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Date of Award

2000

Degree Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

Department

Medical Sciences

Supervisor

R.K. Mishra

Abstract

Central dopaminergic systems have been implicated in CNS disorders such as schizophrenia and Parkinson's disease. The characteristics of dopamine (DA) receptors has been well studied using a variety of pharmacological and biochemical techniques. DA receptor function is known to be modulated by the endogenous tripeptide pro-leu-gly-NH2 (PLG). A combination of novel pharmacological (guanosine-5' -O -(3[ 35 S]thio)triphosphate; [35 S]GTPγS binding) and molecular biological (differential display mRNA by RT-PCR; ddPCR) techniques were used to examine the function of endogenously expressed D2 DA-receptors in the striatum. Studies were undertaken to increase our understanding of the mechanism of action of PLG and its role in D2 receptor regulation and signal transduction. Measurement of antagonist effects in the absence of D2 receptor stimulation revealed that basal [35 S]-GTPγS binding was significantly decreased by haloperidol, butaclamol and chlorpromazine but not clozapine, sulpiride and spiperone. Thus haloperidol, butaclamol and chlorpromazine acted as negative antagonists; while clozapine, sulpiride and spiperone acted as silent antagonists. The role of PLG in D2 receptor stimulation of Gi G-proteins was examined using the [35 S]-GTPγS binding technique. The possible effect of PLG on NPA-stimulated [35 S]GTPγS binding was assessed at both maximal (1 μM NPA) and submaximal (0.1 and 0.03 μM NPA) levels of D2 receptor stimulation. It was found that PLG did not significantly alter [35 S]-GTPγS binding in bovine striatum either in the absence or presence of D2 receptor stimulation; indicating that PLG does not alter the rate of GDP:GTP exchange in the Giα G-protein subunit. Haloperidol and clozapine have distinct pharmacological profiles and have differential effects on many systems in the brain. This likely accounts for the tendency toward the development of extra pyramidal side effects (EPS) with the use of haloperidol but not clozapine. In an attempt to elucidate the mechanism of action of PLG, ddPCR was utilised to discover genes that are regulated by protracted treatment with PLG (20 mg/kg, i.p. for 28 days). In the present study, I have attempted to further the understanding of D2 receptor coupling to G-proteins in the striatum with respect to agonist efficacy and negative versus silent antagonism. I have also determined that PLG does not modulate D2 receptor signal transduction by altering the rate of GDP:GTP exchange in Gi. Furthermore, I have used the [35 S]-GTPγS binding technique to compare the effects of typical and atypical neuroleptic treatment on D2 receptor coupling to Gi. (Abstract shortened by UMI.)

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