Nomura Sachi, Fukaya Masahiro, Tsujioka Takao, Wu Dianqing, Watanabe Masahiko
Department of Anatomy, Hokkaido University School of Medicine, Sapporo 060-8638, Japan.
Eur J Neurosci. 2007 Feb;25(3):659-72. doi: 10.1111/j.1460-9568.2007.05334.x. Epub 2007 Feb 12.
Phospholipase Cbeta3 (PLCbeta3) and PLCbeta4 are the two major isoforms in cerebellar Purkinje cells (PCs), displaying reciprocal expression across the cerebellum. Here, we examined subcellular distribution of PLCbeta3 in the mouse cerebellum by producing specific antibody. PLCbeta3 was detected as a particulate pattern of immunostaining in various PC elements. Like PLCbeta4, PLCbeta3 was richly distributed in somatodendritic compartments, where it was colocalized with molecules constituting the metabotropic glutamate receptor (mGluR1) signalling pathway, i.e. mGluR1alpha, G alpha q/G alpha 11 subunits of G q protein, inositol 1,4,5-trisphosphate receptor IP3R1, Homer1, protein kinase C PKCgamma, and diacylglycerol lipase DAGLalpha. Unlike PLCbeta4, PLCbeta3 was also distributed at low to moderate levels in PC axons, which were intense for IP3R1 and PKCgamma, low for G alpha q/G alpha 11, and negative for mGluR1alpha, Homer1, and DAGLalpha. By immunoelectron microscopy, PLCbeta3 was preferentially localized around the smooth endoplasmic reticulum in spines, dendrites, and axons of PCs, and also accumulated at the perisynapse of parallel fibre-PC synapses. Consistent with the ultrastructural localization, PLCbeta3 was biochemically enriched in the microsomal and postsynaptic density fractions. These results suggest that PLCbeta3 plays a major role in mediating mGluR1-dependent synaptic transmission, plasticity, and integration in PLCbeta3-dominant PCs, through eliciting Ca2+ release, protein phosphorylation, and endocannabinoid production at local somatodendritic compartments. Because PLCbeta3 can be activated by G betagamma subunits liberated from Gi/o and Gs proteins as well, axonal PLCbeta3 seems to modulate the conduction of action potentials through mediating local Ca2+ release and protein phosphorylation upon activation of a variety of G protein-coupled receptors other than mGluR1.
磷脂酶Cβ3（PLCβ3）和磷脂酶Cβ4是小脑浦肯野细胞（PC）中的两种主要亚型，在整个小脑中呈相互表达。在此，我们通过制备特异性抗体来检测小鼠小脑中PLCβ3的亚细胞分布。PLCβ3在各种PC元件中被检测为颗粒状免疫染色模式。与PLCβ4一样，PLCβ3大量分布于体树突区室，在那里它与构成代谢型谷氨酸受体（mGluR1）信号通路的分子共定位，即mGluR1α、Gq蛋白的Gαq/Gα11亚基、肌醇1,4,5-三磷酸受体IP3R1、Homer1、蛋白激酶C PKCγ和二酰甘油脂肪酶DAGLα。与PLCβ4不同，PLCβ3在PC轴突中也有低至中等水平的分布，轴突中IP3R1和PKCγ含量高，Gαq/Gα11含量低，而mGluR1α、Homer1和DAGLα为阴性。通过免疫电子显微镜观察，PLCβ3优先定位于PC的棘突、树突和轴突中的光滑内质网周围，并且也聚集在平行纤维-PC突触的突触周围。与超微结构定位一致，PLCβ3在微粒体和突触后密度组分中生化富集。这些结果表明，PLCβ3在介导mGluR1依赖性突触传递、可塑性和PLCβ3主导的PC中的整合中起主要作用，通过在局部体树突区室引发Ca2+释放、蛋白磷酸化和内源性大麻素产生。由于PLCβ3也可被从Gi/o和Gs蛋白释放的Gβγ亚基激活，轴突PLCβ3似乎通过在激活除mGluR1之外的多种G蛋白偶联受体时介导局部Ca2+释放和蛋白磷酸化来调节动作电位的传导。
