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

Abstract

A hyperpolarization-activated, cyclic nucleotide-gated, (Ihlike) cationic current and HCN gene expression in renal inner medullary collecting duct cells. Am J Physiol Cell Physiol 294: C893-C906, 2008. First published January 16, 2008 doi:10.1152/ajpcell.00616.2006The cation conductancein primary cultures of rat renal inner medullary collecting duct was studied using perforated-patch and conventional whole cell clamp techniques. Hyperpolarizations beyond Ϫ60 mV induced a time-dependent inward nonselective cationic current (Ivti) that resembles the well-known hyperpolarization-activated, cyclic nucleotidegated Ih and If currents. Ivti showed a half-maximal activation around Ϫ102 mV with a slope factor of 25 mV. It had a higher conductance (but, at its reversal potential, not a higher permeability) for K ϩ than for Na ϩ (gK ϩ /gNa ϩ ϭ 1.5), was modulated by cAMP and blocked by external Cd 2ϩ (but not Cs ϩ or ZD-7288), and potentiated by a high extracellular K ϩ concentration. We explored the expression of the Ih 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 ϳ90-and ϳ120-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 ϩ -K ϩ -ATPase at the basolateral membrane of collecting duct cells. This is the first evidence of an Ih-like cationic current and HCN immunoreactivity in either kidney or any other nonexcitable mammalian cells. kidney; hyperpolarization-activated current; nonselective cation channel; sodium transport HYPERPOLARIZATION-ACTIVATED, cyclic nucleotide-gated, cationic nonselective (HCN) currents termed I h , I f , I q , or inward pacemaker currents are found in a wide variety of excitable cells (1,21,32,41). The I h channels activate, with a slow time course, at hyperpolarizing voltages beyond Ϫ60 mV (near the resting potential of most cells) and are permeable to both Na ϩ and K ϩ . They are regulated by cyclic nucleotides (cAMP and cGMP) and external K ϩ and blocked by external Cs ϩ . The genes coding for I h channels form the HCN family, with four members (HCN1 to -4) in mammals. Each HCN isoform is able to form homomeric conducting channels that differ in their kinetics, steady-state voltage dependence, and sensitivity to modulation by cAMP (1,21,41,45). HCN isoforms may also coassemble to form heteromeric channel complexes (54, 27).Although I h and HCN gene expression is observed, almost exclusively, in excitable cells, a Northern blot study performed in mouse has shown the expression in liver and kidney of a splice variant of the HCN3 mRNA expressed in brain (44) while, more recently, the presence of mRNA encoding HCN2 and HCN3 in the kidney-derived transformed HEK293 cell was reported (57). If nonexcitable cells may express HCN genes, then the presence of the ...

Functional properties and cell type specific distribution ofIh channels in leech neurons

“…Comparison with permeability and conductance ratios obtained from other preparations reveals that the values calculated here are within the same range [p Na /p K between 0.2 and 0.4 (for a review, see Pape, 1996); g Na /g K between 0.25 and 0.67 (e.g. Bolívar et al, 2008;Macri et al, 2002)]. …”

The pelvis–kidney junction contains HCN3, a hyperpolarization-activated cation channel that triggers ureter peristalsis

“…14,15 RT-PCR analyses described in this report demonstrate that high levels of HCN3 and barely detectable levels of HCN2 are expressed in the proximal region of the upper urinary tract. Immunohistochemical studies showed that HCN2 expression is restricted to the renal tubules, 20 whereas we show that HCN3 expression is localized to the connective tissue layer underlying the smooth-muscle coat at the PKJ. Consistent with this HCN expression pattern, inhibition of HCN channel activity caused loss of spontaneous electrical activity at the PKJ, desynchronized the electrical propagation in the upper urinary tract, and caused abnormal peristalsis.…”

“…HCN2 expression has been localized to the inner medullary collecting ducts, 20 and to spatially localize HCN3 protein expression we performed chromogenic immunohistochemistry using 80-mm-thick urinary tract sections. HCN3 was highly expressed in the upper urinary tract at the PKJ and in the renal papilla (Figure 5d and e).…”

Effects of hyperpolarization-activated cyclic nucleotide-gated (HCN) channel blockers on the proliferation and cell cycle progression of embryonic stem cells

“…In the brain, they are shown to be related to learning, memory, and epilepsy [4,6,26]. Although there were previous studies showing the molecular expression of HCN isoform transcripts [27,33] or nicely demonstrating the functional expression of HCN currents [7] in non-excitable cells, to our best knowledge, there is no previous report on the function of HCN channels in non-excitable cells. In this study, it is clear that HCN blockers, cesium and ZD7288, decreased the proliferation and affected the cell cycle pattern of mESCs.…”