To study the properties of the Na extrusion mechanism, giant muscle fibers from barnacle (Balanus nubilus) were internally perfused with solutions containing tracer 22Na. In fibers perfused with solutions containing adenosine 5'-triphosphate (ATP) and 30 mM Na, the Na efflux into 10 mM K seawater was approximately 25-30 pmol/cm2.s; 70% of this efflux was blocked by 50-100 microM ouabain, and approximately 30% was blocked by removal of external K. The ouabain-sensitive and K-dependent Na effluxes were abolished by depletion of internal ATP and were sigmoid-shaped functions of the internal Na concentration ([Na]i), with half-maxima at [Na]i approximately or equal to 20 mM. These sigmoid functions fit the Hill equation with Hill coefficients of approximately 3.5. Ouabain depolarized ATP-fueled fibers by 1.5-2 mV ([Na]i greater than or equal to 30 mM) but had very little effect on the membrane potential of ATP-depleted fibers; ATP depletion itself caused a 2-2.5-mV depolarization. When fueled fibers were treated with 3,4-diaminopyridine or Ba2+ (to reduce the K conductance and increase membrane resistance), application of ouabain produced a 4-5 mV depolarization. These results indicate that an electrogenic, ATP-dependent Na-K exchange pump is functional in internally perfused fibers; the internal perfusion technique provides a convenient method for performing transport studies that require good intracellular solute control.
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机译:为了研究Na挤出机制的特性,将藤壶(Balanus nubilus)的巨大肌纤维内部注入了含有示踪剂22Na的溶液。在灌注了含有5'-三磷酸腺苷(ATP)和30 mM Na的溶液的纤维中,Na向10 mM K海水中的流出量约为25-30 pmol / cm2.s。这种外排的70%被50-100 microM哇巴因阻滞,大约30%被外部K的去除阻滞。哇巴因敏感和K依赖的Na外排因内部ATP的消耗而被消除,并且是乙状结肠功能。内部Na浓度([Na]),在[Na]处的半最大值约为或等于20mM。这些S型函数使Hill方程的Hill系数约为3.5。瓦巴因使以ATP为燃料的纤维去极化1.5-2 mV(Na大于或等于30 mM),但对消耗ATP的纤维的膜电位影响很小; ATP耗尽本身会导致2-2.5-mV的去极化。用3,4-二氨基吡啶或Ba2 +处理燃料纤维(以降低K电导并增加膜电阻)时,哇巴因的应用产生了4-5 mV的去极化。这些结果表明,一个电致ATP依赖的Na-K交换泵在内部灌注的纤维中起作用。内部灌注技术为进行需要良好细胞内溶质控制的转运研究提供了一种方便的方法。
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