Localizing the Distribution of the Adenosine A2A Receptor in the Mouse Retina Using a Cre/loxP System

Objective: To localize the distribution of the adenosine A2A receptor (A2AR) in the mouse retina using a Cre/loxP system. Methods: In this experimental study, male A2A-cre mice [B6.FVB (Cg)-Tg (Adora2a-cre)KG139Gsat/Mmucd] were mated with female Ai9 mice [B6.129S6-Gt (ROSA) 26Sortm9 (CAG-tdTomato)Hze/J]. The genotypes of the newborn mice were identified and five A2A-cre+, Ai9 mice were chosen at 4 weeks of age. The Cre enzyme of the mice can knock out the STOP sequence of Ai9 mice, which will cause the expression of the tomato fluorescent protein. The localization of A2AR in the retina can be determined through the double immunostaining of the tomato red fluorescent protein and markers of different retinal cells. Results: Immunofluorescence microscopy showed that Tomato+ cells were mainly distributed in the ganglion cell layer and inner nuclear layer, and not in the outer nuclear layer. In the ganglion cell layer, the minority of Tomato+ cells were ganglion cells while the majority were ectopic amacrine cells. In the inner nuclear layer, Tomato+ cells were mainly Muller and amacrine cells. With further analysis of the amacrine cell subtypes, we discovered that most of the Tomato+ cells were AⅡ amacrine cells, not cholinergic or dopaminergic amacrine cells. Conclusions: This study suggests that A2AR is expressed in ganglion cells, amacrine cells, and Muller cells in 4-week-old mouse retinas. Key words: adenosine A2A receptor; retina; Cre/loxP system; mice