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Bozena Fyk-Kolodziej, Paul D Walker, Tomomi Ichinose; Cell type-specific distribution of dopamine D1a receptors in retina revealed in the Drd1a-tdTomato BAC transgenic mouse.. Invest. Ophthalmol. Vis. Sci. 2014;55(13):2376.
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© ARVO (1962-2015); The Authors (2016-present)
Light-induced dopamine release by amacrine cells regulates transition from rod to cone driven signal flow in the retina. Dopamine acts at different levels through modulation of photoreceptor light responses, voltage-gated channels, and gap junctions. Although dopamine D1 receptors (D1Rs) are involved, it has been difficult to localize D1Rs to specific subtypes of neurons due to inadequate whole-cell staining with specific D1R antibodies. We have overcome this limitation using the BAC transgenic mouse, which shows strong tdTomato fluorescence in whole DR1a receptor-expressing neurons.
Drd1a-tdTomato BAC transgenic mice were used which express tdTomato under the control of DR1a promoter. Specific types of neurons were labeled by markers. Bipolar cells: type1 - NK3R; type 2 and 6 - Syt 2b; type 3 - HCN4; type 4 - Csen; type 5 - HCN1; RBC - PKC α. Type 9 was revealed by contact with genuine S-opsin cone. Horizontal cells: Calbindin. In addition, individual neurons were injected with neurobiotin for subsequent staining with Alexa conjugated streptavidin. tdTomato expression was enhanced by Dsred antibody.
Strong tdTomato fluorescence was observed in entire cells, allowing for morphological subtype identification. The number of cells positive for DR1a within the INL and the GCL was distributed uniformly throughout the retina. Horizontal cells were positively labeled for DR1a. Many, but not all subtypes of bipolar cells (BCs) were positive for DR1a. For OFF BCs, DR1a was positive in subtypes -1 and -4, but was negative in subtypes -2 and -3. In ON BCs, subtypes -5 (partly), -6, and -9 BCs were DR1a positive. Interestingly, terminals of DR1a expressing type-5 BCs were wide-field, located adjacent to the ON ChAT band, and were positive for HCN1. Another subset of subtype -5 with umbrella like terminals were negative for DR1a. Rod BCs did not express DR1a; however, strong fluorescence was seen in close proximity to axon terminals, suggesting the presence of DR1a in AII or A17 amacrine cell (AC). Because A17 ACs were negative for Tomato fluorescence, the AII ACs probably express DR1a.
We have localized DR1a to many subtypes of cone BCs. Since detailed dopaminergic effects on BCs mediated by DR1a have not been studied, these results could lay the groundwork for further physiological experiments to elucidate dopamine action in cone circuitry.
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