recovery, using implantable probes and telemetry hardware (Data Science International, USA). The hardware was adapted and integrated to the Bion-M biosatellite by engineers from Biofizpribor (St. Petersburg, Russia) and the Centre national d'études spatiales (CNES, France). Miniature probes (model PA-C10) weighing 1.4 g were implanted 3 weeks before the start of experiments, following the manufacturer's instructions. The catheter of the probe was implanted into the left carotid so that its tip just reached the aorta [18] . The body of the transmitter was implanted subcutaneously on the flank, briefly described below. Mice were anaesthetized by an intraperitoneal injection of a solution containing
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 19
|
tiletamine/zolazepam (15 mg/kg each) combined with xylazine (3 mg/kg). Depth of anesthesia was assessed regularly by hind leg pinch and observation of respiration rate, and maintained by additional dose(s) of the mixture (about 20% of the initial dose) as needed. A midline incision was made on the ventral neck. The carotid artery was separated and ligated 1–2 mm cranially to its bifurcation. A second ligature was used to suspend the artery 4–5 mm proximal to the site of ligation. Through a small incision in the artery wall, the catheter was advanced 6–7 mm towards the aorta and fixed in place with sutures and acrylic glue. A subcutaneous pocket on the flank of the animal was made with blunt scissors, and the transmitter was inserted there with a small hemostat. The catheter and the transmitter were fixed in place with acrylic glue and the skin incision closed with absorbable sutures. During the recovery period, mice were housed individually; they were under daily veterinarian control and received ibuprofen 4 mg/ml and Bactrim 4 mg/ml in drinking water. Mice for the in vivo studies were trained and tested before the experiments to select homogenous groups and to benefit from repeated measures in experimental
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 20
|
design; the latter consideration was particularly important in light of relatively small number of animals. On the morning of day 0 mice a battery of behavioral and functional tests was administered. First, animals were examined for visible abnormalities in exterior and simple reflexes [19] . The Open field test was performed in a round, 60 cm arena of black plastic with a grey floor and 40 cm high walls [20] . Mouse behavior was video recorded at 25 fps for 10 minutes under a bright white light (200 Lux) followed by 10 min in darkness (IR light). Videos were analyzed for locomotor activity and other behavioral parameters using EthoVision v. 8.5 (Noldus). Grip force was measured using Grass FT-03 force transducer [21] . Analog output was amplified, digitized at 1 kHz sampling rate and analyzed using PowerGraph software (InteropticaS, Russia). RotaRod test was conducted using UgoBasil hardware with rotation speed increased from 6 to 50 rpm in 3 min, 5 trials were performed with each mouse [22] . On the evening of the same day, animals were placed into cages of Phenomaster (TSE Instruments, Germany), an automated data collection system, where they were monitored for 7 days. The system was
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 21
|
equipped with an infrared grid to register animal activity, scales for the water bottle and feeder, a running wheel with unrestricted access and rotation counter, an “operant wall” for positively reinforced operant conditioning [23] . There were 16 such cages, housing mice individually. Measures were read once per minute using TSE software. Blood pressure and heart rate in mice, instrumented with telemetry probes were continuously recorded throughout the tests. Operant conditioning sessions were conducted automatically, once a day and started at 22:30, controlled by IntelliMaze software control. The first session was designed to reinforce nosepokes emitted by an animal while both light stimuli (left and right) were active. This session was used to establish individual baseline tendencies to display nosepokes to the right or left. On subsequent days, mice were trained to direct nosepokes to the side opposite to their initial bias. The number of rewarded responses was limited to 50; each sweetened dry milk pellet weighed 14 mg. To promote learning, mice were maintained at 90% ad lib body weight. Access to food was provided throughout the day from 07:30 until 19:00–20:00. Reward pellets were the alternative source of food. The quality of blood pressure signal (for the operated
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 22
|
mice), probabilities of running wheel use, combined with performance in conditioning task were used as criteria for inclusion into experimental groups. Selected mice were randomly assigned to either flight or ground control or vivarium groups. The groups of mice designated for flight were transported to Cosmodrome Baikonur one week before launch. Transportation was by car and by air in several steps and lasted approximately 24 h. Containers of nontransparent blue plastic 40×60 cm with 4 sub-compartments (each for a group of three mice) equipped with air filters (Animal Breeding Facility - Branch of Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry) were used. Paste food was provided to mice during transportation. One-way ANOVA (transportation bodyweight data) followed by Bonferroni's multiple comparisons was performed with transportation bodyweight data. In order the pre/post flight bodyweight data set met the assumptions of normality and homoscedasticity intrinsic to the ANOVA model, rank transformation was applied to the entire set of observations [24] prior to two-way ANOVA followed by Bonferroni post-hoc comparisons; noteworthy, the results of the test on raw and transformed data were identical. Fisher's exact test (survival), Man-Whitney U-test (home cage food and water consumption, open field test behavioral data) or paired Student's t-test
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 23
|
(metabolic chambers data) was performed using GraphPad Prism software (version 6.00 for MacOS, GraphPad Software, USA).
|
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] |
4136787
|
Mice in Bion-M 1 Space Mission: Training and Selection
|
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4136787/
| 24
|
The study was approved by IACUC of MSU Institute of Mitoengineering (Protocol № 35, 1 November, 2012) and of Biomedical Ethics Commission of IBMP (protocol № 319, 4 April, 2013) and conducted in compliance with the European Convention for the Protection of Vertebrate Animals used for Experimental and Other Scientific Purposes [13] .
|
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