title: Increased expression of hyperpolarization-activated cyclic nucleotide-gated (HCN) channels in reactive astrocytes following ischemia.
abstract: Astrocytes respond to ischemic brain injury by proliferation, the increased expression of intermediate filaments and hypertrophy, which results in glial scar formation. In addition, they alter the expression of ion channels, receptors and transporters that maintain ionic/neurotransmitter homeostasis. Here, we aimed to demonstrate the expression of Hcn1-4 genes encoding hyperpolarization-activated cyclic nucleotide-gated (HCN) channels in reactive astrocytes following focal cerebral ischemia (FCI) or global cerebral ischemia (GCI) and to characterize their functional properties. A permanent occlusion of the middle cerebral artery (MCAo) was employed to induce FCI in adult GFAP/EGFP mice, while GCI was induced by transient bilateral common carotid artery occlusion combined with hypoxia in adult rats. Using FACS, we isolated astrocytes from non-injured or ischemic brains and performed gene expression profiling using single-cell RT-qPCR. We showed that 2 weeks after ischemia reactive astrocytes express high levels of Hcn1-4 transcripts, while immunohistochemical analyses confirmed the presence of HCN1-3 channels in reactive astrocytes 5 weeks after ischemia. Electrophysiological recordings revealed that post-ischemic astrocytes are significantly depolarized, and compared with astrocytes from non-injured brains, they display large hyperpolarization-activated inward currents, the density of which increased 2-3-fold in response to ischemia. Their activation was facilitated by cAMP and their amplitudes were decreased by ZD7288 or low extracellular Na(+) concentration, suggesting that they may belong to the family of HCN channels. Collectively, our results demonstrate that regardless of the type of ischemic injury, reactive astrocytes express HCN channels, which could therefore be an important therapeutic target in poststroke therapy.
authors.affiliation: Department of Cellular Neurophysiology, Institute of Experimental Medicine, Academy of Sciences of the Czech Republic, Prague 4, 14220, Czech Republic.
authors.forename: Pavel
authors.initials: P
authors.lastname: Honsa
authors.forename: Helena
authors.initials: H
authors.lastname: Pivonkova
authors.forename: Lenka
authors.initials: L
authors.lastname: Harantova
authors.forename: Olena
authors.initials: O
authors.lastname: Butenko
authors.forename: Jan
authors.initials: J
authors.lastname: Kriska
authors.forename: David
authors.initials: D
authors.lastname: Dzamba
authors.forename: Vendula
authors.initials: V
authors.lastname: Rusnakova
authors.forename: Lukas
authors.initials: L
authors.lastname: Valihrach
authors.forename: Mikael
authors.initials: M
authors.lastname: Kubista
authors.forename: Miroslava
authors.initials: M
authors.lastname: Anderova
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism*; D001921:Brain / Q000166:cytology; D000242:Cyclic AMP / Q000494:pharmacology; D054815:Cyclic Nucleotide-Gated Cation Channels / Q000235:genetics / Q000378:metabolism*; D004195:Disease Models, Animal; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology*; D005904:Glial Fibrillary Acidic Protein / Q000235:genetics / Q000378:metabolism; D066298:In Vitro Techniques; D007511:Ischemia / Q000473:pathology*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D051379:Mice; D008822:Mice, Transgenic; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D011743:Pyrimidines / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D012964:Sodium / Q000378:metabolism
chemical_list: D054815:Cyclic Nucleotide-Gated Cation Channels; D005904:Glial Fibrillary Acidic Protein; D009419:Nerve Tissue Proteins; D011743:Pyrimidines; C082246:ICI D2788; D012964:Sodium; D000242:Cyclic AMP
keywords: HCN channels; ZD7288; astrocytes; cortex; focal and global cerebral ischemia; hippocampus