Functional consequences of two critical point mutations in the skeletal muscle excitation-contraction coupling complex

Abstract

The control of calcium release in the mammalian skeletal muscle excitation-contraction coupling (e-c coupling) involves two Ca2+ ion channels: the L-type Ca2+ channel (Cav1.1) or dihydropyridine receptor (DHPR) and the ryanodine receptor type 1 (RyR1). The interaction between them is a conformational coupling, while the voltage gated DHPR serves as the voltage sensor of the RyR1. Mutations in these two ion channels can cause alterations or disruptions in e-c coupling.

In this dissertation, two murine models, one carrying a mutation in the pore loop of the DHPR and one carrying a mutation in the RyR1 were studied. Voltage clamp experiments showed that the mutation N617D in the DHPR selectivity filter region eliminated the L-type Ca2+ current. Measurements using a fluorescent indicator demonstrated that Ca2+ release properties, as well as the sarcoplasmic reticulum (SR) Ca2+ content, were unaffected. Interestingly, the mutation altered the voltage dependence of the channel inactivation at low external Ca2+ concentration.

The Y524S mutation in RyR1 is malignant hyperthermia causative and has been shown to cause alterations in Ca2+ release properties and channel inactivation. Here a possible retrograde Ca2+ release effect on Cav1.1 was investigated using SR depletion conditions and strong Ca2+ buffering. Despite experimental conditions that are known to effectively reduce local Ca2+ transients, the previously reported change in the calcium current inactivation persisted. The results indicate that voltage-activated Ca2+ release does not mediate retrograde modulation of inactivation in skeletal muscle e-c coupling of Y524S mice.