演題番号 : P1-a29
清田 恵子 / Keiko Kiyota:1 村松 真 / Makoto Muramatsu:1 山村 寿男 / Hisao Yamamura:1 水谷 浩也 / Hiroya Mizutani:1 大矢 進 / Susumu Ohya:1 今泉 祐治 / Yuji Imaizumi:1
1:名古屋市立大・院薬・細胞分子薬効解析学 / Dept Mol & Cell Pharmacol, Grad Sch Pharmaceut Sci, Nagoya City Univ, Nagoya
Pineal gland regulates circadian rhythm through synthesis and secretion of melatonin. Although melatonin production may be closely associated with cytosolic Ca2+ signals, the regulation of Ca2+ dynamics coupled to melatonin production/secretion, particularly by parasympathetic innervation has not been fully understood. Nicotinic acetylcholine receptor (nAChR) activation is supposed to reduce synthesis and secretion of melatonin via auto/paracrine of glutamate released from pinealocytes. Regulatory mechanisms of the cytoplasmic Ca2+ signaling and melatonin secretion during stimulation of nAChR were examined in rat pinealocytes. We found spontaneous Ca2+ oscillations in 17 % of isolated rat pinealocytes. The application of 1 mM L-glutamate significantly increased the frequency of spontaneous Ca2+ oscillations, but did not affect the amplitude. Moreover, in 20 % of quiescent pinealocytes, Ca2+ oscillations were often evoked after a transient Ca2+ rise in the presence of 1 mM L-glutamate. The glutamate-induced [Ca2+]i elevation was significantly inhibited by 1 μM nifedipine or by 100 μM DL-threo-β-benzyloxyaspartate (TBOA), an inhibitor of excitatory amino acid transporters (EAATs), but not by blockers of ionotropic glutamate receptors, 10 μM 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) or 50 μM DL-2-amino-5-phosphonopentanoic acid (AP-5). The application of 1 mM L-aspartate, which is also transported with EAATs, substantially enhanced spontaneous Ca2+ oscillations. Melatonin secretion from whole pineal gland by 1 μM norepinephrine (NE) was significantly decreased by the co-presence of 100 μM ACh. The inhibitory effect of ACh was attenuated by 1 μM nifedipine. The NE-induced melatonin release was also blocked by 1 mM L-glutamate and the response was partly reversed by 100 μM TBOA. In conclusion, Ca2+ mobilization induced by glutamate, presumably as Ca2+ oscillations at least in part, may be involved in the inhibitory regulation of melatonin secretion in pinealocytes.