CHARACTERIZATION OF VOLTAGE-GATED CALCIUM CURRENTS IN ROD BIPOLAR CELLS OF MUTANT CAV1.X MOUSE MODELS

Cav1.4 voltage-gated calcium channels (VGCCs) are crucial for retinal synaptic transmission and human pathogenic Cav1.4 variants can cause congenital stationary night blindness type-2 (CSNB2). Heterologous systems revealed distinct CSNB2 phenotypes: the truncating R1827X-variant induced calcium-dependent inactivation, but the I745T-variant displayed gain-of-function-characteristics. To determine whether these functional phenotypes also manifest in retinal neurons, we selected rod bipolar cells (RBCs) as our model and first characterized VGCCs in wild-type (WT) RBCs, given that the VGCC composition in RBCs is not yet fully established. qPCR-experiments in FACS-sorted RBCs showed transcripts for β2, α2δ-4, Cav1.3, Cav1.4, Cav3.2 and a Cav1.1 splice variant. Whole-cell patch-clamp recordings in WT-RBCs revealed a sustained high-voltage activated (HVA) L-type and a transient, low-voltage activated T-type calcium current. Despite substantial Cav1.4 transcript being present, a HVA calcium current persisted in Cav1.4KO RBCs, which remained sensitive to the L-type modulators Isradipine and Bay-K. However, analyses of additional mutant Cav1.x mouse models revealed only a minor Cav1.3 and no Cav1.2 contribution. Despite this compensatory HVA current, Cav1.4KO-RBCs exhibited a substantial reduction in overall calcium currents, highlighting the critical role of Cav1.4 channels in RBCs. Thus, our findings enable the functional characterization of its pathogenic variants within these cells. Notably, the pathogenic variants RX and IT both exhibited phenotypes distinct from those previously observed in heterologous expression systems, likely due to compensatory mechanisms involving Cav1.3 channels. The ability of Cav1.3 and Cav1.4 channels to collectively uphold L-type calcium currents in RBCs, suggests that CSNB2-phenotypes may arise solely by mutant Cav1.4 channels in photoreceptors.