Gonadotropin-releasing hormone-1 neuronal activity is independent of hyperpolarization-activated cyclic nucleotide-modulated channels but is sensitive to protein kinase A-dependent phosphorylation

Pulsatile release of GnRH-1 stimulates the anterior pituitary and induces secretion of gonadotropin hormones. GnRH-1 release is modulated by many neurotransmitters that act via G protein-coupled membrane receptors. cAMP is the most ubiquitous effector for these receptors. GnRH-1 neurons express hyperpolarization-activated cyclic nucleotide-modulated (HCN) channel protein in vivo. HCN channels are involved in neuronal pacemaking and can integrate cAMP signals. cAMP-dependent protein kinase (PKA) is also activated by cAMP signals, and PKA-dependent phosphorylation modulates voltage-activated channels. In this report, these two pathways were examined in GnRH-1 neurons as integrators of forskolin (FSK)-induced stimulation. The HCN3 isoform was detected in GnRH-1 neurons obtained from mouse nasal explants. ZD7288, a HCN channel blocker, significantly reduced the efficiency of FSK to stimulate GnRH-1 neurons, whereas blockade of PKA with Rp-adenosine-3',5'-cyclic monophosphorothioate triethylammonium did not attenuate the FSK-induced stimulation. To ensure that disruption of HCN channels on GnRH-1 neurons was responsible for reduction of FSK stimulation, experiments were performed removing gamma-aminobutyric acid (GABA), the major excitatory input to GnRH-1 neurons in nasal explants. Under these conditions, Rp-adenosine-3',5'-cyclic monophosphorothioate triethylammonium, but not ZD7288, altered the FSK-induced response of GnRH-1 neurons. These studies indicate that PKA-dependent phosphorylation is involved in the FSK-induced stimulation of GnRH-1 neurons rather than HCN channels, and HCN channels integrate the FSK-induced stimulation on GABAergic neurons. In addition, blockade of HCN channels did not modify basal GnRH-1 neuronal activity when GABAergic input was intact or removed, negating a role for these channels in basal GABAergic or GnRH-1 neuronal activity.