title: Hyperpolarization-activated cyclic nucleotide-gated channels and cAMP-dependent modulation of exocytosis in cultured rat lactotrophs.
abstract: Hormone and neurotransmitter release from vesicles is mediated by regulated exocytosis, where an aqueous channel-like structure, termed a fusion pore, is formed. It was recently shown that second messenger cAMP modulates the fusion pore, but the detailed mechanisms remain elusive. In this study, we asked whether the hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, which are activated by cAMP, are involved in the regulation of unitary exocytic events. By using the Western blot technique, a real-time PCR, immunocytochemistry in combination with confocal microscopy, and voltage-clamp measurements of hyperpolarizing currents, we show that HCN channels are present in the plasma membrane and in the membrane of secretory vesicles of isolated rat lactotrophs. Single vesicle membrane capacitance measurements of lactotrophs, where HCN channels were either augmented by transfection or blocked with an HCN channel blocker (ZD7288), show modulated fusion pore properties. We suggest that the changes in local cation concentration, mediated through HCN channels, which are located on or near secretory vesicles, have an important role in modulating exocytosis.
authors.affiliation: Centre for Environmental and Marine Studies & Departamento de Biologia, Universidade de Aveiro, 3810-193 Aveiro, Portugal, Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Faculty of Medicine, and.
authors.forename: Ana I
authors.initials: AI
authors.lastname: Calejo
authors.affiliation: Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Faculty of Medicine, and Celica Biomedical Center, 1000 Ljubljana, Slovenia.
authors.forename: Jernej
authors.initials: J
authors.lastname: Jorgačevski
authors.affiliation: Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Faculty of Medicine, and.
authors.forename: Boštjan
authors.identifier: http://orcid.org/0000-0002-1618-3636
authors.initials: B
authors.lastname: Rituper
authors.affiliation: Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Faculty of Medicine, and.
authors.forename: Alenka
authors.identifier: http://orcid.org/0000-0003-4453-1498
authors.initials: A
authors.lastname: Guček
authors.affiliation: Centre for Environmental and Marine Studies & Departamento de Biologia, Universidade de Aveiro, 3810-193 Aveiro, Portugal.
authors.forename: Patrícia M
authors.identifier: http://orcid.org/0000-0002-4828-7632
authors.initials: PM
authors.lastname: Pereira
authors.affiliation: Centre for Environmental and Marine Studies & Departamento de Biologia, Universidade de Aveiro, 3810-193 Aveiro, Portugal.
authors.forename: Manuel A S
authors.initials: MA
authors.lastname: Santos
authors.affiliation: Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Faculty of Medicine, and Celica Biomedical Center, 1000 Ljubljana, Slovenia.
authors.forename: Maja
authors.initials: M
authors.lastname: Potokar
authors.affiliation: Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Faculty of Medicine, and Celica Biomedical Center, 1000 Ljubljana, Slovenia.
authors.forename: Nina
authors.initials: N
authors.lastname: Vardjan
authors.affiliation: Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Faculty of Medicine, and Biotechnical Faculty, University of Ljubljana, 1000 Ljubljana, Slovenia, and Celica Biomedical Center, 1000 Ljubljana, Slovenia.
authors.forename: Marko
authors.initials: M
authors.lastname: Kreft
authors.affiliation: Centre for Environmental and Marine Studies & Departamento de Biologia, Universidade de Aveiro, 3810-193 Aveiro, Portugal.
authors.forename: Paula P
authors.initials: PP
authors.lastname: Gonçalves
authors.affiliation: Laboratory of Neuroendocrinology-Molecular Cell Physiology, Institute of Pathophysiology, Faculty of Medicine, and Celica Biomedical Center, 1000 Ljubljana, Slovenia robert.zorec@mf.uni-lj.si.
authors.forename: Robert
authors.initials: R
authors.lastname: Zorec
mesh_terms: D000818:Animals; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000502:physiology*; D005089:Exocytosis / Q000502:physiology*; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels / Q000037:antagonists & inhibitors / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D052682:Lactotrophs / Q000502:physiology*; D008297:Male; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000235:genetics / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar
chemical_list: C578712:Hcn2 protein, rat; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D015221:Potassium Channels; D000242:Cyclic AMP
keywords: HCN channels; RT-PCR; cAMP; capacitance measurements; exocytosis; lactotrophs