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The effect of dopamine receptor stimulation on protein kinase C isozymes in the rat hippocampus
Dissertation

The effect of dopamine receptor stimulation on protein kinase C isozymes in the rat hippocampus

Karin Adair Yurko-Mauro
Doctor of Philosophy (Ph.D.), Medical College of Pennsylvania
Dec 1994
DOI:
https://doi.org/10.17918/00008446
pdf
Yurko-Mauro_Karin_19946.50 MB
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Abstract

The discovery of multiple dopamine (DA) receptor subtypes has led to the characterization of their signal transduction pathways. In brain, stimulation of a D1-like DA receptor increases phosphoinositide (PI) hydrolysis and accumulation of inositol phosphates. The effects of DA on PI-linked diacylglycerol production and on the activation of protein kinase C (PKC) are not well-known. The present investigation was undertaken to define the effects of selective DA receptor stimulation on specific PKC isozymes in the rat hippocampus, to characterize the receptor and to determine possible mechanisms responsible for DA's effects on PKC. Stimulation of hippocampal slices for 20 min with nanomolar concentrations of DA decreased cytosolic [gamma] PKC by 56 % without altering membrane [gamma] PKC, resulting in decreased total [gamma] isozyme levels, detected by Western blot. The DA-induced decrease was significantly blocked by the D1 antagonist, SCH23390, but not by the D2 antagonist, sulpiride. The D1 agonists, SKF38393 and A77636 mimicked the effect of DA, while the D2/D3 agonist, quinpirole was without effect, indicating that the dopaminergic response is mediated through a D1-like receptor. DA had no effect on [alpha],[delta] or [zeta] PKC isozymes in the same hippocampal preparations. The DA-induced decrease in cytosolic [gamma] PKC was blocked by the Ca⁺⁺-dependent protease inhibitor, N-Acetyl-Leu-Leu-norleucinal, was dependent on extracellular Ca⁺⁺, and was blocked by the inorganic Ca⁺⁺ channel blocker, Co⁺⁺, but not by the dihydropyridine, nifedipine. These results suggest that DA D1-like receptor stimulation may be increasing the proteolysis of [gamma] PKC, a known substrate of the Ca⁺⁺-dependent protease, calpain. Increases in activated calpain I immunoreactivity (76 kDa band) were found in hippocampal membranes after 5 min of DA stimulation, at a time preceding the change in [gamma] PKC. By 20 min of DA incubation, decreases in activated calpain I were seen. These DA-induced changes were also blocked by the calpain inhibitor. The findings of this investigation signify complex regulatory mechanisms of hippocampal D1-like receptor mediated signal transduction whereby DA stimulation activates a Ca⁺⁺-dependent proteolytic path which catalyzes cellular [gamma] PKC.

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