Abundance of Kir2.2 reduces the slope conductance of the inward I_K1 in atrial Kir2.1^E299V channels. A, Western blots for Kir2.1 and Kir2.2 protein levels in atrial and ventricular tissue samples from mice comparing the Kir2.1 (i) and Kir2.2 (ii) expression levels. (iii) Quantification of total Kir2.1 protein levels (top) and Kir.1/Kir2.2 ratio (bottom) in atria vs. ventricles from Kir2.1^WT and Kir2.1^E299V animals (**P < 0.01, ***P < 0.001 and ****P < 0.0001; duplicate experiments in N = 4 animals per condition; Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as loading control for all comparisons). B, IV relationship of I_K1 in HEK-293 cells transfected with dimers expressing Kir2.1^WT-Kir2.2^WT or Kir2.1^E299V-Kir2.2^WT (N = 3 independent transfections; n = 8–9 cells per condition; P < 0.05 for voltages negative to −80 mV). We used a modified external solution (30 mM KCl and 110 mM NaCl) to promote the I_K1 current, shifting the reversal potential towards more positive voltages (from −80 to −30 mV). C, In-silico simulations of Kir2.1^WT-Kir2.1^E299V compared to Kir2.1^E299V-Kir2.2^WT (2.4Å vs. 4.87Å in the extracellular pore of Kir2.1^E299V-Kir2.2^WT and Kir2.1^WT-Kir2.2^WT, respectively; 0.49Å vs. 1.62Å in the cytoplasmic pore for Kir2.1^E299V-Kir2.2^WT and Kir2.1^WT-Kir2.1^E299V, respectively). We applied Welch’s t-test and two-way ANOVA for comparisons.