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A. A. Hirano, C. W. Morgans, N. C. Brecha; Synaptotagmin-4 Expression in Mouse Horizontal Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):2798.
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© ARVO (1962-2015); The Authors (2016-present)
The mechanism underlying transmitter release from retinal horizontal cells is poorly understood. Using immunocytochemistry, we investigated the distribution of synaptotagmin-4, a calcium sensor in vesicle trafficking, in the mammalian retina.
Mouse retinas were fixed for 15-30 min with 4% paraformaldehyde. Cryostat vertical sections were processed for indirect immunofluorescence immunohistochemistry. Antibodies to synaptotagmin-4 (syt-4, Abnova, Taiwan) in combination with antibodies to calbindin, a horizontal cell marker, PKC for rod bipolar cells, VGLUT1 for ribbon synapses, and antibodies to the SNARE protein, SNAP-25, and alpha 1C, an L-type calcium channel, were used.
Syt-4 immunolabeling occurred primarily in the outer plexiform layer (OPL), but also in the deep inner plexiform layer (IPL) of mouse retina. Syt-4 co-localized with calbindin, labeling the horizontal cell (HC) projections that extend distally into photoreceptor terminals. In contrast, syt-4 immunolabeling did not co-localize with PKC-labeled rod bipolar cell dendrites. Furthermore, syt-4 immunoreactivity occurred within photoreceptor terminals, identified by VGLUT-1 immunoreactivity. In the IPL, the syt-4 immunoreactivity appeared distinct from the bipolar cell terminals, also labeled by VGLUT1 antibodies. Consistent with syt-4's role as a calcium sensor, its immunoreactivity co-localized with that of alpha 1C, an L-type calcium channel subunit, in the horizontal cell endings. Finally, syt-4 immunoreactivity localized to the processes and endings of horizontal cells, i.e., the same subcellular compartment, as that of the SNARE protein, SNAP-25.
Syt-4 is expressed in the processes and endings of horizontal cells within the OPL, where it may act as a calcium sensor in vesicle trafficking. The presence of synaptotagmin-4, in addition to SNARE proteins, SNAP-25 and other SNARE-related proteins, e.g. syntaxin-1a and -4 and complexin-I/II, known to occur in horizontal cell processes and tips, suggests that a vesicular mechanism underlies transmitter release from mammalian horizontal cells.
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