The major goal of this proposal is to elucidate the electrophysiological effects of phorbol esters on neurons in the mammalian hippocampus. Phorbol esters are potent tumor promoters and inflammatory agents which are of neurophysiological interest because they can substitute for an endogenous compound, diacylglycerol, in binding to, and activating, protein kinase C. Protein kinase C has been indirectly implicated in mediating the actions of several putative neurotransmitters, but its actual physiological role in the central nervous system in unknown. Protein kinase C may participate, via a new intracellular messenger system, in the control of neuronal excitability. The highest level of specific phorbol ester binding occurs in the brain and, within the brain, in hippocampus. Preliminary experiments have revealed that phorbol esters affect neuronal properties selectively and may block one, or more, potassium potentials important for regulating activity. Hippocampal neurons will be studied, both in the in vitro slice and in an acutely dissociated neuron preparation. Each preparation offers certain distinctive advantages. Using the in vitro slice, we will both determine the neuropharmacological properties of phorbol ester actions and attempt to identify the mechanisms underlying the effects observed thus far, using intracellular electrophysiological techniques. The isolated cell preparation will be used to refine the investigation using whole cell voltage-clamp and patch-clamp methods. The isolated cell also offers the opportunity for studying the effects of diacylglycerol and protein kinase C on ionic currents. Malfunction in the control of normal neuronal excitability is at the root of several neurological diseases, including epilepsy and Huntington's disease. Understanding the neuronal function of protein kinase C may reveal the intracellular mechanism of action of a class of neurotransmitters and shed light on the regulation of neuronal excitability.
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