Raptor ablation in skeletal muscle decreases Cav1.1 expression and affects the function of the excitation-contraction coupling supramolecular complex.

The protein mammalian target of rapamycin (mTOR) is a serine/threonine kinase regulating a number of biochemical pathways controlling cell growth. mTOR exists in two complexes termed mTORC1 and mTORC2. Regulatory associated protein of mTOR (raptor) is associated with mTORC1 and is essential for its function. Ablation of raptor in skeletal muscle results in several phenotypic changes including decreased life expectancy, increased glycogen deposits and alterations of the twitch kinetics of slow fibres. In the present paper, we show that in muscle-specific raptor knockout (RamKO), the bulk of glycogen phosphorylase (GP) is mainly associated in its cAMP-non-stimulated form with sarcoplasmic reticulum (SR) membranes. In addition, 3[H]-ryanodine and 3[H]-PN200-110 equilibrium binding show a ryanodine to dihydropyridine receptors (DHPRs) ratio of 0.79 and 1.35 for wild-type (WT) and raptor KO skeletal muscle membranes respectively. Peak amplitude and time to peak of the global calcium transients evoked by supramaximal field stimulation were not different between WT and raptor KO. However, the increase in the voltage sensor-uncoupled RyRs leads to an increase of both frequency and mass of elementary calcium release events (ECRE) induced by hyper-osmotic shock in flexor digitorum brevis (FDB) fibres from raptor KO. The present study shows that the protein composition and function of the molecular machinery involved in skeletal muscle excitation-contraction (E-C) coupling is affected by mTORC1 signalling.

An RYR1 mutation associated with malignant hyperthermia is also associated with bleeding abnormalities.

Malignant hyperthermia is a potentially fatal hypermetabolic disorder triggered by halogenated anesthetics and the myorelaxant succinylcholine in genetically predisposed individuals. About 50% of susceptible individuals carry dominant, gain-of-function mutations in RYR1 [which encodes ryanodine receptor type 1 (RyR1)], though they have normal muscle function and no overt clinical symptoms. RyR1 is predominantly found in skeletal muscle but also at lower amounts in immune and smooth muscle cells, suggesting that RYR1 mutations may have a wider range of effects than previously suspected. Mild bleeding abnormalities have been described in patients with malignant hyperthermia carrying gain-of-function RYR1 mutations. We sought to determine the frequency and molecular basis for this symptom. We found that some patients with specific RYR1 mutations had abnormally high bleeding scores, whereas their healthy relatives did not. Knock-in mice with the malignant hyperthermia susceptibility RYR1 mutation Y522S (MHS RYR1Y522S) had longer bleeding times than their wild-type littermates. Primary vascular smooth muscle cells from RYR1Y522S knock-in mice exhibited a higher frequency of subplasmalemmal Ca(2+) sparks, leading to a more negative resting membrane potential. The bleeding defect of RYR1Y522S mice and of one patient was reversed by treatment with the RYR1 antagonist dantrolene, and Ca(2+) sparks in primary vascular smooth muscle cells from the MHS RYR1Y522S mice were blocked by ryanodine or dantrolene. Thus, RYR1 mutations may lead to prolonged bleeding by altering vascular smooth muscle cell function. The reversibility of the bleeding phenotype emphasizes the potential therapeutic value of dantrolene in the treatment of such bleeding disorders.