Abstract
Skeletal muscle fibres are large and highly elongated cells specialized for producing the force required for posture and movement. The process of controlling the production of force within the muscle, known as excitation-contraction coupling, requires virtually simultaneous release of large amounts of Ca2+ from the sarcoplasmic reticulum (SR) at the level of every sarcomere within the muscle fibre. Here we imaged Ca2+ movements within the SR, tubular (t-) system and in the cytoplasm to observe that the SR of skeletal muscle is a connected network capable of allowing diffusion of Ca2+ within its lumen to promote the propagation of Ca2+ release throughout the fibre under conditions where inhibition of SR ryanodine receptors (RyRs) was reduced. Reduction of cytoplasmic [Mg2+] ([Mg2+](cyto)) induced a leak of Ca2+ through RyRs, causing a reduction in SR Ca2+ buffering power argued to be due to a breakdown of SR calsequestrin polymers, leading to a local elevation of [Ca2+](SR). The local rise in [Ca2+](SR), an intra-SR Ca2+ transient, induced a local diffusely rising [Ca2+](cyto). A prolonged Ca2+ wave lasting tens of seconds or more was generated from these events. Ca2+ waves were dependent on the diffusion of Ca2+ within the lumen of the SR and ended as [Ca2+](SR) dropped to low levels to inactivate RyRs. Inactivation of RyRs allowed re-accumulation of [Ca2+](SR) and the activation of secondary Ca2+ waves in the persistent presence of low [Mg2+](cyto) if the threshold [Ca2+](SR) for RyR opening could be reached. Secondary Ca2+ waves occurred without an abrupt reduction in SR Ca2+ buffering power. Ca2+ release and wave propagation occurred in the absence of Ca2+-induced Ca2+ release. These observations are consistent with the activation of Ca2+ release through RyRs of lowered cytoplasmic inhibition by [Ca2+](SR) or store overload-induced Ca2+ release. Restitution of SR Ca2+ buffering power to its initially high value required imposing normal resting ionic conditions in the cytoplasm, which re-imposed the normal resting inhibition on the RyRs, allowing [Ca2+](SR) to return to endogenous levels without activation of store overload-induced Ca2+ release. These results are discussed in the context of how pathophysiological Ca2+ release such as that occurring in malignant hyperthermia can be generated.