Timothy J. Ebner - US grants

This proposal is a neurophysiological study of the climbing fiber system's role in the cerebellar cortex and motor behavior. The global hypothesis underlying the grant is that the climbing fiber afferent system, possibly acting as an error signal, increases a Purkinje cell's responsiveness to mossy fiber input. The climbing fiber input to neighboring Purkinje cells is tightly correlated, resulting in an accentuation of the gain of a cerebellar cortical region. The altered Purkinje cell output from a cerebellar cortical region is hypothesized to modify motor behavior. Initial studies in acute cats will evaluate climbing fiber afferent modification of Purkinje cell responsiveness to mossy fiber input. Short and long term changes will be evaluated using analytical techniques designed to remove the simple spike signal from the background activity. These techniques will permit statistical evaluation of the changes in simple spike activity associated with climbing fiber afferent input. Using two models of behavior, locomotion in the premammillary cat and voluntary arm movements in the intact primate, the interaction between climbing fiber afferent input and Purkinje cell discharge will be evaluated. The evoked complex spike activity and associated changes in simple spike activity will be related to alterations in walking or limb movements. Statistical analyses are designed to evaluate the relationships between the complex spike discharge, simple spike activity and the movement kinematics. The inferior olive possesses an inherent tendency to oscillate. Acute cat studies will test whether the rhythmicity represents a brief, but stable limit cycle of a population of inferior olivary neurons. The resetting characteristics of spontaneous as well as evoked complex spike discharge will be evaluated and compared. In the primate movement studies the presence or absence of climbing fiber rhythmicity will be evaluated. Associated rhythmicity in the simple spike discharge, limb kinematics or EMG activity will be evaluated. Overall these studies will increase our understanding of the role of the climbing fiber afferent system in the cerebellar cortex and motor behavior.

The premotor cortex, one of the major areas of the cerebral cortex is suggested to be involved in numerous aspects of motor behavior. One idea is that it has a role in the learning of motor skills. Lesions studies suggest deficits in recalling movements from memory. Other studies show the involvement of this part of the cortex in movement preparation. Dr. Timothy Ebner believes this brain area is concerned with the preparation and visual guidance of the limbs or hands during voluntary movements into various regions of reachable extra personal space. His work proceeds under that hypothesis. In primates trained to make voluntary movements in the horizontal plane which require visually guiding a cursor into target zones, the discharge of single neurons in the premotor cortex will be recorded. Four aspects of the cell's activity in relation to complex movements will be studied. First, is to define the region within two dimensional space in which a cell is modulated during reaching movements. Second, the relationship between the discharge pattern of premotor cortical neurons and movement kinematics will be addressed. Third, the responsiveness of premotor cortex neurons to "errors" in motor performance and corrective movement will be evaluated. Fourth, how the discharge of single cells change in the process of adapting a movement to match externally exposed constraints will be determined. Overall, these studies will define which features of two dimensional, visually guided movements are represented in the premotor cortex.

Project 5, T. Ebner: A striking feature of the cerebellar cortex is its two prominent architectures. One is the parasagittal organization of the olivo-cerebellar and cortico-nuclear project, the other is the transversely oriented parallel fibers that run nearly orthogonal to the parasagittal zones. Understanding the function of these two architecture has been hampered by lack of techniques that permit spatial mapping of the activity in the cerebellar cortex. A novel approach using the pH sensitive dye, neutral red, has been developed to spatially map neuronal activation and will be used to image Crus I and II in the rat cerebellar cortex. Experiments are proposed to test the hypothesis that the parasagittal banding pattern represents a map of body surface and stimulus frequency and that activation of the climbing fiber projection is a primary determinant of this parasagittal banding. Experiments are also proposed to test the hypothesis that parallel fibers are activated by peripheral input and contribute to the spatial patterns of activation. Several hypothesis concerning the functional interactions between these two systems will be examined including whether climbing fiber afferents alter the short and/or long term excitability of the responses to surface stimulation and how parallel fibers alter the subsequent responses to peripheral inputs. A limitation of present in vivo optical imaging methodologies is the lack of cell specificity to address this problem lines of transgenic mice have been generated with the gene for cytoplasmic yellow cameleon-2 coupled to the Purkinje cell specific promoter, PCP-2. Yellow cameleon-2 is a Ca++ sensitive fusion protein consisting of two mutant forms of green fluorescence protein linked to Xenopus calmodulin and a 26 residue calmodulin-binding peptide of myosin light chain kinase. Using these transgenic animals, the calcium fluxes occurring in Purkinje cells will be imaged, in vivo, determining the spatial and temporal patterns of Ca++ fluxes evoked by surface, inferior olive, and peripheral stimulation. Overall, these studies will provide a better understanding of how information is processed spatially in the cerebellar cortex.

DESCRIPTION (Adapted from the Investigator's Abstract): This proposal is to continue a series of single unit recordings from Purkinje cells in the pars intermedia and hemisphere of the cerebellar cortex that are related to arm movements or to smooth tracking. Several questions about the tracking signals represented in the discharge of Purkinje cells will be addressed. Specific aim 1 tests and extends the hypothesis that Purkinje cell discharge is tuned to a preferred velocity and direction. Purkinje cell simple spike and complex spike discharge will be evaluated while the animal generates manual, circular tracking movements. The possibility that a population code predicts movement velocity will be tested by analysis of population responses during more complex tracking tasks (spiral tracking). Specific aim 2 is to determine to what degree Purkinje cell discharge is predictive and to what degree it is dependent on visual feedback. This will be accomplished by brief removal of visual feedback during circular tracking. Specific aim 3 will examine the effects of unpredictable perturbations of target motion on the discharge of Purkinje cells. Animals will perform smooth tracking movements along a straight line, and the direction and speed of the visual target that prompts the movement will be perturbed. The fourth specific aim is to compare the correlation of Purkinje cell discharge with movement dynamics to the correlation with movement kinematics. An attempt will be made to maintain the kinematics of manual tracking constant while manipulating the inertial, viscous or elastic forces that are required to produce smooth tracking arm movements with a two-dimensional robot.

DESCRIPTION (Investigator's Abstract): The long term objective of this word is to define the role of the cerebellar cortex in voluntary motor behavior. Understanding normal function is an absolute requirement for understanding the crippling movement abnormalities that result from - cerebellar disease. This proposal 5 specific goals focus on defining both the kinematic and dynamic parameters - of arm movements encoded in the discharge of Purkinje cells. In visually guided, two-dimensional, multijoint arm movements, the discharge of paravermal and hemispheral Purkinje cells in the rhesus monkey will be studied in various tasks designed to vary different movement parameters while holding others constant. Four specific aims are proposed: 1) Determine the velocity sensitivity of Purkinje cells in 2-dimensional tracking movements. Two series of tracking tasks are proposed, one based on tracking targets moving at constant velocities and the other on tracking targets moving with a bell shaped velocity profile. The first task allows the manipulation of movement velocity in relationship to movement-direction keeping acceleration and position constant. The second tracking task will permit sorting out the relative contributions of three highly coupled parameters: velocity, distance, and movement time. 2) Determine the contribution of multiple parameters of movement, including kinematic and dynamic variables, to the simple-and complex spike discharge of Purkinje cells. Two dimensional, horizontal reaching movements of different direction and distance restricted to the elbow and shoulder will be used to quantify not only the kinematics of the hand, but the underlying joint motions and torques. An emphasis will be placed on whether the purkinje cell discharge is more tightly coupled to the movements in relation to the hand motions or in relationship to the motions of the individual points. 3) Determine if the Purkinje cell discharge evoked in response to a perturbation encodes information about the direction and distance of the error or perturbation. In this task, the animal will move from a start box to a target box. As the animal reaches the target box, the target box will be displaced to a new position. Different directions and distances of this target box perturbation will be evaluated. The simple and complex spikes evoked by the perturbation will be analyzed for both directional and distance dependency. 4) Determine whether Purkirne cell discharge is encoded in a motor or sensory reference frame using a vasomotor dissociation task. Monkeys will be trained to produce movements in which the physical trajectory of the hand and the sensory counterpart of that trajectory are equivalent and when the motor and sensory representations are dissociated. These studies will test directly whether the Purkinje cell responses are organued in one or a combination of these reference frames.

Understanding the role of the central nervous system in the normal and abnormal motor behavior will require deciphering what parameters of movement are represented and where. Psychophysical observations support the hypothesis that reaching movements are planned and/or executed using a kinematic framework. Kinematically the direction, distance, velocity and accuracy of a movement are crucial components to positioning the arm in space. While neuronal correlates of movement direction have been studied and described in several cerebral cortical areas, the central representation of other arm movement parameters has received less attention. This proposal will evaluate five issues focused on the encoding of kinematic parameters in the discharge of primate premotor neurons. Emphasis is placed on determining the contributions and relationships between distance, direction, velocity and spatial accuracy in two and three dimensional arm reaching tasks. In the first Specific Aim the dependence of the activity of premotor neurons on the spatial accuracy of a movement will be evaluated. In the second Specific Aim a two dimensional tracking task will be used to dissect out relative importance and interactions between velocity, distance and movement time to the discharge of premotor cortical cells. In the third Specific Aim the neuronal correlates of movement amplitude will be studied in a three dimensional reaching task. In the fourth Specific Aim we propose to systematically alter the visual feedback, introducing errors into the movement. Using this complex, visually guided task, it will be determined whether the encoding of distance and direction remains invariant and whether the visuomotor "errors" are encoded in these cells' discharge. Lastly, in the fifth Specific Aim the premotor cortex has been hypothesized to play a role in learning the arbitrary associations that couple a movement to a stimulus. Using a learning paradigm which requires the scaling of movement kinematics due to a feedback change, we propose to study how adaptation of movement direction and distance is encoded at the single cell level. Overall, these studies will better define the role the premotor cortex plays in motor behavior.

This study, as originally proposed, uses single neuron recording techniques in behaving primates with the aim of deciphering the movement parameters encoded and the nature of the information represented in the cerebellum. The cerebellar cortex and nuclei play a major role in the production of smooth, coordinated movements. There are a host of hypotheses on the cerebellum's role in motor behavior. Testing the validity of any hypothesis of cerebellar function will require defining the parameters that modulate the functional "activity" in the cerebellum and understanding how these parameters are encoded or represented in the discharge of cerebellar neurons. These are crucial steps in understanding the variables controlled and the information processed by the cerebellum. This conceptual approach forms the basis for this work initially proposed only at the level of neuron recording in behaving monkeys but extended to synergistic studies in humans with the recent exciting developments in functional magnetic resonance imaging. In the last funding year, our aim has been to define in the whole brain areas in the cerebral and cerebellar cortex and subcortical nuclei engaged in the execution of a complex, visually guided movement task. Accurate, visually guided movements require a complex series of integrative steps that transform retinal information into an appropriate motor output. Two major pathways have been implicated in the function of the primate visual system during spatial control of motor movement, the dorsal and the ventral stream, respectively. The dorsal stream connects the striate and extrastriate cortices to areas around the intraparietal sulcus and posterior parietal region. The posterior parietal region connects reciprocally to the motor, premotor and prefrontal cortices. Furthermore, there is evidence of thalamic involvement (e.g. pulvinar) as an important relay station as well as participation of cortico-ponto-cerebellar pathways. The dorsal stream is thought to be involved in the transformation of visual information into motor commands, whereas the ventral stream, projecting from the occipital cortex to the inferotemporal lobe, is proposed to be involved in off-line object recognition. The aim of the present experiments is to examine the hypothesis that in humans, the cortical stream of activation and their subcortical and cortico-cerebellar modulations will correspond to the presumed visuomotor pathways mapped in primates. The paradigm employed for these studies requires subjects to use a joystick to move a hair cross cursor on a rear projection screen. Two sets of eight targets are arranged in circle (45o intervals) around a center start box and appear at two different distances in the same direction. During the task periods, the participants must use the joystick to move the cross-hair from the center start box first to the shorter distance target and then to the target at the greater distance. During the rest periods the volunteers lie motionless looking at a colored box on a lighted screen. Various other tasks are also planned to control for just oculomotor activity, or motor function without visual guidance, etc. Data evaluation relies on first generating individual maps, and then warping each individual map to Talairach coordinates. Subsequently, a composite activation map is generated by superimposing all individual maps and displaying a pixel as "activated" if and only if it was considered "activated" in at least 75% the individual maps. Based on these composite maps a final map is created displaying the results of the comparison between tasks. In a set of completed studies that will be presented as an abstract in the ISMRM meeting in April 1997, we observed a pattern of activation during the visually guided movement that is consistent with the flow of information from striate and extrastriate areas, to the superior and inferior parietal lobe and to frontal motor areas, similar to that of the dorsal stream described in primates.

9870633 Ebner This Integrative Graduate Education and Research Training (IGERT) award will support the establishment of a broadly- based graduate training program that will equip students to use relevant the technologies and insights of computational and physical science in investigation of fundamental problems in neuroscience. The project will be a joint effort of 24 faculty from 8 departments, with collaborative research interests ranging from use of numerical methods in modeling of limb motor control to modeling of synaptic communication in the retina and to 3-dimensional representation of neurophysiological data such as those derived from functional magnetic resonance imaging. NSF support will provide stipends for 13 graduate students each year who will enroll in either the neuroscience or scientific computation Ph.D. programs. Educational opportunities include required six-week summer course in cell and molecular neurobiology, a new year-long course in scientific computation, and a new seminar series in computational neuroscience that will bring 12 speakers from academia and industry to the campus each year. After completing short projects in the research groups of three participating faculty (lab rotations), students will choose two mentors for their thesis research, one from the 12 participating neuroscience faculty, the other from among the 12 computational science faculty. In addition to use of existing and separate recruitment efforts of the two Ph.D. programs, the IGERT-sponsored program will benefit from joint efforts of participating faculty and an ongoing collaborative effort with the Florida A&M University. IGERT is a new, NSF-wide program whose goal is to sponsor the establishment of innovative, research-based graduate programs that will train a diverse group of new scientists and engineers to be well-prepared for varied careers in the private and public sectors. IGERT provides an opportunity for the development of new, well-focuse d multidisciplinary programs that bridge traditional organizational barriers, uniting faculty from several departments or institutions to establish a highly-interactive collaborative environment for both training and research. In its first year, the program will provide support to 17 institutions for new or nascent programs that collectively span all areas of science, engineering and mathematics eligible for support by the NSF.

The global problem addressed by this proposal is how does the central nervous system (CNS) control a structure as complex as the primate hand? To begin to address this question, four Specific Aims are proposed that address three outstanding issues in the neural control of reach-to-grasp. The studies will use chronic single unit recordings in the primary motor cortex (M1) of monkeys trained to perform various reach-to-grasp movements. Extensive monitoring of arm and hand kinematics, grasp forces and the activity of arm and extrinsic/intrinsic muscles of the hand will be done. Statistical and analytical tools including regression analyses and data reduction techniques will be used to extract the neural representation of hand shape/object shape, kinematics, grasp forces and EMG activity. First, findings from psychophysical, lesion, and electrophysiological studies suggest that the CNS, at least in part, reduces the number of degrees of freedom by controlling the hand as a unit. Specific Aims 1 and 2 will address the hypothesis that neurons in the hand area of M1 encode and control hand shape/object properties, reflecting this global control of the hand. Second, early investigations into hand movements categorized prehension into two broad classes, power and precision grip and more elaborate classification systems followed. A prediction of these categorical schemes is that the CNS explicitly controls grasp type. More recent psychophysical studies, however, suggest that a strict division of hand posture into power and precision grasps does not occur. Specific Aim 3 will test the alternative hypothesis that power and precision are part of a continuum of hand postures in which hand shape is primarily controlled. A third major contemporary hypothesis is that reach and grasp are controlled by two independent but coupled channels: a "transport" channel that extracts information about the spatial location of objects to guide the reach and a "manipulation" channel that extracts information about the intrinsic properties of the object such as size and shape to guide hand shape. Although psychophysical results suggest that the two components are coupled, there is virtually no single unit data addressing this question. In Specific Aim 4 the hypothesis is tested that the reach and grasp components are coupled at the neuronal level in M1. In general understanding how the CNS controls prehension is a critical step in understanding human movements. In the future understanding the signals controlling grasp could prove useful for controlling prosthetic devices in patients with brain injury.

The proposed interdisciplinary training program at the University of Minnesota combines graduate training in the computational, chemical, physical, and engineering sciences with graduate training in neuroscience. Neuroscience is a highly interdisciplinary field that uses a variety of experimental approaches to understand the development, structure, and function of the nervous system. As neuroscience matures, the need grows for quantitative modeling, physical and chemical insights, advanced technologies, and state-of-the-art hardware and software that the computational, chemical, physical and engineering sciences can provide. Therefore, interdisciplinary graduate training is needed to take the maximum advantage of these opportunities. The proposal incorporates the strengths, resources, and administrative structures of several existing graduate programs and the University of Minnesota Supercomputing Institute with an interdisciplinary faculty with diverse research interests, to provide a new paradigm in graduate education. We propose to offer 5 three-year Fellowships each year to attract outstanding pre-doctoral students. Enrolling in existing degree programs in Biomedical Engineering, Chemistry, Computer Science, Mathematics, Neuroscience, Physics and Scientific Computation. The Fellows will be trained across disciplines using a variety of tools including special interdisciplinary coursework, research rotations, dual thesis advisors, special seminars and symposia and unique training opportunities. Each trainee's thesis work will cross the disciplines of neuroscience and the physical/computational sciences. An advisory system will help guide students through the program. Also, several mechanisms are proposed to evaluate the effectiveness of the training program. The trainees will receive instructions in the responsible conduct of research. The proposal documents the efforts that the training program and University will make to ensure that traditionally underrepresented students are recruited and included in the program. On completion the trainees will be prepared for research careers in academia, industry and government. The overall goal is to train the next generation of scientists who can bridge the gap between biology and the physical/computational sciences. Broader impacts include advancing our understanding of the brain, cross-fertilization of the disciplines, and establishing a new model for interdisciplinary graduate training.

DESCRIPTION (provided by applicant): The proposed interdisciplinary training program at the University of Minnesota combines graduate training in the computational, chemical, physical, and engineering sciences with graduate training in neuroscience. Neuroscience is a highly interdisciplinary field that uses a variety of experimental approaches to understand the development, structure, and function of the nervous system. As neuroscience matures, the need grows for quantitative modeling, physical and chemical insights, advanced technologies, and state-of-the-art hardware and software that the computational, chemical, physical and engineering sciences can provide. Therefore, interdisciplinary graduate training is needed to take the maximum advantage of these opportunities. The proposal incorporates the strengths, resources, and administrative structures of several existing graduate programs and the University of Minnesota Supercomputing Institute with an interdisciplinary faculty with diverse research interests, to provide a new paradigm in graduate education. We propose to offer 5 three-year Fellowships each year to attract outstanding pre-doctoral students: Enrolling in existing degree programs in Biomedical Engineering, Chemistry, Computer Science, Mathematics, Neuroscience, Physics and Scientific Computation. The Fellows will be trained across disciplines using a variety of tools including special interdisciplinary coursework, research rotations, dual thesis advisors, special seminars and symposia and unique training opportunities. Each trainee's thesis work will cross the disciplines of neuroscience and the physical/computational sciences. An advisory system will help guide students through the program. Also, several mechanisms are proposed to evaluate the effectiveness of the training program. The trainees will receive instructions in the responsible conduct of research. The proposal documents the efforts that the training program and University will make to ensure that traditionally underrepresented students are recruited and included in the program. On completion the trainees will be prepared for research careers in academia, industry and government. The overall goal is to train the next generation of scientists who can bridge the gap between biology and the physical/computational sciences. Broader impacts include advancing our understanding of the brain, cross-fertilization of the disciplines, and establishing a new model for interdisciplinary graduate training.

DESCRIPTION (provided by applicant): The proposed interdisciplinary training program at the University of Minnesota combines graduate training in the computational, chemical, physical, and engineering sciences with graduate training in neuroscience. Neuroscience is a highly interdisciplinary field that uses a variety of experimental approaches to understand the development, structure, and function of the nervous system. As neuroscience matures, the need grows for quantitative modeling, physical and chemical insights, advanced technologies, and state-of-the-art hardware and software that the computational, chemical, physical and engineering sciences can provide. Therefore, interdisciplinary graduate training is needed to take the maximum advantage of these opportunities. The proposal incorporates the strengths, resources, and administrative structures of several existing graduate programs and the University of Minnesota Supercomputing Institute with an interdisciplinary faculty with diverse research interests, to provide a new paradigm in graduate education. We propose to offer 5 three-year Fellowships each year to attract outstanding pre-doctoral students: Enrolling in existing degree programs in Biomedical Engineering, Chemistry, Computer Science, Mathematics, Neuroscience, Physics and Scientific Computation. The Fellows will be trained across disciplines using a variety of tools including special interdisciplinary coursework, research rotations, dual thesis advisors, special seminars and symposia and unique training opportunities. Each trainee's thesis work will cross the disciplines of neuroscience and the physical/computational sciences. An advisory system will help guide students through the program. Also, several mechanisms are proposed to evaluate the effectiveness of the training program. The trainees will receive instructions in the responsible conduct of research. The proposal documents the efforts that the training program and University will make to ensure that traditionally underrepresented students are recruited and included in the program. On completion the trainees will be prepared for research careers in academia, industry and government. The overall goal is to train the next generation of scientists who can bridge the gap between biology and the physical/computational sciences. Broader impacts include advancing our understanding of the brain, cross-fertilization of the disciplines, and establishing a new model for interdisciplinary graduate training.

The proposed interdisciplinary training program at the University of Minnesota combines graduate training in the computational, chemical, physical, and engineering sciences with graduate training in neuroscience. Neuroscience is a highly interdisciplinary field that uses a variety of experimental approaches to understand the development, structure, and function of the nervous system. As neuroscience matures, the need grows for quantitative modeling, physical and chemical insights, advanced technologies, and state-of-the-art hardware and software that the computational, chemical, physical and engineering sciences can provide. Therefore, interdisciplinary graduate training is needed to take the maximum advantage of these opportunities. The proposal incorporates the strengths, resources, and administrative structures of several existing graduate programs and the University of Minnesota Supercomputing Institute with an interdisciplinary faculty with diverse research interests, to provide a new paradigm in graduate education. We propose to offer 5 three-year Fellowships each year to attract outstanding pre-doctoral students. Enrolling in existing degree programs in Biomedical Engineering, Chemistry, Computer Science, Mathematics, Neuroscience, Physics and Scientific Computation. The Fellows will be trained across disciplines using a variety of tools including special interdisciplinary coursework, research rotations, dual thesis advisors, special seminars and symposia and unique training opportunities. Each trainee's thesis work will cross the disciplines of neuroscience and the physical/computational sciences. An advisory system will help guide students through the program. Also, several mechanisms are proposed to evaluate the effectiveness of the training program. The trainees will receive instructions in the responsible conduct of research. The proposal documents the efforts that the training program and University will make to ensure that traditionally underrepresented students are recruited and included in the program. On completion the trainees will be prepared for research careers in academia, industry and government. The overall goal is to train the next generation of scientists who can bridge the gap between biology and the physical/computational sciences. Broader impacts include advancing our understanding of the brain, cross-fertilization of the disciplines, and establishing a new model for interdisciplinary graduate training.

DESCRIPTION (provided by applicant): Multiple forms of activity dependent, synaptic plasticity have been described in the cerebellar cortex. Hypothesized to play a central role in motor learning, the overall goal of this proposal is to address key questions concerning cerebellar synaptic plasticity in the intact animal. The experimental approach is based on optical imaging of activity in the mouse cerebellar cortex to evaluate the spatial and temporal aspects of the long-term changes. Theoretical models and behavioral data require that parallel fiber (PF) activity precede climbing fiber (CF) input for PF-Purkinje cell (PC) long-term depression (LTD) to support learning. Therefore, Specific Aim 1 investigates the optimal timing and stimulation parameters for evoking PF-PC LTD in vivo. Specific Aim 1 also characterizes the signaling requirements, examines whether granular layer and mossy fiber stimulation generates PF-PC LTD, and tests the hypothesis that PF-PC LTD should modify the responses evoked in the cerebellar cortex by peripheral inputs. Long-term potentiation (LTP) has been described at the PF-PC synapse in vitro. Specific Aim 2 tests in vivo whether LTP occurs at the PF-PC synapse, characterizing the stimulation parameters, stimulation at different stages in the circuitry, signaling requirements and spatial characteristics. Specific Aim 2 also examines how PF-PC LTP modifies the responses evoked by peripheral inputs. Central to theories of the cerebellum's role in motor learning is the reversibility of both LTD and LTP. Reversibility has only been demonstrated in vitro with little information on reversibility in vivo. Specific Aim 3 tests whether induction of LTP can reverse PF-PC LTD and conversely whether induction of LTD can reverse PF-PC LTP. Recent evidence suggests that the synaptic connections between the PFs and the molecular layer inhibitory interneurons are modifiable. Using an optical imaging methodology that allows monitoring the off-beam inhibition evoked by PF stimulation, Specific Aim 4 investigates the induction properties, spatial characteristics, and signaling mechanisms of these long-term changes in the molecular layer inhibitory network. Understanding the role of cerebellar synaptic plasticity will require studying LTD and LTP in the awake, behaving animal. Therefore, Specific Aim 5 first characterizes LTD and LTP in the awake animal. Then Specific Aim 5 also tests the hypothesis that LTD and LTP contribute to the changes in cerebellar cortical activity that occur with adaptation to a mechanical perturbation of a forelimb reaching movement. The goal is to directly link the changes in the cerebellar cortex occurring during motor learning to synaptic plasticity at the PF-PC synapses.

The proposed interdisciplinary training program at the University of Minnesota combines graduate training in the computational, chemical, physical, and engineering sciences with graduate training in neuroscience. Neuroscience is a highly interdisciplinary field that uses a variety of experimental approaches to understand the development, structure, and function of the nervous system. As neuroscience matures, the need grows for quantitative modeling, physical and chemical insights, advanced technologies, and state-of-the-art hardware and software that the computational, chemical, physical and engineering sciences can provide. Therefore, interdisciplinary graduate training is needed to take the maximum advantage of these opportunities. The proposal incorporates the strengths, resources, and administrative structures of several existing graduate programs and the University of Minnesota Supercomputing Institute with an interdisciplinary faculty with diverse research interests, to provide a new paradigm in graduate education. We propose to offer 5 three-year Fellowships each year to attract outstanding pre-doctoral students. Enrolling in existing degree programs in Biomedical Engineering, Chemistry, Computer Science, Mathematics, Neuroscience, Physics and Scientific Computation. The Fellows will be trained across disciplines using a variety of tools including special interdisciplinary coursework, research rotations, dual thesis advisors, special seminars and symposia and unique training opportunities. Each trainee's thesis work will cross the disciplines of neuroscience and the physical/computational sciences. An advisory system will help guide students through the program. Also, several mechanisms are proposed to evaluate the effectiveness of the training program. The trainees will receive instructions in the responsible conduct of research. The proposal documents the efforts that the training program and University will make to ensure that traditionally underrepresented students are recruited and included in the program. On completion the trainees will be prepared for research careers in academia, industry and government. The overall goal is to train the next generation of scientists who can bridge the gap between biology and the physical/computational sciences. Broader impacts include advancing our understanding of the brain, cross-fertilization of the disciplines, and establishing a new model for interdisciplinary graduate training.

DESCRIPTION (provided by applicant): The proposed interdisciplinary training program at the University of Minnesota combines graduate training in the computational, chemical, physical, and engineering sciences with graduate training in neuroscience. Neuroscience is a highly interdisciplinary field that uses a variety of experimental approaches to understand the development, structure, and function of the nervous system. As neuroscience matures, the need grows for quantitative modeling, physical and chemical insights, advanced technologies, and state-of-the-art hardware and software that the computational, chemical, physical and engineering sciences can provide. Therefore, interdisciplinary graduate training is needed to take the maximum advantage of these opportunities. The proposal incorporates the strengths, resources, and administrative structures of several existing graduate programs and the University of Minnesota Supercomputing Institute with an interdisciplinary faculty with diverse research interests, to provide a new paradigm in graduate education. We propose to offer 5 three-year Fellowships each year to attract outstanding pre-doctoral students: Enrolling in existing degree programs in Biomedical Engineering, Chemistry, Computer Science, Mathematics, Neuroscience, Physics and Scientific Computation. The Fellows will be trained across disciplines using a variety of tools including special interdisciplinary coursework, research rotations, dual thesis advisors, special seminars and symposia and unique training opportunities. Each trainee's thesis work will cross the disciplines of neuroscience and the physical/computational sciences. An advisory system will help guide students through the program. Also, several mechanisms are proposed to evaluate the effectiveness of the training program. The trainees will receive instructions in the responsible conduct of research. The proposal documents the efforts that the training program and University will make to ensure that traditionally underrepresented students are recruited and included in the program. On completion the trainees will be prepared for research careers in academia, industry and government. The overall goal is to train the next generation of scientists who can bridge the gap between biology and the physical/computational sciences. Broader impacts include advancing our understanding of the brain, cross-fertilization of the disciplines, and establishing a new model for interdisciplinary graduate training.

Overview and Significance: The Steering Committee meets quarterly to review Center operations. During the first year, the meeting schedule would be more frequent (every 4-6 weeks) to ensure the Cores are operating optimally and to facilitate solving problems. The Steering Committee held eight meetings in the last year. In addition to committee memliers, all investigators (primary, new, and other NINDS-funded) are welcome and informed in advance of the agenda by both email and notice on the Center website. The meetings serve as a forum for the faculty to suggest needed capabilities, feedback on how the Cores are functioning, and possibilities for collaborations. Meeting minutes are circulated to Center faculty and posted on the website. Institutional Sharing Plan: The Steering Committee oversees and implements the Institutional Sharing Plan. Prior to using any of the three research Cores, the interested Pl(s) submit a brief description (250 words) for the proposed project to the Core Director. Once the project is approved, the PI meets with the Core Director and Manager to discuss the goals, feasibility, timeline, and to determine the best experimental approach for successful completion of the project. The four priority levels are: 1) Primary investigators with qualifying NINDS grants, 2) NINDS-funded investigators without qualifying grants and new investigators, 3) neuroscience investigators funded by other NIH institutes, and 4) investigators lacking NIH support. Within each level collaborative projects are given the highest priority. The third and fourth priority levels are allowed access to Center resources when demands permits and assures as wide as possible Center impact. For several services and equipment use (e.g. tract tracing, confocal microscope and some behavioral tests), the primary mechanism for prioritization is the advance time for sign-up. NINDS-funded primary investigators (see Table 1 A) have the highest priority. Within this group, top priority goes to projects that involve collaborative efforts between primaiy users with up to 15 days advance sign-up. Primary users not engaged in collaborative projects are able to sign up 12 days in advance. New investigators (Table 3) and NINDS-funded investigators who do not have qualifying grants have the next level of priority, again with the higher access given to projects that are inherently collaborative. Investigators in this priority level engaged in collaborative projects are allowed to sign up 10 days in advance. Investigators working on individual projects are able to sign up 7 days in advance. Other NIH-funded investigators are permitted to reserve core facilities 5 days in advance. Finally, those individuals lacking major external funding are able to sign up (2 days in advance). Several of the services, such as generating a BAC transgenic mouse, engineering a viral vector, performing activity-dependent optical imaging or detailed behavioral phenotyping, are more time intensive and depend on the nature and complexity of the project and the ongoing workload of a Core. Therefore, access to these services cannot rely simply on sign-up sheets. For these services, a monthly deadline for submission of these projects is advertised with priority determined as indicated above. For these services the Core Director then schedules projects based on the order of priority described above and on-going projects. The priority schedule will adjusted if the waiting time for services exceeds demands and waiting times exceed a month. If this occurs, 80% of available time is resen/ed for primary NINDS-funded or new investigators and the remaining 20% reserved for lower-priority investigators. This policy ensures the higher-priority investigators have the greatest access but allows some access for lower-priority users. As needed, these percentages would be adjusted by the Steering Committee. For example, if demand is particulariy great, access would be restricted to the first two priority levels (e.g. 70% for level one and 30% for level two), again ensuring some level of access to NINDS-funded investigators without qualifying grants. On a longer time horizon, the Steering Committee can reallocate resources between Cores based on the patterns of usage and demand. This could, for example, involve reallocating University matching funds among the Cores. The Steering Committee adjudicates sharing issues not resolvable at the Core level. Faculty members are encouraged to provide feedback to the individual Core Directors and/or Steering Committee. At the completion of a project, each Core asks users to complete a survey that includes questions about the quality, timing and added value of the services provided. The survey also requests feedback on other services/instruments needed, ways to improve services, etc. The surveys are used by the Core Directors, Scientific Advisory Committees and the Steering Committee to guide Center development.

This Integrative Graduate Education and Research Traineeship (IGERT) award supports the development of a multi-disciplinary, integrative graduate education and training program in NeuroEngineering (NE) at the University of Minnesota at Twin Cities. Intellectual Merit: The purpose of this program is to train doctoral students to develop the skills to revolutionize technologies for interfacing with the brain and advance the fundamental understanding of neuroscience processes that arise when interfacing with and modulating the brain.

Broader impacts include the development of a multi-disciplinary training program that blurs the boundary between neuroscientists and engineers, thus enabling a new generation of scientists to competently and confidently take on the grand challenges in the interdisciplinary field of NeuroEngineering. The NE program includes major research themes in decoding brain signals, modulating brain dynamics, and bi-directional brain interfacing. The program is a "degree-plus" model in which pre-doctoral students are admitted to one of the participating graduate programs (Biomedical Engineering, Electrical Engineering, Mechanical Engineering, and Neuroscience), and are trained through a series of hands-on, modular neuroengineering courses. All NE Fellows will immerse themselves in a lab outside their major in the summer of their first year, engage in multiple lab rotations, and participate in at least one clinical lab rotation, summer internship at a neurotechnology company, or summer international research experience. NE Fellows will have co-advisors beginning in their first year, one from the engineering sciences and one from the basic or clinical neurosciences. The training program incorporates several outreach efforts to recruit women and underrepresented minorities, provide outreach to K-12 and industry, and train NE Fellows to be effective communicators.

IGERT is an NSF-wide program intended to meet the challenges of educating U.S. Ph.D. scientists and engineers with the interdisciplinary background, deep knowledge in a chosen discipline, and the technical, professional, and personal skills needed for the career demands of the future. The program is intended to catalyze a cultural change in graduate education by establishing innovative new models for graduate education and training in a fertile environment for collaborative research that transcends traditional disciplinary boundaries.

This is a proposal for an NINDS Institutional Center Core grant to support neuroscience research at the University of Minnesota. The mission of the Center and its Cores is to enhance basic and translational research on the nervous system by providing state-of-the art, shared equipment facilities with technical and scientific support. The Center brings together a primary group of 11 NINDS-funded investigators using mouse models to study human neurological disease/disorders and nervous system function. The Center also gives high priority to helping new neuroscience investigators establish their research programs and NINDS-funded investigators without qualifying grants. The Center also offers its services and resources when possible to a large group of investigators, many NIH funded, whose research is focused on the nervous system. The three research Cores provide technological services, training and scientific expertise to increase productivity, promote new research directions and foster collaborations among Center investigators. The Cores were selected to provide access to widely needed, even critical, tools used in the study of mouse models. These tools are best offered in a central facility in which investigators can take advantage of standardized and optimized protocols as well as expert technical assistance. The Cores were also selected to provide access to new, powerful tools for the investigation of nervous system function/dysfunction not generally available to individual investigators. By focusing on the study and evaluation of mouse models, there is a high potential for innovative synergistic effects across the Cores. The Genetic Manipulation Core provides the generation of bacterial artificial chromosome transgenic mice and viral vectors to manipulate gene expression in vitro and in vivo. The Behavioral Phenotyping Core provides facilities and technical support for the evaluation of a spectrum of motor and cognitive behaviors as well as general neurological status in the mouse. The Imaging and Tract Tracing Core provides investigators with the tools and expertise for modern imaging (single and two-photon microscopy), tract tracing/histology, and activity-dependent optical imaging. The fourth Administrative Core with the Executive Committee is responsible for Center oversight, administers the institutional sharing plan, promotes collaborations, informs the neuroscience community, seeks new technologies and evaluates the performance of the Center. The Center and Cores will increase the impact and productivity of NINDS-funded investigators, promote new research directions and foster collaborations among Center researchers.

DESCRIPTION (provided by applicant): The cerebellum is essential for the control of movement and plays an important role in cognitive processes. Within the cerebellum, estrogens have long been demonstrated to affect synaptic circuitry and function. Yet to date, the mechanisms by which estradiol can alter cerebellar output have remained a mystery. Using state-of-the-art optical imaging techniques, we have found estrogen receptor signaling to be essential for the maintenance of the parallel fiber synapses connecting cerebellar granule cells to Purkinje neurons. Further, we have found that estrogen receptor regulation of the parallel fiber synapse to be dependent on locally synthesized estradiol. These effects are observed in both male and female mice. Since parallel fibers dominate the cytoarchitecture of the cerebellar cortex, and are central elements in modulating cerebellar function, we believe these data are the beginnings to our understanding of a new fundamental mechanism of estrogen action. Using the combined expertise and experiences of two complementary investigators at the University of Minnesota, our goal is to characterize this phenomenon. Specific Aim 1 will determine whether electrical activity within the parallel fiber-Purkinje cell network drives local neuroestrogen production. Specific Aim 2 will identify the estrogen receptor(s) responsible for maintaining the efficacy of synaptic neurotransmission between the cerebellar granule neurons and Purkinje cells. This Aim will also determine whether the estrogen receptors are located pre- or postsynaptically. Specific Aim 3 will evaluate the functional impact of diminishing estrogen signaling on cerebellar-mediated locomotor behavior. By characterizing the critical regulatory mechanisms for estrogen receptor support of cerebellar glutamatergic neurotransmission, we will have uncovered a heretofore unknown aspect of nervous system function. In addition, these results will have important implications in the use of estrogen receptor modulators to benefit health and its impact in the treatment of disease. PUBLIC HEALTH RELEVANCE: The goal of this proposal is to characterize the signaling pathway by which locally-synthesized estradiol acts within the cerebellum to support glutamatergic neurotransmission in the parallel fiber network, that is, the connection between cerebellar granule neurons and Purkinje cells. Understanding the requirement of estrogen receptor activation in support of the parallel fiber synapse has important implications not only in cerebellar function, but will impact our perspective regarding the side effects of tamoxifen and aromatase inhibitor therapies. In addition, our work will provide insight into estrogenic support o glutamatergic synaptic function in other brain areas.

The Neuropathology and Functional Imaging Core will provide neuropathological and physiological services focused on the evaluation of both patients with myotonic dystrophy and mouse models of the disease. The Neuropathology component will provide neuropathological characterization of patients who have died with myotonic dystrophy types 1 and 2 (DM1 and DM2). The objective is to define the distribution and nature of the neuropathological alterations associated with DM1 and DM2, with a particular emphasis on correlation with antemortem neuroimaging and/or clinical CNS-related deficits in patients who have been studied in Project 3. The Core will aid investigators in Projects 1 and 2 by providing histological services and tissues with a major focus on correlating the neuropathological findings in the transgenic mouse models with those of the corresponding human diseases to understand pathological mechanisms. The Functional Imaging component of the Core will provide functional assessments of the cerebral cortical circuitry in the mouse models developed by Projects 1 and 2 and test in the mouse models for abnormalities identified in DM patients by Project 3. The Core will use optical imaging and single cell electrophysiological recordings to characterize the cerebral cortical circuitry in vivo. As DM patients have exaggerated sensitivity to sedatives and stimulants, the Core will investigate whether cerebral cortical circuits show altered sensitivity to these drugs. Flavoprotein imaging has revealed correlations in the background fluorescence among different regions in the cerebral cortex that can be used to assess functional connectivity. The alterations in white matter integrity and grey matter in DM is likely to result in changes in functional connectivity. Therefore, Core A will also test whether this functional connectivity in the cerebral cortex is abnormal in the DM mouse models. Finally, Projects 1 and 2 are proposing to develop mouse models with the goal of reversing the disease phenotypes. Having defined specific circuit abnormalities in the cerebral cortex, the Core will evaluate if the abnormalities are corrected in these rescue mouse models.

DESCRIPTION (provided by applicant): Disorders of the nervous system are becoming more prevalent in our society, especially considering the growing number of people with neurological disorders as our population ages. To meet the challenge of developing new and effective therapies to treat these disorders, we need to consistently inspire intelligent and talented undergraduate students to enter careers in neuroscience research. This need is particularly acute among populations of students who are currently underrepresented in the field of neuroscience research. Published analyses have made it clear that making students aware of research fields early in their college careers, especially by involving them directly in the research process, is an extremely effective way of developing a student's interest in research as a future profession. For over 25 years, the University of Minnesota has recognized and met this challenge by offering summer residential research programs in biomedical sciences. This proposal is to fund a neuroscience component of these summer programs in which we will train 8 undergraduate students who have completed their freshman or sophomore years in college. We will recruit students nationally, focusing on students from groups that are underrepresented within the neuroscience research profession. We will provide them with a 10 week intensive research experience that will include professional mentoring (academic survival skills and preparation for graduate school) as well as workshops on research ethics. Our goals are to inspire a new generation of neuroscience researchers as well as to create a national mentoring pool who will accept that responsibility for future generations of students. In turn, we expect these individuals to become part of the research infrastructure dedicated to solving medical problems of nervous system dysfunction.

Project Summary Sensory prediction errors (SPEs) are generated by comparing the sensory consequences of a motor command with the actual sensory feedback. Sensory prediction errors are hypothesized to be the critical error signals for model-based, implicit motor adaptation. However, error processing is not static. Instead, the CNS tunes error sensitivity and sensory feedback to match task demands. Also, updating of movements implies that information about errors occurring during a movement must persist to influence subsequent movements. One possibility is that information about motor errors is retained between movements. In spite of the importance of errors in motor control, very little is known about how SPEs are encoded and processed at the neuronal level. A long standing hypothesis is that the cerebellum implements a forward internal model that predicts the sensory consequences of a motor command and uses SPEs to control movements and update the model. For nearly 50 years, the accepted hypothesis has been that the low frequency complex spike (CS) discharge of Purkinje cells encodes motor errors. However, considerable evidence shows that error signaling in the cerebellum does not rely solely on CS discharge. Recently, our laboratory demonstrated that the high frequency simple spike (SS) discharge of Purkinje cells robustly encodes both predictions and sensory feedback about motor errors. Based on these novel observations, this proposal tests a series of hypotheses on the error signals encoded in the SS discharge of Purkinje cells in Rhesus monkeys during pseudo-random tracking tasks. By controlling the visual feedback available during tracking, Specific Aim 1 tests the hypotheses that the SS error signals at lag times are due to visual feedback and are used to compute SPEs. Furthermore, we will test whether the feedback error signals adapt to match the new feedback conditions. Specific Aim 2 tests the hypotheses that the SS error signals at feedforward times are predictions of the upcoming errors and are also used to generate SPEs by introducing time delays in visual feedback. Specific Aims 1 and 2 also examine how CS modulation changes with altering the visual feedback. We hypothesize that CS modulation is driven by an increase in SPEs. Specific Aim 3 tests the hypothesis that the SS discharge encodes for a working memory of errors. In addition, Aim 3 tests a complementary hypothesis that the SS discharge predicts errors up to several seconds prior to movement. Specific Aim 4 examines how motor errors are represented in the firing of cerebellar nuclear neurons, the target of Purkinje cells and the output stage of the cerebellum. Overall, the experiments will provide a comprehensive examination of error signaling in the firing of Purkinje cells and cerebellar nuclear neurons, including how predictive and feedback signals adapt to match changes in feedback and if these neurons encode a working memory of motor errors. The results have the potential to fundamentally change our view of how the cerebellum encodes and processes motor errors.

Our overall objective is to develop the first sets of Alzheimer?s disease related dementia (ADRD) mouse lines that model the genetics of ADRDs as closely as possible. These models will serve as experimental systems for probing the molecular dysfunctions caused by pathogenic ADRD mutations, identifying quantifiable early-stage endophenotypes directly linked to these mutations, and developing and testing therapeutic interventions for correcting these dysfunctions. To make these models, we have developed Gene Replacement (GR) technology that allows us to replace mouse genes with their full human orthologs up to several hundred kb in size. We used this technology to generate a MAPT-GR line of mice in which we replaced the full mouse Mapt genomic coding and regulatory region (156,547 bp) with the full human MAPT genomic sequence (190,081 bp). We have confirmed that mice homozygous for this MAPT-GR allele express human tau at endogenous levels, and that all expected splice variants are found in the appropriate tissues and in ratios expected for the fully functional human MAPT gene. Our specific aims for the R61 phase of this project are to 1) generate five lines of mice that precisely match our first wt MAPT-GR control line except for the pathogenic frontotemporal dementia with parkinsonism-17 (FTDP-17) mutation that we specifically introduce; 2) identify quantifiable endophenotypes that are significantly different between pathogenic MAPT-GR variant lines and the wt control, with and without external insult (i.e., head trauma); and 3) begin to generate additional sets of GR lines of mice in which other genes involved in the etiology of ADRD have been replaced by their human homologs. Once we have achieved these goals, our specific aims for the R33 phase are to: 1) release the matched set of MAPT- GR lines for distribution without restriction; 2) conduct full longitudinal characterization of identified tau- associated endophenotypes, neuropathology and behavior of the MAPT-GR lines; and 3) generate additional matched sets of ADRD-GR mouse lines similar to the MAPT-GR lines, namely C9orf72-GR (amyotrophic lateral sclerosis- frontotemporal dementia; ALS-FTD), SNCA-GR (dementia with Lewy bodies), MATR3-GR (ALS- FTD), and GRN-GR (FTD). Our contributions here are expected to be: a) sets of full human gene-replacement ADRD mouse models that are completely defined at the genetic level, have precisely matched control lines, and mimic the human genetics of ADRD; and b) identified, quantifiable early-stage endophenotypes closely linked to pathogenic mutations in the human MAPT gene. These contributions will be significant because they will provide new tools to interrogate molecular disease mechanisms, identify therapeutic targets, and develop effective therapies. These precisely matched sets of animal models will allow the research community to evaluate the molecular impact of pathogenic mutations within the context of the human genomic sequence in which they occur in patients, and these mouse lines will contain all potential human therapeutic targets ranging from the full genomic DNA sequences to all RNA transcription variants and protein products that they encode.

PROJECT SUMMARY: Imaging Cells during Behavior Core Addiction is a chronic, relapsing disorder due to perturbations in neural circuits. To better understand the underlying pathophysiological processes occurring in addiction and the actions of new therapies, there are pressing needs to monitor neural activity and structure during addiction and relapse. Visualization of nervous system function and structure in the intact, behaving animal is a powerful approach to meet these needs. Recent advances in genetically encoded Ca2+ indicators and imaging instruments have greatly expanded the opportunities for imaging the nervous system at work. Concurrently, there is a revolution in the use of high-speed video cameras combined with sophisticated computational tools to monitor and quantify behavior. Many labs at the University of Minnesota (UMN) lack the equipment and expertise to perform neural imaging during behavior. Nor is it possible for most investigators to keep up with the rapid changes in optical tools, either in the engineering or application domains. The Imaging Cells during Behavior Core (ICBC) is designed to allow addiction researchers and the wider neuroscience community access to modern techniques to image brain function and structure during behavior. Providing up-to-date imaging techniques, engineering and manufacturing support, as well as computational tools, will not only reduce the time and costs needed to set up experiments and process data, but also lower the entry barriers for both new and established investigators. The ICBC?s engineering, experimental, and analytical expertise will allow labs to be at the forefront of imaging technology. Aim 1 is to provide resources, expertise and training in three in vivo imaging technologies used in behaving animals: 1) head-mounted miniaturized microscopes for cellular level imaging at the surface and in deeper brain structures; 2) wide field-of-view (FOV) imaging of the cerebral cortex at both the mesoscopic and cellular levels; and 3) fiber photometry to monitor the activity of genetically defined elements in neural circuits. UMN investigators deemed these technologies critical to understanding how neural circuits change in addiction. The ICBC will provide sophisticated hardware and software to monitor and analyze behavior in head-fixed and freely moving animals and the engineering and analytical expertise to build and use these in vivo imaging technologies. Aim 2 extends and adds new imaging capabilities. Many UMN addiction researchers use rats, but most neural imaging techniques were developed for mice. Therefore, we will extend our imaging tools to the rat. In addition, Aim 2 will combine fiber photometry with wide FOV cortical imaging, enhance wide FOV imaging to access deeper cortical structures, and make wide FOV imaging compatible with magnetic resonance imaging. ICBC personnel will work closely with investigators to employ these improvements to advance their research programs. In summary, the ICBC will provide the research infrastructure, expertise and training to allow neuroscientists to image neuronal activity and neural circuitry during behavior. Providing these tools will enhance the productivity, quality, and impact of addiction-related and neuroscience research in general at the UMN.