Yoshino M, Matsufuji Y, Yabu H
Department of Physiology, Sapporo Medical University, Japan.
Naunyn Schmiedebergs Arch Pharmacol. 1996 Feb;353(3):334-41. doi: 10.1007/BF00168637.
The suppressive action of caffeine on L-type Ca current (Ica) in smooth muscle cells of the guinea-pig urinary bladder was investigated using the whole-cell patch clamp technique. Caffeine (5-30 mM) suppressed Ica, the effect having two phases: a rapid and transient suppression of Ica, which was followed by a sustained suppression. When intracellular Ca2+ was strongly buffered by the Ca2+ chelator EGTA (20 mM) or BAPTA (5 mM) in the patch pipette, the transient suppression of Ica was abolished, whereas the sustained effect remained. Similarly, inclusion of both 10 mM procaine and 1 mg/ml heparin in the patch pipette blocked the transient suppression of Ica, but did not block the sustained effect. The degree of the sustained effect of caffeine on Ica was dose-dependent with a kd of 20 mM. Application of the cyclic AMP analogue, 8-bromo-cyclic AMP (100 microM) or forskolin (10 microM) to the bath failed to mimick the sustained suppression of Ica, suggesting that inhibition of phosphodiesterase activity was not involved in the caffeine action. The steady-state activation curve remained unchanged by 10 mM caffeine but the steady-state inactivation curve was significantly shifted in the negative direction by 15.6 mV in 1.8 mM Ca2+ solution or by 10 mV in 1.8 mM Ba2+ solution. From these results it appears that caffeine inhibits L-type Ica via two mechanisms: (1) it releases Ca2+ from an internal store causing a transient Ca2+ -mediated inactivation of the Ca channel; (2) it inhibits Ca channel via a mechanism that does not require such a Ca2+ release. It is possible that caffeine suppresses Ica through a preferential binding to the inactivated state of L-type Ca channel.
采用全细胞膜片钳技术研究了咖啡因对豚鼠膀胱平滑肌细胞L型钙电流（Ica）的抑制作用。咖啡因（5 - 30 mM）抑制Ica，其作用有两个阶段：Ica的快速短暂抑制，随后是持续抑制。当膜片钳微管中的Ca2+螯合剂EGTA（20 mM）或BAPTA（5 mM）强烈缓冲细胞内Ca2+时，Ica的短暂抑制被消除，而持续作用仍然存在。同样，膜片钳微管中加入10 mM普鲁卡因和1 mg/ml肝素可阻断Ica的短暂抑制，但不阻断持续作用。咖啡因对Ica持续作用的程度呈剂量依赖性，解离常数为20 mM。向浴槽中施加环磷酸腺苷类似物8 - 溴环磷酸腺苷（100 microM）或福斯高林（10 microM）未能模拟Ica的持续抑制，表明磷酸二酯酶活性的抑制不参与咖啡因的作用。10 mM咖啡因使稳态激活曲线保持不变，但在1.8 mM Ca2+溶液中，稳态失活曲线显著向负向移动15.6 mV，在1.8 mM Ba2+溶液中向负向移动10 mV。从这些结果看来，咖啡因通过两种机制抑制L型Ica：（1）它从内部储存库释放Ca2+，导致Ca通道的短暂Ca2+介导的失活；（2）它通过一种不需要这种Ca2+释放的机制抑制Ca通道。咖啡因可能通过优先结合L型钙通道的失活状态来抑制Ica。
