D1R-induced increase in CaV2.2 currents requires Gs activation. (A) Real-time cAMP time course recorded from HEK293T cells transfected with D1R (+D1R; n = 3), D1R plus D1R loop 2 (+D1R + loop 2; n = 3), and empty plasmid (−D1R; n = 3) with acute application of 1 µM dopamine (DA; black circles), 1 µM chlorpromazine (CPZ; white circles), and DMEM (gray triangles). Bars on the right represent R/Ro/540 s averaged values in the different conditions. (B) Basal measurement of cAMP values in HEK293T cells transfected with D1R (+D1R; n = 6), D1R plus D1R loop 2 (+D1R + loop 2; n = 3), and empty plasmid (−D1R; n = 3). (C) Representative traces and averaged percentage of inhibition of CaV2.2 currents (ICav2.2) by dopamine in HEK293T cells cotransfected with CaV2.2, CaVα2δ1, CaVβ3, and D1R preincubated or not with 500 ng/ml ChTx (+D1R −ChTx; n = 6) and (+D1R +ChTx; n = 5). (D) Representative traces and averaged ICav2.2 from HEK293T cells cotransfected with CaV2.2, CaVα2δ1, CaVβ3, and either D1R (+D1R −ChTx; n = 17) or empty plasmid (−D1R −ChTx; n = 12), preincubated or not with ChTx (−D1R +ChTx; n = 11) and (+D1R +ChTx; n = 20). One-way ANOVA with Dunn’s post-test versus −D1R (A), versus +D1R (B), and versus −ChTx (C) and Student’s unpaired t test (C). AUC, area under the curve; n.s., nonstatistically significant. Data were expressed as mean ± SEM, and dots represent individual data points.