Kinetic and ionic properties of the human HCN2 pacemaker channel

Abstract.

Human cDNA coding for the hyperpolarization-activated "pacemaker" channel HCN2 was expressed in Phoenix cells and yielded an inward current (I _hHCN2) activated on hyperpolarization. The average I _hHCN2 was half-activated at –83.1 mV and its kinetics could be described by second-order Hodgkin–Huxley gating. The time constant curve was bell-shaped and peaked at –82.2 mV. With 115 mM external Na⁺ and 30 mM external K⁺, I _hHCN2 reversed at –17.1 mV, and had a mean conductance of 5.6 nS. Reducing the external K⁺ or Na⁺ concentration led to a concentration-dependent reduction of the I _hHCN2 conductance and to a hyperpolarizing shift of reversal potential. External Cs⁺ ions (5 mM) blocked I _hHCN2 in a voltage-dependent way according to a Woodhull-type block model, at an electrical distance of 0.66 from the external membrane surface, and with a dissociation constant of 15 mM at 0 mV. Increasing cytoplasmic cAMP using forskolin increased I _hHCN2 by shifting the current activation curve to more positive voltages (11.7 mV). Exposure of the intracellular side of inside-out macro-patches to cAMP led to a depolarizing shift of the channel open probability curve (15.2 mV with 10 µM cAMP). These results indicate that although hHCN2 channels share several properties with native cardiac f-channels, differences also exist in permeability and block properties, suggesting that native channels may not be composed simply of homomeric constructs.