Figure 4.

A₁R-dependent intracellular Ca²⁺mobilization. (A) Determination of A₁R-mediated intracellular calcium accumulation by means of a luciferase reporter assay system. HEK-293T cells were transiently transfected with the firefly luciferase-encoding plasmid (pGL4-NFAT-luc2p) and the cDNAs encoding the A₁R^SNAP and the YFP. Thirty-six hours after transfection, cells were treated 6 h with the A₁R agonist R-PIA (PIA, 50 nM) in the absence or presence of DPCPX (500 nM) or guanosine (Guo, 100 μM). Light emission is presented as the percentage increase over basal levels. The data are expressed as the mean ± SEM of three independent experiments performed in triplicate. The asterisks indicate statistically significant differences *** p < 0.001, one-way ANOVA followed by Dunnett’s post-hoc test when compared to control. (B) Guanosine modulation of R-PIA-mediated intracellular Ca²⁺mobilization (PIA-mediated NFAT-Luc induction) in cells expressing A₁R^SNAP (red bars) or A₁R^SNAP plus A_2AR^SNAP (blue dashed bars). The dotted line represents the Ca²⁺ mobilization induced by R-PIA in the absence of guanosine within each cell transfection group. The data are expressed as the mean ± SEM of three independent experiments performed in triplicate.