Blockade of chloride conductance antagonizes PMA-induced ramification in the murine microglial cell line, BV-2.
In microglial cells, activation of ion channels and ion transporters is associated with the transformation from an amoeboid to a ramified phenotype and vice versa. In the present study, we evaluated the contributions of protein kinase C (PKC) activity and ion conductance to the phorbol 12-myristate 13-acetate (PMA)-dependent ramification in the murine microglial cell line, BV-2. In a first set of experiments, we showed that PMA, a commonly used activator of PKC, but not the bioinactive analog 4 alpha-phorbol 12,13-didecanoate (4 alpha-PDD), induces ramification in BV-2 cells. Surprisingly, the PKC inhibitors calphostin C, chelerythrine, or bisindolylmaleimide II did not antagonize PMA-induced ramification. In a further set of experiments, we found that 4,4'-diisocyanatostilbene-2,2' disulfonic acid (DIDS), 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS), which block chloride channels and K-Cl cotransporters, and SKF 96365, a non-selective ion channel blocker, consistently suppressed PMA-induced ramification in BV-2 cells. Additional ion channel blockers, including lanthanides, amiloride, Ba2+, 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB), and flufenamic acid did not affect PMA-induced ramification in BV-2 cells. Cs+ accentuated the PMA-dependent ramification in BV-2 cells. Thus, our results indicate (1) that a PMA-binding protein, excluding PKC isoforms, is critical in structural remodeling of microglial cells and (2) that chloride conductance plays a pivotal role in induction of ramification in microglial cells.
Zierler
Susanna
S
Division of Animal Physiology, Department of Cellular Biology, University of Salzburg, Hellbrunnerstr. 34, 5020 Salzburg, Austria.
Kerschbaum
Hubert H
HH
D000818:Animals; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D002273:Carcinogens / Q000494:pharmacology; D002352:Carrier Proteins / Q000187:drug effects / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D048430:Cell Shape / Q000187:drug effects* / Q000502:physiology; D022081:Cell Surface Extensions / Q000187:drug effects* / Q000201:enzymology; D018118:Chloride Channels / Q000187:drug effects / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D051379:Mice; D017628:Microglia / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology; D015221:Potassium Channels / Q000187:drug effects / Q000378:metabolism; D011493:Protein Kinase C / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D015220:Calcium Channels; D002273:Carcinogens; D002352:Carrier Proteins; D018118:Chloride Channels; D004791:Enzyme Inhibitors; D015221:Potassium Channels; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate