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Maria Soledad Soledad Velasco Dalesio, Ian Thompson; Characterization of SMI-32 positive RGC in the mouse retina. Invest. Ophthalmol. Vis. Sci. 2018;59(9):5496.
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© ARVO (1962-2015); The Authors (2016-present)
Our understanding of how different classes of retinal ganglion cells (RGC) innervate their nuclear targets in the mouse brain has been hindered by the scarcity of molecular markers available known to label specific types of RGC. In this experimental research, we used the antibody SMI-32 in the mouse retina and the dorsal lateral geniculate nucleus (dLGN) to evaluate if it effectively labels one subclass of RGC as suggested by Lin et al (2004) and Huberman et al (2008), or more than one subclass as reported by Coombs et al (2006). It was hypothesized that if the first case was true, these cells would be distributed non-randomly across the retina forming a regular mosaic and its reticulogeniculate terminations would differ between wild-type and β2-/- mice.
The animals studied were wild type and nAChR-β2-/- mutant mice. The retinas were extracted, fixed and whole mounted. The brains were fixed and embedded in gelatine to obtain coronal sections. The retinas were incubated in primary antibody (1:5,000 mouse SMI-32 antibody) for 48 hr and then in secondary antibody (1:1,000 goat anti-mouse) for 2 hrs. The protocol used to stain brain tissue was similar but with double antibody concentration to optimise penetration and staining. An upright microscope with camara lucida and confocal microscope were used for photographing the retinas and brain sections. Population density profile of SMI-32 positive cells was calculated by analysing one wild-type retina.
Huberman et al (2008) suggested that SMI-32-labelled RGCs formed a specific retinotopic mosaic of transient OFF-αRGC, following a minimum distance rule of about 100 µm. However, nearest neighbour analysis revealed that these cells do not form a regular mosaic (fig.1). In the dLNG, anatomical and physiological studies have suggested that ON/OFF-centre cell projections are significantly more organized in the β2-/- than in wild type dLGN. However, there was no difference in the overall architecture of labelled terminals that could be observed (fig.2).
In view that in our experiment SMI-32 positive RGC did not form a regular mosaic in the retina and their terminals formed a similar architecture in wild-type and β2 -/- dLGN, we concluded SMI-32 labels more than one subclass of RGC.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
Density profile of SMI-32 positive RGCs a function of distance from neighbouring equals.
Comparison between wild-type (A) and β2 -/- (B) dLGN brain sections (d=dorsal; l=lateral).
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