Changes in cytoplasmic free calcium caused by halothane. Role of the plasma membrane and intracellular Ca2+stores

Abstract

Malignant hyperthermia is a muscle disease characterized by an abnormal response to anaesthetics, stress, and exercise. It is typified by muscle contracture and a dramatic elevation in body temperature. A defect in the regulation of the concentration of cytoplasmic free calcium, [Ca²]_i, is thought to underlie this disease, but the actual [Ca²⁺]_i was not measurable until recently. We have shown that the anaesthetic halothane increases [Ca²⁺]_i in isolated lymphocytes from malignant hyperthermia-susceptible humans and pigs but not in the normal counterparts. In this report we extend these observations to a larger number of cases and analyze the molecular mechanisms responsible for the increase. The halothane-mediated rise in [Ca²⁺]_i required external Ca²⁺ and was prevented by nifedipine, an inhibitor of the voltage-sensitive Ca²⁺ channels of the cell membrane. In addition, the effect of halothane on the releasable Ca²⁺ from intracellular stores was determined by measuring the size of the releasable pool before and after addition of the anaesthetic. After addition of halothane, about 73% of this Ca²⁺ pool was still available for release by the Ca²⁺ ionophore ionomycin in cells from normal humans and pigs. In contrast, only about 45% of the free Ca²⁺ in intracellular stores was left after treatment with halothane in cells from malignant hyperthermia-susceptible humans and swine. These results indicate that halothane acts both at the cell membrane and at intracellular organelles, and that this action results in a net increase in [Ca²⁺]_i in malignant hyperthermia, but not in normal cells. The action at the cell membrane appears to be on the voltage-sensitive Ca²⁺ channels. Further, it is suggested that the halothane-mediated increase in [Ca²⁺]_i could be used as the basis for a noninvasive test of malignant hyperthermia.