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Date of Award

8-1998

Degree Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

Department

Medical Sciences

Supervisor

Professor Ram K. Mishra

Abstract

There is growing evidence that factors other than cell-surface recetors are involved in regulating the sensitivity of cells to external signals. In particular, G proteins have been implicated in the increased sensitivity of numerous receptor systems under a variety of conditions (Mishra et al., 1997). The goal of this research project was to determine the role of G proteins in mediating dopamine receptor supersensitivity in Parkinson's disease. Prelimary studies of G protein levels in human post-mortem brain tissue proved inconclusive, due to the limited availability and variability of tissue samples. Subsequent studies in the 6-hydroxydopamine (6-OHDA) rat lesion model of Parkinson's disease revealed that stimulatory G protein levels are persistently elevated following denervation (Marcotte et al., 1994). These G proteins are presumably coupled to dopamine D₁ receptors, which show clear evidence of supersensitivity despite apparently normal receptor levels. This result supports the hypothesis that G proteins are involved in the maintenance of dopamine receptor supersensitivity (Marcotte and Mishra, 1997). Stimulatory G proteins acutely following MPTP mouse model, with decreased stimulatory G proteins acutely following MPTP treatment, and increased stimulatory G proteins after long-term recovery (Marcotte et al., 1998a). Although the significance of these findings is unclear, they provide additional support for the hypothesis that G proteins are modulated in response to dopaminergic denervation. Attempts to measure functional changes in stimulatory G protein activity in the rat striatum proved unsuccessful, consistent with the available literature. Specifically, neither the GTPase nor a specific GTP binding assay was able to consistently detect stimulatory G protein activity following dopamine D₁ receptor stimulation. To provide direct evidence for the role of Golf in mediating dopamine receptor supersensitivity, Golf antisense oligonucleotides were administered to 6-OHDA lesioned rats. Intrastriatal infusion of Gold antisense, but not control sense oligonucleotides, specifically reduced apomorphine-induced rotational behaviour and Gold levels. The effects of Golf antisense infusion were at least partially reversible, supporting a specific antisense mechanism of action. However, one of the control oligonucleotides, Golf missense, consistently reduced rotational behaviour and G protein levels in a non-specific fashion. This effect was dose- and sequence-dependent, and may be due to a non-specific binding to other nucleotides or proteins (Marcotte and Mishra, 1998). Taken together, these studies support the hypothesis that stimulatory G proteins are involved in mediating dopamine D₁ receptor supersensitivity. Further characterization of the effects of in vivo antisense oligonucleotides may provide more definitive conclusions regarding the role of G proteins in mediating this phenomenon.

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