Immunohistochemical Localization of Hyperpolarization Activated Current Channel Subunits in Rat Suprachiasmatic Nucleus
Background: The hyperpolarization-activated current (Ih) is a mixed Na+/K+ inward current that is believed to regulate a wide variety of physiological function in both the central nervous system (CNS) and in the heart. The mammalian Ih is encoded by four members of the Hyperpolarization-activated and Cyclic Nucleotide-gated nonselective cation channel (HCN) gene family (HCN1-4). There was a conflict in previous reports regarding the fraction of suprachiasmatic nucleus (SCN) neurons that exhibited Ih. We first tested the hypothesis that channel subunits are found within the SCN. Methods: Using immunohistochemistry, the distribution of HCN1, HCN2, and HCN4 in the SCN was studied. Both HCN1 and HCN2 subunits were present in the SCN but with different patterns of localization. HCN4 was not detected in the SCN. Ih was also recorded from SCN neurons using whole cell voltage clamp. Results: The results show that Ih is functionally well expressed in SCN neurons. 84% of SCN neurons exhibited Ih. Ih recorded had the activation constant (τ) of 236±2 ms and amplitude of 25±1 pA at -40 mV step. At -60 mV step, τ was significantly reduced to 167±3 ms (p<0.5) while the current amplitude was significantly increased to 34±2 pA (p<0.01). In conclusion, Ih channel subunits are abundant in the SCN; the kinetic properties of the recorded Ih resemble those of HCN2 homomers. Conclusion: Our results show that Ih plays a major role in the SCN excitability and therefore may regulate its circadian function.
