Connexins and electrical synapses in the retina

视网膜中的连接蛋白和电突触

• 批准号: 6927113
• 负责人: DAVID L PAUL
• 金额: $ 25.43万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2008-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6927113
• 关键词: amacrine cells; cone cell; gap junctions; gene expression; genetically modified animals; horizontal cell; immunocytochemistry; in situ hybridization; laboratory mouse; neural transmission; polymerase chain reaction; retina; retinal bipolar neuron; retinal ganglion; rod cell; synapses

DESCRIPTION (provided by applicant): In the retina, gap junctions are established by many cell types and may participate in retinal circuitry in a variety of ways. Gap junctions are composed of connexins, encoded by a >20-member family of highly related genes. Only one of these, connexin36 (Cx36), has been conclusively demonstrated in retinal neurons. Using a Cx36 knockout (KO) mouse incorporating a histochemical reporter, we found Cx36 to be present in most or all All amacrine cells, many photoreceptors, subsets of ON and OFF cone bipolar cells and a low number of ganglion cells. However, horizontal cells and many ganglion cells establish gap junctions yet do not express Cx36. Using RT-PCR, we have found Cx45 and Cx57 to be prominently expressed in mouse retina. We propose to characterize the cellular distribution of these connexins and determine if other gap junction channel-forming proteins may be present in retinal neurons. Many lines of evidence indicate that rod photoreceptor signals utilize multiple pathways to reach ganglion cells. In our Cx36 KO, ON ganglion cell responses to scotopic stimuli are completely eliminated, indicating that Cx36 is required in all pathways contributing to rod ON signaling. We showed that gap junctional coupling between AII and cone ON bipolar cells, the presumed 'primary' pathway for rod photoreceptor signaling, was abolished in the KO. Since a proposed 'alternative' pathway involved gap junction between rods and cones, and since we found Cx36 expression in photoreceptors, our findings were consistent with the proposed 'alternative' pathway. However, recent studies of retinas from mice genetically altered to lack cones suggest that rod-cone coupling does not contribute to rod photoreceptor signaling. In addition, it has not been technically feasible to measure junctional coupling between photoreceptors in mouse retinas. Therefore, to resolve the discrepancy between current models of the 'alternative' pathway, we will 1) develop a method to directly assess rod-cone coupling in WT and Cx36 KO mice and 2) produce conditional knockouts that eliminate Cx36 specifically from either cones or from AII amacrine cells. Then, ganglion cell responses to light can be assessed in retinas where either primary or alternative pathways are ablated.

描述（由申请人提供）：在视网膜中，差距连接是由许多细胞类型建立的，可能会以多种方式参与视网膜电路。间隙连接由连接蛋白组成，由> 20人的高度相关基因家族编码。在视网膜神经元中最终证明了其中一个，即connexin36（CX36）。使用结合了组织化学记者的CX36敲除（KO）小鼠，我们发现CX36存在于大多数或所有无链氨酸细胞中，许多光感受器，On和Off锥双极细胞的子集以及较少的神经节细胞。然而，水平细胞和许多神经节细胞会建立间隙连接，但未表达CX36。使用RT-PCR，我们发现CX45和CX57在小鼠视网膜中突出表达。我们建议表征这些连接素的细胞分布，并确定视网膜神经元中是否可能存在其他间隙连接通道形成蛋白。 许多证据表明，杆感光器信号利用多个途径到达神经节细胞。在我们的CX36 KO中，完全消除了对神经节细胞对Scotopic刺激的反应，这表明在所有促成杆的信号传导的途径中都需要CX36。我们表明，在KO中废除了双极细胞上AII和锥之间的间隙连接耦合，即假定的杆光感受器信号传导的“主要”途径。由于提出的“替代”途径涉及杆和锥之间的间隙连接，并且由于我们在光感受器中发现CX36表达，因此我们的发现与所提出的“替代”途径一致。然而，对缺乏锥体的小鼠的视网膜的最新研究表明，杆孔耦合并不促进杆感光受体信号传导。此外，在小鼠视网膜中测量光感受器之间的连接耦合在技术上是不可行的。因此，为了解决“替代”途径的当前模型之间的差异，我们将开发一种直接评估WT和CX36 KO小鼠中的杆锥耦合的方法，以及2）产生有条件的敲除，以从锥形或AII amacrine细胞中取消CX36，以消除CX36。然后，可以在烧毁一级或替代途径的视网膜中评估神经节细胞对光的反应。