Light microscopic localization of putative glycinergic neurons in the larval tiger salamander retina by immunocytochemical and autoradiographical methods

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Abstract
Putative glycinergic neurons in the larval tiger salamander retina were localized by a comparative analysis of high affinity 3H‐glycine uptake and glycine‐like immunoreactivity (Gly‐IR) at the light microscopic level. Commonly labeled neurons include at least three types of amacrine cell (Type IAd, Type IAb, Type IIAd; distinquished by soma location and dendritic ramification), cell bodies in the ganglion cell layer (GCL), and rarely observed Type II (inner) bipolar cells. With the increased resolution provided by Gly‐IR, we identified a Type IAa amacrine cell, two types of Type IIAd amacrine cells, and Gly‐IR interplexiform cells. Gly‐IR axons in longitudinal sections of the optic nerve indicate the presence of Gly‐IR ganglion cells. The percentage of labeled somas in the inner nuclear layer (INL) compared to all cells in each layer was similar for the two methods: 30‐40% in INL 2 (middle layer of somas), 30–40% in INL 3 (inner layer of somas), and about 5% in the GCL. Labeled processes were found throughout the full thickness of the inner plexiform layer (IPL), but with a much denser band in the proximal half (sublamina b). The only major difference between the two methods (3H‐glycine uptake vs. Gly‐IR) was that Type I (outer) bipolar cells were labeled only by 3H‐glycine uptake; these cells were more lightly labeled with silver grains than cell bodies in either INL 2 or INL 3. Postembed labeling of 1 μm Durcupan plastic sections for Gly‐IR showed the same pattern, but with much higher resolution, as obtained with 10 μm cryostat sections. This study indicates extensive colocalization of labeling by both probes in INL 2, INL 3, the IPL, and the GCL. We conclude that Gly‐IR can serve as a valid and reliable marker for glycine‐containing neurons in this retina and suggest that glycine serves as a transmitter for several morphologically distinct types of amacrine cell, an interplexiform cell, and perhaps a small percentage of Type II bipolar cells and ganglion cells.