Heidelberger Ruth, Wang Meng M, Sherry David M
Department of Neurobiology and Anatomy and the W.M. Keck Center for the Neurobiology of Learning and Memory, University of Texas-Houston Medical School, Houston, USA.
Vis Neurosci. 2003 Jan-Feb;20(1):37-49. doi: 10.1017/s095252380320105x.
Synaptotagmin I is the leading candidate for the calcium sensor that triggers exocytosis at conventional synapses. However, physiological characterization of the calcium sensor for phasic release at the ribbon-style synapses of the goldfish Mb1 bipolar cell demonstrates a lower than predicted affinity for calcium, suggesting that a modified or different sensor triggers exocytosis at this synapse. We examined synaptotagmin immunolabeling in goldfish retina using two different antibodies directed against synaptotagmin epitopes that specifically labeled the expected 65-kDa protein on western blots of goldfish and mouse retinal membranes. The first antiserum strongly labeled conventional synapses in the inner plexiform layer (IPL), but did not label the ribbon-style synapse-containing synaptic terminals of goldfish Mb1 bipolar cells or photoreceptors. The second antibody also specifically labeled the expected 65-kDa protein on western blots but did not label any synapses in the goldfish retina. A third synaptotagmin antibody that performed poorly on western blots selectively labeled goldfish photoreceptor terminals. These results suggest that synaptotagmin may exist in at least three distinct "forms" in goldfish retinal synapses. These forms, which are differentially localized to conventional synapses, bipolar cell, and photoreceptor terminals, may represent differences in isoform, posttranslational modifications, epitope availability, and protein-binding partners. Labeling with these antibodies in the salamander and mouse retina revealed species-specific differences, indicating that synaptotagmin epitopes can vary across species as well as among synapses.
突触结合蛋白I是传统突触中触发胞吐作用的钙传感器的主要候选者。然而，对金鱼Mb1双极细胞带状突触处相位释放的钙传感器的生理学特征分析表明，其对钙的亲和力低于预期，这表明在该突触处可能是一种经过修饰或不同的传感器触发了胞吐作用。我们使用两种针对突触结合蛋白表位的不同抗体，在金鱼视网膜中检测了突触结合蛋白的免疫标记情况，这两种抗体在金鱼和小鼠视网膜膜的蛋白质印迹上能特异性标记预期的65 kDa蛋白。第一种抗血清强烈标记了内网状层（IPL）中的传统突触，但未标记金鱼Mb1双极细胞或光感受器中含有带状突触的突触终末。第二种抗体在蛋白质印迹上也特异性标记了预期的65 kDa蛋白，但未标记金鱼视网膜中的任何突触。第三种在蛋白质印迹上表现不佳的突触结合蛋白抗体选择性地标记了金鱼光感受器终末。这些结果表明，突触结合蛋白在金鱼视网膜突触中可能至少以三种不同的“形式”存在。这些形式在传统突触、双极细胞和光感受器终末中定位不同，可能代表了异构体、翻译后修饰、表位可用性和蛋白质结合伴侣方面的差异。在蝾螈和小鼠视网膜中用这些抗体进行标记显示出物种特异性差异，表明突触结合蛋白表位在不同物种以及不同突触之间可能存在差异。
