hHCN4 serine 1158 alone does not directly contribute to AMPK-mediated channel modulation. (A and B) In A and B, upper panels are typical current records, and lower panels are fully activated I/V relations, normalized to cell capacitance, from HEK293F cells in control conditions (black) and after AICAR treatment (red). Data from cells transfected with wild-type hHCN4 (left in A and B) are compared with data from cells transfected with S1158A or S1158D mutant channels (right in A and B, respectively). Linear fitting of I/V curves yielded the following normalized conductance values (nS/pF): (A) 1.08, 0.70 (left) and 1.27, 0.80 (right); (B) 1.10, 0.50 (left) and 1.02, 0.59 (right) for wild-type and AICAR-treated cells, respectively. Statistical analysis and data comparisons are shown in Tables 1 and S2.