Research in our laboratory aims to unveil the detailed mechanisms of excitation-contraction (EC) coupling in vertebrate skeletal muscle and their pathological alterations in diseases. To this end, the laboratory uses a combination of advanced optical and electrophysiological approaches. We have shown, with a localized detection method and pulse laser imaging that one of the earliest manifestations of the EC coupling process in amphibian skeletal muscle is a massive localized increase of the intracellular Ca2+ concentration in microscopic regions of the muscle fibers where the sarcoplasmic reticulum (SR) apposes the T-tubule (triadic junction). We have recently found that Ca2+ domains in mammalian muscle fibers are doubled peaked, as expected from the existence of two triadic junctions per sarcomere. The large, rapidly dissipating, Ca2+ gradients are not restricted to action potential- evoked Ca2+ release in muscle fibers; they also occur as a triggering mechanism in synaptic transmission and other neurosecretory processes. We have extended the use of spot detection techniques to demonstrate the existence of Ca2+ domains in active zones of presynaptic terminals in a co-culture neuromuscular junction preparation. New imaging and flash-photolysis techniques, with improved time and spatial resolution, are currently under development in our Cell Physiology Imaging Group. These methodologies will allow us to obtain new insights on the role of the membrane potential in initiating the cascade of events that lead to EC coupling in skeletal muscle fibers, and to the release of chemical transmitters in presynaptic terminals.