Al-Ubaidi M R, White T W, Ripps H, Poras I, Avner P, Gomès D, Bruzzone R
Lions of Illinois Eye Research Institute, Department of Ophthalmology and Visual Sciences, University of Illinois College of Medicine, Chicago, IL 6012, USA.
J Neurosci Res. 2000 Mar 15;59(6):813-26. doi: 10.1002/(SICI)1097-4547(20000315)59:6<813::AID-JNR14>3.0.CO;2-#.
Retinal neurons of virtually every type are coupled by gap-junctional channels whose pharmacological and gating properties have been studied extensively. We have begun to identify the molecular composition and functional properties of the connexins that form these 'electrical synapses,' and have cloned several that constitute a new subclass (gamma) of the connexin family expressed predominantly in retina and brain. In this paper, we present a series of experiments characterizing connexin36 (Cx36), a member of the gamma subclass that was cloned from a mouse retinal cDNA library. Cx36 has been localized to mouse chromosome 2, in a region syntenic to human chromosome 5, and immunocytochemistry showed strong labeling in the ganglion cell and inner nuclear layers of the mouse retina. Comparison of the developmental time course of Cx36 expression in mouse retina with the genesis of the various classes of retinal cells suggests that the expression of Cx36 occurs primarily after cellular differentiation is complete. Because photic stimulation can affect the gap-junctional coupling between retinal neurons, we determined whether lighting conditions might influence the steady state levels of Cx36 transcript in the mouse retina. Steady-state levels of Cx36 transcript were significantly higher in animals reared under typical cyclic-light conditions; exposure either to constant darkness or to continuous illumination reduced the steady-state level of mRNA approximately 40%. Injection of Cx36 cRNA into pairs of Xenopus oocytes induced intercellular conductances that were relatively insensitive to transjunctional voltage, a property shared with other members of the gamma subclass of connexins. Like skate Cx35, mouse Cx36 was unable to form heterotypic gap-junctional channels when paired with two other rodent connexins. In addition, mouse Cx36 failed to form voltage-activated hemichannels, whereas both skate and perch Cx35 displayed quinine-sensitive hemichannel activity. The conservation of intercellular channel gating contrasts with the failure of Cx36 to make hemichannels, suggesting that the voltage-gating mechanisms of hemichannels may be distinct from those of intact intercellular channels.
几乎每一种类型的视网膜神经元都通过间隙连接通道相互耦联，人们已对其药理学和门控特性进行了广泛研究。我们已开始确定形成这些“电突触”的连接蛋白的分子组成和功能特性，并克隆了几种连接蛋白，它们构成了连接蛋白家族中的一个新亚类（γ），主要在视网膜和大脑中表达。在本文中，我们展示了一系列表征连接蛋白36（Cx36）的实验，Cx36是从小鼠视网膜cDNA文库中克隆出来的γ亚类成员。Cx36已定位到小鼠的2号染色体上，该区域与人的5号染色体同线，免疫细胞化学显示在小鼠视网膜的神经节细胞层和内核层中有强烈标记。将小鼠视网膜中Cx36表达的发育时间进程与各类视网膜细胞的发生过程进行比较表明，Cx36的表达主要在细胞分化完成后出现。由于光刺激可影响视网膜神经元之间的间隙连接耦联，我们确定光照条件是否可能影响小鼠视网膜中Cx36转录本的稳态水平。在典型的循环光照条件下饲养的动物中，Cx36转录本的稳态水平显著更高；暴露于持续黑暗或持续光照下会使mRNA的稳态水平降低约40%。将Cx36的cRNA注射到成对的非洲爪蟾卵母细胞中可诱导细胞间电导，该电导对跨连接电压相对不敏感，这是连接蛋白γ亚类其他成员共有的特性。与鳐鱼的Cx35一样，小鼠的Cx36与另外两种啮齿动物连接蛋白配对时无法形成异型间隙连接通道。此外，小鼠的Cx36无法形成电压激活的半通道，而鳐鱼和鲈鱼的Cx35都表现出奎宁敏感的半通道活性。细胞间通道门控的保守性与Cx36无法形成半通道形成对比，这表明半通道的电压门控机制可能与完整的细胞间通道不同。
