Non-equilibrium behavior of HCN channels: insights into the role of HCN channels in native and engineered pacemakers.
OBJECTIVE
I(f), encoded by the hyperpolarization-activated, cyclic nucleotide-modulated (HCN) channel gene family, modulates cardiac pacing. During cardiac pacing, changes in membrane potential are rapid, preventing the very slow HCN channels from reaching equilibrium. Here, we examined the properties of HCN channels under non-equilibrium conditions to shed insight into how different HCN isoforms contribute to cardiac pacing.


RESULTS
HCN1, 2 and 4 channels were heterologously expressed in Xenopus laevis oocytes or mammalian Cos7 cells and subjected to voltage clamp. We found that HCN1 channel activation (V1/2) depended strongly on the holding potential (V(H)) for short (100 ms; V1/2=-118 mV, -78 mV and -19 mV for V(H)= +70, -75 and -140 mV, respectively, in Xenopus oocytes) but not long (300-ms) test-pulses, hinting that shifts of V1/2 under non-equilibrium conditions may alter the impact of I(f) in different phases of the cardiac circle. Consistent with this notion, when a train of SA nodal-like action potentials was applied in voltage-clamp experiments, HCN1 exhibited pronounced current-voltage (IV)-hysteresis. Using computational modeling, we demonstrate that the intrinsically sluggish HCN1 activation kinetics underlie their IV-hysteretic behavior and do not hinder the ability to modulate cardiac pacing. By contrast, HCN4 did not exhibit IV-hysteresis. This difference can be attributed to the relatively large activation time constant and markedly delayed onsets of time-dependent HCN4 currents. Indeed, HCN2 channels, which have intermediate activation time constants and delays, displayed and intermediate hysteretic phenotype.


CONCLUSIONS
We conclude that non-equilibrium properties of HCN channels contribute to cardiac pacing. These results provide insight for tuning the firing rate of endogenous and induced pacemakers using engineered HCN constructs with distinct gating phenotypes.
Azene
Ezana M
EM
Department of Medicine, Johns Hopkins University, 720 Rutland Avenue/Ross 1165, Baltimore MD 21205, USA.
Xue
Tian
T
Marbán
Eduardo
E
Tomaselli
Gordon F
GF
Li
Ronald A
RA
D000200:Action Potentials; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D003198:Computer Simulation; D054815:Cyclic Nucleotide-Gated Cation Channels; D004548:Elasticity; D005260:Female; D006329:Heart Conduction System / Q000502:physiology*; D006801:Humans; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D015640:Ion Channel Gating / Q000502:physiology*; D007473:Ion Channels / Q000235:genetics / Q000378:metabolism*; D008955:Models, Cardiovascular; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism; D009865:Oocytes; D018408:Patch-Clamp Techniques; D015221:Potassium Channels; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D014981:Xenopus
D054815:Cyclic Nucleotide-Gated Cation Channels; C578665:HCN1 protein, human; C578710:HCN2 protein, human; C119587:HCN4 protein, human; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D007473:Ion Channels; D009124:Muscle Proteins; D015221:Potassium Channels; D020033:Protein Isoforms