Global Cooperation Enhances Space Flight Research

Before the April 17 launch of Neurolab, the 16-day space shuttle Columbia flight during which 26 studies of the nervous system would be conducted, researchers differed in opinion concerning the microneurography experiment. Either the thin needle placed in a nerve just below the knee of an astronaut would show that electrochemical signals were being transmitted normally from brain to blood vessels via the autonomic nervous system, or the nerve activity would be greater in microgravity than on Earth, or it would decrease during space flight and the experiment wouldn't work at all. Astronaut James A. Pawelczyk, an assistant professor of kinesiology and physiology at Pennsylvania State University and a coinvestigator for this particular experiment, would be the one to insert the needle. PLAYING CATCH: A spring-loaded apparatus measures the anticipatory contraction of James A. Pawelczyk's muscles as he prepares to catch a ball descending in microgravity. Back at the National Aeronautics and Space Administration (NASA) Johnson Space Center in Houston after the mission, Pawelczyk recalls his concern that decreased nerve activity might make it impossible to move the needle into the precise area necessary to get a measurement. His first test subject was payload specialist Jay Clark Buckey Jr ., an associate professor of medicine at Dartmouth Medical School. "I remember very distinctly the first time I put the needle in Jay's knee, I heard a burst of electrical activity, just like you would if you were on the ground," Pawelczyk says. "As I understand it, the ground crew broke into wild applause." He adds that preliminary analyses of data indicate that this sympathetic nervous system activity actually increases in microgravity. Assessment of such findings is the current task of Neurolab's researchers. They must determine the implications of their data for future missions, for living in space on the $21 billion International Space Station--the first section of which is due to be launched this summer--and for various possible therapeutic applications on Earth. "I felt the mission was tremendously successful," asserts Mary Anne Frey, NASA's Neurolab program scientist in Washington, D.C. "We were able to see many of the experiments from the ground, and we got a lot of data down." She says the quality of the data obtained often was comparable to that of Earth-based laboratories, even though some of the hardware was greatly miniaturized for the flight. For example, physiological measurements taken of sleeping astronauts were recorded on equipment about half the size of a loaf of bread, although Frey says the hardware for such recordings normally is about as big as a large washer-dryer combination. However, the mission was not without glitches. More than half of the 96 baby rats that went up in the shuttle did not survive, dying from failed interaction between the mothers and pups. Frey says a panel will be formed to examine specific causes of the deaths. "The investigators shared tissue, and all of them will be able to accommodate their primary research objectives," she adds. She attributes this cooperative outcome to the four years that the scientific teams worked together prior to the mission, and to the early organization of the scientists into eight teams, which also helped to conserve resources and focus the work. Crew members were subjects in a total of 11 studies conducted by four teams: autonomic nervous system; sensory motor performance; vestibular [equilibrium]; and sleep. Animals were used in 15 projects conducted by the other four teams: mammalian development; neuronal plasticity; neurobiology; and aquatics. "Other missions have had many international experiments, but those countries really put their own experiments together," Frey notes. "On this mission, we all worked together from the start." Neurolab involved close cooperation among NASA and the national space agencies of Canada, France, Germany, Europe, and Japan. The peer-review system for selection of participating investigators and their projects was led by the National Institutes of Health and included representatives from NASA, the National Science Foundation, the Office of Naval Research, and the overseas space agencies. Meetings were held around the world to solicit expert opinion on the most important areas of neuroscientific research that would benefit from experiments conducted in microgravity. Guidelines for submissions were drawn up that included such general research areas as development of the nervous system, studies in cellular and molecular biology, and vestibular system function. The review team chose the best 26 of 174 proposals received worldwide. The autonomic nervous system team that devised the microneurography experiment was an example of strong national and international scientific cooperation. C. Gunnar Blomqvist, a professor of internal medicine at the University of Texas Southwestern Medical Center, was principal investigator for a neural cardiovascular control study that integrated techniques and expertise of scientists at his institution as well as at Virginia Commonwealth University, Vanderbilt University, and the Institute of Aerospace Medicine in Germany. "We had a unique development of this experiment, because we started out as four separate experiments that all dealt with the basic problem in one way or another," says Blomqvist. "But we had a peaceful development of a joint experiment." Each of the participating scientists was interested in different aspects of cardiovascular system function that might play a causative role in "orthostatic intolerance," a lightheadedness caused by insufficient blood flow to the brain when a person is standing. Tests by Blomqvist and colleagues during previous Spacelab flights indicated that after returning from orbit, most astronauts could not stand upright for 10 minutes without feeling faint (J.S. Buckey Jr. et al., Journal of Applied Physiology, 81:7-18, 1996). The general cause of this condition had been traced to inadequate vasoconstriction, an autonomic function that normally increases blood pressure by constricting the vessel. JOINT DEVELOPMENT: C. Gunnar Blomqvist of the University of Texas Southwestern Medical Center was principal investigator for a study that involved four institutions. "With that background, the sympathetic nervous system becomes crucial to orthostatic hypotension," Blomqvist notes. "The mechanisms that mediate vasoconstrictor regulation are clearly the major factors." Although a tendency to orthostatic intolerance may be inherited, Blomqvist says, the Neurolab autonomic nervous system team's focus was on examining the specific mediating mechanisms by applying a battery of physiological stimuli to the astronauts, both during and after the flight. They did isometric exercises and dipped their hands in icy cold gel to raise blood pressure. They were strapped into a Lower Body Negative Pressure device created by the German Space Agency to emulate in orbit the stress placed on the cardiovascular system while standing in Earth's gravity, which helps to show changes in the body's fluid distribution during weightlessness. They did the microneurography tests, led by a Vanderbilt professor of medicine, pharmacology, and neurology, David Robertson. They conducted tests of the baroreflex, a collection of sensory nerve endings primarily in the carotid sinuses and the aortic arch that monitors changes in blood pressure, a special interest of Virginia Commonwealth University professor of cardiology and physiology Dwain L. Eckberg. Among tests supervised by Blomqvist's colleague at Southwestern Medical Center, associate professor of internal medicine and cardiology Benjamin D. Levine, were transcranial Doppler measurements that provided continuous, noninvasive monitoring of cerebral artery blood flow velocity. Other studies examined the effects of controlled breathing on blood pressure control. Blomqvist says it's still too early to determine what all the collected data suggest about the specific causes of orthostatic intolerance, but he cautiously suggests, "In general, there seems to be less degradation of [blood pressure] control mechanisms in orbit than we had expected." He adds, "This may mean that changes in fluid distribution may be the primary problem." The same autonomic nervous system team that worked on Neurolab also examined two cosmonauts prior to their current flight in the Russians' Mir space station. Blood pressure control tests are being conducted on them in flight. After the cosmonauts land in August, the investigators intend to do more testing. Blomqvist says a matrix has been designed to apportion all data analysis among team members. One of the most interesting scientific innovations on Neurolab was the off-axis rotator, a chair developed by the European Space Agency that rotates at 45 m.p.h. to simulate Earth's gravity and stimulates the vestibular apparatus of the inner ear with both spinning and tilting sensations. Infrared video cameras recorded the astronauts' eye movements during the exercise, the results of which Pawelczyk describes as "completely unexpected." On the first day of flight, he says, "We all thought the experiment was a bust, because it didn't feel any different from being on the ground." But as they repeated the experiment in succeeding days, it felt increasingly as if they were tilting to one side. In the chair, the astronauts watched vertical stripes moving horizontally, but Pawelczyk and one or two others eventually began to perceive the stripes as horizontal and moving vertically. He says this result has intrigued the vestibular team, which is now trying to determine the reason for the brain's reinterpretation of the vestibular system data. Unlike many astronauts, Pawelczyk didn't have trouble sleeping in orbit, but he found it comforting to strap a pillow to his head--a needless thing in microgravity. "Also, I can't understand why you want to roll over in space, but I still found myself tossing and turning at night," he says. He slept tied to the ceiling in his bag and when he awoke, he had to mentally rearrange the shuttle to avoid perceiving it as upside down. Using a spring-loaded apparatus developed by the French Space Agency, the astronauts took turns sitting and catching a ball propelled downward from overhead. In microgravity, a ball has a constant velocity, rather than accelerating with descent. "We were all surprised at how slowly the ball came down in flight," Pawelczyk reports. "You could grab lunch before it came down and still catch it." The anticipatory contraction of the astronauts' muscles was recorded, which he says could be useful in helping people to relearn simple motor tasks after debilitating traumas such as stroke. NASA's Virtual Environment Generator (VEG) headgear was worn to study how the balance between the visual and vestibular system shifts toward visual cues in a weightless environment. The astronauts watched computer-generated, three dimensional images of movement, at varying speeds, down an endless corridor. The effect, Pawelczyk says, is like involuntarily stepping back when you see a car from the corner of your eye. "I was sucked right into it," he marvels. Much of the hardware devised and contributed by the participating countries worked well, but two malfunctions threatened to ruin one of the most ambitious experiments. The objective was to examine the effects of space travel on the vestibular system of the oyster toadfish (Opsanus tau). In both humans and fish, masses of calcium carbonate particles called otoliths or otolithic stones move within the fluid-filled canals of the inner ear whenever the head moves, pressing against hair cells that then send sensory information about both gravity and acceleration to the brain. This provides the vertical orientation required by vertebrates to function in Earth's gravitational field. Lack of gravity is thought to alter nerve impulses from the inner ear, leading to the "space adaptation syndrome" often experienced by astronauts, a nausea similar to motion sickness. FEELING QUEASY: Fish and humans have similar inner-ear mechanisms to sense gravity and acceleration. The oyster toadfish (Opsanus tau) was used in experiments to determine whether weightlessness alters nerve impulses from the inner ear, leading to the nausea often experienced by astronauts. Principal investigator Stephen M. Highstein, a professor of otorhinolaryngology and anatomy and neurobiology at Washington University in St. Louis, has been studying the toadfish for about two decades and has published widely on its vestibular system (R. Boyle and S.M. Highstein, Journal of Neuroscience, 10:1570-82, 1990). The fish's broad, flat head makes its neural network relatively easy to measure by instrumentation. Before the flight, Highstein and his team cut the nerve in four toadfish that detects gravity signals. They then inserted in the nerve's path a "wafer electrode assembly," through the pores of which the nerve could regrow. Some of the pores were lined with electrodes developed by David J. Anderson, professor of otorhinolaryngology and electrical and computer science at the University of Michigan. These electrodes would pick up electrochemical signals from the regrown nerve, sending the signals to a wireless telemeter created by the National Space Development Agency of Japan (NASDA) and surgically mounted on the fishes' heads. The telemeters, powered by a magnetic field outside the fish tanks, sent nerve activity information to a recorder. LONG PLANNING: NASA's program scientist on Neurolab, Mary Anne Frey, attributes the success of the mission in large part to four years of cooperation among the scientific teams. However, a bacterial filter failed on the fish tanks, which also were designed by NASDA. Ammonia levels rose, and two of the fish perished on the 10th day of the mission. What's more, the wafer assembly functioned properly on just one fish, although it was one of the two surviving animals. Highstein says there will be consultation with the Japanese about the tanks. Of the wafer assembly problems, he explains, "It's a complicated process, there are a lot of variables, and we're still studying it." Indeed, assembly and implantation of the device consists of about 24 steps involving groups across the country that perform such tasks as ultrasonically bonding the materials, adhesively coating the wafer, spinning biopolymer guide tubes for the electrodes, and ensuring that the cut axons are close enough to the wafer to grow through the 1-millimeter-diameter pores. Eventually, Highstein says, the invention might function as an interface or junction box for the human nervous system, enabling the brain to communicate with external prostheses or data processors. He muses, "Such devices might allow anyone to speak any language immediately, understand complex mathematical equations, et cetera." Meanwhile, he's studying the approximately four gigabytes of data yielded by the toadfish experiment. "We taped data on the response of the nerves to the impulsive input of linear acceleration and to microgravity, and we also have recorded extensively in the fish postflight," he says. Early indications are that the fish's vestibular system remained sensitive to impulsive input, when an astronaut would move the tank laterally on specially built rails, but the animal's static or gravitational sensitivity was lacking. How microgravity affects the development of mammals was the focus of another study team. One of the team's experiments analyzed the effects of reduced gravity on the developing nervous systems of mice. It was devised by Richard S. Nowakowski, associate professor of neuroscience and cell biology at the University of Medicine and Dentistry, New Jersey, with his wife and coinvestigator, Nancy L. Hayes, an assistant professor of neuroscience and cell biology at the same university. The flight crew, led in this experiment by Pawelczyk, labeled the proliferating cells in the cerebral cortex of mouse fetuses with two DNA markers. The first marker was thymidine, one of the four DNA base pairs. The second marker, administered two hours later to follow the cells as they proliferated in the developing brain, was an analog of thymidine similar enough to become assimilated into the DNA. All cells in the DNA synthesis phase were marked each time, but during the two-hour window between the different markers, some cells would have stopped synthesizing DNA and continued on in the cell cycle. By counting the difference between cells carrying the first marker and those carrying the second marker, a sensitive measure could be obtained of how many cells had progressed. The crew worked with 18 pregnant mice at three developmental stages, marking their embryonic cells on different days of the flight, then removing the fetuses and placing them in fixatives for analysis back on Earth. About 150 fetuses were obtained, which will be compared with two ground-based control groups, making a total cohort of about 500 fetuses. Nowakowski and colleagues previously had identified and precisely measured the lengths of 11 cell cycles in the six-day neurogenetic period of mice, providing a well-established baseline of data during normal development (V.S. Caviness et al, Trends in Neurosciences, 18:379-83, 1995). The Neurolab experiment examines whether gravity, ever-present in the evolution of mammals, plays a key role in that development. The answer could be useful not only to the health of developing embryos but to adults, in whom four physiological systems maintain a high volume of cell proliferation: the skin, the gut, the immune system, and the bone marrow. None of those systems functions with complete normalcy in space. If cell proliferation in microgravity is abnormal, it could be that the various adverse effects of reduced gravity on astronauts are occuring at the cellular level rather than the systems level. Nowakowski and Hayes think the specific mechanism for the influence of microgravity on individual cells could be the absence of small convection currents within the cells that normally would bring in new metabolites as the cell exhausted the metabolites around it. But convection currents require gravity, because they are set up by buoyancy differences, and buoyancy is proportional to weight, not to mass. Without convection, the only way for the cells to get new metabolites for continued synthesis is diffusion, which is not dependent on gravity but works slower than convection. Hence, cells in space would synthesize DNA more slowly than on Earth. "There is no evidence one way or the other that convection currents are working in cells, or whether or not they are significant," Nowakowski points out. "If we get no differences in cell proliferation parameters, then the significance will be that buoyancy differences are relatively unimportant. But if we get differences, we will want to attribute them to buoyancy." The couple already is planning ways to observe convection currents inside living cells, using an ordinary light microscope. And they suggest that the importance of knockout mice to human genomic research makes mice a logical study animal on the International Space Station. Conception, early and later embryonic development, and birthing all remain to be studied in space. "Is the complete life cycle compatible with microgravity?" Nowakowski asks. It's a feasibility question, he adds--but one that he and many others are eager to address in scientific terms.

Global Cooperation Enhances Space Flight Research

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