Immunocytochemical evidence for SNARE protein‐dependent transmitter release from guinea pig horizontal cells

Abstract

Horizontal cells are lateral interneurons that participate in visual processing in the outer retina but the cellular mechanisms underlying transmitter release from these cells are not fully understood. In non-mammalian horizontal cells, GABA release has been shown to occur by a non-vesicular mechanism. However, recent evidence in mammalian horizontal cells favors a vesicular mechanism as they lack plasmalemmal GABA transporters and some soluble NSF attachment protein receptor (SNARE) core proteins have been identified in rodent horizontal cells. Moreover, immunoreactivity for GABA and the molecular machinery to synthesize GABA have been found in guinea pig horizontal cells, suggesting that if components of the SNARE complex are expressed they could contribute to the vesicular release of GABA. In this study we investigated whether these vesicular and synaptic proteins are expressed by guinea pig horizontal cells using immunohistochemistry with well-characterized antibodies to evaluate their cellular distribution. Components of synaptic vesicles including vesicular GABA transporter, synapsin I and synaptic vesicle protein 2A were localized to horizontal cell processes and endings, along with the SNARE core complex proteins, syntaxin-1a, syntaxin-4 and synaptosomal-associated protein 25 (SNAP-25). Complexin I/II, a cytosolic protein that stabilizes the activated SNARE fusion core, strongly immunostained horizontal cell soma and processes. In addition, the vesicular Ca2+-sensor, synaptotagmin-2, which is essential for Ca2+-mediated vesicular release, was also localized to horizontal cell processes and somata. These morphological findings from guinea pig horizontal cells suggest that mammalian horizontal cells have the capacity to utilize a regulated Ca2+-dependent vesicular pathway to release neurotransmitter, and that this mechanism may be shared among many mammalian species.

Cytochrome P450 17α-hydroxylase/C(17,20)-lyase immunoreactivity and molecular expression in the cerebellar nuclei of adult male rats

“…5C). The labelling patterns of the antibodies detected by double immunofluorescence procedures were identical to those observed by other authors (Herzog et al, 2006;Zhou et al, 2007;Grønborg et al, 2010;Lee and Brecha, 2010).…”

Specialized synaptic pathway for chromatic signals beneath S-cone photoreceptors is common to human, Old and New World primates

“…Specificity of this antibody has been shown by Western blots of mouse retina and brain, where it recognized a single band at the appropriate molecular weight [4]. Immunostaining with syntaxin-4 shows the same staining pattern in the outer plexiform layer of guinea pig [5] mouse, rat, rabbit [3, 4] macaque, and ground squirrel ([1, 2] and this study). Preadsorption of the antibody with the corresponding control peptide blocked all staining in rabbit retina [3].…”

“…Syntaxin-4 was used as an indicator for vesicular GABA release via horizontal cells [5]. Therefore, our data points to enhanced GABA release via HII horizontal cells beneath S-cones.…”

“…The question addressed was if primate horizontal cells may employ a feed-forward pathway from horizontal cells directly to cone bipolar cells. Syntaxin-4 has been shown to be expressed by horizontal cells in different mammals and it was used as an indicator for vesicular GABA release in horizontal cells [3–5]. In macaque monkeys, syntaxin-4 was found in HI and HII horizontal cells postsynaptic to all photoreceptor types, but highly enriched beneath short (S-) wavelength sensitive cones, co-localized with dendritic processes of HII horizontal cells.…”

“…It is known that mammalian horizontal cells express the neurotransmitter gamma-aminobutyric acid (GABA), the GABA-synthesizing enzyme glutamic acid decarboxylase (GAD), and the vesicular GABA-transporter vGAT (e.g. [5, 21–32]. Ultrastructural studies revealed synaptic contacts between horizontal and bipolar cell dendrites in cat and rabbit [33–35] and GABA receptors have been shown to be expressed on bipolar cell dendrites at the corresponding sites in the outer plexiform layer (OPL) [25, 36].…”