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Irene E. Whitney, Mary Raven, Benjamin E. Reese; Genetic Modulation of the Ratio of Cholinergic Amacrine Cells in the GCL and INL of the Mouse Retina. Invest. Ophthalmol. Vis. Sci. 2012;53(14):6327.
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© ARVO (1962-2015); The Authors (2016-present)
Cholinergic amacrine cells are positioned within the ganglion cell layer (GCL) and inner nuclear layer (INL) of the mature retina. Initially, however, these cells form a single stratum that separates into the two populations as development proceeds. We are seeking to identify the potential mechanisms that control the positioning of these cells.
Immunofluorescence was used to identify cholinergic amacrine cells in the GCL and INL of two inbred laboratory strains, C57BL/6J (B6/J) and A/J, and 25 recombinant inbred (RI) strains derived from them. Total estimates of cells in the GCL and INL were generated for a minimum of three mice per strain. Interval mapping was performed at GeneNetwork.org to identify quantitative trait loci (QTLs), followed by composite interval mapping to identify the presence of any secondary QTLs. The intervals underlying QTLs were assessed to identify possible causal genes based on multiple criteria including: retinal expression, regulatory or coding variants, and known function.
The A/J strain has approximately 12,400 cholinergic amacrine cells in the GCL and 14,500 in the INL. The B6/J strain has 34% more cells in both layers, with about 16,700 and 19,500 in the GCL and INL, respectively. This difference spans much of the range seen across the RI strains, although four of them had numbers exceeding those found in B6/J. Interestingly, the ratio of cells in the GCL to those in the INL is nearly identical between the parental strains, being 0.86 for A/J and 0.85 for B6/J. These ratios fall in the middle of the range found across the RI strains, which spans from 0.76 to 0.95. We failed to map any QTLs of interest for GCL number, INL number, or total number. However, the GCL:INL ratio mapped a QTL to the proximal end of chromosome (Chr) 3, with a likelihood ratio score (LRS) of 15.7. At this locus, the presence of two A alleles is associated with an additive effect of 0.07 for the trait. Composite interval mapping confirmed the presence of a secondary QTL on Chr 12, with an LRS of about 12.5, and an additive effect for two A alleles of 0.04. Taken together, these two QTLs account for 55% of the variance in ratio across the RI strains. 15 genes at the Chr 3 locus are expressed in the retina and have possible causal variants between B6/J and A/J. For the Chr 12 locus, there are three genes that meet these two criteria.
The GCL:INL ratio of cholinergic amacrine cells is modulated independently from the determination of their total numbers. Polymorphic genes at two genomic loci contribute roughly half of this trait variation. Identifying promising candidate genes may provide insight into the mechanisms controlling the positioning of these cells in one layer or the other during development.
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