Search

The role of the Earth's gravitational and magnetic fields in the evolution and maintenance of normal processes of various animal species remains unclear. The aim of this work was to determine the effect of simulated microgravity and hypomagnetic conditions for 1, 3, and 6 h on the sperm motility of the fruit fly Drosophila melanogaster . In addition to the usual diet, the groups were administered oral essential phospholipids at a dosage of 500 mg/kg in medium. The speed of the sperm tails was determined by video recording and analysis of the obtained video files, protein content by western blotting, and cell respiration by polarography. The results indicated an increase in the speed of movement of the sperm tails after 6 h in simulated microgravity. The levels of proteins that form the axoneme of the sperm tail did not change, but cellular respiration was altered. A similar effect occurred with the administration of essential phospholipids. These results may be due to a change in the level of phosphorylation of motor proteins. Exposure to hypomagnetic conditions led to a decrease in motility after 6 h against a background of a decrease in the rate of cellular respiration due to complex I of the respiratory chain. This effect was not observed in the flies that received essential phospholipids. However, after 1 h under hypomagnetic conditions, the rate of cellular respiration also increased due to complex I, including that in the sperm of flies receiving essential phospholipids.

The paper presents experimental data on the influence of a 1000-fold weakening of the Earth's magnetic field on the embryonic and postembryonic development of the Japanese quail in three generations. It has been shown that the weakening of the earth's magnetic field by a factor of 1000 affects the formation of blood vessels in Japanese quail embryos, in particular, causing a decrease in angiogenesis in seven-day-old embryos of both the first generation (F 1 ) and the next two ones (F 2 and F 3 ). Pathological and anatomical studies of embryos of different ages in three generations have revealed various pathologies associated with vascular system disorders, as well as disorders in the development of the beak and eyes. In the ontogenesis of F 3 quails, there is a decrease in the hatchability of chicks.

The paper presents the results of experiments with spore-forming bacteria and microscopic fungi performed in the framework of the Russian Research Program outside the International Space Station. It has been found that microorganisms not only survive in this extreme environment, but also retain reproductive ability. Moreover, most microorganisms exhibit an increase in biochemical activity and resistance to antimicrobial agents, specifically antibiotics. These findings are of obvious interest to the developers of both planetary quarantine methods and biomedical safety systems for manned space exploration missions. In addition, they demonstrate the necessity of experiments on the exposure of bio-objects to simulated environmental factors beyond Earth's magnetosphere.

To analyze the effect of gravity on the structure of germinal tissues, we examined tissues of the testes and duct deferens of mice that were exposed to space flight conditions for 21-24 days (experiment Rodent Research-4, SpaceX-10 mission, February 2017, USA). We evaluated the levels of cytoskeletal proteins, sperm-specific proteins, and epigenetic events; in particular, we evaluated levels of 5-hydroxymethylcytosine and of enzymes that regulate DNA methylation/demethylation. We did not detect changes in the levels of cytoskeletal proteins, sperm-specific proteins, DNA-methylases, DNA demethylases, DNA acetylases, or histone deacetylases. However, there were changes at the gene expression level. In particular, there was an increase in the demethylase Tet2 and a decrease in the histone deacetylase Hdac1. These gene expression changes may be of key importance during the early period of readaptation since they could lead to an increase in the expression of target genes.

The ontogenesis and reproduction of plants cultivated aboard a spacecraft occur inside the unique closed ecological system wherein plants are subjected to serious abiotic stresses. For the first time, a comparative molecular cytogenetic analysis of Pisum sativum L. ( Fabaceae ) grown on board the RS ISS during the Expedition-14 and Expedition-16 and also plants of their succeeding (F1 and F2) generations cultivated on Earth was performed in order to reveal possible structural chromosome changes in the pea genome. The karyotypes of these plants were studied by multicolour fluorescence in situ hybridization (FISH) with five different repeated DNA sequences (45S rDNA, 5S rDNA, PisTR-B/1, microsatellite motifs (AG) 12 , and (GAA) 9 ) as probes. A chromosome aberration was revealed in one F1 plant. Significant changes in distribution of the examined repeated DNAs in karyotypes of the "space grown" pea plants as well as in F1 and F2 plants cultivated on Earth were not observed if compared with control plants. Additional oligo-(GAA) 9 sites were detected on chromosomes 6 and 7 in karyotypes of F1 and F2 plants. The detected changes might be related to intraspecific genomic polymorphism or plant cell adaptive responses to spaceflight-related stress factors. Our findings suggest that, despite gradual total trace contamination of the atmosphere on board the ISS associated with the extension of the space station operating life, exposure to the space environment did not induce serious chromosome reorganizations in genomes of the "space grown" pea plants and generations of these plants cultivated on Earth.

The purpose of this work was to evaluate the protein and mRNA expression levels of multiple cytoskeletal proteins in the cardiac and lung tissue of mice that were euthanized onboard the United States Orbital Segment of the International Space Station 37 days after the start of the SpaceX-4 mission (September 2014, USA). The results showed no changes in the cytoskeletal protein content in the cardiac and lung tissue of the mice, but there were significant changes in the mRNA expression levels of the associated genes, which may be due to an increase in total genome methylation. The mRNA expression levels of DNA methylases, the cytosine demethylases Tet1 and Tet3, histone acetylase and histone deacetylase did not change, and the mRNA expression level of cytosine demethylase Tet2 was significantly decreased.

Unlabelled: Mice were exposed to 1 month of space flight on the Russian biosatellite BION-M1 to determine its effect on the expression of genes involved in the maintenance of the mouse brain dopamine system. The current article focuses on the genes encoding glial cell line-derived neurotrophic factor (GDNF) and cerebral dopamine neurotrophic factor (CDNF). Space flight reduced expression of the GDNF gene in the striatum and hypothalamus but increased it in the frontal cortex and raphe nuclei area. At the same time, actual space flight reduced expression of the gene encoding CDNF in the substantia nigra but increased it in the raphe nuclei area. To separate the effects of space flight from environmental stress contribution, we analyzed expression of the investigated genes in mice housed for 1 month on Earth in the same shuttle cabins that were used for space flight and in mice of the vivarium control group. Shuttle cabin housing failed to alter the expression of the GDNF and CDNF genes in the brain structures investigated. Thus, actual long-term space flight produced dysregulation in genetic control of GDNF and CDNF genes. These changes may be related to downregulation of the dopamine system after space flight, which we have shown earlier. © 2015 Wiley Periodicals, Inc., Significance: Our results provide the first evidence of microgravity effects on expression of the GDNF and CDNF neurotrophic factor genes. A considerable decrease in mRNA level of GDNF and CDNF in the nigrostriatal dopamine system was found. Because both GDNF and CDNF play a significant role in maintenance and survival of brain dopaminergic neurons, we can assume that this dysregulation in genetic control of GDNF and CDNF genes in substantia nigra could be among the reasons for the deleterious effects of space flight on the dopamine system., (© 2015 Wiley Periodicals, Inc.)

Mice of C57BL/6J strain were exposed to 1-month spaceflight on Russian biosatellite Bion-M1 to determine the effect of long-term actual spaceflight on the expression of genes involved in the processes of neurogenesis and apoptosis. Specifically, we focused on the genes encoding proapoptotic factor BAX, antiapoptotic factor BCL-XL, brain-derived neurotrophic factor (BDNF) and BDNF receptors TrkB and p75. Spaceflight reduced the expression of the antiapoptotic BCL-XL gene in the striatum and hypothalamus, but increased it in the hippocampus. To estimate environmental stress contribution into spaceflight effects we analyzed spaceflight-responsive genes in mice housed for 1 month on Earth in the same shuttle cabins that were used for spaceflight, and in mice of the laboratory control group. It was shown that 1-month shuttle cabin housing decreased BCL-XL gene expression in the striatum but failed to alter BCL-XL mRNA levels in the hippocampus or hypothalamus. Spaceflight failed to alter the expression of the proapoptotic BAX gene in all investigated brain structures, although the insignificant increase of the BAX mRNA level in the hippocampus of spaceflight mice was found. At the same time, shuttle cabin housing produced insignificant decrease in BAX gene expression in the hippocampus. In contrast to the BCL-XL gene, genes encoding BAX, BDNF as well as TrkB and p75 receptors did not respond to 30-day spaceflight. Thus, long-term spaceflight (1) did not affect the expression of genes encoding BDNF as well as TrkB and p75 receptors, (2) produced dysregulation in genetic control of the neuronal apoptosis, (3) implicated BCL-XL as the risk factor for spaceflight-induced behavioral abnormalities., (Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.)

Background: Spaceflight environment have been shown to generate reactive oxygen species (ROS) and induce oxidative stress in plants, but little is known about the gene expression of the ROS gene network in plants grown in long-term spaceflight. The molecular response and adaptation to the spaceflight environment of Mizuna plants harvested after 27 days of cultivation onboard the International Space Station (ISS) were measured using genome-wide mRNA expression analysis (mRNA-Seq)., Results: Total reads of transcripts from the Mizuna grown in the ISS as well as on the ground by mRNA-Seq showed 8,258 and 14,170 transcripts up-regulated and down-regulated, respectively, in the space-grown Mizuna when compared with those from the ground-grown Mizuna. A total of 20 in 32 ROS oxidative marker genes were up-regulated, including high expression of four hallmarks, and preferentially expressed genes associated with ROS-scavenging including thioredoxin, glutaredoxin, and alternative oxidase genes. In the transcription factors of the ROS gene network, MEKK1-MKK4-MPK3, OXI1-MKK4-MPK3, and OXI1-MPK3 of MAP cascades, induction of WRKY22 by MEKK1-MKK4-MPK3 cascade, induction of WRKY25 and repression of Zat7 by Zat12 were suggested. RbohD and RbohF genes were up-regulated preferentially in NADPH oxidase genes, which produce ROS., Conclusions: This large-scale transcriptome analysis revealed that the spaceflight environment induced oxidative stress and the ROS gene network activation in the space-grown Mizuna. Among transcripts altered in expression by space conditions, some were common genes response to abiotic and biotic stress. Furthermore, certain genes were exclusively up-regulated in Mizuna grown on the ISS. Surprisingly, Mizuna grew in space normally, as well as on the ground, demonstrating that plants can acclimate to long-term exposure in the spaceflight environment by reprogramming the expression of the ROS gene network.

Purpose of the BION-M1 project was laying the evidence and technological basis for addressing the medical issues of future remote space exploration missions by humans. The program of researches with the use of mice was focused on elicitation of cellular and molecular mechanisms of the muscular, cardiovascular and immune reactions to extended exposure in microgravity. The comprehensive studies combined lifetime measurements with investigations of mice tissues and cells by dint of the cutting-edge morphological, biochemical and molecular biology techniques. Males of mice C57/BL6 aged 4 to 5 months were chosen as the object of studies. They were distributed into the flight, ground control and two vivarium (laboratory control) groups and investigated immediately on return and after 7 days of readaptation. Some of the physiological functions were recorded throughout the flight. To ensure wellbeing of the animals in the experiments and to enhance data quality, prior to launch the mice were specially trained so as to accommodate to the group living, eating space food, and in-flight stress factors. Those of the mice that were designated for lifetime investigations were tested and received training pre-launch.

One of the crucial issues of handling animals in space flight is availability of food and water supply systems. However, water supply for animal experiments onboard unmanned spacecrafts is a particularly formidable problem. To maintain laboratory mice in space flight, a paste-type food was formulated on the basis of the standard extruded combined food for laboratory rodents (PK-120) with a 18.3% wet protein content and moisture raised to 68%. Food manufacturing technology for rodents reproduces essentially the one for quails that had shown good advantages in an experiment with adult birds aboard orbital complex Mir. The proposed food was tested with white laboratory mice (males and females) of mature 37-d age. According to the data of blood clinical analysis and visceral organs morphology investigations, feeding with the paste-type food without additional water over 21 days did not change the life weight of mice or food digestibility and availability. These biological test results gave go to feed mice in the Bion-M1 and synchronous ground experiments with the paste-type food.

Biosatellite BION-M1 was launched on April 19 and landed on May 19, 2013. The mission program was largely a continuation of the earlier flown 11 BION projects, FOTON-M2 and FOTON-M3. The biosatellite was inhabited by a great variety of living organisms used for experiments and studies in gravitational physiology, gravitational biology, biotechnology, astrobiology and radiation biology, dosimetry and spectrometry. This was the first time in the history of national biology and physiology when male mice C57bl/6 were chosen for a long-term space experiment focused upon molecular biology investigations. Unfortunately, because of technical failures during the flight a part of the animals were lost. However, the major objectives were attained through reconsideration of biomaterial division among investigators and completion of virtually the total scope of investigations.

The article reports the results of comparative histological studies of skeleton development in 14- and 16-day Japanese quail embryos grown in space flight and in the 1-g gravity. The investigation revealed retardation of cartilage replacement by bone in the femur and tibia in space embryos as compared with their controls. Perichondral ossification metaphysis was reached by day 14 both in the space and control embryos. Destruction of cartilaginous diaphysis advanced onto the proximal and distal parts including the periphery. Tibia and femur cartilaginous cores in space embryos were destructed worse than in the controls in consequence of insufficient minerals supply. Perichondral ossification in the 16-day space and control embryos was alike close to completion spreading as far as the epiphysis. Long bones metaphysis was abundant in monomorphic cells as a growth bank existing during and 7 days post hatching. However, absence, in contrast to the controls, of osteogensis sites in long bones epiphysis, suggested retardation of chondrocytes calcification in these areas, as well as of ossification in space embryos.

Results of studying spinal marrow histogenesis in age varying Japanese quail embryos and chicklings whose embryonic development took place in microgravity are presented. Histological comparison of spinal cords showed retardation of morphogenetic development, lumber spine especially, in the group of flight embryos manifested as incomplete proliferative activity during migration processes and nervous cell differentiation. Like on earth, in space microgravity chicklings also hatched on day 17 of embryogenesis. Multiform marrow changes in space chicklings were a nervous cells reaction to the functional state of organism in the micro-g environment.

Over the past 22 years, the Institute of Biomedical Problems has stubbornly continued the investigations with higher plants aimed at the development of cultivation technologies suitable for the conditions of space flight. Analysis of the results of 24 plant experiments performed aboard orbital complex MIR and the ISS Russian segment evidenced the ability of higher plants to grow, develop and reproduce inside spacecraft living compartments. Space crops were normal as compared with the laboratory controls. Microbial contamination of the plants was within the normal limits; no pathogen has been detected on plant surfaces. Plants did not change genetically, at least in four space generations. It should be noted that the presence of greenhouse on board the ISS also has a marked positive effect on wellbeing of people living in the close environment and isolation from Earth's biosphere. In the context of the above, the higher plants might become a secure and beneficial part of the life support system for crews on space exploration missions.

The article presents the results of a comparative histological investigation of skeletal bones genesis in Japanese quail embryos developed in the spaceflight microgravity (space group) and laboratory (control group). Total preparations of 4-day-old embryos from both groups demonstrated clearly that the cartilaginous anlage of the femoral bone had central, dyaphisial, 2 epiphysial and 2 proliferation zones. By day 7 of embryogenesis, cartilaginous anlages had grown in size in both groups due to intensive chondrocytes multiplication and gain in the intercellular substance mass. Tibial cuff in space embryos measured half and femoral cuff was 2.3 times smaller in comparison with these parameters in the control group. In addition, intensity of chondrocyte multiplication was reduced Histological profiles of the femur and tibia in 10-day old embryos of the control pointed to enhancement of osteogenesis. The metaphysis zone contained distinct mitosis figures on different stages of division. Bone deposition could be seen below the peristoma. The osteogenesis cuff spread up to the femoral anlage metaphysis; cartilage was calcined. Space embryos display retard osteogenesis. There were ingrown blood vessels in the region of cartilage destruction; however, vessels grown in the periosteum were less in number as compared with the laboratory control. Also, the perichondral ossification layer was considerably thinner, whereas the osseous cuff was 1.3 and 1.45 times shorter in the femur and tibia, respectively. To sum up, the histological investigation of bones from 4-, 7- and 10- day old Japanese quail embryos demonstrated retardation of osteogenesis in the conditions of microgravity.

Tibia and femur osteogenesis was studied in embryonic lower limbs developed in microgravity. The maximal difference in ash content of shell in the flight and control groups was registered on days 4, 10 and 14 amounting to 10.27; 9.56 and 12.95% respectively. Shell analysis for calcium showed the largest difference between the groups on day 4 (8.94 mg). However, this difference was not seen already on days 14 and 16. Hence, according to the results of the investigation, although shell calcium utilization and osteogenesis in flight embryos of the Japanese quail were retarded, by the time of hatching the morphological and chemical parameters of the lower limb bones were essentially same as of the ground controls.

The article reports the results of histogenetic studies of the liver extracted from Japanese quail embryos and hatchlings that had passed the embryonic and postnatal development in microgravity. Comparison of the liver from the flight and laboratory embryos showed identity of organogenesis and histogenesis processes in space and on Earth. However, it should be noted that early histogenetic development of the space embryo liver had been retarded which was concluded from an enlarged lumen in bile capillaries and loose arrangement of epithelium girders as compared with the control On day-10 post flight, the flight embryos exhibited liver induration through narrowing of sinusoids lumen. Haemopoietic zones became less numerous. By the end of embryonic development these histological differences of the flight and control liver disappeared essentially. Organogenesis and histogenesis of the liver obtained from the hatchlings that had spent 5 days in microgravity did not depart from the norm. However, inability of the birds to adapt to microgravity and ensuing starvation was the reason for dystrophic changes in the liver, venous congestion and increased histoleucocytic activity. All the liver changes faded away soon alter the hatchlings were settled in favorable life conditions

Transcriptome of barley Hordeum vulgare grown aboard International Space Station (ISS) was analyzed by means of microarray. It was revealed 500 genes with mRNA level, changed more than two folds in space environment. Among them are genes encoding stress response proteins, videlicet Heat Shock Proteins (HSP), Pathogenesis-Related Proteins (PR) and Antioxidant Proteins. Further analysis of these genes by real time PCR showed enhanced transcription level of Reactive oxygen Species (ROS) scavenging genes. The mRNA level of superoxide dismutase (sod) was 6 folds higher in space environment when compare to Earth conditions. Glutamyl transferase gene expression was enhanced 24 times in space. Transcription of catalase gene (cat) was increased 18 times and of ascorbate peroxidase was increased 3 times in space in comparison with ground control. For the first time it was shown that space flight environment may induce oxidative stress in plants.

The work was performed in order to evaluate possible negative effect of high temperatures on biological properties of seeds in space experiment BIORISK-MSN-2. Laboratory experiments showed that seeds possess high enough resistance to extended exposure to high temperatures. Results of studying effects of positive temperatures in the region of 40 to 100 degrees C on biological properties of seeds of 19 species and varieties of higher plants belonging to three different families lead to the conclusion that heat stability of seeds depends on temperature value, period of exposure, species, variety and individual peculiarities of plants. These data can be useful in future interpretation of results of experiment BIORISK-MSN-2.

The gene expression and enzyme activity of superoxide dismutase, catalase, and ascorbate peroxidase in the space-grown barley were not significantly different from those of the ground-grown barley. Cu2+ reducing and radical scavenging activities in an extract of the space-grown barley were lower than those of the ground-grown barley by 0.7 fold, suggesting that the space environment does not induce oxidative stress, and reduces antioxidant capacity in plants.

The paper reports the results of organogenic and histogenic investigations of the visceral organs of embryonic Japanese quails incubated in the microgravity aboard orbital complex Mir. Investigations of the gastrointestinal tract failed to reveal macrodeviations in the organs' structure, differentiation of layers or cells along the full length of the entodermal canal. However, comparison with the ground controls exhibited poor development of stromal connective tissues in the flight embryos evidencing loose arrangement and small number of fibers. Local hyperplasia in the duodenal epithelium was due to the proliferation rather than differentiation processes; it could affect food intake and parietal digestion in the flight chicks. Though the Japanese quail embryos developed in space microgravity had some deviations, their digestic apparatus was mature to uptake and assimilate food.

One of the challenges for space biology and medicine is resolution of lots of problems of biomedical support of humans in the extreme environment of space flight. These problems include also designing of robust and efficiently functioning life support systems (LSS). The paper gives an overview of the investigations of ground-based BLSS with human subjects conducted in Russia and other countries. Also, it contains the basic data of studying the BLSS photoautotrophic components (higher plants) in the series of experiments with the total duration of 630 days fulfilled on orbital complex Mir and the series of experiments with the total duration of 820 days in the ISS Russian segment. Analysis of the results from the BLSS investigations on Earth and in space flights drives to the conclusion that some of the BLSS components, greenhouses specifically, can be integrated even now into the currently used systems of space crew life support.

Experimental data on the impact of argon-containing hypoxic breathing mixtures on the early embryonic growth of Japanese quail (Coturnix coturnix japonica dom.) are presented. The results showed that after 4-day incubation of eggs in an oxygen-nitrogen breathing mixture (13.8-15.0% O2, 86.2-85.0% N2) only 16.7% embryos reached the stage of 3.5-d development. Introduction in the hypoxic mixture of argon (55% of the total volume) increased the number of embryos on the stage of 3.5-4 day of normal morphological development to 60%. It became evident from the analysis that embryos developed in the O2-Ar-N2 hypoxic mixture had 25% less pathologies as compared with embryos developed in a largely nitrogen-containing gas mixture. Hence, we received the experimental evidence that 55% of argon in a hypoxic breathing mixture with up to 15% of oxygen is favorable to the early embryonic growth of Japanese quail stimulating metabolic processes in germ organism.

Results of studies on growth and development of offspring of two genetically marked dwarf pea lines planted during the whole ontogenesis cycle in the Lada space greenhouse on board of Russian Segment of International Space Station (RS ISS) are presented. The offspring of M1 and M2 plants grown from seeds formed during space flight was examined under conditions of Earth-based. Cultivation. It had been shown that growth and developmental characteristics, frequency of chromosome aberrations in primary root meristem and level of molecular polymorphism revealed in individual plants via RAPD method show no significant differences between offspring of "space-grown" and control seeds.

Effects of a prenatal rhythmic tone on behavior of Japanese quail chicks were studied on the next days after hatching. The acoustically stimulated chicks demonstrated a significantly more frequent following response, feeding reaction and comfortable behavior when hearing the tone as compared with their controls. Bandwidths stimulating this or another behavior were determined.

There are several experimental models of biological life support systems (BLSS) designed to incorporate a chlorella pool. These BLSS can be optimized if populated by algal associations that could take up more functions within the closed cycling system than a single alga species. Introduction of a Spirulina and Chlamydomonas poly-culture with differing in gas exchange and biochemical composition resulted in a tighter closure of linkages within the system. The factors determining the size of a species population in intensive continuous poly-cultures are, first and foremost, pH and suspension flow rate. Experimental testing of this supposition brought us to the conclusion that parametric control of alga productivity and species composition dynamics makes it possible to create a steady intensive poly-culture as part of the LSS for humans. Flow rate and pH can be the parameters for control of the Spirulina and Chlamydomonas populations during continuous cultivation of this poly-culture.

Time of the eating reaction (response to video signals) and eating behavior shortly after hatching were studied in Japanese quail chicks whose embryonic development took place under normal or changed gravity. Chicks partially incubated in a changed gravity showed a much slower eating reaction when compared with the chicks the prenatal development of which occurred under the normal gravity. In the chicks incubated at 1 g and placed in individual cages immediately after dominating afferentation for the eating behavior was visual Observations in the study will be used as a basis for designing a technology for handling and maintenance of hatchlings of the Japanese quail as a potential heterotrophic component of space life support systems.

In the period between March 2003 and April 2005 five crops of genetically marked dwarf pea were cultivated in greenhouse LADA on the ISS Russian segment to study morphology and genetics in consecutive generations. Results of the first space experiment were analyzed and characteristics of ground plants grown from space seeds were studied. It was shown that parameters of growth, development and genetic status of pea plants that completed the ontogenetic cycle in LADA did not differ substantially from the laboratory control and that plants cultivated in space flight do not incur losses in the reproductive function and form viable seeds. Genetic analysis of plants from the first generation of space and ground seeds using the methods of RAPD-primers (10 markers) and chromosomal aberration analysis failed to reveal genetic polymorphism which means that the spaceflight factors had no effect on the genetic apparatus of the first generation of space-grown plants.

Five ground-based experiments at RF SRC-IBMP had the purpose to make a thorough investigation of a model of the human-unicellular algae-mineralization life support system. The system measured 15 m3 and contained 45 liters of alga suspension; the dry alga density was 10 to 12 g/l and water volume (including the alga suspension) amounted to 59 l. More sophisticated LSS models where algae were substituted by higher plants (crop area in the greenhouse equaled 15 m2) were investigated in three experiments from 1.5 mos. to 2 mos. in duration. It was found that the alga containing LSS was able to fulfill not only the macrofunction (air and water regeneration) but also several additional functions (air purification, establishment of microbial cenosis etc.) providing an adequate human environment. This polyfunctionality of the biological regenerative processes is a weighty argument for their integration into space LSSs. Another important aspect is that the unicellular algae containing systems are highly reliable owing to a huge number of species-cells which will be quickly recovered in case of the death of a part of the population and, consequently, functionality of the LSS autotrophic component will be restored before long. For an extended period of time the Martian crew will have no communication with the Earth's biosphere which implies that LSS should be absolutely reliable and redundant. Redundancy can be achieved through installation aboard the vehicle of two systems constructed on different principles of regeneration, i.e. physical-chemical and biological. Each of the LSSs should have the power to satisfy all needs of the crew. The best option is when two systems are functioning in parallel sharing the responsibility for the human environment. Redundancy in this case will mean that in the event of failure or a drastic decrease in performance of one system the other one will make up for the loss by increasing its share in the overall regeneration process.

Ground-based experiments at RF SSC-IBMP RAS (State Science Center of Russian Federation--Institute of Biomedical Problems of Russian Academia of Science) were aimed at overall studies of a human-unicellular algae-mineralization LSS (life support system) model. The system was 15 m3 in volume. It contained 45 L of algal suspension with a dry substance density of 10-12 g per liter; water volume, including the algal suspension, was 59 L. More sophisticated model systems with partial substitution of unicellular algae with higher plates (crop area of 15 m2) were tested in three experiments from 1.5 to 2 months in duration. The experiments demonstrated that LSS employing the unicellular algae play not only a macrofunction (regeneration of atmosphere and water) but also carry some other functions (purification of atmosphere, formation of microbial cenosis etc.) providing an adequate human environment. It is also important that functional reliability of the algal regenerative subsystem is secured by a huge number of cells able, in the event of death of a part of population, to recover in the shortest possible time the size of population and, hence, functionality of the LSS autotrophic component. For a long period of time a Martian crew will be detached from Earth's biosphere and for this reason LSS of their vehicle must be highly reliable, robust and redundant. One of the approaches to LSS redundancy is installation of two systems with different but equally efficient regeneration technologies, i.e. physical-chemical and biological. At best, these two systems should operate in parallel sharing the function of regeneration of the human environment. In case of failure or a sharp deterioration in performance of one system the other will, by way of redundancy, increase its throughput to make up for the loss. This LSS design will enable simultaneous handling of a number of critical problems including adequate satisfaction of human environmental needs., (c2003 COSPAR. Published by Elsevier Science Ltd. All rights reserved.)

This work is a part of comprehensive research into the effects of space flight on Japanese quail ontogenesis. Analysis of eye morphogenesis in the embryonic Japanese quails developed in microgravity discovered considerable deviations and abnormalities. Ocular abnormalities in the embryonic quail were mainly micro-ophthalmic and associated with disproportional growth of the pigmental epithelium and neural retina which resulted in plication and a broken sandwich structure of the retina.

The purpose was to study characteristic features of growth and development of several plant generations in space flight in experiment GREENHOUSE-3 as a part of the Russian-US space research program MIR/NASA in 1997. The experiment consisted of cultivation of Brassica rapa L. in board greenhouse Svet. Two vegetative cycles were fully completed and the third vegetation was terminated on day 13 on the phase of budding. The total duration of the space experiment was 122 days, i.e. same as in the ground controls. In the experiment with Brassica rapa L. viable seeds produced by the first crop were planted in space flight and yielded next crop. Crops raised from the ground and space seeds were found to differ in height and number of buds. Both parameters were lowered in the plants grown from the space seeds. The prime course for smaller size and reduced organogenic potential of plantTs reproductive system seems to be a less content of nutrients in seeds that had matured in the space flight. Experiment GREENHOUSE-3 demonstrated principle feasibility of plant reproduction in space greenhouse from seeds developed in microgravity.

Since 1990, the orbital complex MIR has witnessed several incubator experiments for determination of spaceflight effects on embryogenesis of Japanese quail. First viable chicks who had completed the whole embryological cycle in MIR microgravity hatched out in 1990; it became clear that newborns would not be able to adapt to microgravity unaided. There were 8 successful incubations of chicks in the period from 1990 to 1999. In 1995-1997 the MIR-NASA space science program united Russian and US investigators. As a result, experiments Greenhouse-1 and 2 were performed with an effort to grow super dwarf wheat from seed to seed, and experiment Greenhouse-3 aimed at receiving two successive generations of Brassica rapa. But results of these experiments could not be used for definitive conclusions concerning effects of spaceflight on plant ontogenesis and, therefore, experiments Greenhouse-4 and 5 were staged within the framework of the Russian national space program. The experiments finally yielded wheat seeds. Some of the seeds was left on the space station and, being planted, gave viable seedlings which, in their turn, produced the second crop of space seeds., (c 2001. COSPAR. Published by Elsevier Science Ltd. All rights reserved.)

Results of the experiment aimed at harvesting a second space generation of wheat var. Apogee in Mir greenhouse Svet (experiment GREENHOUSE-5) are presented. In space flight, germination rate of space seeds from the first crop made up 89% against 100% of the ground seeds. The full biological ripeness was observed in 20 plants grown from the ground seeds and one plant grown from the space seeds following 80- to 90-d vegetation. The plant of the second space generation was morphologically different neither from the species in the first space crop nor from the ground controls. To study the biological characteristics of Apogee seeds gathered in the first and second crops in spaceflight experiment GREENHOUSE-5, the seeds were planted on their return to the laboratory. Morphometric analysis showed that they were essentially similar to the controls. Hence, the space experiments in Mir greenhouse Svet performed during 1998-1999 gave proof that plants cultivated in microgravity can pass the ontogenetic cycle more than once. However, initial results of the investigations into growth and development of plants through several generations are still in-sufficient to speak of possible delayed effects of the spaceflight factors (microgravity, multicomponent radiation, harmful trace contaminants etc.).

Laboratory experiments were performed to study cytoembryologic development of regenerative organs of wheat cult. 20-1-Shortstem and cult. Apogee in atmosphere with elevated concentration of ethylene. Development of wheat regenerative organs at a concentration of air ethylene of 1 mg/m3 resulted in peculiar changes in floret morphogenesis and partial or complete male barrenness. Cult. Apogee was found to be least vulnerable and, therefore, appears to be suitable for cultivation in greenhouses mounted in pressurized habitats including space station modules. Vulnerability of male reproductive organs in short-stem wheat to ethylene correlates convincingly with the sign of dwarfishness demonstrated by hybrid forms of one of praparents.

Data of the cytoembryological analysis of plants of super-dwarf wheat grown in laboratory experiment with elevated ethylene content in atmosphere were compared with results of space experiment and the ground control with ambient ethylene concentration. It was shown that crop barrenness in both the space and ground experiments was caused by exposure to increased ethylene throughout the whole ontogenesis which made pollen nonviable. The cytogenetic analysis revealed that male sterility of the wheat flowerlets in space flight evolved, as a rule, earlier and was deeper by character, i.e. pollen did not progress farther than the one-nucleus phase, whereas in the laboratory experiment with ethylene the pollen grains reached sometimes even the three-nucleus phase.

We plan to perform space experiments on development of a technology for cultivation of leaf vegetables that might be a component of future life support systems for space crews. For this purpose, we are going to fly research greenhouses with the crop area from 0.03 up to 0.1 m2 inside the universal docking module of the ISS Russian segment. To prepare for future space experiments, ground investigations were made in order to compare crop capacity of various artificial soils used to grow leaf vegetables in greenhouse Svet. Useful life of root module Svet can be extended with a new technology based on resupply of fibrous substrate with nutrients. The most effective volume of soil per a plant was determined which sustains high productivity of leaf vegetables in Svet. To select leaf vegetables for in-space cultivation, we conducted investigations of productivity, morphometric and biochemical characteristics, and palatability of 18 cultures including alternative greens highly popular in Japan and China which have been earlier tested neither in laboratory nor in space within the closed eco-system projects. We would prioritize mizuna, pak choi, tatsoi, rapina or broccoli raab, and red giant mustard as objects for in-space investigations.

The hypothesis being tested is that Super Dwarf wheat, Triticum aestivum L., plants in the Svet Greenhouse onboard the Russian Space Station Mir will complete a life cycle in spaceflight, providing that the environmental conditions necessary for adequate growth on Earth are supplied. Twenty six seeds of wheat were planted in each of 2 rows of 2 root compartments for a total of 104 seeds in Svet. Germination rate at 7 d was 56 and 73% on Mir and 75 and 90% in ground-based controls. Plants were grown throughout the whole cycle of ontogenesis (123 d) with samples gathered at different times to validate the morphological and reproductive stages of the plants. Young plants showed vigorous early seedling growth, with large biomass production, including the formation of 280 floral spikes. Upon return to Earth, comparative analyses showed that the number of tillers and flowers per spikelet were 63.2% and 40% greater, respectively, in Mir-grown plants than in the controls. By contrast, the stem length (52.4%), spike mass (49.2%) and length (23.1%), awn length (75.7%), number of spikelets per spike (42.8%) and number of seeds per spike (100% sterile) from Mir-grown plants were substantially less than the controls. Distribution of moisture and roots throughout the substrate was very good. All florets on Mir-grown spikes ceased development at the same stage of ontogeny. Lack of caryopses formation was attributed to male sterility occurring at different stages of staminal development. Anthers failed to dehisce and pollen grains were smaller and shriveled compared to the controls, suggesting a chronic stress had occurred in the Svet growth chamber. Recent ground-based studies indicated that ethylene, which was measured at 0.3 to 1.8 mg kg-1 in the Mir, almost certainly could have induced male sterility in the wheat plants grown on the Mir.

Growth of higher plants in the microgravity environment of orbital platforms has been problematic. Plants typically developed more slowly in space and often failed at the reproductive phase. Short-duration experiments on the Space Shuttle showed that early stages in the reproductive process could occur normally in microgravity, so we sought a long-duration opportunity to test gravity's role throughout the complete life cycle. During a 122-d opportunity on the Mir space station, full life cycles were completed in microgravity with Brassica rapa L. in a series of three experiments in the Svet greenhouse. Plant material was preserved in space by chemical fixation, freezing, and drying, and then compared to material preserved in the same way during a high-fidelity ground control. At sampling times 13 d after planting, plants on Mir were the same size and had the same number of flower buds as ground control plants. Following hand-pollination of the flowers by the astronaut, siliques formed. In microgravity, siliques ripened basipetally and contained smaller seeds with less than 20% of the cotyledon cells found in the seeds harvested from the ground control. Cytochemical localization of storage reserves in the mature embryos showed that starch was retained in the spaceflight material, whereas protein and lipid were the primary storage reserves in the ground control seeds. While these successful seed-to-seed cycles show that gravity is not absolutely required for any step in the plant life cycle, seed quality in Brassica is compromised by development in microgravity.

In our earlier space experiment with super dwarf wheat we found the spikes developed in space to be barren. The cause of the full crop sterility was sensitivity of this wheat species to the ethylene concentration of 0.3-0.8 mg/m3 during the experiment. The follow-up ground experiments were made to identify species of dwarf wheat that could be raised in space greenhouse Svet and are distinguished by partial tolerance of their reproductive organs to elevated ethylene in air. The choice fell on the USU-Apogee cultivar specially developed for planting in growth chambers as an integral part of various bioregenerative life support systems, including the space ones. An experiment with wheat Apogee was performed in greenhouse Svet on board MIR. The period of the full crop vegetation cycle was not significantly altered under the spaceflight conditions. The experiment yielded 508 seeds from 12 plants, i.e. by 38% less than in laboratory experiments and by 69% more as compared with results of growing crops in ethylene-contaminated atmosphere (1 mg/m3). Mass of the space seeds was low if compared with the laboratory crops. This was the first time when the feasibility of gathering seeds from wheat that had passed the whole vegetation cycle in space flight was demonstrated. The experiment will give mightly impetus to the advancement of research on space biological LSS and gravitational biology.

Metabolism of sLt during Russian-US experiment GREENHOUSE-2 (July 9, 1996-January 17, 1997) within the MIR/NASA space research program and in laboratory Svet experiments in 1995-1996 was studied. Chemical, biochemical and pigment analyses of the flight and laboratory plants were made after the first (dry biomass) and second vegetation (photosynthetically active 41-d old plants). Data on the composition of leaves and stems of ground and flight wheat do not attest any biologically significant shifts in plant metabolism. There were slight changes in accumulation and migration of several macro- and microelements, protein nitrogen and phosphororganic compounds in microgravity. Lowered content of lignin, a critical supportive element for cellular walls was observed only during early stages of vegetation. In the Mir experiment, concentrations of photosynthetically active pigments also decreased a little but the chlorophyills-carotenoids balance was not upset.

In 1996-1997 an experiment with super dwarf wheat (Greenhouse-2) was made aboard the orbital complex MIR as a part of the MIR-NASA space science program. The article deals with the main production and morphometric characteristics of plants that completed their vegetation cycle in the space flight. Lengths of the whole cycle of vegetation and its individual stages were essentially same as in ground control experiments. Dry mass of one plants equal, the number of headed shoots was in 2.7 times less in the flight harvest as compared with the control. The height of shoots was reduced by one half. No seeds were found in the heads formed in space. The architecture of heads was substantially different from what had been observed in the preceeding ground control experiments: mass of the heads was halved and lengths of inflorescence and palea awn shortened. The number of spikelets in a head reduced up to 8-10 vs. 13-14 in the controls, whereas the number of florets per a spikelet averaged 5 vs. 3 in the controls. The experiments showed that mainly the most profound changes in the productive and morphometric parameters of the super dwarf wheat plants were largely caused by the phytotoxic effects of ethylene rather than spaceflight specific factors as its concentrations in the MIR air amount to 0.3-1.8 mg/m3.

From 1989 till 1998 twelve experiments were performed by Bulgarian, Russian, Slovak, and US researchers and engineers on the effects of space flight on the model of ecosystem "algae-fishes-bacteria", and ontogenesis of birds (Japanese quail) and higher plants. For the first time several viable chicks were hatched and passed the whole cycle of their embryonic development in the MIR microgravity. The length of the plant ontogenetic cycle as a whole and its specific stages appeared to be same as on Earth. Seeds of Brassica rapa gathered and planted in greenhouse Svet on board MIR yielded robust shoots. Photosynthesis and dark respiration of plants growing in spaceflight were successfully measured.

The cytoembryologic analysis of wet and dry samples of super dwarf wheat cultivated in greenhouse SVET aboard the MIR station over the whole cycle of vegetation was made with the use of light microscopy. Characteristic features of wheat development in space flight are plentiful early tillering, and formation and rapid growth of side shoots. Elementary spikelets in the composite head were more numerous but the top spikelets were rudimentary and, therefore, the ripe head contained less of these spikelets as compared with the ground controls (9-13 and 14, respectively). The number of florets in a spike was also higher reaching 14-16 vs. 7-8 in the control. Typically, no more than 4 to 5 florets vs. 3 in the control were fully differentiated while the others died off earlier in development. The fact that there were no caryopses found in the flight crop is explained by absolute male sterility appearing at different stages of staminal development: before archesporium formation, on the stage of differentiated archesporium, during meiosis, on the stage of microspores or uninucleate "pollen". The female generative system developed mainly without abnormalities. An assumption was made that elevated ethylene concentrations in the MIR atmosphere at the time of the wheat experiment were the cause for abnormal development of the male generative system which led to barrenness of the super dwarf wheat crop.