title: Chloride channel blockers suppress formation of engulfment pseudopodia in microglial cells.
abstract: OBJECTIVE
  Phagocytosis depends on the formation of engulfment pseudopodia surrounding the target. We tested in microglia, monocyte-derived cells in the brain, whether a swelling-activated Cl(-)-current (I(Cl,swell)), required for global cell volume (CV) regulation, also contributes to local expansion and retraction of engulfment pseudopodia.
  
  
  METHODS
  We used scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) to visualize and quantify the uptake of polystyrene microbeads (MBs) by microglial cells. Flow cytometry was used for cell volume measurments and I(Cl,swell) was measured by whole-cell patch clamp.
  
  
  RESULTS
  We found that exposure of microglial BV-2 cells to MBs in Cl(-)-free extracellular solution attenuated MB uptake and that the Cl(-)-channel blockers DIOA, flufenamic acid, NPPB and DCPIB suppressed the uptake of MBs in BV-2 cells and in primary microglial cells. Microglial cells exposed to MBs in the presence of Cl(-) channel blockers failed to extend engulfment pseudopodia. We observed that cells containing at least three MBs revealed an about twofold increase in current density of I(Cl, swell) compared to cells without MB. Osmotic challenges to stimulate global CV regulation before exposure to MBs modulated phagocytosis. Pre-conditioning of cells in hypo- or hypertonic medium for 12-16 hours caused a decrease in MB uptake.
  
  
  CONCLUSIONS
  These findings indicate that I(Cl,swell) contributes to formation of engulfment pseudopodia and participates in engulfment and particle uptake in microglial cells.
authors.affiliation: Department of Cell Biology, University of Salzburg, Salzburg, Austria.
authors.forename: Barbara
authors.initials: B
authors.lastname: Harl
authors.forename: Judith
authors.initials: J
authors.lastname: Schmölzer
authors.forename: Martin
authors.initials: M
authors.lastname: Jakab
authors.forename: Markus
authors.initials: M
authors.lastname: Ritter
authors.forename: Hubert H
authors.initials: HH
authors.lastname: Kerschbaum
mesh_terms: D000085:Acetates / Q000494:pharmacology; D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D048429:Cell Size; D002478:Cells, Cultured; D018118:Chloride Channels / Q000037:antagonists & inhibitors* / Q000378:metabolism; D003517:Cyclopentanes / Q000494:pharmacology; D005434:Flow Cytometry; D005439:Flufenamic Acid / Q000494:pharmacology; D007189:Indans / Q000494:pharmacology; D007192:Indenes / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D017628:Microglia / Q000166:cytology / Q000378:metabolism; D018613:Microscopy, Confocal; D008855:Microscopy, Electron, Scanning; D008863:Microspheres; D009579:Nitrobenzoates / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D010587:Phagocytosis / Q000187:drug effects; D011137:Polystyrenes / Q000737:chemistry; D011554:Pseudopodia / Q000187:drug effects* / Q000502:physiology
chemical_list: C443139:4-(2-butyl-6,7-dichlor-2-cyclopentyl-indan-1-one-5-yl)oxybutyric acid; D000085:Acetates; D002121:Calcium Channel Blockers; D018118:Chloride Channels; D003517:Cyclopentanes; D007189:Indans; D007192:Indenes; D009579:Nitrobenzoates; D011137:Polystyrenes; C095651:((2-n-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden-5-yl)oxy)acetic acid; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; D005439:Flufenamic Acid