Justin S. Rhodes - US grants

The major goal of the proposed project is to investigate the neural basis of hyperactivity in a novel mouse model. Through artificial selection we have produced lines of mice that are internally motivated to run faster on running wheels than randombred controls, resulting in a 26-fold increase in wheel revolutions in a 24 hour period. Wheel-running in the hyperactive animals is composed of short bursts of activity separated by frequent short rests. High wheel-running mice are also more active in their home cages when deprived of wheels. The high-running mice may, therefore, be a good animal model for attention deficit hyperactivity disorder (ADHD). An association between deficiencies in dopamine transmission and ADHD has been suggested, and preliminary neuropharmacological results are consistent with the hypothesis that the hyperactive wheel running is caused by diminished dopaminergic function. To further explore the neural basis of genetic hyperactivity, I propose to examine whether: 1) apomorphine, and Ritalin(TM) attenuate wheel-running behavior in the high-running mice, 2) dopamine metabolism is slower in the high-running mice compared to controls at a given speed of running, and 3) the high-running mice contain fewer or smaller dopamine neurons in the ventral tegmental area. Together the proposed studies will help clarify the underlying neurochemistry and neuroanatomy of genetic hyperactivity.

DESCRIPTION (provided by applicant): It is widely accepted that genes interact with environment to affect brain function and behavior. Exercise is a potent environmental factor with known benefits for physical health such as strength and stamina. The recent discovery that exercise can also enhance cognitive performance has generated much enthusiasm and interest, but mechanisms are far less understood. A growing body of evidence in rodent animal models suggests that exercise is a natural generator of neural plasticity. One example is growth of new neurons in the hippocampus, which is strongly regulated by exercise along with many other changes (e.g., trophic factors, growth factors, capillaries). Harnessing mechanisms of the natural generator could be useful for treating a wide range of neurological problems such as cognitive aging, neurodegenerative disease, stroke, or head trauma. One approach to find a mechanism is to systematically block changes in each hypothesized substrate (e.g., new neurons). Another is unbiased exploration of genetic mechanisms. We propose both. Preliminary data suggest that exercise induced changes in hippocampal neurogenesis and learning vary depending on genotype in mice. Aims 1 and 2 are to identify sets of genotypes that display larger versus smaller benefits. This crucial information will be contributed to a database of phenotypic and genetic information on these strains, and used in the future to identify mechanisms for cognitive benefits of exercise at multiple levels of biological organization from genes to physiology to behavior. In addition to laying the groundwork for the genetic analysis, we also propose an innovative method in aim 3 to reduce neurogenesis in exercising mice, focal gamma radiation, to directly test the hypothesis that new neurons are required for enhanced learning from exercise in predisposed genotypes. For this, we propose to use the genotype C57BL/6J, as proof of principal, because a strong correlation between exercise, neurogenesis, and learning is well established for this strain. PUBLIC HEALTH RELEVANCE: The goal of this project is to discover mechanisms for pro cognitive effects of exercise at multiple levels of biological organization from genes to physiology to behavior.

DESCRIPTION (provided by applicant): The interaction between aerobic exercise and drug abuse is relatively unexplored. It deserves attention because recent data suggest neuroadaptations from exercise promote learning in circuits that overlap with drug abuse. The hippocampus is an important point of intersection because it is a major locus for change from aerobic exercise and it plays a central role in contextual conditioning. Specifically, contextual cues paired with drugs trigger emotional responses related to craving and relapse. The long-term goal of this research program is to identify molecular and physiological mechanisms underlying the influence of exercise on drug-related behaviors in mice. The overall objective of this application is to develop a mouse model to study the effects of wheel running exercise on extinction of cocaine conditioned place preference, and to test one hypothesized mechanism for how exercise can influence Pavlovian drug associations. The central hypothesis is that new neurons from exercise can cause drug associations to persist if the drug is administered at a critical period in the development of the new neurons when they are preferentially recruited into learning networks. This is supported by the Preliminary Studies that show resistance to extinction of conditioned place preference for cocaine in runners as compared to sedentary animals after re-exposure to cocaine in context. The objective of this application will be accomplished by pursuing two specific aims. Aim 1 is to identify the impact of aerobic exercise on extinction of conditioned place preference for cocaine. Based on Preliminary Studies, the working hypothesis is that exercise will either facilitate or delay extinction of conditioned place preference depending on whether drug exposure occurs before or after exercise training, in parallel with increased adult hippocampal neurogenesis in the dentate gyrus. To accomplish this aim, the order of conditioning and exercise treatments will be manipulated, and then conditioned place preference will be measured repeatedly until extinction. Animals will be injected with BrdU to label dividing cells, and the number of BrdU cells co-labeled with neuronal nuclear marker, NeuN, in the granule layer of the dentate gyrus will be used to measure neurogenesis. Aim 2 is to determine the extent to which new neurons from exercise causally contribute to persistence of conditioned place preference for cocaine. Based on Preliminary Studies, the working hypothesis is that new neurons from exercise will function to enhance Pavlovian conditioning. This hypothesis will be directly tested by reducing neurogenesis using 2 separate methods, a transgenic mouse model and focal gamma irradiation, to determine whether new neurons are required for exercise to delay extinction of place preference. The extent to which new neurons are preferentially recruited into circuits involved in cocaine conditioning will also be determined by measuring the proportion of BrdU positive versus negative cells expressing c-Fos in response to the preference test. The project will discover mechanisms for interactions between exercise and drug abuse. This will be useful for evaluating the benefits or risks of incorporating exercise in treatment of drug abuse. PUBLIC HEALTH RELEVANCE: This proposal will discover the impact of enhanced neuroplasticity from aerobic exercise on extinction of cocaine conditioned behavior in a mouse model. The project will provide useful evidence for evaluating the benefits or risks of incorporating exercise in treatment of drug abuse.

DESCRIPTION (provided by applicant): It is widely accepted that genes interact with environment to affect brain function and behavior. Exercise is a potent environmental factor with known benefits for physical health such as strength and stamina. The recent discovery that exercise can also enhance cognitive performance has generated much enthusiasm and interest, but mechanisms are far less understood. A growing body of evidence in rodent animal models suggests that exercise is a natural generator of neural plasticity. One example is growth of new neurons in the hippocampus, which is strongly regulated by exercise along with many other changes (e.g., trophic factors, growth factors, capillaries). Harnessing mechanisms of the natural generator could be useful for treating a wide range of neurological problems such as cognitive aging, neurodegenerative disease, stroke, or head trauma. One approach to find a mechanism is to systematically block changes in each hypothesized substrate (e.g., new neurons). Another is unbiased exploration of genetic mechanisms. We propose both. Preliminary data suggest that exercise induced changes in hippocampal neurogenesis and learning vary depending on genotype in mice. Aims 1 and 2 are to identify sets of genotypes that display larger versus smaller benefits. This crucial information will be contributed to a database of phenotypic and genetic information on these strains, and used in the future to identify mechanisms for cognitive benefits of exercise at multiple levels of biological organization from genes to physiology to behavior. In addition to laying the groundwork for the genetic analysis, we also propose an innovative method in aim 3 to reduce neurogenesis in exercising mice, focal gamma radiation, to directly test the hypothesis that new neurons are required for enhanced learning from exercise in predisposed genotypes. For this, we propose to use the genotype C57BL/6J, as proof of principal, because a strong correlation between exercise, neurogenesis, and learning is well established for this strain. PUBLIC HEALTH RELEVANCE: The goal of this project is to discover mechanisms for pro cognitive effects of exercise at multiple levels of biological organization from genes to physiology to behavior.

PI: Boppart, Stephen A.
Proposal: 1450829
Title: BRAIN EAGER: Spatially-Resolved In Vivo Optogenetic Stimulation and Imaging Platform

Significance
The successful outcome of this research project will have a broad impact in neuroscience in addition to optical science and engineering. The PI will use implanted imaging fiber bundles that will enable
in vivo imaging as well as spatially-controlled optical stimulation and optical feedback of
large-area neural circuits. Current fibers only indiscriminately illuminate large-areas. Optogenetics is expected to make a broad impact in neuroscience, as well as medical science and clinical medicine in the future. This proposed research offers the potential to have an even greater impact by controlling the light stimulus and enhancing specificity in the control of neural circuits. The results of this project will be shared widely amongst the scientific and engineering communities, and also across wide segments of society in outreach activities. The new imaging and visualization capabilities will inspire K-12 students to think about how technology can be used to see things one cannot normally see, and how we can invent new ways of seeing the world around us and discovering new knowledge. Outreach activities will include demos of these imaging fiber bundles and novel light sources to K-12 and community groups through
annual Engineering Open House events, as well as integration of these technological methods in Prof. Boppart?s undergraduate ECE/BioE 467 Biophotonics and ECE/BioE 380 Biomedical Imaging
courses.

Technical Description
Optogenetics is a rapidly developing field with an ever-expanding toolkit of molecular biology
techniques to enable light-activated switching and control of cells, most commonly neurons.
Equally significant advances have occurred in optical science and engineering. By understanding
and exploiting physics-based principles of how light interacts in photonic crystal fibers (PCFs) and within imaging fiber bundles, it is possible to generate, control, and optimize a wide range of new optical parameters for in vivo optogenetic stimulation. Traditionally in in vivo optogenetic applications, light has been sent down single multi-mode optical fibers to diffusely illuminate the brain, relying on the molecular biology of optogenetically-modified neurons for cell and circuit specificity. This EAGER project will uniquely develop and demonstrate the use of imaging fiber bundles, and the generation of specific light pulse parameters to enable spatially-resolved optogenetic stimulation and imaging of neural circuits in vivo. These novel neurotechnologies will enable new investigations underlying behavior and cognition.

Summary Exercise robustly enhances cognitive performance across the lifespan but the mechanisms are not well understood. The long-term goal of this research program is to elucidate the neurological mechanisms by which exercise improves cognition. The objective of this application is to determine the origin of exercise-induced hippocampal neurogenesis and enhanced behavioral performance, whether from contracting muscles or acute activation of the hippocampus during physical exertion. Recent work has emphasized the importance of the muscle-brain axis and has assumed the main signals originate from skeletal muscle. On the other hand, there is a large and well established literature illustrating a close correlation between neural activation of the hippocampus and the speed of movement. Results will provide crucial information for deciding whether to focus on the muscle secretome or mechanisms within the brain for recapitulating pro-cognitive effects of exercise. The rationale is to develop better strategies for neuronal regeneration and repair and for ameliorating cognitive deficits associated with neurological disorders. The central hypothesis is that both muscle contractions and hippocampal neuronal activation each independently enhance neurogenesis and related behaviors. The hypothesis is supported by preliminary studies showing that both repeated electrical contractions of the hindlimb muscles and activation of hippocampal neurons increases hippocampal neurogenesis in mice. One of the PIs has a productive research program on exercise induced-neurogenesis and measuring behavioral performance in mice, and the other has expertise on muscle physiology and electrical stimulation. Moreover, the PIs have developed multiple innovative methods for powerful hypothesis testing. The objectives of this application will be accomplished by pursuing 2 specific aims: 1) Determine the extent to which repeated electrical contraction of the hindlimb muscles is sufficient to increase adult hippocampal neurogenesis and enhance learning and memory. 2) Determine the extent to which optogenetic activation of the dentate gyrus in a pattern that mimics running is sufficient for exercise-induced neurogenesis and enhanced behavioral performance. An established procedure for electrically contracting the hindlimb muscles will be used to repeatedly contract the muscles while the mouse is anesthetized. State-of-the-art optogenetic methods will be used to instantaneously activate dentate gyrus granule neurons that were acutely and transiently activated in response to running and to measure the long-term effects on neurogenesis and behavior. Elucidating and unequivocally establishing mechanisms underlying pro-cognitive effects of exercise holds the key to discover novel and more efficient ways to maintain, promote and improve cognitive performance. The proposed research is highly innovative, it addresses pressing questions in the field using very novel strategies and state-of-art optogenetics technology that will allow us to generate causal, mechanistic data on the origin of exercise's effects on neurogenesis and cognitive performance.

Summary Exercise robustly enhances cognitive performance across the lifespan but mechanisms are not well understood. The long-term goal of this research program is to elucidate the neurological mechanisms by which exercise improves cognition. The objective of this application is to use an in vitro model to identify factors released from contracting primary muscle fibers that increase the connectivity and synchronous activation of primary hippocampal neuron cultures. A recent line of investigation has presented muscle-released circulating factors produced during physical activity as potential causal agents driving changes in hippocampal plasticity. However, the full complement of factors, including exosomes, which might contribute to long distance communication between muscles and hippocampal neurons has not been well characterized. Identifying the factors would be useful for therapeutic applications aiming to recapitulate effects of exercise for neuronal regeneration and repair. The central hypothesis is that muscle fiber contractions release factors which have the capability to enhance the rate of maturation of hippocampal neuronal circuits. The hypothesis is supported by preliminary studies showing neuronal cultures exposed to the media from contracting muscle fibers display more rapid maturation of neuronal connections and synchronous activation patterns as compared to neuronal cultures exposed to control media. One of the PIs has a productive research program on exercise-brain interactions, and the other on mechanical micro-environment effects on cell functionality. The PIs have developed multiple innovative methods for powerful hypothesis testing and exploration. The objectives of this application will be accomplished by pursuing 3 specific aims: 1) Determine the extent to which factors released from primary skeletal muscle cells subjected to prescribed range of contraction regimens accelerate synchronous firing of cultured primary hippocampal neurons. 2) Identify novel compounds released from contracting muscle fibers and explore whether exosomes are also released. 3) Determine the extent to which cross-talk between hippocampal neurons and contracting muscle cells affects maturation and connectivity of cultured hippocampal neuronal circuits. A novel platform which allows cross-talk between hippocampal neurons and muscle-motor neuron units but prevents physical contact will be used. State-of-the-art peptidomics methods will lead to the discovery of new molecules released by contracting muscle fibers that influence plasticity of neurons, and new imaging methods will be used to explore whether exosomes are also released. Elucidating and unequivocally establishing mechanisms underlying pro-cognitive effects of exercise holds the key to discover novel and more efficient ways to maintain, promote and improve cognitive performance. The proposed research is highly innovative, it addresses pressing questions in the field using very novel strategies and state-of-art technologies that will allow us to generate causal, mechanistic data on how muscles communicate with hippocampal neurons.