Several immunological processes can be affected by space flight. However, there is little evidence to suggest that flight-induced immunological deficits lead to illness. Therefore, one of our goals has been to define models to examine host resistance during space flight. Our working hypothesis is that space flight crews will come from a heterogeneous population; the immune response gene make-up will be quite varied. It is unknown how much the immune response gene variation contributes to the potential threat from infectious organisms, allergic responses or other long term health problems (e.g. cancer). This article details recent efforts of the Kansas State University gravitational immunology group to assess how population heterogeneity impacts host health, either in laboratory experimental situations and/or using the skeletal unloading model of space-flight stress. This paper details our use of several mouse strains with several different genotypes. In particular, mice with varying MHCII allotypes and mice on the C57BL background with different genetic defects have been particularly useful tools with which to study infections by Staphylococcus aureus, Salmonella typhimurium, Pasteurella pneumotropica and Ehrlichia chaffeensis. We propose that some of these experimental challenge models will be useful to assess the effects of space flight on host resistance to infection.
The levels of environmental light experienced by organisms during the behavioral activity phase deeply influence the performance of important ecological tasks. As a result, their shape and coloring may experience a light-driven selection process via the day-night rhythmic behavior. In this study, we tested the phenotypic and genetic variability of the western Mediterranean squat lobster (Munida tenuimana). We sampled at depths with different photic conditions and potentially, different burrow emergence rhythms. We performed day-night hauling at different depths, above and below the twilight zone end (i.e., 700 m, 1200 m, 1350 m, and 1500 m), to portray the occurrence of any burrow emergence rhythmicity. Collected animals were screened for shape and size (by geometric morphometry), spectrum and color variation (by photometric analysis), as well as for sequence variation at the mitochondrial DNA gene encoding for the NADH dehydrogenase subunit I. We found that a weak genetic structuring and shape homogeneity occurred together with significant variations in size, with the smaller individuals living at the twilight zone inferior limit and the larger individuals above and below. The infra-red wavelengths of spectral reflectance varied significantly with depth while the blue–green ones were size-dependent and expressed in smaller animals, which has a very small spectral reflectance. The effects of solar and bioluminescence lighting are discussed as depth-dependent evolutionary forces likely influencing the behavioral rhythms and coloring of M. tenuimana.
The hindlimb unloading rodent model is used extensively to study the response of many physiological systems to certain aspects of space flight, as well as to disuse and recovery from disuse for Earth benefits. This chapter describes the evolution of hindlimb unloading, and is divided into three sections. The first section examines the characteristics of 1064 articles using or reviewing the hindlimb unloading model, published between 1976 and April 1, 2004. The characteristics include number of publications, journals, countries, major physiological systems, method modifications, species, gender, genetic strains and ages of rodents, experiment duration, and countermeasures. The second section provides a comparison of results between space flown and hindlimb unloading animals from the 14-day Cosmos 2044 mission. The final section describes modifications to hindlimb unloading required by different experimental paradigms and a method to protect the tail harness for long duration studies. Hindlimb unloading in rodents has enabled improved understanding of the responses of the musculoskeletal, cardiovascular, immune, renal, neural, metabolic, and reproductive systems to unloading and/or to reloading on Earth with implications for both long-duration human space flight and disuse on Earth.
In response to hypergravity, it appears that the larger the animal, the greater the response, if present. Therefore, the response of a rat exceeds that of a mouse in the same hypergravity environment. When investigated in the microgravity environment of space flight, this appears to hold true. The lack of definitive data obtained in space for either species makes the extrapolation of the continuum to levels below Earth-gravity problematic. However, in systems where responses are detected for both space flight and acceleration by centrifugation, a gravitational continuum is present supporting the "principle of continuity". For those and similar systems, it appears that the use of hypergravity could be used to predict responses to space flight.
Experiments were performed while monkeys flew in space in the ‘‘Cosmos/ Bion’’ Missions to determine the effect of microgravity on the oculomotor and vestibular systems. Eye-head coordination during gaze shifts to lateral targets (gaze fixation reaction, GFR) and multiunit activity in the medial vestibular nuclei (MVN) and cerebellar flocculus were studied in rhesus monkeys in the Bion 6 (Cosmos 1514) through Bion 11 projects. In the first few days of space flight, gaze displacement onto lateral targets became hypermetric, and the amplitude of head movements decreased. This was compensated for by 112
Animal research has been critical to the initiation and progress of space exploration. Animals were the original explorers of "space" two centuries ago and have played a crucial role by demonstrating that the space environment, with precautions, is compatible with human survival. Studies of mammals have yielded much of our knowledge of space physiology. As spaceflights to other planets are anticipated, animal research will continue to be essential to further reveal space physiology and to enable the longer missions. Much of the physiology data collected from space was obtained from the Cosmos (Bion) spaceflights, a series of Russian (Soviet)-International collaborative flights, over a 22 year period, which employed unmanned, free flyer biosatellites. Begun as a Soviet-only program, after the second flight the Russians invited American and other foreign scientists to participate. This program filled the 10 year hiatus between the last US biosatellite and the first animal experiments on the shuttles. Of the 11 flights in the Cosmos program nine of them were international; the flights continued over the years regardless of political differences between the Soviet Union and the Western world. The science evolved from sharing tissues to joint international planning and development, and from rat postmortem tissue analysis to in vivo measurements of a host of monkey physiological parameters during flight. Many types of biological specimens were carried on the modified Vostok spacecraft, but only the mammalian studies are discussed herein. The types of studies done encompass the full range of physiology and have begun to answer "critical" questions of space physiology posed by various ad hoc committees. The studies have not only yielded a prodigious and significant body of data, they have also introduced some new perspectives in physiology. A number of the physiological insights gained are relevant to physiology on Earth. The Cosmos flights also added significantly to flight-related technology, some of which also has application on our planet. In summary, the Cosmos biosatellite flights were extremely productive and of low cost. The Bion vehicles are versatile in that they can be placed into a variety of orbits and altitudes, and can carry radiation sources or other hazardous material which cannot be carried on manned vehicles. With recent advances in sensor, robotic, and data processing technology, future free flyers will be even more productive, and will largely preclude the need to fly animal experiments on manned vehicles. Currently, mammalian researchers do not have access to space for an unknown time, seriously impeding the advancement and understanding of space physiology during long duration missions. Initiation of a new, international program of free flyer biosatellites is critical to our further understanding of space physiology, and essential to continued human exploration of space.
Adaptation is a central precept of biology; it provides a framework for identifying functional significance. We equate mammalian development with adaptation, by viewing the developmental sequence as a series of adaptations to a stereotyped sequence of habitats. In this way development is adaptation. The Norway rat is used as a mammalian model, and the sequence of habitats that is used to define its adaptive-developmental sequence is (a) the uterus, (b) the mother's body, (c) the huddle, and (d) the coterie of pups as they gain independence. Then, within this framework and in relation to each of the habitats, we consider problems of organismal responses to altered gravitational forces (micro-g to hyper-g), especially those encountered during space flight and centrifugation. This approach enables a clearer identification of simple "effects" and active "responses" with respect to gravity. It focuses our attention on functional systems and brings to the fore the manner in which experience shapes somatic adaptation. We argue that this basic developmental approach is not only central to basic issues in gravitational biology, but that it provides a natural tool for understanding the underlying processes that are vital to astronaut health and well-being during long duration flights that will involve adaptation to space flight conditions and eventual re-adaptation to Earth's gravity.
1. Overview (G. Sonnenfeld). 2. The hindlimb unloading rat model: literature overview, technique update, and comparison with space flight data (E. Morey-Holton, R.K. Globus, A. Kaplansky). 3. The international collaboration on Russian spacecraft and the case for free flyer biosatellites (R.E. Grindeland et al.). 4. Mouse infection models for space flight immunology (S.K. Chapes, R.R. Ganta). 5. Vestibular experiments in space (B. Cohen et al.) 6. Effect of space flight on circadian rhythms (G. Tosini, J. Aguzzi). 7. Development as adaptation: a paradigm for gravitational and space biology (J.R. Alberts, A.E. Ronca). 8. The use of animal models to study skeletal effects of space flight (S.B. Doty, T. Wronski, E. Morey-Holton). 9. Responses across the gravity continuum: hypergravity to microgravity (C.E. Wade). 10. Aquatic animals: their contribution to an understanding of gravity effects on life processes (E.R. Horn). 11. Primates in space flight (T.M. Hoban-Higgins, E.L. Robinson, C.A. Fuller).
Gravity related behavior and the underlying neuronal networks are the most suitable model systems to study basic effects of altered gravitational input on the development of neuronal systems. A feature of sensory and motor systems is their susceptibility to modifications of their adequate physical and/or chemical stimuli during development. This discovery led to the formulation about critical periods, which defines the period of susceptibility during post-embryonal development. Critical periods can be determined by long-lasting modifications of the stimulus input for the gravity sensory system (GSS). Techniques include: (1) destruction of the gravity sense organ so that the gravity cannot be detected any longer and the central neuronal network of the GSS is deprived of gravity related information, (2) loading or deloading of parts of the body by weights or counterweights, respectively, which compensates for the gravitational pull, and (3) absence or augmentation of the gravitational environment per se by the exposure of organisms to microgravity during spaceflights or to hypergravity by centrifugation. Most data came from studies on compensatory eye or head movements in the clawed toad Xenopus laevis, the cichlid fish Oreochromis mossambicus, and crickets (Acheta domesticus, Gryllus bimaculatus). The responses are induced by a roll or pitch stimulation of the gravity sense organs, but are also affected by sensory inputs from proprioreceptors and eyes. The development of these compensatory eye and head responses reveals species-specific time courses. Based on experiments using spaceflights, centrifugation, lesion and loading or deloading, all species revealed a significant susceptibility to modifications of the gravity sensory input during development. Behavioral responses were depressed (Xenopus) or augmented (Xenopus, Oreochronis) by microgravity, and depressed by hypergravity except in crickets. In Acheta, however, the sensitivity of its position sensitive neuron PSI was reduced by microgravity. After termination of the period of modified gravity sensory input, all behavioral and physiological modifications disappeared, in some preparations such as the PSI of Acheta or the eye response in Xenopus, however, delayed after exposure to hypergravity. Irreversible modifications were rare; one example were malformations of the body of Xenopus tadpoles caused by lesion induced deprivation. Several periods of life such as the period of hatching or first appearance of gravity related reflexes revealed a specific sensitivity to altered gravity. Although all studies gave clear evidences for a basic sensitivity of developing GSSs to long-lasting modifications of the gravity sensory input, clear arguments for the existence of a critical period in the development of the sense of gravity are still missing. It has to take into consideration that during long-term exposures, adaptation processes take place which are guided by central physiological and genetically determined set points. The International Space Station (ISS) is the necessary platform of excellence if biological research is focussed on the analysis of long-term space effects on organisms.
Fish are the most likely candidates to be the first vertebrate to live their life cycle aboard the International Space Station (ISS). In the space-shuttle experiment using medaka, the fry born in space had the same number of germ cells as the ground control fish, and these germ cells later developed to produce the offspring on the ground. Fry hatched in space did not exhibit any looping behavior regardless of their strain, visual acuity, etc. The aquatic habitat (AQH) is a space habitat designed for long-term breeding of medaka, zebrafish and Xenopus, and recent advancements in this hardware also support fish life-cycle experiments. From the crosses between two strains, fish having good eyesight and less sensitivity to gravity were obtained, and their tolerance to microgravity was tested by parabolic flight using an airplane. The fish exhibited less looping and no differences in degree of looping between light and dark conditions. These are possible candidates for the first adult medaka (parent fish) to start a life cycle aboard ISS. Embryos were treated with a three-dimensional clinostat. Such simulated microgravity caused no differences in tissue architecture or in gene expression within the retina, nor in formation of cartilage (head skeleton). Otolith formation in embryos and fry was investigated for wild-type and mutant (ha) medaka. The ha embryos could not form utricular otoliths. They formed saccular otoliths but with a delay. Fry of the mutant fish lacking the utricular otoliths are highly light-dependent at the time of hatching, showing a perfect dorsal-light response (DLR). As they grow, they eventually shift from being light dependent to gravity dependent. Continuous treatment of the fry with altered light direction suppressed this shift to gravity dependence. Being less dependent on gravity, these fish can serve as model fish in studying the differences expected for the fish that have experienced a life cycle in microgravity.
Among the urodele amphibians, only Cynops pyrrhogaster and Pleurodeles waltl, two species of the Salamandridae family, were used in space experiments. The advantages for using urodeles reside (i) in reproduction: a few months after natural breeding, females can lay eggs in absence of males after a hormonal treatment, because spermatozoa were preserved in the cloacal pelvic glands of matted females, (ii) in the rate of development which is slower in Cynops and Pleurodeles than in the anuran Xenopus, (iii) in their physiological properties: they can live in a closed water container or in a moisturized environment, and they can fast during several days. Moreover, urodeles have an important phylogenetic interest. Many biological phenomena differ from those of anurans, such as fertilization events, the germ cell origin and the migration toward the differentiating gonads, and their regeneration capabilities. The main goals of the space experiments were to answer the following questions. On the one hand, does fertilization occur normally in microgravity? Is subsequent embryonic development normal in microgravity? Is further development and reproduction normal after return to Earth? On the other hand, does microgravity affect the organs in adult animals? Does microgravity affect the regeneration of organs? Fertilization in space is clearly demonstrated. However, subsequent embryonic development appears to be altered in microgravity. In Pleurodeles, abnormalities such as cortical cytoplasmic movements, decrease of cell adhesion, and loss of cells were observed. Although, early development was not strictly normal as a consequence of embryological regulation phenomena, young hatching larvae had normal morphological phenotypes and swimming behavior. After landing, no differences were observed between born-in-space animals to standard ones during the embryonic development to adulthood. The analyses of their offspring demonstrated that the percentages of fertilization and development were in accordance with the control animals. No genetic abnormalities were detected during the analysis of the offspring. The development of their progenies were also without characteristic differences compared to control Pleurodeles. Microgravity seems to have effects on the morphological and histological structures of organs of flight adults. However, as was the case in several experiments the number of analyzed adults was low, and it is too early to conclude on specific effects of microgravity. Moreover, in certain flights the temperature was not regulated, and an increase in temperature occurred. Conditions of these space flights had certainly influenced the samples, and consequently the interpretations of results. Space flights have clear effects on organs in regeneration. But more specifically, they have long term effects that last several weeks after the return of the animals to Earth. A similar result was also obtained for otoconia several months after landing. So far, however, no clear hypothesis could be proposed to interpret these observations.
During the entire evolution of life on Earth, the phylogenetic as well as the individual development of all organisms took place under constant gravity conditions, against which they achieved specific countermeasures for compensation and adaptation. On the one side, gravity represents a factor of physical restriction, which compelled the ancestors of all extant living beings to develop basic achievements to counter the gravitational force (e.g., elements of statics like any kind of skeleton--from actin to bone--to overcome gravity enforced size limits or to keep form). On the other side, already early forms of life possibly used gravity as an appropriate cue for orientation and postural control, since it is continuously present and has a fixed direction. Due to such a thorough adaptation to the Earthly gravity vector, both orientation behaviour as well as the ontogenetic development of animals is impaired, when they have to experience altered gravity (delta g; i.e., hyper- or microgravity). On this background, it is still an open question to which extent delta g affects the normal individual development, either on the systemic level of the whole organism or on the level of individual organs or even single cells. The present review provides information on these questions, focusing on developing fish as model systems. Special emphasis is being laid on the effect of delta g on the developing brain and vestibular system, comprising investigations on behaviour and plastic reactivities of the brain and inner ear. Moreover, clues and insights into the possible basic causes of space motion sickness-phenomena (SMS; a kinetosis) are provided. Overall, the results speak in favour of the following concept: short-term altered gravity (< or = 1 day) can induce transitional aberrant behaviour due to malfunctions of the inner ear, originating from asymmetric otoliths or, generally, from a mismatch between canal and otolith afferents. The vanishing aberrant behaviour is due to a reweighing of sensory inputs and neurovestibular compensation, probably on bioelectrical basis. During long-term altered gravity (several days and more), step by step neuroplastic reactivities on molecular basis (i.e., molecular facilitation) in the brain and inner ears obviously activate feedback mechanisms between the CNS and the vestibular organs for the regain of normal behaviour. Mainly, the following areas of research with animals at altered gravity need to be addressed in the future: (1) Maintenance of animals through two complete life cycles in the space environment (developmental deficiencies?). (2) Investigation of the peripheral and central vestibular system by ground-based studies (mutants, hypergravity experiments...), focusing on plasticity in developing animals as well as in adults. (3) Investigation of the effect of microgravity during critical developmental periods (imprinting phase for graviperception?). Answers to these questions may be of crucial interest for basic gravitational research.
Three new ESA facilities will be available for biological experiments in Space, Biopack on the Space Shuttle and two instruments on the International Space Station (ISS): BIOLAB in the European "Columbus" Laboratory and the European Modular Cultivation System (EMCS) in the US Lab "Destiny". The experiments are housed in standard Experiment Containers, allowing either research in microgravity or acceleration studies with variable g-levels, if mounted on the centrifuges. While Biopack provides only thermal control, BIOLAB and EMCS supply each container with a dedicated atmosphere (controlled CO2, O2 concentration and relative humidity, trace gas removal): EMCS contains also fresh and wastewater reservoirs on its rotors. Power and data lines are available in all the described facilities. Highly automated systems, like BIOLAB's Handling Mechanism and Analysis Instruments, support the telescience concept and help reducing crew time in orbit. A BioGlovebox with its support instruments allows unique research possibilities in Space. The feasibility of experiment hardware inside the containers has been studied by ESA for several kinds of Experiment Support Equipment with potential use for research in Developmental Biology: design concepts for experiments with small eggs, cells and tissues, with small aquatic animals, with insects and with plants are described in this article.
Life on Earth, and thus the reproductive and ontogenetic processes of all extant species and their ancestors, evolved under the constant influence of the Earth's l g gravitational field. These considerations raise important questions about the ability of mammals to reproduce and develop in space. In this chapter, I review the current state of our knowledge of spaceflight effects on developing mammals. Recent studies are revealing the first insights into how the space environment affects critical phases of mammalian reproduction and development, viz., those events surrounding fertilization, embryogenesis, pregnancy, birth, postnatal maturation and parental care. This review emphasizes fetal and early postnatal life, the developmental epochs for which the greatest amounts of mammalian spaceflight data have been amassed. The maternal-offspring system, the coordinated aggregate of mother and young comprising mammalian development, is of primary importance during these early, formative developmental phases. The existing research supports the view that biologically meaningful interactions between mothers and offspring are changed in the weightlessness of space. These changes may, in turn, cloud interpretations of spaceflight effects on developing offspring. Whereas studies of mid-pregnant rats in space have been extraordinarily successful, studies of young rat litters launched at 9 days of postnatal age or earlier, have been encumbered with problems related to the design of in-flight caging and compromised maternal-offspring interactions. Possibilities for mammalian birth in space, an event that has not yet transpired, are considered. In the aggregate, the results indicate a strong need for new studies of mammalian reproduction and development in space. Habitat development and systematic ground-based testing are important prerequisites to future research with young postnatal rodents in space. Together, the findings support the view that the environment within which young mammals develop, comprised of its mother and siblings, is of paramount importance in interpreting spaceflight effects.
Bdelloid rotifers are suitable model systems for space experiments. Due to their developmental pattern they appear adequate to investigate the role of the cytoskeleton during oogenesis and during early developmental stages, and to reflect the effects of disturbances in the spatial arrangement of cytoskeletal components. The effect of weightlessness on the developmental pattern of a bdelloid rotifer will be studied in the International Space Station: in preparation for it we are performing ground-based experiments on the development of rotifer embryos under either increased or decreased gravity. The model studied is Macrotrachela quadricornifera, a species of rotifers belonging to the Bdelloidea class. Samples exposed to gravity disturbance were analyzed for morphology and fitness-related parameters. Rotifers were exposed over several days to altered gravity conditions and the morphology of eggs laid during this period were investigated using a confocal laser microscope. A subset of eggs was allowed to hatch to determine newborn developmental time and age at maturity. High (up to 20g) gravity was obtained in a slow centrifuge suitable for animal cultivation over several days. To produce low (simulated 0.0001g) gravity a Random Positioning Machine equipped with a 'rotifer bioreactor' was used. Under all conditions the rotifer retained normal life-history traits, and did not show permanent changes in embryo morphology, regardless to the stresses to which it was exposed. Only some modification of the shape of early embryos, experiencing 20g, has been noted, but later developmental stages appeared unaffected, and normal juveniles hatched. Whether this result indicates any capacity to repair damage during embryogenesis of these Spiralia experiencing 20g is an open question. The significance of the result as well as the use of instruments to simulate gravity perturbations are discussed.
Space exploration, especially its future phase involving the International Space Station (ISS) makes possible the study of the effects on living systems of long-term expositions to such a strange environment. This phase is being initiated when Biological Sciences are crossing a no-return line into a new territory where the connection between phenotype and genotype may be finally made. We briefly review the paradoxical results obtained in Space experiments performed during the last third of the XX Century. They reveal that simple unicellular systems sense the absence of gravity changing their cytoskeletal organization and the signal transduction pathways, while animal development proceeds unaltered in these conditions, in spite of the fact that these processes are heavily involved in embryogenesis. Longer-term experiments possible in the ISS may solve this apparent contradiction. On the other hand, the current constraints on the scientific use of the ISS makes necessary the development of new hardware and the modification of current techniques to start taking advantage of this extraordinary technological facility. We discuss our advances in this direction using one of the current key biological model systems, Drosophila melanogaster. In addition, the future phase of Space exploration, possibly leading to the exploration and, may be, the colonization of another planet, will provide the means of performing interesting evolutionary experiments, studying how the terrestrial biological systems will change in their long-term adaptation to new, very different environments. In this way, Biological Research in Space may contribute to the advancement of the new Biology, in particular to the branch known as "Evo-Devo". On the other hand, as much as the Space Adventure will continue involving human beings as the main actors in the play, long-term multi-generation experiments using a fast reproducing species, such as Drosophila melanogaster, capable of producing more than 300 generations in 15 years, the useful life foreseen for ISS, will be important. Among other useful information, they will help in detecting the possible changes that a biological species may undergo in such an environment, preventing the uncontrolled occurrence of irreversible deleterious effects with catastrophic consequences on the living beings participating in this endeavour.
Fertilization and subsequent embryogenesis of newts occurred normally under microgravity in two Astronewt flight experiments. By accumulation of the results from the amphibian flight experiments including 'Astronewt', it is considered that gravity has rather small effects on the early development of amphibian eggs. However, some temporary abnormalities, which recover in the course of the further developmental process, have been observed. Some regulations may occur in whole embryos. For a thorough knowledge about the role of gravity in morphogenesis, we need to investigate the gravitational effects on a single cell in a whole embryo. We propose a new experimental system with sea urchin embryos and micromeres for further studies at a cellular level of the effects of gravity on morphogenesis.
