CA Movements in Excitation-Contraction Coupling

Rios, Eduardo

Abstract

Calcium movements determine both the onset and the termination of the mechanochemical reactions that underlie contractile movement and force generation in muscle. The process whereby Ca is released intracellularly is believed to involve a pathway in the Sarcoplasmic Reticulum (SR), ill-defined in regard to location, gating and permeation properties. Ca diffuses in the myoplasm, binds to specific sites in troponin C and initiates contraction. Many other sites compete to remove Ca from the myoplasm and a """"""""pump"""""""" eventually returns it to the SR. We have developed an optic technique involving a Ca-sensitive dye, to measure the rapid [Ca] transient of the skeletal muscle cell under voltage clamp. With this technique, taking advantage of the accessibility of the intracellular space in our cut fiber preparation, we shall further characterize the release pathway. Be chelating most of the release Ca with a fast proportional buffer we shall prevent the removal processes and study the release in isolation. This should provide insights to the gating of this channel and interactions involved in the gating, like Ca-induced Ca release. Specific interference with the flow of counteriones across the SR membrane will probe the electrogenic aspects of the release and help understand the function of a recently described K-channel of the SR. Combining our optic technique for measuring Ca with fast monitoring of the Fraunhofer diffraction pattern the location of the Ca release pathway will be explored; evidence indicating that the release channels are close to the triadic junctions of SR and transverse tubules (TT) will suport the idea that the SR Ca channel gating is directly controlled by the TT membrane voltage. We have found that an increase in transparency recorded during an activating clamp pulse has striking correlations with the Ca transient; it probably corresponds to changes in the SR or the myofilaments upon Ca binding. Effects of this kind should be better seen as changes in the diffraction pattern. These will be recorded, and ascribed either to events at the myofilaments or the SR on the basis of formal analysis and experiment. Release, diffusion and uptake of Ca are mutually related. The interactions of light with intrinsic and extrinsic components of the cell seem to be effective tools to understand quantitatively those calcium movements.