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Ultrastructural observation of altered chloroplast morphology in space-grown Brassica rapa cotyledons.

Photosynthesis will be indispensable in a bioregenerative life-support systems for long space missions. It is critical understand the effects of space on this complex process, especially the loss of gravity. Past has noted changes in plant growth and development; differences about cell size, shape, division, and differentiation; and plastid distribution and structure alterations. The amyloplast-containing columelar cells in root tips were carefully examined since they are likely gravity-sensing sites. Changes on photosynthetic physiology and chloroplast structure have been reported. Both increases and decreases of chlorophyll and carotenoid contents were reported. Structural changes of thylakoid membranes in chloroplasts were observed in pea and Arabidopsis grown in space or clinorotation. Recently, a decrease of CO2 assimilation rate and of electron transport rate of both PSI and PSII on thylakoid membranes were reported in space-grown wheat. These imply an overall decrease of photosynthetic activities, and implicate thylakoid-old structural changes. For example, PSI activity, and its reaction center subunits (PsaA, PsaB, and PsaC) and the LHCIs, were decreased under microgravity. Here, we further examined cellular morphology and ultrastructural features of the chloroplast and its thylakoid membranes by electron microscopy and in situ immunolocalization.

Brassica↗

Site selection for Mars exobiology.

The selection of sites on Mars that have a high priority for exobiological research is fundamental for planning future exploration. The most immediate need is to identify targets for high resolution orbital imaging during the Mars Observer and Mars '94/'96 missions that can be used to refine site priorities for surface exploration. We present an objective approach to site selection whereby individual sites are selected and scored, based on the presence of key geological features which indicate high priority environments. Prime sites are those that show evidence for the prolonged activity of liquid water and which have sedimentary deposits that are likely to have accumulated in environments favorable for life. High priority areas include fluvio-lacustrine (stream-fed lake systems), springs, and periglacial environments. Sites where mineralization may have occurred in the presence of organisms (e.g. springs) are given high priority in the search for a fossil record on Mars. A systematic review of Viking data for 83 sites in the Mars Landing Site Catalog resulted in the selection of 13 as being of exobiological interest. The descriptions of these sites were expanded to address exobiological concerns. An additional five sites were identified for inclusion in the second edition of the MLSC. We plan to broaden our site selection activities to include a systematic global reconnaissance of Mars using Viking data, and will continue to refine site priorities for exobiological research based on data from future missions in order to define strategies for surface exploration.

Exobiology↗

Percutaneous aspiration of fluid for management of peritonitis in space.

BACKGROUND: As a medical emergency that can affect even well-screened, healthy individuals, peritonitis developing during a long-duration space exploration mission may dictate deviation from traditional clinical practice due to the absence of otherwise indicated surgical capabilities. Medical management can treat many intra-abdominal processes, but treatment failures are inevitable. In these circumstances, percutaneous aspiration under sonographic guidance could provide a "rescue" strategy. HYPOTHESIS: Sonographically guided percutaneous aspiration of intra-peritoneal fluid can be performed in microgravity. METHODS: Investigations were conducted in the microgravity environment of NASA's KC-135 research aircraft (0 G). The subjects were anesthetized female Yorkshire pigs weighing 50 kg. The procedures were rehearsed in a terrestrial animal lab (1 G). Colored saline (500 mL) was introduced through an intra-peritoneal catheter during flight. A high-definition ultrasound system (HDI-5000, ATL, Bothell, WA) was used to guide a 16-gauge needle into the peritoneal cavity to aspirate fluid. RESULTS: Intra-peritoneal fluid collections were easily identified, distinct from surrounding viscera, and on occasion became more obvious during weightless conditions. Subjectively, with adequate restraint of the subject and operators, the procedure was no more demanding than during the 1-G rehearsals. CONCLUSIONS: Sonographically guided percutaneous aspiration of intra-peritoneal fluid collections is feasible in weightlessness. Treatment of intra-abdominal inflammatory conditions in spaceflight might rely on pharmacological options, backed by sonographically guided percutaneous aspiration for the "rescue" of treatment failures. While this risk mitigation strategy cannot guarantee success, it may be the most practical option given severe resource limitations.

Animals↗

On the possibility of chemosynthetic ecosystems in subsurface habitats on Mars.

We have reexamined the question of extant microbial life on Mars in light of the most recent information about the planet and recently discovered nonphotosynthetic microbial ecosystems on Earth--deep sea hydrothermal vent communities and deep subsurface aquifer communities. On Mars, protected subsurface niches associated with hydrothermal activity could have continued to support life even after surface conditions became inhospitable. Geochemical evidence from the SNC meteorites and geomorphological evidence for recent volcanism suggest that such habitats could persist to the present time. There are also extensive geological features that attest to the ubiquitous nature of volcano-ground ice interactions on Mars. We suggest a possible deep subsurface microbial ecology similar to those discovered to depths of several kilometers below the surface of the Earth. We focus on anaerobic systems utilizing CO2 as the primary source of carbon. Liquid water could be provided by the heat of geothermal or volcanic activity melting permafrost or other subsurface water sources. Gases from volcanic activity deep in the planet could provide reducing power (as CH4, H2, or H2S) percolating up from below and enabling the development of a microbial community based upon chemolithoautotrophy. We suggest a methanogen (or acetogen) and sulfur-based microbial ecology as one possible basis for microbial primary production. Our hypothetical ecosystem is neither supported, nor excluded, by current observations of Mars. Tests for such a subsurface system involve locating active geothermal areas associated with ground ice or detecting trace quantities of reduced atmospheric gases that would leak from such a system.

Carbon Dioxide↗

Mineral catalysis of the formation of the phosphodiester bond in aqueous solution: the possible role of montmorillonite clays.

The binding of adenosine to Na(+)-montmorillonite 22A is greater than 5'-AMP, at neutral pH. Adenine derivatives bind more strongly to the clay than the corresponding uracil derivatives. These data are consistent with the protonation of the adenine by the acidic clay surface and a cationic binding of the protonated ring to the anionic clay surface. Other forces must be operative in the binding of uracil derivatives to the clay since the uracil ring system is not basic. The reaction of the 5'-AMP with water soluble carbodiimide in the presence of Na(+)-montmorillonite results in the formation of 2',5'-pApA (18.9%), 3',5'-pApA (11%), and AppA (4.8%). When poly(U) is used in place of the clay the product yields are 2',5'-pApA (15.5%), 3',5'-pApA (3.7%) and AppA (14.9%). The cyclic nucleotide, c(pA)2 is also formed when poly(U) is used. AppA is the principal reaction product when neither clay nor poly(U) is present in the reaction mixture. When 2'-deoxy-5'-AMP reacts with carbodiimide in the presence of Na(+)-montmorillonite 22A the products are dpApA (4.8%), dAppApA (4.5%) and dAppA (17.4%). Cyclic 3',5'-dAMP is the main product (14%) of the reaction of 2'-deoxy-3'-AMP.

Adenine↗

How long did it take for life to begin and evolve to cyanobacteria?

There is convincing paleontological evidence showing that stromatolite-building phototactic prokaryotes were already in existence 3.5 x 10(9) years ago. Late accretion impacts may have killed off life on our planet as late as 3.8 x 10(9) years ago. This leaves only 300 million years to go from the prebiotic soup to the RNA world and to cyanobacteria. However, 300 million years should be more than sufficient time. All known prebiotic reactions take place in geologically rapid time scales, and very slow prebiotic reactions are not feasible because the intermediate compounds would have been destroyed due to the passage of the entire ocean through deep-sea vents every 10(7) years or in even less time. Therefore, it is likely that self-replicating systems capable of undergoing Darwinian evolution emerged in a period shorter than the destruction rates of its components (<5 million years). The time for evolution from the first DNA/protein organisms to cyanobacteria is usually thought to be very long. However, the similarities of many enzymatic reactions, together with the analysis of the available sequence data, suggest that a significant number of the components involved in basic biological processes are the result of ancient gene duplication events. Assuming that the rate of gene duplication of ancient prokaryotes was comparable to today's present values, the development of a filamentous cyanobacterial-like genome would require approximately 7 x 10(6) years--or perhaps much less. Thus, in spite of the many uncertainties involved in the estimates of time for life to arise and evolve to cyanobacteria, we see no compelling reason to assume that this process, from the beginning of the primitive soup to cyanobacteria, took more than 10 million years.

Biological Evolution↗

Entropy and charge in molecular evolution--the case of phosphate.

Biopoesis, the creation of life, implies molecular evolution from simple components, randomly distributed and in a dilute state, to form highly organized, concentrated systems capable of metabolism, replication and mutation. This chain of events must involve environmental processes that can locally lower entropy in several steps; by specific selection from an indiscriminate mixture, by concentration from dilute solution, and in the case of the mineral-induced processes, by particular effectiveness in ordering and selective reaction, directed toward formation of functional biomolecules. Numerous circumstances provide support for the notion that negatively charged molecules were functionally required and geochemically available for biopoesis. Sulfite ion may have been important in bisulfite complex formation with simple aldehydes, facilitating the initial concentration by sorption of aldehydes in positively charged surface active minerals. Borate ion may have played a similar, albeit less investigated role in forming charged sugar complexes. Among anionic species, oligophosphate ions and charged phosphate esters are likely to have been of even more wide ranging importance, reflected in the continued need for phosphate in a proposed RNA world, and extending its central role to evolved biochemistry. Phosphorylation is shown to result in selective concentration by surface sorption of compounds, otherwise too dilute to support condensation reactions. It provides protection against rapid hydrolysis of sugars and, by selective concentration, induces the oligomerization of aldehydes. As a manifestation of life arisen, phosphate already appears in an organic context in the oldest preserved sedimentary record.

Animals↗

Transgene expression patterns indicate that spaceflight affects stress signal perception and transduction in arabidopsis.

The use of plants as integral components of life support systems remains a cornerstone of strategies for long-term human habitation of space and extraterrestrial colonization. Spaceflight experiments over the past few decades have refined the hardware required to grow plants in low-earth orbit and have illuminated fundamental issues regarding spaceflight effects on plant growth and development. Potential incipient hypoxia, resulting from the lack of convection-driven gas movement, has emerged as a possible major impact of microgravity. We developed transgenic Arabidopsis containing the alcohol dehydrogenase (Adh) gene promoter linked to the beta-glucuronidase (GUS) reporter gene to address specifically the possibility that spaceflight induces the plant hypoxia response and to assess whether any spaceflight response was similar to control terrestrial hypoxia-induced gene expression patterns. The staining patterns resulting from a 5-d mission on the orbiter Columbia during mission STS-93 indicate that the Adh/GUS reporter gene was activated in roots during the flight. However, the patterns of expression were not identical to terrestrial control inductions. Moreover, although terrestrial hypoxia induces Adh/GUS expression in the shoot apex, no apex staining was observed in the spaceflight plants. This indicates that either the normal hypoxia response signaling is impaired in spaceflight or that spaceflight inappropriately induces Adh/GUS activity for reasons other than hypoxia.

Alcohol Dehydrogenase↗

Human skeletal muscle protein breakdown during spaceflight.

Human spaceflight is associated with a loss of body protein. Excretion of 3-methylhistidine (3-MH) in the urine is a useful measurement of myofibrillar protein breakdown. Bed rest, particularly with 6 degrees head-down tilt, is an accepted ground-based model for human spaceflight. The objectives of this report were to compare 3-MH excretion from two Life Sciences shuttle missions (duration 9.5 and 15 days, n = 9) and from 17 days of bed rest (n = 7) with 6 degrees head-down tilt. The bed rest study was designed to mimic an actual Life Sciences spaceflight and so incorporated an extensive battery of physiological tests focused on the musculoskeletal system. Results showed that nitrogen retention, based on excretion of nitrogen in the urine, was reduced during both bed rest [from 22 +/- 1 to 1 +/- 5 mg N x kg(-1) x day(-1) (n = 7; P < 0.05)] and spaceflight [from 57 +/- 9 to 19 +/- 3 mg N x kg(-1) x day(-1) (n = 9; P < 0.05)]. 3-MH excretion was unchanged with either bed rest [pre-bed rest 5.30 +/- 0.29 vs. bed rest 5.71 +/- 0.30 micromol 3-MH x kg(-1) x day(-1), n = 7; P = not significant (NS)] or spaceflight [preflight 4.98 +/- 0.37 vs. 4.59 +/- 0.39 micromol 3-MH x kg(-1) x day(-1) in-flight, n = 9; P = NS]. We conclude that 1) 3-MH excretion was unaffected by spaceflight on the shuttle or with bed rest plus exercise, and 2) because protein breakdown (elevated 3-MH) was increased on Skylab but not on the shuttle, it follows that muscle protein breakdown is not an inevitable consequence of spaceflight.

Bed Rest↗

Changes in Arabidopsis leaf ultrastructure, chlorophyll and carbohydrate content during spaceflight depend on ventilation.

Leaf structure and function under spaceflight conditions have received little study despite their important implications for biological life support systems using plants. Previous reports described disruption of the membrane apparatus for photosynthesis and a general decrease in carbohydrate content in foliage. During a series of three short-duration experiments (Chromex-03, -04, -05) on the US space shuttle (STS-54, STS-51, STS-68), we examined Arabidopsis thaliana leaves. The plants were at the rosette stage at the time of loading onto the space shuttle, and received the same light, temperature, carbon dioxide and humidity regimes in the orbiter as in ground controls. The experiments differed according to the regime provided in the headspace around the plants: this was either sealed (on mission STS-54); sealed with high levels of carbon dioxide (on mission STS-51) or vented to the cabin air through a filtration system (on mission STS-68). Immediately post-flight, leaf materials were fixed for microscopy or frozen in liquid nitrogen for subsequent analyses of chlorophyll and foliar carbohydrates. At the ultrastructural level, no aberrations in membrane structure were observed in any of the experiments. When air-flow was provided, plastids developed large starch grains in both spaceflight and ground controls. In the experiments with sealed chambers, spaceflight plants differed from ground controls with regard to measured concentrations of carbohydrate and chlorophyll, but the addition of airflow eliminated these differences. The results point to the crucial importance of consideration of the foliage microenvironment when spaceflight effects on leaf structure and metabolism are studied.

Arabidopsis↗

Ceratopteris richardii: a productive model for revealing secrets of signaling and development.

Ceratopteris richardii is an aquatic fern grown in tropical and subtropical regions of the world. It is proven to be a productive model system for studies in the genetics, biochemistry, and cell biology of basic biologic processes that occur in early gametophytic development. It provides several advantages to biologists, especially those interested in gravitational biology, polarity development, and in the genetics of sexual development. It is easy to culture, has a relatively short life cycle, and offers an array of attractive features that facilitate genetic studies. The germination and early development of large populations of genetically identical spores are easy to synchronize, and both the direction of polarity development and cell-level gravity responses can be measured and readily manipulated within the first 24 h of spore development. Although there is no reliable transformation system available yet in Ceratopteris, recent studies suggest that the technique of RNA interference can be used to block translation of specific genes in a related fern, Marsilea, and current studies will soon reveal the applicability of this approach, as well as of other transformation approaches, in Ceratopteris. A recently completed expressed sequence tag (EST) sequencing project makes available the partial sequence of more than 2000 cDNAs, representing a significant percentage of the genes being expressed during the first 24 h of spore germination, when many developmentally interesting processes are occurring. A microarray of these ESTs is being constructed, so especially for those scientists interested in basic cellular phenomena that occur early in spore germination, the availability of the ESTs and of the microarray will make Ceratopteris an even more attractive model system.

Base Sequence↗

Dynamic posture analysis of Spacelab-1 crew members.

Dynamic posture testing was conducted on the science crew of the Spacelab-1 mission on a single axis linear motion platform. Tests took place in pre- and post-flight sessions lasting approximately 20 min each. The pre-flight tests were widely spaced over the several months prior to the mission while the post-flight tests were conducted over the first, second, fourth, and sixth days after landing. Two of the crew members were also tested on the day of landing. Consistent with previous postural testing conducted on flight crews, these crew members were able to complete simple postural tasks to an acceptable level even in the first few hours after landing. Our tests were designed to induce dynamic postural responses using a variety of stimuli and from these responses, evaluate subtle changes in the postural control system which had occurred over the duration of the flight. Periodic sampling post-flight allowed us to observe the time course of readaptation to terrestrial life. Our observations of hip and shoulder position, when subjected to careful analysis, indicated modification of the postural response from pre- to post-flight and that demonstrable adjustments in the dynamic control of their postural systems were taking place in the first few days after flight. For transient stimuli where the platform on which they were asked to stand quickly moved a few centimeters fore or aft then stopped, ballistic or open loop 'programs' would closely characterize the response. During these responses the desired target position was not always achieved and of equal importance not always properly corrected some 15 seconds after the platform ceased to move. The persistent observation was that the subjects had a much stronger dependence on visual stabilization post-flight than pre-flight. This was best illustrated by a slow or only partial recovery to an upward posture after a transient base-of-support movement with eyes open. Postural responses to persistent wideband pseudorandom base-of-support translation were modeled as time invarient linear systems arrived at by Kalman adaptive filter techniques. Derived model parameters such as damping factor and fundamental frequency of the closed loop system showed significant modification between pre- and post-flight. This phenomenon is best characterized by movement of the poles toward increasing stability. While pre-flight data tended to show shoulders and hips moving in phase with each other, post-flight data showed a more disjoint behavior.(ABSTRACT TRUNCATED AT 400 WORDS)

Adaptation, Physiological↗

Template-directed oligonucleotide ligation on hydroxylapatite.

Bernal, and subsequently other authors, have suggested that the prebiotic synthesis of the precursors of biopolymers could have occurred on a solid surface such as that provided by clay or some other mineral. The separation of products from the other components of the reaction mixture in such a system is reminiscent of modern solid-phase synthesis of polypeptides and polynucleotides. One such scheme envisages that growing polymers were localized by adsorption to a mineral surface where an activating agent or activated monomers were supplied continuously or cyclically. We have been trying to test this scheme using reactions which we believe may be related to those that occurred during prebiotic evolution. We have already reported that oligonucleotides adsorbed onto hydroxylapatite provide suitable templates for the oligomerization of (guanosine 5'-phosphor)-2-methylimidazolide (2-MeImpG). However, this is not a suitable test reaction, as 2-MeImpG oligomerization proceeds almost to completion in a single step. Here we report that a sequence of reactions in which initially formed oligo(G)s are reactivated by conversion to phosphorimidazolides in the presence of poly(C) and then allowed to ligate is ideal, in that repeated cycles can be carried out on the surface of hydroxylapatite, whereas in the liquid phase the cycle could be achieved only with considerable difficulty.

Adsorption↗

Thin filament diversity and physiological properties of fast and slow fiber types in astronaut leg muscles.

Slow type I fibers in soleus and fast white (IIa/IIx, IIx), fast red (IIa), and slow red (I) fibers in gastrocnemius were examined electron microscopically and physiologically from pre- and postflight biopsies of four astronauts from the 17-day, Life and Microgravity Sciences Spacelab Shuttle Transport System-78 mission. At 2.5-microm sarcomere length, thick filament density is approximately 1,012 filaments/microm(2) in all fiber types and unchanged by spaceflight. In preflight aldehyde-fixed biopsies, gastrocnemius fibers possess higher percentages (approximately 23%) of short thin filaments than soleus (9%). In type I fibers, spaceflight increases short, thin filament content from 9 to 24% in soleus and from 26 to 31% in gastrocnemius. Thick and thin filament spacing is wider at short sarcomere lengths. The Z-band lattice is also expanded, except for soleus type I fibers with presumably stiffer Z bands. Thin filament packing density correlates directly with specific tension for gastrocnemius fibers but not soleus. Thin filament density is inversely related to shortening velocity in all fibers. Thin filament structural variation contributes to the functional diversity of normal and spaceflight-unloaded muscles.

Adult↗

Instrumentation and methodology for quantifying GFP fluorescence in intact plant organs.

The General Fluorescence Plant Meter (GFP-Meter) is a portable spectrofluorometer that utilizes a fiber-optic cable and a leaf clip to gather spectrofluorescence data. In contrast to traditional analytical systems, this instrument allows for the rapid detection and fluorescence measurement of proteins under field conditions with no damage to plant tissue. Here we discuss the methodology of gathering and standardizing spectrofluorescence data from tobacco and canola plants expressing GFP. Furthermore, we demonstrate the accuracy and effectiveness of the GFP-Meter. We first compared GFP fluorescence measurements taken by the GFP-Meter to those taken by a standard laboratory-based spectrofluorometer, the FluoroMax-2. Spectrofluorescence measurements were taken from the same location on intact leaves. When these measurements were tested by simple linear regression analysis, we found that there was a positive functional relationship between instruments. Finally, to exhibit that the GFP-Meter recorded accurate measurements over a span of time, we completed a time-course analysis of GFP fluorescence measurements. We found that only initial measurements were accurate; however, subsequent measurements could be used for qualitative purposes.

Brassica↗

3He lung imaging in an open access, very-low-field human magnetic resonance imaging system.

The human lung and its functions are extremely sensitive to gravity; however, the conventional high-field magnets used for most laser-polarized (3)He MRI of the human lung restrict subjects to lying horizontally. Imaging of human lungs using inhaled laser-polarized (3)He gas is demonstrated in an open-access very-low-magnetic-field (<5 mT) MRI instrument. This prototype device employs a simple, low-cost electromagnet, with an open geometry that allows variation of the orientation of the imaging subject in a two-dimensional plane. As a demonstration, two-dimensional lung images were acquired with 4-mm in-plane resolution from a subject in two orientations: lying supine and sitting in a vertical position with one arm raised. Experience with this prototype device will guide optimization of a second-generation very-low-field imager to enable studies of human pulmonary physiology as a function of subject orientation.

Administration, Inhalation↗

High-LET radiation induces inflammation and persistent changes in markers of hippocampal neurogenesis.

Exposure to heavy-ion radiation is considered a potential health risk in long-term space travel. It may result in the loss of critical cellular components in complex systems like the central nervous system (CNS), which could lead to performance decrements that ultimately could compromise mission goals and long-term quality of life. Specific hippocampal-dependent cognitive impairment occurs after whole-body 56Fe-particle irradiation, and while the pathogenesis of this effect is not yet clear, it may involve damage to neural precursor cells in the hippocampal dentate gyrus. We irradiated mice with 1-3 Gy of 12C or 56Fe ions and 9 months later quantified proliferating cells and immature neurons in the dentate subgranular zone (SGZ). Our results showed that reductions in these cells were dependent on the dose and LET. When compared with data for mice that were studied 3 months after 56Fe-particle irradiation, our current data suggest that these changes are not only persistent but may worsen with time. Loss of precursor cells was also associated with altered neurogenesis and a robust inflammatory response. These results indicate that high-LET radiation has a significant and long-lasting effect on the neurogenic population in the hippocampus that involves cell loss and changes in the microenvironment.

Animals↗

Confined-pyrolysis as an experimental method for hydrothermal organic synthesis.

A closed pyrolysis system has been developed as a tool for studying the reactions of organic compounds under extreme hydrothermal conditions. Small high pressure stainless steel vessels in which the ratio of sediment or sample to water has been adjusted to eliminate the headspace at peak experimental conditions confines the organic components to the bulk solid matrix and eliminates the partitioning of the organic compounds away from the inorganic components during the experiment. Confined pyrolysis experiments were performed to simulate thermally driven catagenetic changes in sedimentary organic matter using a solids to water ratio of 3.4 to 1. The extent of alteration was measured by monitoring the steroid and triterpenoid biomarkers and polycyclic aromatic hydrocarbon distributions. These pyrolysis experiments duplicated the hydrothermal transformations observed in nature. Molecular probe experiments using alkadienes, alkenes and alkanes in H2O and D2O elucidated the isomerization and hydrogenation reactions of aliphatic compounds and competing oxidative reactions occurring under hydrothermal conditions. This confined pyrolysis technique is being applied to test experiments on organic synthesis of relevance to chemical evolution for the origin of life.

Alkanes↗