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C. E. Strang, K. T. Keyser; Subcellular Distribution of Homomeric and Heteromeric Nicotinic Receptors in Rabbit Retina. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1223. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To compare the expression of α7 and α3ß2 nicotinic acetylcholine receptors (nAChRs) on dendritic processes in the inner plexiform layer (IPL) of retina.
Retinas isolated from adult rabbits were sectioned into 200 µm slices and incubated with alexa 594-conjugated α-bungarotoxin (αBGT) in Ames medium. The retinas were then fixed and processed for immunohistochemistry using antibodies against α7 nAChRs (SC-1447, Santa Cruz), α3ß2 nAChRs (mAB210, the generous gift of Dr. Jon Lindstrom) and/or ChAT (Chemicon) in a whole mount protocol. Confocal images of labeled retinas were obtained and processed using Leica software. Brightness and contrast were adjusted using Adobe Photoshop. In some cases cytofluorochrome masking was used to provide a semi-quantitative assessment of colocalization.
Alexa 594-conjugated α-BGT labeled processes were distributed throughout the IPL. This labeling was decreased by pre-incubation of the retina in unlabeled αBGT, and corresponded with immunoreactivity to SC-1447, an antibody against α7 nAChRs. The dendritic processes of cells expressing α3ß2 nAChRs were also distributed throughout the IPL. The α3ß2-bearing processes were closely apposed to, and displayed some receptor colocalization with, αBGT-labeled processes.
The incomplete colocalization of homomeric and heteromeric nAChRs suggests that only subsets of cells express more than one nAChR subtype. However, the similar patterns of distribution together with the apparent proximity of the different receptor subtypes on some dendrites suggests that the effects of ACh may be mediated in a cooperative fashion.
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