Lledo P M, Legendre P, Israel J M, Vincent J D
INSERM U.176, 33077 Bordeaux, France.
Endocrinology. 1990 Sep;127(3):990-1001. doi: 10.1210/endo-127-3-990.
The effects of dopamine (DA) on voltage-dependent Ca2+ currents were investigated in cultured rat lactotroph cells using the patch clamp recording technique. Each recorded cell was identified by the reverse hemolytic plaque assay. In the whole-cell configuration, two types of Ca2+ currents, L and T, were characterized on the basis of their kinetics, voltage sensitivity, and pharmacology. The L component had a threshold of -25 mV, showed little inactivation during a 150-msec voltage step, and was maximal at +10 mV. Cadmium ions (100 microM) significantly reduced its amplitude (75%). The T component was activated at a membrane potential close to -50 mV, was maximal at -10 mV, and showed a voltage-dependent inactivation between -90 and -30 mV. It was quickly inactivated during a maintained depolarization (time constant, 27 ms at -30 mV) and was strongly reduced (80%) by nickel ions (100 microM). Bath application of DA (10 nM) caused a markedly general depression of inward Ca2+ currents, acting differently on the T- and L-type currents. DA application shifted the voltage-dependence of the L-type current activation toward depolarization values (8 mV) without modifying its time- and voltage-dependent inactivation. In contrast, DA enhanced the inactivation of the T-type current by accelerating its time-dependent inactivation (25% decrease in the time constant of inactivation) and by shifting the voltage-dependence of the T-type current inactivation toward hyperpolarizing values (-63 mV in control vs. -77 mV in the presence of DA). These effects of DA were dose-dependent and involved the activation of a D2 receptor type. They were mimicked by bromocriptine application (10 nM), whereas sulpiride (100 nM) blocked the DA-evoked response. The D1 antagonist SCH 23390 was ineffective up to 100 microM. All of these DA-induced modifications in Ca2+ currents were abolished using a GTP-free pipette solution or after pretreatment of cells with pertussis toxin, suggesting that DA can regulate the function of Ca2+ channels through GTP-binding proteins (G-proteins). Our results show that DA acts simultaneously by reducing both voltage-dependent Ca2+ currents on lactotroph cells. Thus, DA reduces the entry of Ca2+ ions across the surface membrane and thereby influences electrical activity and the cytosolic free Ca2+ concentration involved in both basal and evoked PRL release.
采用膜片钳记录技术，在培养的大鼠促乳素细胞中研究了多巴胺（DA）对电压依赖性Ca2+电流的影响。通过反向溶血空斑试验鉴定每一个记录的细胞。在全细胞模式下，根据其动力学、电压敏感性和药理学特性，对两种类型的Ca2+电流，即L型和T型电流进行了表征。L型电流成分的阈值为-25 mV，在150毫秒的电压阶跃期间几乎没有失活，在+10 mV时达到最大值。镉离子（100 microM）显著降低其幅度（75%）。T型电流成分在接近-50 mV的膜电位时被激活，在-10 mV时达到最大值，并在-90至-30 mV之间表现出电压依赖性失活。在持续去极化过程中它迅速失活（在-30 mV时时间常数为27毫秒），并被镍离子（100 microM）强烈降低（80%）。浴槽中加入DA（10 nM）导致内向Ca2+电流明显普遍降低，对T型和L型电流的作用不同。施加DA使L型电流激活的电压依赖性向去极化值偏移（8 mV），而不改变其时间和电压依赖性失活。相反，DA通过加速其时间依赖性失活（失活时间常数降低25%）并将T型电流失活的电压依赖性向超极化值偏移（对照中为-63 mV，存在DA时为-77 mV）来增强T型电流的失活。DA的这些作用呈剂量依赖性，涉及D2型受体的激活。它们可被施加溴隐亭（10 nM）模拟，而舒必利（100 nM）可阻断DA诱发的反应。D1拮抗剂SCH 23390在高达100 microM时无效。使用无GTP的移液管溶液或在用百日咳毒素预处理细胞后，所有这些DA诱导的Ca2+电流变化均被消除，这表明DA可通过GTP结合蛋白（G蛋白）调节Ca2+通道的功能。我们的结果表明，DA通过降低促乳素细胞上的电压依赖性Ca2+电流同时发挥作用。因此DA减少Ca2+离子跨表面膜的内流，从而影响基础和诱发的PRL释放中涉及的电活动和胞质游离Ca2+浓度。
