NINCDS support is requested for the study of voltage-dependent Ca2+ channel regulation by protein kinase C. Phorbol ester tumor promoters, which activate protein kinase C, inhibit Ca2+ influx through voltage-dependent Ca2+ channels in the cultured neural cell line, PC12. The interaction between phorbol esters and Ca2+ channels will be examined in detail by studying K+-stimulated 45Ca2+ uptake and 3H-Ca2+ channel antagonist binding in PC12 cells. Muscarinic agonists also activate protein kinase C, by stimulating phosphoinositide hydrolysis and diacylglycerol production, and the effect of these agents on K+-stimulated 45Ca2+ uptake will be studied as well. Ca2+ channel antagonist receptors, which are functionally linked to Ca2+ channels in PC12, will be isolated by extensive purification or affinity labeling, and receptors will be phosphorylated by protein kinase C in vitro. In vivo phosphorylation by protein kinase C will be studied by treating 32P-labeled cells with phorbol esters or muscarinic agonists, and by measuring 32P incorporation into Ca2+ channel antagonist receptors isolated from these cells. Finally, the autoradiographic distribution of phorbol ester and Ca2+ channel antagonist receptors will be studied in brain slices from epileptic mutant mice to investigate changes in Ca2+ channel density or protein kinase C content that may predispose to epilepsy. Seizure-induced changes in Ca2+ channel number and protein kinase C levels will be examined by autoradiography of brain slices from mice subjected to chemically- or electrically-induced convulsions. The ultimate goal of this proposal is to delineate molecular events that underlie regulation of voltage-dependent Ca2+ channels by protein kinase C and to investigate whether disturbances in regulation might be involved in susceptibility to epilepsy, or in neuronal injury following repeated seizures. These studies may reveal information that will lead to better understanding and more effective treatment of epileptic disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Clinical Investigator Award (CIA) (K08)
Project #
5K08NS001151-03
Application #
3083890
Study Section
Neurological Disorders Program Project Review B Committee (NSPB)
Project Start
1986-07-01
Project End
1991-06-30
Budget Start
1988-07-01
Budget End
1989-06-30
Support Year
3
Fiscal Year
1988
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Type
Schools of Medicine
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143