Blocker state dependence and trapping in hyperpolarization-activated cation channels: evidence for an intracellular activation gate.

摘要: Hyperpolarization-activated cation currents (Ih) are key determinants of repetitive electrical activity in heart and nerve cells. The bradycardic agent ZD7288 is a selective blocker these currents. We studied the mechanism for blockade cloned Ih channels excised inside-out patches. mammalian mHCN1 channel appeared to require opening channel, but strong hyperpolarization disfavored blockade. steepness this voltage-dependent effect (an apparent valence ∼4) makes it unlikely arise solely from direct voltage on binding. Instead, probably indicates differential affinity different conformations. Similar properties were seen sea urchin homologue (SPIH), some was irreversible. To explore molecular basis difference reversibility, we constructed chimeric SPIH localized structural determinant reversibility three residues S6 region likely line pore. Using triple point mutant S6, also revealed trapping by closing channel. Overall, observations led us hypothesize that responsible block located pore lining, guarded an intracellular activation gate