INTRACELLULAR CA2+ CHANNELS OF STRIATED MUSCLE

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Ca2+ channels of intracellular Ca2+ stores, such as ryanodine and IP3 receptors, are ultimately responsible for the generation of Ca2+ signals within cells when the surface membrane is stimulated by voltage, neurotransmitters, or hormones. The kinetics of activation and inactivation of intracellular Ca2+ channels is responsible for the ubiquitous intracellular Ca2+ release mechanism known as Ca2+-induced Ca2+ release. The proposal focuses on the molecular and kinetic properties of ryanodine receptors and IP3 receptors of cardiac and skeletal muscle sarcoplasmic reticulum with the interest in understanding the contribution of these channels to the changes in sarcoplasmic reticulum Ca2+ permeability during stimulus-contraction coupling. The specific aims of the proposal are 1) to study the block or activation ryanodine receptors by peptide toxins purified from Buthotus judaicus and Heloderma horridium horridium venoms; 2) to establish the size of the Ca2 pools controlled by, and rates of Ca2+ release mediated by, ryanodine and IP3 receptors in cardiac muscle; and 3) to determine the molecular structure of ryanodine receptor-specific toxins by molecular cloning techniques. The specific aims are drawn from the experience gained by this group on the functional reconstitution of Ca2+ channels of intracellular and surface origin using a combination of 45Ca2+ fluxes and planar bilayer recording. The strength of the proposal resides in the resolving power of these techniques when used within a quantitative framework.