Don Gash - US grants

Over thirty million American suffer from debilitating neurological disorders. The ability to treat many of these diseases and dysfunctions of the central nervous system is quite limited. The studies conducted on the present grant critically examine a promising new approach of using neural transplants to replace dead or damaged neural tissue. The first six years of research have focused on elucidating factors important in neural transplant survival and function. Studies over the next three years will continue this line of investigation. An additional series of experiments will be initiated to address issues relevant to the possible clinical application of neural transplants. The ultimate goal of the studies conducted on this grant is to provide a better understanding of the factors important in the response of the brain to injury and recovery of function. It is hoped that these experiments will suggest better methods of treating neural dysfunctions in man. The specific goals for the next three grant years are: 1. To quantify the effects of physical and chemical lesions on transplant survival and function. 2. To critically analyze the influences of steroid hormones on grafted peptidergic neurons. 3. To evaluate alternatives to using fetal tissue as donor tissue in neural transplantation. 4. To determine if graft behavor in rodents is a good predictor of graft behavior in nonhuman primates. The studies proposed require a multidisciplinary research team approach. The basic and clinical scientists participating in this research program have six years of experience in working together. The techniques necessary to accomplish the goals outlined are in use in our laboratories.

In the past decade neural transplants have been demonstrated to be effective in ameliorating neurological deficits in rodent model systems of Parkinson's disease, Huntington's disease, neuroendocrine disorders and cognitive deficits. This success with transplants in rodents has led to suggestions that neural grafts might be of therapeutic value in treating human neurological disorders including Parkinson's disease. The purpose of the present program project is to address the pre-clinical issue which must be resolved in order to develop a rational clinical approach for using neural transplants to treat Parkinsonism. Our studies begin with the premise that practical sources of donor tissue must be identified if clinical trials are to proceed. Project 1 focuses on the characterization in vitro of potential nonfetal donor tissues including cell lines which might be used for transplantation . An important component of this project is to create cell hybrids of dopaminergic neurons which can be used for implantation. Project 2 conducts the in vivo analysis of the prospective donor tissues identified in Project 1 utilizing rodent model systems of Parkinsonism. Project 3 is also committed to the in vivo analysis of donor tissues in rodent model systems but will concentrate on paraneural tissues and will conduct correlative ultrastructural studies for both Projects 1 and 2. Project 4 will assess the development of MPTP induced Parkinsonism in nonhuman primates and the efficacy of transplants in reversing the functional deficits. Core A is the Laboratory Animal Medical Facility which will be responsible for all nonhuman primates used in these studies. Collectively these projects represent a systematic investigation of potential donor tissues from neonfetal sources which can be utilized clinically for the treatment of Parkinson's disease. The results from these studies should provide the necessary pre-clinical data needed to proceed with clinical investigations.

Over thirty million American suffer from debilitating neurological disorders. The ability to treat many of these diseases and dysfunctions of the central nervous system is quite limited. The studies conducted on the present grant critically examine a promising new approach of using neural transplants to replace dead or damaged neural tissue. The first six years of research have focused on elucidating factors important in neural transplant survival and function. Studies over the next three years will continue this line of investigation. An additional series of experiments will be initiated to address issues relevant to the possible clinical application of neural transplants. The ultimate goal of the studies conducted on this grant is to provide a better understanding of the factors important in the response of the brain to injury and recovery of function. It is hoped that these experiments will suggest better methods of treating neural dysfunctions in man. The specific goals for the next three grant years are: 1. To quantify the effects of physical and chemical lesions on transplant survival and function. 2. To critically analyze the influences of steroid hormones on grafted peptidergic neurons. 3. To evaluate alternatives to using fetal tissue as donor tissue in neural transplantation. 4. To determine if graft behavor in rodents is a good predictor of graft behavior in nonhuman primates. The studies proposed require a multidisciplinary research team approach. The basic and clinical scientists participating in this research program have six years of experience in working together. The techniques necessary to accomplish the goals outlined are in use in our laboratories.

Parkinson's disease is a relentlessly progressive neurodegenerative disorder afflicting over 500,000 Americans. Patients with parkinsonism can expect to live a normal lifespan due to levodopa and other drug therapies, but most experience a steadily eroding quality of life in the later stages of the disease. This program project was organized to (i) systematically investigative adrenal chromaffin cell transplantation as an approach to treating Parkinson's disease, and (ii) better understand the mechanisms underlying functional recovery from parkinsonism in rodent and nonhuman primate models of Parkinson's disease. The present program consists of three projects and two core facilities. In Project 1, Dr. Gash will critically test the hypothesis that increasing chromaffin cell survival in intrastriatal grafts significantly increases behavioral recovery in parkinsonian primates. In Project 2, Dr. Notter will characterize in depth the properties of primate Schwann cells and chromaffin cells by bioassays, immunoblasts and in situ hybridization for mRNA. She will also evaluate the response of primate chromaffin cells to factors they will be exposed to in implant sites including a aFGF, cytokines, macrophage-derived factors and levodopa. Dr. Hansen, in Project 3, using a selective lesion rat model will test the hypothesis that a host sprouting response is important for restoring striatal dopamine levels and may be the single most important event in ameliorating functional deficits that result from dopamine depletion in the striatum. Collectively, these studies should not only increase our understanding of the processes important for functional recovery from parkinsonism but should also be of significant benefit in designing new therapeutic approaches to the treatment of Parkinson's disease.

This award supports summer research by undergraduates at the University of Kentucky College of Medicine. Students pursue research projects of their own choosing during the 10-week program, which also includes an intensive short course that covers anatomical, physiological, behavioral and developmental aspects of modern neuroscience. Other student activities provide a better understanding of the elements of graduate training and the various career opportunities in neuroscience.

DESCRIPTION (provided by applicant): Behavioral slowing is one of the cardinal features of human aging, contributing to the debilitating deterioration of motor functions in senescence. Our principal hypothesis for the past five years of research on this Program Project has been that changes in central dopaminergic pathways constitute a fundamental component of age-associated motoric declines. Converging evidence from our studies and others are providing strong support for this hypothesis. Our experimental plan for the next five years is designed to further our understanding of CNS processes underlying behavioral slowing and analyze therapeutic approaches for intervention. Specifically, our studies focus on the dopamine (DA) neurons in the substantia nigra (SN) and their projections to the caudate nucleus, putamen and globus pallidus of the basal ganglia. The proposed studies will analyze key junctions in the neural circuitry regulating motor functions in the basal ganglia, using behaviorally characterized female rhesus monkeys ranging in age from young adulthood to old age (5-25years+) as a model of human aging. Collectively, the three Projects and three supporting Cores in this Program will critically test the following hypotheses: Hypothesis 1 - That while changes in dopaminergic functions occur throughout the basal ganglia, alterations in neural processing in the SN is a principal component of age-associated motor declines. Hypothesis 2 - That functional changes in the basal ganglia dopaminergic system, including in tyrosine hydroxylase (TH), dopamine transporters (DAT) and DA receptors, are closely associated with age-associated motoric declines. Hypothesis 3 - That anatomical changes in normal aging in the basal ganglia are less predictive than functional changes of age-associated declines in motoric performance. Hypothesis 4 - That local administration of the potent dopaminergic trophic factor GDNF (glial cell line-derived neurotrophic factor) into the SN significantly repairs and restores age-associated declines in SN dopaminergic functions.

Increasing problems with rigidity, bradykinesia, tremor and gait disturbances are seen with advancing age. These movement dysfunctions are associated with a significant increase in morbidity and mortality in the elderly. Our studies in rhesus monkeys will initially focus on delineating the relationship between age-associated changes in the nigrostriatal system and the decline in motor functions during normal aging processes. Our research program project has been designed to critically test four hypotheses. Hypothesis 1: that rhesus monkeys undergo declines in motor functions which closely model those seen in humans. Hypothesis 2: that there is a gradual, continuous change in nigrostriatal dopaminergic function in rhesus monkeys with age. Hypothesis 3: that there is a gradual continuous loss of midbrain dopamine neurons in rhesus monkeys in aging. Hypothesis 4: that upregulation of the nigrostriatal dopaminergic system in older rhesus monkeys by intracerebral administrations of the potent dopaminergic trophic factor GDNF improves motor functions. The program project is organized into three research projects which are supported by two core units. Projects 1-3 are carefully integrated and coordinated to analyze motor function and the nigrostriatal dopaminergic system in rhesus monkeys. An Administrative Core supports all three projects, coordinating collection, pooling and analysis of data. A Primate Core Facility supervises and coordinates the use of all rhesus monkeys in these studies.

All three projects of this program focus on critically testing the central hypothesis that chronic infusions of GDNF promote the regeneration and functional recovery of the nigrostriatal dopamine system. Project 3 utilizes quantitative morphometric procedures, immunocytochemistry, in stu hybridization, immunoautoradiographic procedures and retrograde tract tracing to analyze the recovery of the nigrostriatal dopamine system in parkinsonian rhesus monkeys receiving GDNF. The effects of chronic infusions into two CNS sites are critically tested, the lateral ventricle and the putamen. The analysis includes determining the consequences of interrupting trophic factor for four months and then reinstating GDNF infusion for eight weeks. The results from these studies, combined with the neurochemical analyses in Project 1 and the behavioral and functional MRI measures of Project 2, should advance our understanding of the factors underlying successful regeneration in the parkinsonian brain. In addition, these studies significantly contribute to developing the methodology for chronically delivering other tropic factors and therapeutic compounds into the CNS.

DESCRIPTION (provided by applicant): The goal of this Training Program is to prepare promising graduate students and postdoctoral fellows for successful careers in the neurobiology of aging. We will provide broad-based training in modern concepts in the neurobiology of aging with an emphasis on cellular and molecular mechanisms underlying aging processes and possible therapeutic interventions. The unifying focus of the training faculty is our interest in understanding both normal and pathological mechanisms by which the nervous system responds to changes that occur with age and emerging concepts in therapeutic intervention. This Training Program provides the formal framework for faculty in different departments, who share a common interest in the molecular - cellular basis of brain aging and translational research, to provide in-depth training in aging. The emphasis of the Training Faculty complement each other: some focus on processes that occur in the brain during normal aging, others emphasize the neuropathological processes of diseases and injuries that predominate in the aging brain, and there is a combined interest in applying this knowledge to treating neural disorders of aging. Thus, trainees will learn from faculty who utilize a broad spectrum of state-of-the-art methodological approaches to probe critical questions in aging research. Our successes attest to our ability to identify, attract, train and place promising young investigators in the area of the neurobiology of aging. Now more than ever before, we are particularly qualified to train graduate students and postdoctoral fellows. The national need for training new investigators in the area of aging is clear due to our changing demography and the increasing average lifespan. Normal aging or pathology of the nervous system underlies much age-associated morbidity. Only if we better understand the basic mechanisms that regulate the aging of the brain and apply this to the treatment of the elderly, can we hope to improve the quality of life during the latter half of the lifespan. Our ability to attract and train graduate students and postdoctoral fellows to meet the challenges in the area of the neurobiology of aging will be greatly enhanced by this Training Program.

Core A is the data management and administrative core unit for the Program Project. The purpose of this core is to provide integration and administrative support for the other two cores and the three research projects. In addition, this core will maintain fiscal accountability for the program project and facilitate data entry into the central data base. Core A, in collaboration with Core C (the Data Analysis Core), will manage the computer network linking all investigators for collecting, pooling, collating, analyzing and sharing data between projects.

Behavioral slowing is one of the cardinal features of human aging, contributing to the debilitating deterioration of motor functions in senescence. Our principal hypothesis is that functional changes in central dopaminergic pathways contribute significantly to age-associated declines in motor functions. Our experimental plan for the next five years is designed to further our understanding of CNS processes underlying behavioral slowing and analyze therapeutic approaches for intervention. Specifically, our studies focus on the dopamine (DA) neurons in the substantia nigra (SN) and their projections to the caudate nucleus, putamen and globus pallidus of the basal ganglia. The proposed studies will analyze key junctions in the neural circuitry regulating motor functions in the basal ganglia, using behaviorally characterized female rhesus monkeys ranging in age from young adulthood to old age (5-25years+) as a model of human aging. The studies and methodology in this Project are designed to quantify age-associated declines in motoric and cognitive functions in rhesus monkeys from 5 -25+ years old. The ability of specific DA receptor agonists and GDNF to improve motor and cognitive performance levels of aged animals will be tested. The behavioral results are crucial for analyzing the functional neurochemical studies in Project by Gerhardt and the fMRI and anatomical MRI studies in Project by Zhang. In addition, this Project will analyze mRNA expression levels of four important markers of DA neurons, TH, DAT, GFRalpha-1 and Ret. GFRalpha-1 and Ret form a receptor complex for GRNF that is found at high levels in the SN. Expression levels of GDNF mRNA in DA neuron target sites, the caudate nucleus and putamen, will also be evaluated. These data will be collated with parallel Western blot analyses of protein levels of the same markers in the Project by Gerhardt to better understand molecular changes underlying functional changes in central DA neurons.