Novel CACNA1S mutations for malignant hyperthermia enhance depolarization-induced Ca²⁺ release

Abstract

Malignant hyperthermia (MH) is a life-threatening genetic disorder in which general anesthetics cause massive Ca²⁺release from the sarcoplasmic reticulum in skeletal muscle. While most MH mutations have been found in the ryanodine receptor type 1 (RyR1), other genes are also associated with MH. We have recently identified several novel mutations in CACNA1S, the α1S subunit of the dihydropyridine receptor, from MH-susceptible patients. In this study, we performed a functional analysis of MH mutants using the reconstituted depolarization-induced Ca²⁺ release (DICR) platform. HEK293 cells expressing RyR1 were infected with baculovirus containing genes essential for DICR (CACNA1S, β1a, JP2, Stac3, and Kir2.1). DICR was induced by depolarization with different concentrations of potassium (K⁺) solutions. Cells expressing two CACNA1S mutants showed greater sensitivity to K⁺ than WT cells. Consistently, charge movements were shifted to more hyperpolarizing potentials. These results suggest that CACNA1S MH mutations shift the DICR to a more hyperpolarizing potential. The two mutant cells showed greater sensitivity to caffeine than WT cells. Interestingly, the enhanced caffeine sensitivity was abolished by hyperpolarizing the resting membrane potential or by removing Stac3. Possible mechanisms of MH pathogenesis are discussed.