HCN channels promote Na/K-ATPase activity during slow afterhyperpolarization after seizure-like events in vitro.

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are strongly involved in the regulation of neuronal excitability, with their precise role being determined by their subcellular localization and interaction with other ion channels and transporters. Their role in causing epileptic seizures is not fully understood. Using whole-cell patch-clamp recordings of rat brain slices, we show that HCN channels constitute a substantial fraction of the membrane conductance of deep entorhinal principal neurons. Using the 4-aminopyridine model of epileptic seizures in vitro, we show that HCN channel blockade with ZD-7288 increases the frequency of seizure-like events (SLEs) and alters the time course of afterhyperpolarization after SLEs (post-SLE AHP), promoting its faster onset and making it more transient. Simultaneous whole-cell patch-clamp and K ⁺ ion-selective electrode recordings revealed that the time course of changes in neuronal membrane potential and extracellular K ⁺ concentration after SLEs in the presence of ZD-7288 differed from that in the control, which can be explained by altered Na/K-ATPase [sodium-potassium adenosine triphosphatase (sodium-potassium pump)] activity after SLEs. To confirm this hypothesis, we demonstrated the ouabain sensitivity of post-SLE AHP and showed that loading neurons with high intracellular Na ⁺ concentration prevented the effect of HCN channel blockade on post-SLE AHP. Taken together, the results obtained suggest that during post-SLE AHP, the influx of Na ⁺ through HCN channels helps to maintain Na/K-ATPase hyperactivity, resulting in the longer pauses between SLEs. Mathematical modelling confirmed the feasibility of the proposed mechanism. Such an interplay between Na/K-ATPase and HCN channels may be crucial for the regulation of seizure termination in epilepsy. KEY POINTS: HCN channels constitute a significant fraction of the resting membrane conductance of deep entorhinal principal neurons. HCN channels modulate the seizure-like events (SLEs) in the entorhinal cortex. The blockade of HCN channels increases the frequency of SLEs and reduces the duration of the afterhyperpolarization that follows them. The results suggest that HCN channels affect intracellular sodium ion concentration dynamics, prolonging the activity of the Na/K-ATPase [sodium-potassium adenosine triphosphatase (sodium-potassium) pump] after SLEs, which in turn results in longer pauses between them.
(© 2025 The Authors. The Journal of Physiology © 2025 The Physiological Society.)