Zimov Sarah, Yazulla Stephen
Department of Neurobiology and Behavior, Stony Brook University, Stony Brook, NY 11794-5230, USA.
J Neurocytol. 2004 Jul;33(4):441-52. doi: 10.1023/B:NEUR.0000046574.72380.e8.
The vanilloid receptor type 1 (TRPV1/VR1) is a non-specific calcium-permeable ionotropic cation channel expressed in the peripheral sensory system as well as in the central nervous system. An endogenous ligand for TRPV1 is arachidonoyl ethanolamide (anandamide), which also activates the metabotropic cannabinoid receptor 1 (CB1). Previous studies in this laboratory reported CB1 receptors and CB1-mediated effects on voltage-gated currents in goldfish cones and bipolar cells. In this study, we show TRPV1-like-immunoreactivity (TRPV1-L-IR) by immunoblot analysis of goldfish retina and rat brain homogenates with a guinea pig polyclonal antibody against the C-terminus of the rat TRPV1. Light-level immunocytochemistry showed restriction of the guinea pig-TRPV1 antibody to a very narrow band in the outer plexiform layer in goldfish and zebrafish retinas. However, no immunoreactivity was detected using rabbit-polyclonal antibodies against the C or N-termini of the rat TRPV1. Pre and post-embedding electron microscopy (EM) immunocytochemistry revealed that TRPV1-L-IR (using the guinea pig antibody) was restricted to synaptic ribbons of all cones and many rods, but never was observed at the synaptic ribbons of bipolar cells. While pre-embedded tissue showed diffuse label associated only with photoreceptor-synaptic ribbons, analysis of post-embedded tissue showed label tightly restricted to synaptic ribbons of all cones and many rods. Oblique sections showed that immunogold particles were confined to the outer electron dense region of the ribbons, with few or no gold particles in the ribbon core or associated with tethers or vesicles. Although TRPV1-L-IR described here, does not necessarily represent TRPV1 antigen associated with synaptic ribbons, these data provide an unequivocal label with which to study the functional dynamics of ribbon formation and degradation in teleost photoreceptors.
1型香草酸受体（TRPV1/VR1）是一种非特异性的钙通透性离子型阳离子通道，在外周感觉系统以及中枢神经系统中均有表达。TRPV1的内源性配体是花生四烯酸乙醇酰胺（阿南达米德），它也能激活代谢型大麻素受体1（CB1）。本实验室之前的研究报道了CB1受体以及CB1对金鱼视锥细胞和双极细胞电压门控电流的影响。在本研究中，我们用豚鼠抗大鼠TRPV1 C端的多克隆抗体对金鱼视网膜和大鼠脑匀浆进行免疫印迹分析，显示出TRPV1样免疫反应性（TRPV1-L-IR）。光镜水平的免疫细胞化学显示，豚鼠TRPV1抗体在金鱼和斑马鱼视网膜的外网状层中局限于一条非常窄的带。然而，用抗大鼠TRPV1 C端或N端的兔多克隆抗体未检测到免疫反应性。包埋前和包埋后电子显微镜（EM）免疫细胞化学显示，TRPV1-L-IR（使用豚鼠抗体）局限于所有视锥细胞和许多视杆细胞的突触带，但在双极细胞的突触带中从未观察到。虽然包埋前的组织显示弥漫性标记仅与光感受器突触带相关，但包埋后组织的分析显示标记紧密局限于所有视锥细胞和许多视杆细胞的突触带。斜切片显示免疫金颗粒局限于突触带的外电子致密区，在突触带核心或与系绳或囊泡相关的区域几乎没有或没有金颗粒。虽然这里描述的TRPV
