Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya, Japan; and.
Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya, Japan; and Department of Pharmacology, Division of Pathological Sciences, Kyoto Pharmaceutical University, Kyoto, Japan.
Am J Physiol Cell Physiol. 2014 Jun 1;306(11):C1008-16. doi: 10.1152/ajpcell.00014.2014. Epub 2014 Apr 2.
The pineal gland regulates circadian rhythm through the synthesis and secretion of melatonin. The rise of intracellular Ca(2+) concentration ([Ca(2+)]i) following nicotinic acetylcholine receptor (nAChR) stimulation due to parasympathetic nerve activity downregulates melatonin production. Important characteristics and roles of Ca(2+) mobilization due to nAChR stimulation remain to be clarified. We report here that spontaneous Ca(2+) oscillations can be observed in ∼15% of the pinealocytes in slice preparations from rat pineal glands when this dissociation procedure is done within 6 h from a dark-to-light change. The frequency and half-life of [Ca(2+)]i rise were 0.86 min(-1) and 19 s, respectively. Similar spontaneous Ca(2+) oscillations were recorded in 17% of rat pinealocytes that were primary cultured for several days. Simultaneous measurement of [Ca(2+)]i and membrane potential revealed that spontaneous Ca(2+) oscillations were triggered by periodic membrane depolarizations. Spontaneous Ca(2+) oscillations in cultured pinealocytes were abolished by extracellular Ca(2+) removal or application of nifedipine, a blocker of voltage-dependent Ca(2+) channel (VDCC). In contrast, blockers of intracellular Ca(2+)-release channels, 2-aminoethoxydiphenylborate and ryanodine, have no effect. Our results also reveal that, in 23% quiescent pinealocytes, Ca(2+) oscillations were observed following the withdrawal of nicotine. Norepinephrine-induced melatonin secretion from whole pineal glands was significantly decreased by the coapplication of acetylcholine (ACh). This inhibitory effect of ACh was attenuated by nifedipine. In conclusion, both spontaneous and evoked Ca(2+) oscillations are due to membrane depolarization following activation of VDCCs. This consists of VDCC α1F subunit, and the associated Ca(2+) influx can strongly regulate melatonin secretion in pineal glands.
松果腺通过合成和分泌褪黑素来调节昼夜节律。由于副交感神经活动引起的烟碱型乙酰胆碱受体 (nAChR) 刺激导致细胞内 Ca(2+) 浓度 ([Ca(2+)]i) 升高，从而下调褪黑素的产生。nAChR 刺激引起的 Ca(2+) 动员的重要特征和作用仍有待阐明。我们在这里报告，当在从黑暗到光照变化后 6 小时内进行这种分离程序时，可以在来自大鼠松果腺的切片制剂中观察到约 15%的松果体细胞中自发的 Ca(2+) 振荡。[Ca(2+)]i 上升的频率和半衰期分别为 0.86 min(-1)和 19 s。在培养数天的大鼠松果体细胞中也记录到了类似的自发 Ca(2+) 振荡。同时测量 [Ca(2+)]i 和膜电位表明，自发 Ca(2+) 振荡是由周期性的膜去极化触发的。在培养的松果体细胞中，通过去除细胞外 Ca(2+) 或应用硝苯地平（一种电压依赖性 Ca(2+) 通道 (VDCC) 阻断剂）消除自发的 Ca(2+) 振荡。相比之下，细胞内 Ca(2+) 释放通道的阻断剂 2-氨基乙氧基二苯硼酸盐和钌红没有作用。我们的结果还表明，在 23% 的静止松果体细胞中，在尼古丁撤出后观察到 Ca(2+) 振荡。在整个松果腺中，去甲肾上腺素诱导的褪黑素分泌由于乙酰胆碱 (ACh) 的共同应用而显著减少。ACh 的这种抑制作用被硝苯地平减弱。总之，自发和诱发的 Ca(2+) 振荡都是由于 VDCC 激活后膜去极化引起的。这包括 VDCC α1F 亚基，并且相关的 Ca(2+) 内流可以强烈调节松果腺中的褪黑素分泌。
