The work contained in this proposal probes the effects of acute and chronic alcohol exposure on the functioning of nerve cell membranes. We use two model systems, the marine invertebrate, Aplysia, and a mammalian neuroblastoma cell line. Measurement techniques are predominantly electrophysiological, but also incorporate fluorescent techniques to enable us to correlate membrane lipid properties with electrophysiological properties. We will study the effects of alcohol on calcium channels within the membrane, using single channel recording techniques to allow resolution of the behavior of individual channel proteins. Ethanol affects calcium currents in Aplysia neurons at physiologically relevant concentrations below 50 mM. Temperature will be used as a variable to examine the relationship between lipid state and channel function, further illuminating the mechanisms of ethanol action. The mobility of a range of membrane lipid probes will be examined under conditions of ethanol exposure and temperature in which electrophysiological function is altered, in an attempt to identify changes in the lipid matrix which might be responsible for the electrophysiological effects of alcohol. We will use the advantages of the model Aplysia system to examine the effects of ethanol on different classes of synapses. By determining the hierarchical sensitivities of voltage-dependent channels and synapses within a single model nervous system, we hope to be able to predict which elements of more complex nervous systems will be the most sensitive to ethanol action. Moreover, physical accessibility to the elements at the synapse is good, allowing voltage clamping of the presynaptic cell and pressure ejection of pharmacological agents onto the postsynaptic membrane. Once sensitivity is determined, the site of action within the functional linkage between receptor binding and channel opening will be examined. Experiments will be performed to determine the effects of chronic alcohol exposure on the subsequent response to alcohol challenge in cultured Aplysia cells. Finally, we will examine the alcohol sensitivity of membrane channels from different species of Aplysia in an attempt to gain insights into the basis for genetic and environmental influences on development of alcoholism and alcohol-related disorders.
| Levine, John B; Martin, Gilles; Wilson, Andrew et al. (2003) Clozapine inhibits isolated N-methyl-D-aspartate receptors expressed in xenopus oocytes in a subunit specific manner. Neurosci Lett 346:125-8 |
