Mechanism underlying polarized subcellular distribution of Ca channels and its neurobiological significance.

两个机制连接突触前缺陷突触后方面的神经元分化和成熟已经揭示。首先，在n型Ca^<2+>通道缺陷小鼠中，自动神经系统的阳性调节被消除。考虑到野生型小鼠突触前末端的去甲肾上腺素释放主要由n型通道控制，这是合理的。然而，有趣的是，心脏收缩、血管收缩和受体对激动剂的敏感性在突变小鼠中上调。其次，我们比较了Ca^<2+>通道亚型在携带P/ q型Ca^<2+>通道α_<1A> (Cav2.l)亚基基因突变的野生型对照、摇摇（tg）和滚动名古屋（tg^<rol>）中小脑兴奋性突触传递的基本特性和Ca^<2+>通道亚型的作用。攀爬纤维-浦肯野细胞（CF-PC）突触的EPSC振幅在tg中保持不变，甚至在tg^< 1 / 3 - 1 / 3中增加，这与突触后谷氨酸受体的特性改变有关。在突变小鼠中，攀爬纤维介导的EPSC更依赖于其他Ca^<2+>通道亚型，这表明这种代偿机制有助于维持CF-PC突触传递几乎完整。因此，突触前Ca^<2+>通道控制突触后兴奋性和功能，从而调节突触的分化和成熟。我们发现G蛋白偶联、代谢性谷氨酸受体亚型1 （mGluR1）和P/ q型Cav2.1共定位于小脑浦肯野神经元的树突，并在脑和异源表达的COS-7细胞中形成异质组装。mGluR1分别以不依赖配体的方式抑制含有辅助α 2/ δ亚基和β 1亚基的表达Cav2.1的HEK 293细胞中Cav2.1介导的[Ca^<2+>]i升高和Ba2+电流，并通过mGluR1的预激活以依赖配体的方式增强。此外，在二氢吡啶（DHP）敏感的l型Ca^<2+>通道Cav1.2 （α1c）中，Ser1142是DHP激动剂BAYk8644和被okadaic酸抑制的磷酸酶的识别位点。这表明DHP激动剂和丝氨酸磷酸化对l型通道的上调是通过涉及丝氨酸1142的共同机制介导的。最后，我们利用β4亚基进行双杂交筛选，分离出一个编码突触囊泡融合调节蛋白的克隆。蛋白质的功能和分子特性正在进行中。少

Two mechanisms that link presynaptic defects to postsynaptic aspects of neuronal differentiation and maturation have been revealed. Firstly, in N-type, Ca^<2+> channel-deficient mice we created, positive, regulation by automic nervous system was ablated. This is reasonable, considering that noradrenalin release from presynaptic terminal is predominantly controlled by N-type channels in wild-type mice. However, interestingly, cardiac contraction, vascular constriction, and sensitivity of receptors to their agonists were upregulated in the mutant mice. Secondly, we compared fundamental properties of excitatory synaptic transmission in the cerebellum and roles of Ca^<2+> channel subtypes among wild-type control, tottering (tg) and rolling Nagoya (tg^<rol>) that carry mutations in the P/Q-type Ca^<2+> channel α_<1A> (Cav2.l) subunit gene. The EPSC amplitude of the climbing fiber-Purkinje cell (CF-PC) synapses was preserved in tg, and it was even increased in tg^<rol>, which was associated … More with altered properties of the postsynaptic glutamate receptors. The climbing fiber mediated EPSC was more dependent on other Ca^<2+> channel subtypes in mutant mice, suggesting that such compensatory mechanisms contribute to maintaining the CF-PC synaptic transmission virtually intact. Thus, presynaptic Ca^<2+> channels control postsynaptic excitability and function and thereby regulate differentiation and maturation synapses. We showed that G protein-coupled, metabotropic glutamate receptor subtype 1 (mGluR1) and P/Q-type Cav2.1 are colocalized at dendrites of cerebellar Purkinje neurons and form the heteromeric assembly in both the brain and heterologously expressing COS-7 cells. mGluR1 inhibited Cav2.1-mediated [Ca^<2+>]i increases and Ba2+ currents in HEK 293 cells expressing Cav2.1 with auxiliary alpha2/delta and beta1 subunits, respectively, in a ligand-independent manner and was enhanced by pre activation of mGluR1 in a ligand-dependent manner. Furthermore, in dihdropyridine (DHP)-sensitive L-type Ca^<2+> channel Cav1.2 (α1c), Ser1142 was the recognition site for both the DHP agonist BAYk8644 and Phosphatase inhibited by okadaic acid. This suggests that upregulation of L-type channels by DHP agonist and that by Ser phophorylation are mediated by a common mechanism involving Ser1142. Finally, we carried out two-hybrid screening using the β4 subunit as a bate to isolate a clone encoding the protein regulating fusion of synaptic vesicles. Functional and molecular characterization of the protein is under way. Less