The mechanisms underlying the central actions of cocaine that predispose to abuse of this drug are complex and varied. The two major actions of cocaine generally fit the profiles of psychomotor stimulants and local anesthetics. Based upon animal studies, the [after activity may be correlated with binding to and blockade of voltage-gated sodium channels. The role of sodium channel blockade in the rewarding or euphorigenic aspects of cocaine abuse remains speculative. In analogy with pharmacologic kindling models in animals, however, cocaine-induced blockade of human brain sodium channels could contribute substantially to cocaine toxicity and cocaine-related seizures. Given this potential involvement of brain sodium channels in the the phenomena/epiphenomena of cocaine abuse, along with the fact that there is currently only limited knowledge of these ion channels in humans, the goal of our research is to characterize human brain voltage-gated sodium channels in molecular biological, electrophysiological and pharmacological terms. These studies are prerequisite to the eventual clarification of the role of sodium channel blockade in human cocaine abuse and toxicity. To accomplish this goal, the project has three specific aims. First, genes for 3 human brain sodium channel subtypes will be cloned and sequenced in their entirety using primer extension strategies (e.g. the RACE protocol) and standard library screening techniques starting with partial clones already identified in this laboratory. A new gene mapping strategy will be used to define the chromosomal location of each subtype, and the relative distribution of the respective mRNA's will be determined in various brain regions with the ligase detection reaction (LDR). Second, the electrophysiological characteristics of each subtype will be studied with standard two-electrode voltage clamp or patch clamp procedures following transient expression of full length clones in frog oocytes and mammalian cell lines. Finally, the pharmacologic profiles of the individual human brain sodium channel subtypes with respect to standard sodium channel neurotoxins and cocaine plus cocaine-related drugs will be analyzed by radioligand binding techniques in transfected mammalian cell lines. This research should help provide the foundation to understand more fully the role of cocaine's local anesthetic effects in the human brain.
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