DOPAMINE 1 RECEPTOR ACTIVATION MODULATES OPTOGENETIC ELICITED GLYCINERGIC BUT NOT GABAergic INHIBITION ON OFF BIPOLAR CELLS

Abstract

Purpose : Light evoked inhibition on OFF cone bipolar cells (BCs) has been shown to be modulated by Dopamine Receptor 1 (D1R) agonist SKF. It is not known where in the light signaling cascade dopamine is modulating inhibition on the OFF BCs. The goal of this study was to 1) determine what contributions GABA_AR, GABA_CR and GlycineR make on OFF BCs when optogeneticly activating the inhibitory neurons. 2) determine which of these contributions if any are modulated by agonizing D1R agonist SKF.

Methods : Retinas were isolated from 8-11 week old B6.Cg-Tg(Slc32al-COP4*H134R/EYFP) male and female mice that contain ChR2 under the VGAT promoter in the inhibitory amacrine and horizontal cells and cut into 250 µm thick slices. Whole-cell voltage clamp recordings of light-evoked Channel Rhodopsin 2 (ChR2) currents were measured on OFF BCs (n=19). CHR2 expressing cells were optogenetically activated by a 1s full-field light stimulus projected through a 4x or 60x objective (λ = 470 nm). Photoreceptor inputs were pharmacologically blocked with CNQX (25 µM), APV (50 µM), ACET (1 μM), and L-AP4 (50 µM). GABA_ARs, GABA_CRs and glycineR were antagonized using TPMPA (50 µM) and gabazine (20 µM) or strychnine (1 µM), respectively. D1Rs were agonized with SKF-38393 (20 µM). All experiments were performed under ambient room light. The peak amplitude, decay Tau, time to first peak, frequency and charge transfer (Q) were measured for all evoked responses and analyzed by paired t-tests.

Results : While measuring optogenetically evoked inhibition on OFF BCs, application of a D1R agonist did not significantly change total inhibition or GABAergic inhibition as compared to control, normalized peak amplitude (p= 0.17, 0.88), charge transfer (p=0.97,0.50), or decay tau (p= 0.39, 0.14) respectively (n=7). In contrast, glycinergic inhibition was significantly reduced (n=6) peak amplitude (p<0.01), charge transfer (p<0.01), and decay tau (p<0.05) after application of SKF-38393.

Conclusions : Direct activation of amacrine cells through optogenetics elicits GABA_AR, GABA_CR, and glycineR currents in OFF BCs. Dopamine dependent reductions in inhibition appear to be driven by a reduction in glycine currents and not GABA currents. D1R activation has similar effects on light-evoked OFF BC currents, suggesting that the D1R mechanisms are in the presynaptic amacrine cells.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.