Programme Contents

Voltage-dependent and -independent Ca²⁺ entry into skeletal muscle during excitation-contraction coupling

Excitation-contraction coupling (EC coupling) is the process that links the activation of the surface membrane of a muscle to the force response produced. This process is reliant on a rapid and large change in cytoplasmic [Ca²⁺] ([Ca²⁺]_cyto) due to release activated from the internal Ca²⁺ store, the sarcoplasmic reticulum (SR). The change in [Ca²⁺]_cyto regulates the movement of the contractile proteins in the muscle to produce force. Also, during EC coupling there can be an input of Ca²⁺ across the surface membrane, or from the invagination of this membrane, the tubular (t-) system. In skeletal muscle, this influx of Ca²⁺ is not strictly required for a contracture to occur, but its importance may be in the longer-term regulation of function or may indeed have more subtle, immediate consequences that are not clear from present measurements. Several pathways can be responsible for an influx of Ca²⁺ into skeletal muscle fibres during EC coupling. These can be voltage-dependent, including L-type Ca²⁺ current, action potential-activated Ca²⁺ current (APACC) and excitation-coupled Ca²⁺ entry (ECCE); or voltage-independent, including store-operated Ca²⁺ entry (SOCE), stretch-activated Ca²⁺ entry or otherwise activated to enter through transient receptor protein (TRP) channels. It should be noted that the conditions that prevail under bouts of EC coupling are the conditions that activate voltage-dependent and –independent Ca²⁺ entry during EC coupling in skeletal muscle (Allen et al. 2005; Launikonis et al., 2010).

The nature of these voltage-dependent and -independent pathways in skeletal muscle have been examined by a number of groups using either fully differentiated fibres or myotubes. A problem that does arise here is that properties derived on myotubes are often assumed to be very similar to that in the fully differentiated muscle. There are significant differences in myotube and adult fibre physiology and membrane ultrastructure, making extrapolation of results derived on one cell to other not a simple matter. The examination of Ca²⁺ currents in myotubes may have been necessary in some instances because conventional electrophysiological methods are difficult or do not have the sensitivity to measure such tiny Ca²⁺ currents in adult fibres. Furthermore, the use of pharmacological agents commonly used to block Ca²⁺ entry pathways are, unfortunately, non-specific in most cases. A combination of these factors may have lead to a misrepresentation of certain pathways existing in myotubes or adult fibres.

In adult, mammalian skeletal muscle fibres it has been possible to image Ca²⁺ in the lumen of the t-system using a low-affinity Ca²⁺-sensitive fluorescent dye. The dye is trapped in the t-system of skinned fibres, where the surface membrane has been removed by microdissection. With this preparation, Ca²⁺ movement across the t-system membrane is visualized as a net change in the Ca²⁺-dependent fluorescence signal emitted from the t-system lumen. This can be monitored in conjunction with a spectrally separate Ca²⁺-sensitive dye in the cytoplasm to monitor changes in [Ca²⁺]_cyto, which will be predominately due to fluxes across the SR membrane. This preparation has allowed the identification and characterization of SOCE during voltage-independent Ca²⁺ release and also APACC, which has been identified and observed to occur during single and trains of action potentials in adult mammalian skeletal muscle fibres.

Allen DG, Whitehead NP, Yeung EW. (2005) Journal of Physiology, 567: 723-735.

Launikonis BS, Murphy RM, Edwards JN. (2010) Pflügers Archiv, 460: 813-823.