Oba T, Yamaguchi M
Department of Physiology, Nagoya City University Medical School, Japan.
Am J Physiol. 1990 Nov;259(5 Pt 1):C709-14. doi: 10.1152/ajpcell.1990.259.5.C709.
To examine the molecular mechanism underlying contractile activation, we studied effects of a sulfhydryl reagent, N-(7-dimethylamino-4-methylcoumarinyl)maleimide (DACM), on twitch, Ag(+)-induced contraction, and K+ and caffeine contractures in single toe muscle fibers of frog. DACM suppressed twitch and Ag(+)-induced contraction, dose dependently, but not caffeine contracture. K+ contracture also was decreased appreciably by exposure to 40 microM DACM for 10 min. DACM elicited no shift of the mechanical threshold or inhibition of resting potential but slightly inhibited action potential. Increase of the fluorescence intensity produced by binding of 10 microM DACM to sulfhydryl groups was depressed by brief pretreatment with 100 microM Ag+. When exposed to 1 mM dithiothreitol (DTT) within 5 s of the rising phase of 5 microM Ag(+)-induced contraction, the fiber rapidly decreased the tension to the resting level. In this case, reapplication of 5 microM Ag+ after washing out DTT elicited a new contraction similar to the first Ag(+)-induced contraction. The second contraction amplitude depended on the time between the onset of the first Ag(+)-induced contraction and DTT application. If DTT was applied after more than 16 s, tension no longer developed on the second exposure to Ag+ or K+. The experiments provide evidence that crucial sulfhydryl groups participate in muscle activation. The possible role of the sulfhydryl group on the transverse tubular membrane in tension development is discussed.
为了研究收缩激活的分子机制，我们研究了一种巯基试剂N-(7-二甲基氨基-4-甲基香豆素基)马来酰亚胺（DACM）对青蛙单趾肌纤维的单收缩、银离子（Ag⁺）诱导的收缩以及钾离子（K⁺）和咖啡因诱导的挛缩的影响。DACM剂量依赖性地抑制单收缩和Ag⁺诱导的收缩，但不抑制咖啡因诱导的挛缩。暴露于40微摩尔/升的DACM 10分钟后，K⁺挛缩也明显降低。DACM没有引起机械阈值的变化或静息电位的抑制，但对动作电位有轻微抑制作用。100微摩尔/升的Ag⁺预处理可抑制10微摩尔/升的DACM与巯基结合产生的荧光强度增加。在5微摩尔/升的Ag⁺诱导收缩的上升期5秒内暴露于1毫摩尔/升的二硫苏糖醇（DTT）时，纤维迅速将张力降至静息水平。在这种情况下，洗去DTT后重新施加5微摩尔/升的Ag⁺会引发类似于第一次Ag⁺诱导收缩的新收缩。第二次收缩幅度取决于第一次Ag⁺诱导收缩开始与DTT施加之间的时间。如果在超过16秒后施加DTT，第二次暴露于Ag⁺或K⁺时不再产生张力。这些实验提供了关键巯基参与肌肉激活的证据。讨论了横管膜上巯基在张力产生中的可能作用。
