We will examine the mechanism by which the Ca2+ binding protein, calmodulin (CaM) and GTP regulate dopamine (DA) receptor-coupled (D-1 and D-2) adenylate cyclase activity. We will investigate the contributions of these coupling agents to supersensitivity of DA receptors (D-1 and D-2) in rat striatum resulting from chronic treatment with psychoactive drugs. We have shown, in striatum and retina, that CaM increases DA-stimulated adenylate cyclase activity in membrane preparations. We have also shown that when dopaminergic supersensitivity is developed by pharmacological or physiological means that the sensitivity of adenylate cyclase to CaM is increased in parallel with that of DA. GTP couples DA stimulatory (D-1) and inhibitory (D-2) receptors to adenylate cyclase by binding to distinct GTP-binding proteins. We have shown that CaM can interact with guanyl nucleotides but its mechanism in affecting coupling with D-1 or D-2 activity is unknown. There are three specific aims of this proposal: 1. To investigate whether activation of basal and neurotransmitter-stimulated adenylate cyclase activity by CaM involves an interaction with a stimulatory or inhbitory GTP-binding protein. This will be performed in the model system of C6 glioma cells by examining the interaction of CaM with toxins known to affect selective GTP-binding proteins. 2. To examine whether Ca2+ and CaM can affect binding and release of [3H]GppNHp to GTP-binding proteins mediated by D-1 and D-2 agonists. This will indicate effects of CaM on receptor coupling and possibly D-2 activity since it is difficult to directly measure this in striatum. 3. To determine whether there are selective alterations in GTP and CaM-related components in rat striatum as a result of chronic treatment with psychoactive drugs that elicit dopaminergic supersensitivity such as haloperidol, sulpiride and amphetamine. We will examine whether D-1 and D-2 receptor coupling is altered by these treatments. Techniques used will be adenylate cyclase activity measurement, [3H]GppNhp and DA receptor binding studies and [125I]labeled CaM binding to gels to measure changes in specific CaM-binding proteins resulting from chronic drug treatment. These studies will further delineate the intramembrane role of GTP, Ca2+ and CaM in modulating DA receptor activity and further examine their role in dopaminergic supersensitivity which is important in behavioral abnormalities of Parkinsons disease, tardive dyskinesias, amphetamine psychosis and schizophrenia.
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