Expression and function of calcium channels in skeletal muscle

Skeletal muscle excitation-contraction coupling results from voltage-dependent interactions between two different calcium channels: voltage-gated dihydropyridine receptors (DHPR) and ligand-gatged ryanodine receptors (RYR). The basic constituents of these calcium channels have been known for some time, yet the function of the different subunit isoforms that make up each channel molecules remains poorly understood. In this thesis, I have tried to uncover some of the physiological roles for these proteins in skeletal muscle function.;I employed biophysical and pharmacological methods to study the physiological role of type-3 ryanodine receptors (RYR3) in developing skeletal muscle. I showed that RYR3 is essential for IPTXA-mediated enhancement of the calcium transient in skeletal myotubes. IPTXA increased the amplitude and the rate of rise of the calcium transient in wild-type, but not RYR3-null skeletal muscle, indicating that RYR3 participates in CICR in skeletal muscle cells.;Skeletal muscle expresses a variety of alpha2/delta subunit transcripts. The expression of most of these transcripts is stable during differentiation. However, the expression of two isoforms changes during differentiation. During in vitro differentiation, expression of the alpha2/delta-1a and alpha2/delta-1e isoforms changed. Calcium channel kinetics and amplitude strongly correlated with alpha2/delta-1a expression. These findings suggest that the alpha2/delta-1 subunit may play a key role in regulating calcium channel activity in skeletal muscle.;I examined cellular targeting of the alpha2/delta-1a subunit during differentiation. In two-day old myotubes, alpha2/delta-1a is diffusely expressed throughout the myotube. Few alpha2/delta-1a protein co-localized with alpha1S. In contrast, many distinct foci containing both alpha2/delta-1a and alpha1S were found. This suggests that the sorting of the alpha2/delta-1a protein may be controlled by several different biochemical signals.;In conclusion, I have investigated the physiological role of several different calcium channel subunits in skeletal muscle excitation-contraction coupling. My results suggest that differential expression of calcium channel subunits may be an important mechanism muscle cells employ to shape intracellular calcium signaling.