Ryanodine receptor type 1 (RyR1) possessing malignant hyperthermia mutation R615C exhibits heightened sensitivity to dysregulation by non-coplanar 2,2′,3,5′,6-pentachlorobiphenyl (PCB 95)

Abstract

Malignant hyperthermia (MH) susceptibility is conferred by inheriting one of >60 missense mutations within the highly regulated microsomal Ca 2+ channel known as ryanodine receptor type 1 (RyR1). Although MH susceptible patients lack overt clinical signs, a potentially lethal MH syndrome can be triggered by exposure to halogenated alkane anesthetics. This study compares how non-coplanar 2,2′,3,5′,6-pentachlorobiphenyl (PCB 95), a congener identified in environmental and human samples, alters the binding properties of [ 3H]ryanodine to RyR1 in vitro. Junctional sarcoplasmic reticulum (SR) was isolated from skeletal muscle dissected from wild type pigs ( WtRyR1) and pigs homozygous for MH mutation R615C ( MHRyR1), a mutation also found in humans. Although the level of WtRyR1 and MHRyR1 expression is the same, MHRyR1 shows heightened sensitivity to activation and altered regulation by physiological cations. We report here that MHRyR1 shows more pronounced activation by Ca 2+, and is less sensitive to channel inhibition by Ca 2+ and Mg 2+, compared to WtRyR1. In a buffer containing 100 nM free Ca 2+, conditions typically found in resting cells, PCB 95 (50–1000 nM) enhances the activity of MHRyR1 whereas it has no detectable effect on WtRyR1. PCB 95 (2 μM) decreases channel inhibition by Mg 2+ to a greater extent in MHRyR1 (IC 50 increased nine-fold) compared to WtRyR1 (IC 50 increased by 2.5-fold). PCB95 reduces inhibition by Ca 2+ two-fold more with MHRyR1 than WtRyR1. Our data suggest that non-coplanar PCBs are more potent and efficacious toward MHRyR1 than WtRyR1, and have more profound effects on its cation regulation. Considering the important roles of Ca 2+ and Mg 2+ in regulating Ca 2+ signals involving RyR channels, these data provide the first mechanistic evidence that a genetic mutation known to confer susceptibility to pharmacological agents also enhances sensitivity to an environmental contaminant.