Molecular identity, ontogeny, and cAMP modulation of the hyperpolarization-activated current in vestibular ganglion neurons

Abstract

Properties, developmental regulation, and cAMP modulation of the hyperpolarization-activated current (I(h)) were investigated by the whole cell patch-clamp technique in vestibular ganglion neurons of the rat at two postnatal stages (P7-10 and P25-28). In addition, by RT-PCR and immunohistochemistry the identity and distribution of hyperpolarization-activated and cyclic nucleotide-gated channel (HCN) isoforms that generate I(h) were investigated. I(h) current density was larger in P25-28 than P7-10 rats, increasing 410% for small cells (<30 pF) and 200% for larger cells (>30 pF). The half-maximum activation voltage (V(1/2)) of I(h) was -102 mV in P7-10 rats and in P25-28 rats shifted 7 mV toward positive voltages. At both ages, intracellular cAMP increased I(h) current density, decreased its activation time constant (τ), and resulted in a rightward shift of V(1/2) by 9 mV. Perfusion of 8-BrcAMP increased I(h) amplitude and speed up its activation kinetics. I(h) was blocked by Cs(+), zatebradine, and ZD7288. As expected, these drugs also reduced the voltage sag caused with hyperpolarizing pulses and prevented the postpulse action potential generation without changes in the resting potential. RT-PCR analysis showed that HCN1 and HCN2 subunits were predominantly amplified in vestibular ganglia and end organs and HCN3 and HCN4 to a lesser extent. Immunohistochemistry showed that the four HCN subunits were differentially expressed (HCN1 > HCN2 > HCN3 ≥ HCN4) in ganglion slices and in cultured neurons at both P7-10 and P25-28 stages. Developmental changes shifted V(1/2) of I(h) closer to the resting membrane potential, increasing its functional role. Modulation of I(h) by cAMP-mediated signaling pathway constitutes a potentially relevant control mechanism for the modulation of afferent neuron discharge.

Developmental increase in hyperpolarization-activated current regulates intrinsic firing properties in rat vestibular ganglion cells

“…In the present study, we have shown a developmental increase in I h current density, a decrease in its activation time constant, and a positive shift of V 1/2 in VGCs obtained from P3 to P14 rats. These results are consistent with those of Almanza et al (2012), who compared properties of I h in VGCs obtained from rats between P7-10 and P25-28 (Almanza et al, 2012). In the present study, we also showed that the expression of HCN1 protein as well as its mRNA increases developmentally while the other HCN isoforms did not change during the same developmental period.…”

“…Our results suggest that HCN channels as well as Kv1 channels contribute to control the pattern of spike discharges in a complementary way. The HCN channel blocker ZD7288 changed the shape of APs, showing decrease in the AP spike height, increase in the AP spike width, decrease in the amplitude of AHP, and increase in the width of AHP, consistent with the results reported by Almanza et al (2012). The sculpting of APs is probably due to persistent activity of I h at the resting membrane potential since I h kinetics are considerably slower than those of the APs.…”

“…Developmental changes in the properties of I h have been observed in several kinds of neurons, such as hippocampal neurons (Surges et al, 2006), cardiac neurons (Hogg et al, 2001), trigeminal neurons (Cho et al, 2011), and VGCs (Almanza et al, 2012;Horwitz et al, 2014). In rat hippocampal neurons, I h amplitude increased progressively and its activation kinetics became faster during the first postnatal week (Surges et al, 2006).…”

“…The HCN channels are present in many excitable cells (Pape, 1996;Santoro et al, 2000) and all subtypes are present in VGCs (Almanza et al, 2012;Horwitz et al, 2014). The HCN channels have been implicated in maintaining the resting potential and input resistance (Surges et al, 2004), rhythmogenesis (McCormick andPape, 1990), and control of synaptic transmitter release (Beaumont and Zucker, 2000).…”

“…In vestibular ganglion neurons, it has recently been reported that the current density of I h increases and activation kinetics become faster during postnatal periods (Almanza et al, 2012). However, the functional role of the developmental increase of I h in the firing properties of VGCs remains unclear.…”