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

最近有报道称苯二氮卓类药物抑制Na^+，K^+- atp酶的活性，但细节尚不清楚。我们旨在阐明苯二氮卓类药物对Na^+，K^+- atp酶的抑制机制。采用兔脑纯化的Na^+、K^+- atp酶检测苯二氮卓类药物（咪达唑仑、地西泮和氟硝西泮）对Na^+、K^+- atp酶和Na^+- atp酶活性以及Na^+、K^+- atp酶磷酸化中间体（EP）的影响。得到了以下结果：三种苯二氮卓类药物均能抑制Na^+、K^+- atp酶和Na^+ - atp酶活性和EP形成，且呈剂量依赖性，对Na^+、K^+ atp酶活性的半最大抑制浓度（Ki 0.5）分别为0.6、0.42和0.25 mM，对地西泮、咪达唑仑和氟硝西泮均有抑制作用。所有苯二氮卓类药物对Na^+、K^+-ATPase活性的抑制顺序为EP形成> Na^+、K^+-ATPase活性> Na^+-ATPase活性，表明苯二氮卓类药物对Na^+、K^+-ATPase的抑制主要是通过抑制EP形成后的反应引起的。所有苯二氮卓类药物通过稀释其浓度部分恢复活性。氟马西尼不影响苯二氮卓类药物对Na^+、K^+- atp酶活性和EP形成的抑制作用，也不影响抑制的恢复试验，说明氟马西尼不与苯二氮卓类药物对Na^+、K^+- atp酶和mat的作用形成竞争。苯二氮卓类药物的Na^+、K^+- atp酶与GABA_A受体的结合位点结构不同。在其他实验中，我们发现局麻药利多卡因、普鲁卡因和二布卡因通过抑制EP形成来抑制Na^+、K^+- atp酶活性，局麻药的某些方面可能与Na^+、K^+- atp酶活性的抑制有关。我们还发现，不同种类的麻醉药对Na^+，K^+- atp酶活性的抑制机制不同，异氟醚通过降低EP对钾离子的敏感性来抑制活性。少

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