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A.A. Hirano, J.H. Brandstätter, N.C. Brecha; Cellular Distribution and Subcellular Localization of Molecular Components of Vesicular Transmitter Release In Horizontal Cells of Rabbit Retina . Invest. Ophthalmol. Vis. Sci. 2005;46(13):4535.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: The mechanism underlying transmitter release from retinal horizontal cells is poorly understood. Using immunocytochemistry, we investigated the possibility of vesicular transmitter release from mammalian horizontal cells by examining the expression of synaptic proteins, complexin I/II (CPX I/II), syntaxin 1, and synapsin I, which participate in vesicular transmitter release at chemical synapses in rabbit retina. Methods: Rabbit retinas were fixed for 15–30 min with 4% paraformaldehyde (PFA) or with 2% PLP. Cryostat vertical sections were processed for indirect immunofluorescence immunohistochemistry. Antibodies used include well–characterized antibodies to CPX I/II, syntaxin–1/HPC–1 and synapsin I (P610) in combination with antibodies to calbindin, a horizontal cell marker, GABA and glycine, markers for amacrine cells, and VGLUT1 for ribbon synapses. Confocal microscopy was used to assess co–localization. Subcellular localization of CPX I/II, syntaxin–1 and synapsin I was determined using pre–embedding immunoelectron microscopy, on retina fixed for 50 min in 4% PFA. Results: Strong labeling for CPX I/II was found in both synaptic layers and the nerve fiber layer of the retina. Co–localization with calbindin indicated that CPX I/II occurs in somata, processes and endings of both A– and B–type horizontal cells. Further, subpopulations of GABA– and glycinergic amacrine cells and ganglion cells expressed CPX I/II. Immunoelectron microscopy demonstrated the presence of CPX I/II in horizontal cell processes postsynaptic to rod and cone ribbon synapses. Syntaxin–1 and synapsin I were also present in the horizontal cells and their processes at photoreceptor synapses. Photoreceptors and bipolar cells did not express any of these proteins at their axon terminals. Conclusions: The presence of CPX I/II, syntaxin–1, a binding partner of complexins, and synapsin I, a protein involved in the calcium–dependent recruitment of synaptic vesicles, in rabbit horizontal cell processes and tips suggests that a vesicular mechanism may underlie transmitter release from mammalian horizontal cells.
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