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Imaging tubular system, sarcoplasmic reticulum and myoplasmic calcium with novel fluorescence methods

B.S. Launikonis, School of Biomedical Sciences, The University of Queensland, St Lucia, QLD 4072, Australia.

The precise control of contraction in skeletal muscle critically depends on the rapid and precise delivery of Ca2+ from the specialized internal store, the sarcoplasmic reticulum (SR), which is under voltage control from the tightly apposed tubular (t-) system. Regulation of Ca2+ in the SR and cytoplasm during periods of muscle work is complex and involves influxes of Ca2+ from the t-system. A major problem is that measurements of voltage and store-dependent Ca2+ fluxes from the t-system of skeletal muscle are not possible with standard electrophysiological techniques. Methods to simultaneously image Ca2+ in two subcellular compartments of single muscle fibres using laser scanning confocal microscopy have recently been developed. This has largely involved trapping one Ca2+ sensitive dye in the t-system of a mechanically skinned fibre and introducing a second spectrally separate Ca2+ indicator to the cytoplasm. By stimulating release of Ca2+ directly from SR or via t-system depolarization, voltage dependent and independent fluxes across the t-system can be spatiotemporally resolved against the release flux of Ca2+ from SR. These measurements have identified an action potential-induced Ca2+ flux across the t-system and an ultra-rapid store-operated Ca2+ entry (SOCE) mechanism in skeletal muscle. This has led to the development of a working model for SOCE that is relevant within the large Ca2+ release fluxes initiated by excitation-contraction coupling. Potential roles of the action potential-induced Ca2+ flux in skeletal muscle remain speculative.