The initial phase of research will investigate the calcium channels in T- lymphocytes. Studies have shown that activation of T lymphocytes involves an increase of intracellular free calcium. While the initial rise in intracellular calcium is due to a release of intracellular stores mediated by inositol 1,4,5 trisphosphate (InsP3), evidence suggest that the sustained rise in calcium is due to a transmembrane influx of calcium. The voltage insensitive calcium channel responsible for this sustained influx is not yet fully characterized and is the subject of this proposal. Patch clamp techniques will be utilized to demonstrate regulation by InsP3. Single channel recordings of cell-attached and inside-out patches will be compared to whole cell studies. Monovalent and divalent cation permeability will be determined. The effect of specific blockers on the calcium channel current will also be studied. During the second phase of research, the properties determined by single channel recordings will be compared to purified cerebellar InsP3 receptors reconstituted in lipid bilayers.
