title: Quantification of the functional expression of the Ca2+-activated K+channel KCa3.1 on microglia from adult human neocortical tissue.
abstract: The KCa3.1 channel (KCNN4) is an important modulator of microglia responses in rodents, but no information exists on functional expression on microglia from human adults. We isolated and cultured microglia (max 1% astrocytes, no neurons or oligodendrocytes) from neocortex surgically removed from epilepsy patients and employed electrophysiological whole-cell measurements and selective pharmacological tools to elucidate functional expression of KCa3.1. The channel expression was demonstrated as a significant increase in the voltage-independent current by NS309, a KCa3.1/KCa2 activator, followed by full inhibition upon co-application with NS6180, a highly selective KCa3.1 inhibitor. A major fraction (79%) of unstimulated human microglia expressed KCa3.1, and the difference in current between full activation and inhibition (ΔKCa3.1) was estimated at 292 ± 48 pA at -40 mV (n = 75), which equals at least 585 channels per cell. Serial KCa3.1 activation/inhibition significantly hyperpolarized/depolarized the membrane potential. The isolated human microglia were potently activated by lipopolysaccharide (LPS) shown as a prominent increase in TNF-α production. However, incubation with LPS neither changed the KCa3.1 current nor the fraction of KCa3.1 expressing cells. In contrast, the anti-inflammatory cytokine IL-4 slightly increased the KCa3.1 current per cell, but as the membrane area also increased, there was no significant change in channel density. A large fraction of the microglia also expressed a voltage-dependent current sensitive to the KCa1.1 modulators NS1619 and Paxilline and an inward-rectifying current with the characteristics of a Kirchannel. The high functional expression of KCa3.1 in microglia from epilepsy patients accentuates the need for further investigations of its role in neuropathological processes. GLIA 2016;64:2065-2078.
authors.affiliation: Saniona A/S, Baltorpvej 154, 2750, Ballerup, Denmark.|Neurobiology Research Unit, Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark.
authors.forename: Linda V
authors.identifier: 0000-0002-7611-9740
authors.initials: LV
authors.lastname: Blomster
authors.affiliation: Saniona A/S, Baltorpvej 154, 2750, Ballerup, Denmark.
authors.forename: Dorte
authors.initials: D
authors.lastname: Strøbaek
authors.affiliation: Saniona A/S, Baltorpvej 154, 2750, Ballerup, Denmark.
authors.forename: Charlotte
authors.initials: C
authors.lastname: Hougaard
authors.affiliation: Saniona A/S, Baltorpvej 154, 2750, Ballerup, Denmark.
authors.forename: Jessica
authors.initials: J
authors.lastname: Klein
authors.affiliation: Neurobiology Research Unit, Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark.|Epilepsy Clinic, Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark.
authors.forename: Lars H
authors.initials: LH
authors.lastname: Pinborg
authors.affiliation: Neurobiology Research Unit, Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark.
authors.forename: Jens D
authors.initials: JD
authors.lastname: Mikkelsen
authors.affiliation: Saniona A/S, Baltorpvej 154, 2750, Ballerup, Denmark. pc@saniona.com.
authors.forename: Palle
authors.initials: P
authors.lastname: Christophersen
mesh_terms: D001562:Benzimidazoles / Q000494:pharmacology; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004827:Epilepsy / Q000473:pathology; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D015847:Interleukin-4 / Q000494:pharmacology; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D017628:Microglia / Q000187:drug effects / Q000378:metabolism*; D019579:Neocortex / Q000473:pathology*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D010091:Oximes / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers / Q000494:pharmacology; D013997:Time Factors
chemical_list: C493327:6,7-dichloro-1H-indole-2,3-dione 3-oxime; D001562:Benzimidazoles; D016207:Cytokines; D007211:Indoles; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels; C496923:KCNN4 protein, human; D008070:Lipopolysaccharides; D009419:Nerve Tissue Proteins; D010091:Oximes; D026902:Potassium Channel Blockers; C086491:NS 1619; D015847:Interleukin-4
keywords: BK channel; IK channel; glial cell; neuroinflammation; patch clamp; potassium channel