Rusin K I, Giovannucci D R, Stuenkel E L, Moises H C
Department of Physiology, University of Michigan Medical School, Ann Arbor, Michigan 48109-0622, USA.
J Neurosci. 1997 Sep 1;17(17):6565-74. doi: 10.1523/JNEUROSCI.17-17-06565.1997.
Whole-cell patch-clamp recordings were performed together with time-resolved measurements of membrane capacitance (Cm) in nerve terminals acutely dissociated from neurohypophysis of adult rats to investigate modulation of Ca2+ currents and secretion by activation of opioid receptors. Bath superfusion of the kappa-opioid agonists U69,593 (0.3-1 microM), dynorphin A (1 microM), or U50,488H (1-3 microM) reversibly suppressed the peak amplitude of Ca2+ currents 32. 7 +/- 2.7% (in 41 of 56 terminals), 37.4 +/- 5.3% (in 5 of 8 terminals), and 33.5 +/- 8.1% (in 5 of 10 terminals), respectively. In contrast, tests in 11 terminals revealed no effect of the mu-opioid agonist [D-Pen2,5]-enkephalin (1-3 microM; n = 7) or of the delta-agonist Tyr-D-Ala-Gly-N-Me-Phe-Gly-ol (1 microM; n = 4) on Ca2+ currents. Three components of high-threshold current were distinguished on the basis of their sensitivity to blockade by omega-conotoxin GVIA, nicardipine, and omega-conotoxin MVIIC: N-, L-, and P/Q-type current, respectively. Administration of U69,593 inhibited N-type current in these nerve terminals on average 32%, whereas L-type current was reduced 64%, and P/Q-type current was inhibited 28%. Monitoring of changes in Cm in response to brief depolarizing steps revealed that the kappa-opioid-induced reductions in N-, L-, or P/Q-type currents were accompanied by attenuations in two kinetically distinct components of Ca2+-dependent exocytotic release. These data provide strong evidence of a functional linkage between blockade of Ca2+ influx through voltage-dependent Ca2+ channels and inhibitory modulation of release by presynaptic opioid receptors in mammalian central nerve endings.
采用全细胞膜片钳记录技术，并同步对成年大鼠神经垂体急性分离出的神经末梢进行膜电容（Cm）的时间分辨测量，以研究阿片受体激活对Ca2+电流和分泌的调节作用。用κ-阿片受体激动剂U69,593（0.3 - 1微摩尔）、强啡肽A（1微摩尔）或U50,488H（1 - 3微摩尔）对浴槽进行灌流，可分别可逆性地抑制Ca2+电流的峰值幅度，抑制率分别为32.7±2.7%（56个神经末梢中的41个）、37.4±5.3%（8个神经末梢中的５个）和33.5±8.1%（10个神经末梢中的5个）。相比之下，在11个神经末梢中进行的测试显示，μ-阿片受体激动剂[D-Pen2,5]-脑啡肽（1 - 3微摩尔；n = 7）或δ-阿片受体激动剂Tyr-D-Ala-Gly-N-Me-Phe-Gly-ol（1微摩尔；n = 4）对Ca2+电流没有影响。根据高阈值电流对ω-芋螺毒素GVIA、尼卡地平以及ω-芋螺毒素MVIIC阻断作用的敏感性，区分出三种高阈值电流成分，分别为N型、L型和P/Q型电流。给予U69,593后，这些神经末梢中的N型电流平均被抑制32%，L型电流减少64%，P/Q型电流被抑制28%。监测对短暂去极化步骤的反应中Cm的变化发现，κ-阿片受体诱导的N型、L型或P/Q型电流减少伴随着Ca2+依赖性胞吐释放的两个动力学上不同的成分的衰减。这些数据为哺乳动物中枢神经末梢中通过电压依赖性Ca2+通道阻断Ca2+内流与突触前阿片受体对释放的抑制性调节之间的功能联系提供了有力证据。
