D455F and D455F/H28R channels exhibit long single-channel opening events. (A) Representative examples of excised outside-out membrane patches containing a large number of WT hASIC1a channels activated by external pH 6.5. (B) hASIC1a with D455F substitution. (C) Representative example of an excised outside-out membrane patch containing four or five channels of hASIC1a with two substitutions D455F/H28R activated by external pH 7.1. (D) Representative examples of excised outside-out membrane patches containing multiple (D455F; left panels) or one to three (D455F/H28R; right panels) channels. Holding potential was −60 mV. External solution was 100 mM Na⁺. Pipette solution was 100 mM K⁺. Dashed blue lines indicate current levels when 0, 1, 2, 3, or 4 channels were open.