Study for the action and side effects of anesthetics using plasma membrane Ca-ATPase as a model

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Recently it was reported that the activity of Na^+,K^+-ATPase was inhibited by benzodiazepines, but detail was not yet clear. We aimed to clarify the inhibition mechanism of benzodiazepines for Na^+,K^+-ATPase. The effects of benzodiazepines (midazolam, diazepam and flunitrazepam), on Na^+,K^+-ATPase and Na^+-ATPase activities, and phosphointemiediate (EP) of Na^+,K^+-ATPase were tested using Na^+,K^+-ATPase purified from rabbit brain. The following results were obtained. All three benzodiazepines inhibited Na^+,K^+-ATPase and Na^+ -ATPase activities, and EP formation in a dose-dependent manner and the half maximal inhibition concentrations (Ki 0.5) for Na^+,K^+ATPase activity were 0.6, 0.42 and 0.25 mM, for diazepam, midazolam and flunitrazepam, respectively. Tlie orders of Ki 0.5 values were EP formation > Na^+,K^+-ATPase activity > Na^+-ATPase activity for all benzodiazepines, suggesting that inhibition of Na^+,K^+-ATPase by benzodiazepines was mainly caused by inhibiting the reacti … More on sequence after EP formation. The activities were partially recovered for all benzodiazepines by dilution of their concentrations. Flumazenil did not affect the inhibition of Na^+,K^+-ATPase activity and EP formation by benzodiazepines, and recovery test of inhibition, suggesting that flumazenil did not compete with the reactions of benzodiazepines on Na^+,K^+-ATPase and mat the.structure of binding sites of Na^+,K^+-ATPase for benzodiazepines are different from those of GABA_A receptor. In other experiments, we showed that local anesthetics, lidocaine, procaine and dibucaine, inhibited Na^+,K^+-ATPase activity by inhibition of EP formation and some aspects of local anesthesia may relate to the inhibition of Na^+,K^+-ATPase activity. We also showed that the inhibitory mechanisms of general anesthetics against Na^+,K^+-ATPase activity are diverse among the different categories of anesthetics and that isoflurane inhibit the activity by decrease in the sensitivity of EP to potassium ions. Less