HyNaC5 increases the apparent affinity for amiloride and amiloride analogs. A, concentration response curve for the inhibition of HyNaC currents by amiloride. Curves were determined with 0.3 μm (HyNaC2/3/5) and 30 μm Hydra-RFamide II (HyNaC2/3 and HyNaC2/3/5), respectively (n = five oocytes for HyNaC2/3 and n = 6 oocytes for HyNaC2/3/5). B, left, representative current traces of whole oocytes either expressing HyNaC2 and -3 or HyNaC2, -3, and -5. Channels were activated in the presence of different blockers (100 μm). Right, bar graphs illustrating the current remaining in the presence of the respective blockers; black bars represent HyNaC2/3/5, white bars HyNaC2/3. Before and after application of the blockers, channels were activated without blockers; the mean value of these two measurements was used to normalize the currents obtained in the presence of the blockers. Error bars represent S.E. (six oocytes); HyNaCs were activated with 30 μm (HyNaC2/3) or 1 μm Hydra-RFamide I (HyNaC2/3/5), respectively. In the presence of HyNaC5, amiloride (Aml) and benzamil (Benza) blocked significantly more current, suggesting an increased affinity for these blockers. *, p < 0.05; ***, p < 0.001, two-tailed t test. Phena, phenamil.