A hyperpolarization-activated, cyclic nucleotide-gated, (Ih-like) cationic current and HCN gene expression in renal inner medullary collecting duct cells

Abstract

The cation conductancein primary cultures of rat renal inner medullary collecting duct was studied using perforated-patch and conventional whole cell clamp techniques. Hyperpolarizations beyond -60 mV induced a time-dependent inward nonselective cationic current (I(vti)) that resembles the well-known hyperpolarization-activated, cyclic nucleotide-gated I(h) and I(f) currents. I(vti) showed a half-maximal activation around -102 mV with a slope factor of 25 mV. It had a higher conductance (but, at its reversal potential, not a higher permeability) for K(+) than for Na(+) (gK(+)/gNa(+) = 1.5), was modulated by cAMP and blocked by external Cd(2+) (but not Cs(+) or ZD-7288), and potentiated by a high extracellular K(+) concentration. We explored the expression of the I(h) channel genes (HCN1 to -4) by RT-PCR. The presence of transcripts corresponding to the HCN1, -2, and -4 genes was observed in both the cultured cells and kidney inner medulla. Western blot analysis with HCN2 antibody showed labeling of approximately 90- and approximately 120-kDa proteins in samples from inner medulla and cultured cells. Immunocytochemical analysis of cell cultures and inner medulla showed the presence of HCN immunoreactivity partially colocalized with the Na(+)-K(+)-ATPase at the basolateral membrane of collecting duct cells. This is the first evidence of an I(h)-like cationic current and HCN immunoreactivity in either kidney or any other nonexcitable mammalian cells.