Pharmacologic modulation has previously shown that transient receptor potential canonical (TRPC) channels play an important role in the pathogenesis of malignant hyperthermia. This study tested the hypothesis that genetically suppressing the function of TRPC6 can partially ameliorate muscle cation dyshomeostasis and the response to halothane in a mouse model relevant to malignant hyperthermia.
This study examined the effect of overexpressing a muscle-specific nonconducting dominant-negative TRPC6 channel in 20 RYR1-p.R163C and 20 wild-type mice and an equal number of nonexpressing controls, using calcium- and sodium-selective microelectrodes and Western blots.
RYR1-p.R163C mouse muscles have chronically elevated intracellular calcium and sodium levels compared to wild-type muscles. Transgenic expression of the nonconducting TRPC6 channel reduced intracellular calcium from 331 ± 34 nM (mean ± SD) to 190 ± 27 nM (P < 0.0001) and sodium from 15 ± 1 mM to 11 ± 1 mM (P < 0.0001). Its expression lowered the increase in intracellular Ca2+ of the TRPC6-specific activator hyperforin in RYR1-p.R163C muscle fibers from 52% (348 ± 37 nM to 537 ± 70 nM) to 14% (185 ± 11 nM to 210 ± 44 nM). Western blot analysis of TRPC3 and TRPC6 expression showed the expected increase in TRPC6 caused by overexpression of its dominant-negative transgene and a compensatory increase in expression of TRPC3. Although expression of the muscle-specific dominant-negative TRPC6 was able to modulate the increase in intracellular calcium during halothane exposure and prolonged life (35 ± 5 min vs. 15 ± 3 min; P < 0.0001), a slow, steady increase in calcium began after 20 min of halothane exposure, which eventually led to death.
These data support previous findings that TRPC channels play an important role in causing the intracellular calcium and sodium dyshomeostasis associated with RYR1 variants that are pathogenic for malignant hyperthermia. However, they also show that modulating TRPC channels alone is not sufficient to prevent the lethal effect of exposure to volatile anesthetic malignant hyperthermia–triggering agents.
The type 1 ryanodine receptor (RYR1) gene encoding the skeletal muscle sarcoplasmic reticulum Ca2+ release channel is the primary locus for malignant hyperthermia susceptibility
Secondary loci have been identified in other proteins involved with RyR1 in excitation–contraction coupling that are thought to sensitize the RyR1 channel
Transient receptor potential canonical channels constitute a large and functionally versatile superfamily of cation channel proteins expressed in many cell types that control influxes of Ca2+ and other cations to regulate diverse cellular processes
Muscle-specific overexpression of the nonconducting transient receptor potential canonical 6 channel both reduced intracellular Ca2+ concentration in RYR1-p.R163C mice at rest and reduced the absolute maximum levels of intracellular Ca2+ concentration reached during exposure to halothane
Despite this, its overexpression did not restore intracellular Ca2+ concentration to wild-type levels, and although its overexpression increased the length of survival after halothane exposure, it was unable to rescue the lethal phenotype