This project focuses on the role of intracellular calcium in the control of basic neuronal function. Questions addressed involve issues in fundamental neuroscience having to do with the cellular physiology of single neurons. The mechanism of transmitter release at neuronal synapses by calcium ions, and the regulation by calcium ions of membrane channels involved in electrical activity, are the neuronal processes which will be studied in detail.
Specific aims i nclude the following: 1) determining the calcium-dependence of three calcium-regulated membrane channels -- calcium-activated potassium channels, calcium-activated nonspecific cation channels, and calcium-dependent inactivation of calcium channels; 2) determining the relative importance of surface extrusion pumps and uptake into organelles in the regulation of cytoplasmic calcium following calcium influx during electrical activity; 3) determining the quantitative dependence of transmitter release on intracellular calcium activity, and the role of calcium in post-tetanic potentiation; 4) and determining whether presynaptic potential can modulate the dependence of transmitter release upon internal calcium. Experimental preparations will include the large identified central neurons in the abdominal ganglion of the sea slug Aplysia, neuromuscular junctions in crayfish and lobster, and the giant synapse in the squid stellate ganglion. Experimental methods include measurement of membrane currents by voltage clamp and transmitter release by postsynaptic intracellular recording, measurement of intracellular calcium by arsenazo III absorbance microspectrophotometry and fura-2 fluorescence spectroscopy, and control of intracellular calcium with photoisomerizable calcium chelators.
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