Translational Neuroscience Facility and Department of Physiology, School of Medical Sciences, The University of New South Wales, Sydney, New South Wales 2052, Australia.
J Neurosci. 2012 Aug 15;32(33):11414-23. doi: 10.1523/JNEUROSCI.6446-11.2012.
Canonical transient receptor potential (TRPC3) nonselective cation channels are effectors of G-protein-coupled receptors (GPCRs), activated via phospholipase C-diacylglycerol signaling. In cerebellar Purkinje cells, TRPC3 channels cause the metabotropic glutamate receptor (mGluR)-mediated slow EPSC (sEPSC). TRPC3 channels also provide negative feedback regulation of cytosolic Ca(2+), mediated by a C terminus "calmodulin and inositol trisphosphate receptor binding" (CIRB) domain. Here we report the alternative splicing of the TRPC3 mRNA transcript (designated TRPC3c), resulting in omission of exon 9 (approximately half of the CIRB domain) in mice, rats, and guinea pigs. TRPC3c expression is brain region specific, with prevalence in the cerebellum and brainstem. The TRPC3c channels expressed in HEK293 cells exhibit increased basal and GPCR-activated channel currents, and increased Ca(2+) fluorescence responses, compared with the previously characterized (TRPC3b) isoform when activated via either the endogenous M3 muscarinic acetylcholine receptor, or via coexpressed mGluR1. GPCR-induced TRPC3c channel opening rate (cell-attached patch) matched the maximum activation achieved with inside-out patches with zero cytosolic Ca(2+), whereas the GPCR-induced TRPC3b activation frequency was significantly less. Both TRPC3 channel isoforms were blocked with 2 mm Ca(2+), attributable to CIRB domain regulation. In addition, genistein blocked Purkinje cell (S)-2-amino-2-(3,5-dihydroxyphenyl) acetic acid (mGluR1)-activated TPRC3 current as for recombinant TRPC3c current. This novel TRPC3c ion channel therefore has enhanced efficacy as a neuronal GPCR-Ca(2+) signaling effector, and is associated with sensorimotor coordination, neuronal development, and brain injury.
经典瞬时受体电位 (TRPC3) 非选择性阳离子通道是 G 蛋白偶联受体 (GPCR) 的效应器，通过磷脂酶 C-二酰基甘油信号转导激活。在小脑浦肯野细胞中，TRPC3 通道引起代谢型谷氨酸受体 (mGluR) 介导的慢 EPSC (sEPSC)。TRPC3 通道还通过 C 末端“钙调蛋白和肌醇三磷酸受体结合”(CIRB) 结构域提供细胞溶质 Ca(2+) 的负反馈调节。在这里，我们报告了 TRPC3 mRNA 转录本的选择性剪接（命名为 TRPC3c），导致小鼠、大鼠和豚鼠中第 9 外显子缺失（约一半的 CIRB 结构域）。TRPC3c 表达具有脑区特异性，在小脑和脑干中普遍存在。与先前表征的 (TRPC3b) 同工型相比，在 HEK293 细胞中表达的 TRPC3c 通道表现出增加的基础和 GPCR 激活的通道电流，以及增加的 Ca(2+) 荧光反应，当通过内源性 M3 毒蕈碱乙酰胆碱受体或共表达的 mGluR1 激活时。GPCR 诱导的 TRPC3c 通道开放率（细胞附着式斑片）与零细胞溶质 Ca(2+) 下用内面向外斑片达到的最大激活相匹配，而 GPCR 诱导的 TRPC3b 激活频率明显较低。两种 TRPC3 通道同工型均被 2mm Ca(2+) 阻断，归因于 CIRB 结构域调节。此外，金雀异黄素阻断浦肯野细胞（S）-2-氨基-2-（3,5-二羟基苯基）乙酸（mGluR1）激活的 TPRC3 电流，与重组 TRPC3c 电流相同。因此，这种新型 TRPC3c 离子通道作为神经元 GPCR-Ca(2+) 信号效应物具有更高的功效，与感觉运动协调、神经元发育和脑损伤有关。
