Robert Henry Roth - US grants

DESCRIPTION: (provided by applicant) Knowledge concerning the regulatory control of mesotelencephalic dopamine (DA) neurons is crucial for obtaining a better understanding of their role in both normal and abnormal behavior, and the mechanisms by which antipsychotic drugs (APD) partially ameliorate psychotic symptoms. DA depletion, as well as DA excess, can produce profound deficits in prefrontal cortex (PFC) function. This proposal is directed at studying the regulatory control, adaptation, and pharmacology PFC DA system under normal, hypoactive and hyperactive states. The hypoactive state will be induced by subchronic treatment with phencyclidine (PCP) and/or delta9-tetrahydrocannabinol (ThC). Long-term abuse of PCP or marijuana is linked with deficits of frontal lobe function providing relevance for our findings in animals. Our studies with PCP(rat and monkey) and THC (rat) have demonstrated that subchronic treatment with these two different classes of psychotomimetic agents produce a decrease in DA release and turnover in the PFC and impair spatial working memory. The atypical APD, clozapine, is effective in reversing the cognitive impairment and normalizing PFC DA turnover in the PCP dysregulated state. This proposal will study the regulatory control of PFC DA in the PCP dysfunctional state in the rat and examine the receptor mechanisms involved in the clozapine reversal of the deficit. The ability of other atypical APD to reverse this PFC DA deficit and cognitive impairment will also be studied. Our rat data indicate that subchronic exposure to THC produces a prolonged dysregulation of PFC dopaminergic transmission and altered PFC dependent behavior. The neurochemical and anatomical specificity of this deficit, and its endurance and reversal by atypical APD, will be examined in both rat and monkey. Acute administration of THC in rats elicits a rapid increase in PFC DA turnover and release, and impairs PFC-dependent functions. These cognitive deficits can be reversed or prevented by drugs or conditions that attenuate the DA increase or block its action. Because of the considerable differences in the anatomy and regulatory control of cortica] DA systems between rodents and primates, the acute and chronic effects of THC on PFC DA function will be studied in the monkey. We will also determine if the pharmacological reversal of the THC-induced DA dysregulation and cognitive deficits observed in rodents is conserved in primates. The generation of critical neurochemical, behavioral and pharmacological data on 2 perturbed conditions of the PFC DA system in both rodents and primates may provide important new insights concerning the neural systems relevant to the PFC cognitive dysfunction in schizophrenia and their modulation by APD and other novel agents.

The euphoric properties of psychomotor stimulants impart a powerful reinforcing action to these agents which may underly their strong potential for abuse in humans. The rewarding properties of CNS stimulants in general and cocaine in particular appear to be due, in part, to their ability to increase neurotransmission at dopaminergic (DA) synapses. Much of what is currently known about the mechanism of action of cocaine on DA transmission is derived from pharmacological studies evaluating cocaine's actions on DA systems not directly involved in the process of stimulant self administration. Since it is now appreciated that midbrain DA systems are not homogenous and possess a number of different biochemical, physiological and pharmacological properties, the relevance of prior studies investigating the interaction of cocaine with DA systems to the larger issue of cocaine abuse is unclear. The proposed project involves a combined electrophysiological and biochemical approach to evaluate the effects of cocaine on mesotelencephalic DA neurons in rodents. In the first group of experiments, extracellular single unit recording and microiontophoretic techniques will be used to determine whether antidromically identified DA neurons differ in their response to locally or parenterally administered cocaine. Biochemical studies will be conducted in parallel to assess alterations in DA metabolism in and release from selected mesotelencephalic systems. Findings will be correlated with alterations in neuronal firing evaluated by single cell recording. In a second group of experiments, biochemical and electrophysiological techniques will be employed to study the effects of cocaine on DA target neurons in regions thought to participate in stimulant self-administration (e.g. nucleus accumbens, prefrontal cortex) and compared with the non-rewarding DA projections to the DA receptive neurons in striatum. In both groups of experiments, comparisons will be made between cocaine and other drugs exhibiting similar pharmacological properties but differing in their ability to sustain self-administration. If cocaine is observed to elicit significant and selective effects on the functioning of certain DA systems or postsynaptic follower neurons, attempts will be made to prevent or reverse these electrophysiological and neurochemical effects pharmacologically. By more clearly understanding the pharmacological actions of cocaine on specific DA neurons and their follower cells, we may gain further insight into the neural mechanisms underlying cocaine's reinforcing properties and identify strategies for reversing or preventing these actions.

stress; antipsychotic agents; mental disorders; neuroanatomy; molecular pathology; biomedical facility; haloperidol; neurotransmitter metabolism; Tourette's syndrome; genetic transcription; clozapine; mitochondria; gene expression; histopathology; neuropharmacology; cytochrome oxidase; northern blottings; high performance liquid chromatography; Primates; laboratory rat; immunocytochemistry;

Although some promising data have been obtained in studies attempting to ameliorate experimental or clinical Parkinsonism, the technique of neural transplantation is still in its infancy. Controversy surrounds many issues in the field. This Core Unit is well situated to tackle some of these important questions. Biochemical and pharmacological techniques, including HPLC, GC-MS, autoradiography, and radioimmunoassay will be used to study; the factors that may influence the survival of fetal dopamine cells destined for transplantation, the functioning and growth of graft fibers following transplantation, the integration of the grafted cells with the host brain, the mechanism underlying the behavioral effects of transplantation, and ways to improve both the model of Parkinson's disease and the cells that are implanted. In addition, new approaches and techniques will be implemented as they become available. The Core pioneered the potential of novel cocaine analogs as highly sensitive markers for dopamine neurons. Quantitative autoradiography using two such analogs has yielded new insights into graft function in the striatum of the MPTP-treated monkey and contributed a useful ligand for imaging studies also now in progress.

DESCRIPTION: (Applicant's Abstract) The alarming increase in cocaine abuse that has occurred over the last decade has been reflected in the prevalence of cocaine use during pregnancy, estimated to be as high as 8% to 30% in some regions. Consequently, there is now a sizable population of children that have been exposed to the effects of cocaine during the initial stages of their development. Cocaine use during pregnancy appears to be associated with a variety of neurobehavioral abnormalities in the progeny. While some of the effects of fetal exposure to cocaine appear transiently in neonates, there are indications that some long-lasting deficits may occur. In particular, the incidence of attention deficit hyperactivity disorder (ADDH) appears to be markedly increased following prenatal exposure to cocaine. While these clinical studies are extremely suggestive of harmful effects of gestational exposure to cocaine, controlled studies in laboratory animals are needed in order to understand more fully the effect that cocaine use during pregnancy has on the offspring, and to find ways of reversing the deficit. The overall hypothesis of this proposal, for which strong preliminary data are presented, is that prenatal exposure to cocaine can disturb the function of the dopamine input to the prefrontal cortex of the progeny, and that such a deficit is reflected in cognitive deficits that are most apparent under stressful conditions. We propose to pursue this hypothesis using sensitive biochemical and behavioral measurements to study the offspring of rats given cocaine during pregnancy. Biochemical techniques will include in vitro and in vivo measures of neuronal activity pre- and postsynaptic to the dopaminergic terminals in the prefrontal cortex. Stress-induced alterations in behavioral measures of spatial working memory will be assessed by performance in a delayed alternation task. Stress will be induced by exposing rats to the effects of either an anxiogenic B-carboline drug or a stressful olfactory cue. Abnormalities induced by prenatal exposure to cocaine will be investigated at 3 postnatal ages. The effect will be pursued in both male and female progeny, as our preliminary data indicate some gender specific effects of gestational exposure to cocaine. While we anticipate that such studies will provide valuable scientific insights into the impact of prenatal cocaine on the function of the prefrontal cortex, we are hopeful that the studies in which the deficits are reversed pharmacologically will have positive social and economical repercussions by providing leads to the logical choices of drugs, and development of new drugs, to treat the detrimental effects of prenatal cocaine exposure.

DESCRIPTION (Adapted from applicant's abstract): No convincing animal-model includes evidence of both negative and positive symptoms of schizophrenia, selective pharmacological reversal of these deficits, or provides evidence of its pathophysiological circuits. We have exciting new data in an Old World primate which suggests the possibility of such a model as well as strategies for more effective and selective treatments. PCP is known to induce an enduring schizophrenic-like syndrome with repeated use in humans. Our pilot data in rats and monkeys indicate that subchronic PCP exposure induces a DA deficiency in the PFC and that this deficiency is manifest in cognitive impairment and behavioral abnormalities in the monkey. Further, this cognitive impairment is ameliorated by administration of the atypical antipsychotic drug clozapine, an agent with high efficacy in treating negative symptoms in schizophrenics. Using in vivo and ex-vivo techniques in rats and monkeys, this project will examine the mechanisms responsible for the neurobiological changes induced by repeated PCP administration on the anatomical integrity, neurotransmitter regulation and behavioral functions associated with the PFC. The research plan will investigate the following, hypotheses: (1) Subchronic treatment with PCP induces enduring changes in PFC DA function that persist for more than a month and demonstrates neurochemical and anatomical specificity. (2) The responsivity of the mesoPFC DA system to stress and psychomotor stimulants is blunted suggesting a global functional inhibition of these neurons. (3) Dysregulation of specific auto- or afferent-regulatory control of midbrain DA neurons is responsible for or associated with the observed inhibition of meso-cortical DA function induced by PCP. (4) The behavioral/cognitive deficits induced by repeated PCP exposure are directly related to PFC dysfunction. (5) Atypical will be more effective than typical antipsychotic drugs in reversing,the cognitive deficits, and this difference is dependent on a DA D4 receptor mechanism. The generation of critical neurochemical and behavioral data in the monkey will provide important new insight concerning the neural systems relevant to schizophrenia and aid in the development of novel strategies for ameliorating the neurochemical and behavioral effects in this new, potential animal model of the disorder.

DESCRIPTION: (Adapted from applicant's abstract) The cognitive deficits that occur in schizophrenia are arguably the most debilitating of the symptoms, and the most resistant to pharmacological treatment. While the atypical antipsychotic drug, clozapine, is the one of the few drugs with any success in treating the negative and cognitive symptoms of schizophrenia, its mechanism of action is not fully understood and this has hindered development of other agents that are more effective than clozapine and lack its dangerous side effects. In man, repeated use of phencyclidine (PCP) can often induce an enduring schizophrenic-like syndrome. In the monkey, we have found that subchronic exposure to PCP induces a decrease in doparnine function in the prefrontal cortex (PFC), which persists for more than a month. Demonstrates, neurochemical and anatomical specificity. This PCP-induced PFC dopamine deficiency correlates with cognitive impairments in the monkey, which resemble those occurring in schizophrenia Furthermore, these cognitive deficits are partially ameliorated by administration of clozapine Using in vivo and ex vivo techniques in rats and monkeys, this project will examine the mechanisms responsible for the neurobiological changes induced by repeated PCP administration on the anatomical integrity, neurotransmitter regulation and behavioral functions associated with the PFC. In addition, the mechanisms involved in the pharmacological reversal of the cognitive deficits produced by subchronic exposure to PCP will be evaluated. The research plan will address the following four specific aims: (1) What neurobiological changes; responsible for the reduction in dopamine function in the PFC following repeated PCP administration, (2) clozapine's ability to reverse the PCP-induced cognitive impairment mediated by a preferential increase dopamine turnover in the PFC, and what receptors are essential for this action? (3) Determine if atypical antipsychotic drugs (or receptor specific agents) that reverse the PFC dopamine deficit in PCP-treated monkey attenuate the cognitive impairments, and (4) Determine whether there are regionally specific adaptive changes the parvalbumin GABAergic interneurons of the PFC in the monkey repeatedly treated with PCP. The generation of critical neurochemical and behavioral data in this monkey model of PFC dopamine deficiency and impaired cognition will provide important new insights concerning the neural systems relevant the frontal cortical cognitive dysfunction in schizophrenia. These data will aid in the development of novel strategies for ameliorating the neurochemical and behavioral deficits in this potential animal model, and in the cognitive dysfunctions associated with schizophrenia and other psychiatric disorders.

PROJECT 2. ENHANCING FUNCTION OF GRAFTED PRIMATE DOPAMINE NEURONS WITH NEUROTROPHIC FACTORS The death of the vast majority of transplanted dopamine neurons within the first few days appears to limit the success of this treatment strategy for Parkinson's disease. The inadequacy of critical growth factors in the adult parkinsonian brain may be a major cause of early cell death and restricted outgrowth of grafts placed into the striatum. This project proposes studies to determine whether chronically delivered neurotrophic factors can increase the survival and outgrowth of fetal mesencephatic dopamine neurons, and improve the biochemical and functional outcome of neural grafting in parkinsonian primates. Factors tested will include glial cell line-derived neurotrophic factor (GDNF), delivered biologically by macroencapsulated cells. Fetal striatum releases several neurotrophic factors besides GDNF, and this milieu provides critical support to dopamine neurons during their normal development. Thus, the effect of factors released from fetal striatum enriched in oligodendrocyte-type-2 astrocyte (SO2A) progenitor cells will also be investigated on dopamine neuron survival. Experiments will utilize implantation of GDNF-releasing capsules, or cografting of SO2A cells at increasing distances from ventral mesencephalic grafts placed into the caudate in the MPTP-treated monkey, a model that reproduces all the hallmark behavioral features of Parkinson's disease. Defined dissections of donor tissue and pre-selection of MPTP-treated monkeys will limit variability in the studies. The main end-point measures will be improvement in parkinsonism, correlated with dopamine neuron survival and soma size, and dopamine fiber outgrowth, determined by biochemical measures and light and electron microscopy. Methods for these studies are in place, and key pilot studies strongly support feasibility and the hypotheses. These studies will increase understanding of the interaction of primate fetal dopamine neurons with neurotrophic factors, and will lead to improved and more reproducible methods of cellular replacement which may benefit patients with Parkinson's disease and possibly other neurodegenerative disorders.

PROJECT 2. ENHANCING FUNCTION OF GRAFTED PRIMATE DOPAMINE NEURONS WITH NEUROTROPHIC FACTORS The death of the vast majority of transplanted dopamine neurons within the first few days appears to limit the success of this treatment strategy for Parkinson's disease. The inadequacy of critical growth factors in the adult parkinsonian brain may be a major cause of early cell death and restricted outgrowth of grafts placed into the striatum. This project proposes studies to determine whether chronically delivered neurotrophic factors can increase the survival and outgrowth of fetal mesencephatic dopamine neurons, and improve the biochemical and functional outcome of neural grafting in parkinsonian primates. Factors tested will include glial cell line-derived neurotrophic factor (GDNF), delivered biologically by macroencapsulated cells. Fetal striatum releases several neurotrophic factors besides GDNF, and this milieu provides critical support to dopamine neurons during their normal development. Thus, the effect of factors released from fetal striatum enriched in oligodendrocyte-type-2 astrocyte (SO2A) progenitor cells will also be investigated on dopamine neuron survival. Experiments will utilize implantation of GDNF-releasing capsules, or cografting of SO2A cells at increasing distances from ventral mesencephalic grafts placed into the caudate in the MPTP-treated monkey, a model that reproduces all the hallmark behavioral features of Parkinson's disease. Defined dissections of donor tissue and pre-selection of MPTP-treated monkeys will limit variability in the studies. The main end-point measures will be improvement in parkinsonism, correlated with dopamine neuron survival and soma size, and dopamine fiber outgrowth, determined by biochemical measures and light and electron microscopy. Methods for these studies are in place, and key pilot studies strongly support feasibility and the hypotheses. These studies will increase understanding of the interaction of primate fetal dopamine neurons with neurotrophic factors, and will lead to improved and more reproducible methods of cellular replacement which may benefit patients with Parkinson's disease and possibly other neurodegenerative disorders.

[unreadable] DESCRIPTION (provided by applicant): The cognitive deficits that occur in schizophrenia are arguably the most debilitating of the symptoms, and the most resistant to pharmacological treatment. While the atypical antipsychotic drug (APD), clozapine, is one of the few drugs with any success in treating the negative and cognitive symptoms of schizophrenia, it's mechanism of action is not fully understood, and this has hindered development of other agents that are more effective than clozapine and lack its dangerous side effects. In man, repeated use of phencyclidine (PCP) can often induce an enduring schizophrenic-like syndrome. In the monkey, we have found that subchronic exposure to PCP induces a decrease in dopamine function in the prefrontal cortex (PFC) which persists for more than a month, and demonstrates neurochemical and anatomical specificity. This PCP- induced PFC dopamine deficiency correlates with cognitive impairments in the monkey, which resemble those occurring in schizophrenia. Furthermore, these cognitive deficits are partially ameliorated by administration of clozapine. PCP also causes a decrease in the number of spine synapses and density of dendritic spines in layer V of rat PFC. Using in vivo and ex vivo techniques in rats and monkeys, this project will examine the mechanisms responsible for the neurobiological changes induced by repeated PCP administration on the anatomical integrity, neurotransmitter regulation and behavioral functions associated with the PFC. In addition, the mechanisms involved in the pharmacological reversal of the cognitive deficits produced by subchronic exposure to PCP will be evaluated. The research plan will address the following: Does dopamine play a critical role in the PCP induced decrease in dendritic spine density and spine synapse loss observed in the PFC of rodents? Can these anatomical changes be reversed by atypical APDs? Is the loss in the number of spine synapses and decrease in spine density and the dopamine modulation of this effect observed in the rat conserved in monkeys? Can the PCP and MPTP induced loss of dendritic spine synapses in the PFC and the ensuing cognitive deficits be reversed by chronic administration of atypical APDs? What receptors are essential for clozapine's ability to normalize dopamine turnover in the PFC of PCP treated monkeys? Do atypical APDs (or receptor specific agents) that reverse the PFC dopamine deficit in PCP-treated monkeys attenuate the cognitive impairments? The generation of critical neurochemical, anatomical and behavioral data in this monkey model of PFC dopamine deficiency and impaired cognition will provide important new insights concerning the neural systems relevant to the frontal cortical cognitive dysfunction in schizophrenia. These data will aid in the development of novel strategies for ameliorating the neurochemical, anatomical and behavioral deficits in this potential animal model, and hopefully in the cognitive dysfunctions associated with schizophrenia and other psychiatric disorders. [unreadable] [unreadable] [unreadable]

? DESCRIPTION (provided by applicant): Cognitive deficits are an intrinsic part of schizophrenia, occurring independently of positive symptoms, and often persisting even when psychotic symptoms of schizophrenia have been successfully treated. Cognitive functioning is moderately to severely impaired in patients with schizophrenia and is typically present even in the prodromal phase of the disorder, in young drug-naïve patients. The deficits are in the domain of executive function largely controlled by the prefrontal cortex (PFC). However, there is only a fragmentary understanding of biochemical dysfunctions in brain that leads to cognitive impairment in schizophrenia. Furthermore, even though atypical antipsychotic drugs can improve certain aspects of cognition, many patients do not achieve remission. Therefore, the development of new therapeutic drugs for cognitive impairment remains imperative. However, the design of new agents for treating cognitive deficits in schizophrenia is hampered by not knowing how the more successful antipsychotic drugs exert their benefit on cognitive function. The goals of the application are to understand the biochemical and synaptic changes in PFC that underlie schizophrenia-like cognitive deficits and identify the beneficial alterations produce by drugs that have demonstrated efficacy in combating these symptoms. These goals will be addressed using non-human primate models that have face, construct and predictive validity for the cognitive deficits of schizophrenia, together with the sex bias that characterizes the risk for schizophrenia. Our hypothesis is that dopamine, brain derived neurotrophic factor (BDNF) and estradiol are important factors that interact in the PFC to modify excitatory spine synapses on dendrites of pyramidal neurons, which in turn critically modulates executive function. This project will use a novel developmental phencyclidine (PCP) model and an adult primate PCP model that have resonance to the dopamine, glutamate and GABA hypotheses of schizophrenia, in addition to being relevant to the theories of developmental origins for the illness. The application has 3 specific aims: 1) Identify to what extent estradiol contributes to resilience of the peri-adolescent monkey PFC to the detrimental biochemical, synaptic and cognitive effects of PCP, 2) Identify the potential of local over-expression of BDNF to initiate recovery of excitatory spine synapses from the impact of PCP on primate PFC, and 3) Identify biochemical, synaptic and cognitive consequences of acute and repeated novel atypical antipsychotic drug administration in PFC of PCP-exposed monkeys. This current project contains refinement of previous theoretical concepts and approaches, and tests the impact of new interventions, utilizing improvements in current methodology. This work is expected to lead to new strategies in treating cognitive deficits in schizophrenia, which is the prime driver of significant disabilities in occupational, social, and economic functioning in patients, and imposes a heavy emotional burden on the family and an economic toll on the healthcare system.