Ay, Binnaz

Background Volatile anesthetics produce bronchodilation in part by depleting sarcoplasmic reticulum Ca stores in airway smooth muscle (ASM). Other bronchodilatory drugs are known to act via cyclic nucleotides (cyclic adenosine 3',5'-cyclic monophosphate, cyclic guanosine 3',5'-cyclic monophosphate). Intracellular Ca regulation in ASM involves plasma membrane Ca influx, including that triggered by sarcoplasmic reticulum Ca depletion (store-operated Ca entry [SOCE]). The authors hypothesized that anesthetics and bronchodilatory agents interact in inhibiting SOCE, thus enhancing ASM relaxation. Methods In enzymatically dissociated porcine ASM cells imaged using fluorescence microscopy, sarcoplasmic reticulum Ca was depleted by 1 microm cyclopiazonic acid in 0 extracellular Ca, nifedipine, and potassium chloride (preventing Ca influx through L-type channels and SOCE). Extracellular Ca was rapidly reintroduced to selectively activate SOCE in the presence or absence of 1 minimum alveolar concentration (MAC) halothane, isoflurane, or sevoflurane. Anesthetic interference with SOCE regulation by cyclic nucleotides was examined by activating SOCE in the presence of (1) 1 microm acetylcholine, (2) 100 microm dibutryl cyclic adenosine 3',5'-cyclic monophosphate, or (3) 100 microm 8-bromo-cyclic guanosine 3',5'-cyclic monophosphate. Results SOCE was enhanced by acetylcholine, whereas volatile anesthetics and both cyclic nucleotides partially inhibited Ca influx. Preexposure to 1 or 2 MAC anesthetic (halothane > isoflurane > sevoflurane) inhibited SOCE. Only halothane and isoflurane inhibited acetylcholine-induced augmentation of Ca influx, and significantly potentiated cyclic nucleotide inhibition such that no influx was observed in the presence of anesthetics and cyclic nucleotides. Conclusions These data indicate that volatile anesthetics prevent sarcoplasmic reticulum refilling by inhibiting SOCE and enhancing cyclic nucleotide blunting of Ca influx in ASM. Such interactions likely result in substantial airway relaxation in the presence of both anesthetics and bronchodilatory agents such as beta agonists or nitric oxide.

Background Ca2+ influx is a key component of neuronal intracellular Ca2+ ([Ca2+]i) regulation. The authors hypothesized that volatile anesthetic inhibition of neuronal activity is mediated by inhibition of Ca2+ influx via two major mechanisms: plasma membrane Na+-Ca2+ exchange (NCX) and the novel mechanism of Ca2+ influx triggered by endoplasmic reticulum Ca2+ depletion (store-operated Ca2+ channels [SOCCs]). Methods Differentiated rat pheochromocytoma cells loaded with the Ca2+ indicator fura-2 were Na+-loaded with 0 Ca2+, 145 mm Na+ Tyrode's and 5 microm cyclopiazonic acid plus 10 microm ryanodine (functionally isolating plasma membrane). Influx-mode NCX was rapidly reactivated by 0 Na+ and 2.5 mm Ca2+. The protocol was repeated in the presence of volatile anesthetics (0.5-1.5 minimum alveolar concentration [MAC] halothane, isoflurane, or sevoflurane) or other drugs to characterize NCX. To examine SOCCs, endoplasmic reticulum Ca2+ was depleted by cyclopiazonic acid in 0 extracellular Ca2+, and Ca2+ influx was triggered by rapid reintroduction of extracellular Ca2+. The protocol was repeated in the presence of anesthetics or other drugs to characterize SOCCs. Results Influx via NCX was not inhibited by voltage-gated Ca2+ channel blockers but was sensitive to NCX inhibitors. Halothane and isoflurane (0.5-1.5 MAC) significantly inhibited NCX (P < 0.05; paired comparisons), whereas sevoflurane at less than 1.5 MAC did not inhibit NCX. SOCC-mediated Ca2+ influx was insensitive to a variety of Ca2+ channel blockers but was inhibited by Ni2+. Such influx was sensitive only to halothane at greater than 1 MAC but not isoflurane or sevoflurane. Conclusions These data indicate that volatile anesthetics, especially halothane and isoflurane, interfere with neuronal [Ca2+]i regulation by inhibiting NCX but not SOCC-mediated Ca2+ influx (except high concentrations of halothane).

Background In airway smooth muscle (ASM), volatile anesthetics deplete sarcoplasmic reticulum (SR) Ca(2+) stores by increasing Ca(2+) "leak." Accordingly, SR replenishment becomes dependent on Ca(2+) influx. Depletion of SR Ca(2+) stores triggers Ca(2+) influx via specific plasma membrane channels, store-operated Ca(2+) channels (SOCC). We hypothesized that anesthetics inhibit SOCC triggered by increased SR Ca(2+) "leak," preventing SR replenishment and enhancing ASM relaxation. Methods In porcine ASM cells, SR Ca was depleted by cyclopiazonic acid or caffeine in 0 extracellular Ca(2+), nifedipine and KCl (preventing Ca(2+) influx through L-type and SOCC channels). Extracellular Ca(2+) was rapidly introduced to selectively activate SOCC. After SOCC activation, SR was replenished and the protocol repeated in the presence of 1 or 2 minimum alveolar concentration halothane, isoflurane, or sevoflurane. In other cells, characteristics of SOCC and interactions between acetylcholine (Ach) and volatile anesthetics were examined. Results Cyclopiazonic acid produced slow SR leak, whereas the caffeine response was transient in ASM cells. Reintroduction of extracellular Ca(2+) rapidly increased [Ca(2+)]i. This influx was insensitive to nifedipine, SKF-96365, and KBR-7943, inhibited by Ni and blockade of inositol 1,4,5-triphosphate-induced SR Ca(2+) release, and enhanced by ACh. Preexposure to 1 or 2 minimum alveolar concentration halothane completely inhibited Ca(2+) influx when extracellular Ca(2+) was reintroduced, whereas isoflurane and sevoflurane produced less inhibition. Only halothane and isoflurane inhibited ACh-induced augmentation of Ca(2+) influx. Conclusion Volatile anesthetics inhibit a Ni/La-sensitive store-operated Ca(2+) influx mechanism in porcine ASM cells, which likely helps maintain anesthetic-induced bronchodilation.