胆碱能受体激动剂引起豚鼠离体前庭毛细胞内钙离子浓度升高

摘要

Abstract:Objective To better understand the cholinergic receptors in vestibular hair cells (VHC) and their subtypes, and to investigate the effects of cholinergic agonists on intracellular calcium concentration ([Ca2+]i ) in guinea pig VHCs.Methods VHCs were isolated from guinea pig crista ampullaris by enzymatic and mechanical methods. The effect of cholinergic agonists on [Ca2+]i was examined using laser scanning confocal microscopy and the Ca2+ sensitive dye Fluo-3.Results The results showed that the addition of acetylcholine (ACh) and carbachol (CCh), muscarnic and nicotinic agonists, induced [Ca2+]i increases in all the VHCs, whereas acetylcholine bromide (ACh-Br), a nicotinic agonist, induced the [Ca2+]i increase in only a small percentage of VHCs. The ACh or CCh-induced Ca2+ response could be partially suppressed by atropine. In the presence of 0.1?mmol/L atropine, the amplitudes of ACh or CCh-induced [Ca2+]i responses became significantly smaller than those in atropine free medium (P<0.01). Conclusions The results suggest the existence of cholinergic receptors in guinea pig VHCs. It is the muscarnic agonists rather than nicontic receptors that dominate [Ca2+]i variation. Atropine can suppress muscarnic agonist-induced Ca2+ responses.%目的通过观察胆碱能受体激动剂对离体前庭毛细胞内钙离子浓度的影响,以探讨前庭毛细胞膜所存在的胆碱能受体及其分型.方法用胶原酶消化后机械分离法,分离豚鼠前庭毛细胞(VHC), 钙敏荧光探针Fluo-3染色,用激光扫描共聚焦显微镜记录VHC的钙荧光图象及细胞内游离钙离子浓度([Ca2+]i)的动态变化.结果胆碱能M和N型受体激动剂乙酰胆碱(ACh)、氨甲酰胆碱(CCh)均可引起离体VHC内[Ca2+]i的升高;N型受体激动剂溴化乙酰胆碱(ACh-Br)仅在高浓度(10?mmol/L)时引起部分(4/5个)离体VHC内[Ca2+]i升高; 阿托品对ACh、CCh引起的VHC内[Ca2+]i的升高有抑制作用;加入0.1?mmol/L阿托品可使1?mmol/L Ach或CCh引起的VHC内[Ca2+]i升高的峰值明显减小(t检验均有显著差异P<0.01).结论豚鼠壶腹嵴前庭毛细胞膜上存在M型和N型两种受体,M型受体激动剂引起VHC内[Ca2+]i的升高较N型明显.阿托品对M型受体激动剂引起的[Ca2+]i升高有抑制作用.