ABSTRACT
The delayed rectifying outward K+ (IK) current was measured in lipopolysaccharide (LPS)-activated cultured rat microglial cells by using whole-cell patch clamp method. The current showed 'window current' where channels were available for activation but never fully inactivated. At near resting membrane potential some part of the current was able to be activated by depolarization. Among the several K+ channel blockers tested, only 4-aminopyridine (4-AP) was able to block most of the current and depolarize the membrane potential reversibly. These results suggest that 4-AP sensitive IK current plays a direct role of setting the resting membrane potential in LPS-activated microglia.
Subject(s)
Microglia/chemistry , Microglia/physiology , Potassium Channels, Voltage-Gated , Potassium Channels/physiology , 4-Aminopyridine/pharmacology , Animals , Astrocytes/cytology , Cells, Cultured , Delayed Rectifier Potassium Channels , Ion Channel Gating/physiology , Lipopolysaccharides , Membrane Potentials/physiology , Microglia/drug effects , Patch-Clamp Techniques , Rats , Rats, Sprague-DawleyABSTRACT
Among G-proteins, Gh is unique in structural differences in the GTP-binding domain and possessing transglutaminase activity. We have studied the role of G protein in modulation of large conductance Ca2+-activated K+ (Maxi-K+) channel by the inside-out mode of patch clamp in smooth muscle cells from superior mesenteric artery of the rabbit. When the non-hydrolyzable GTP analogue, GTPgammaS, was applied, the channel activity was increased about 2.5-fold. Addition of GDPbetaS resulted in reversal of the GTPgammaS effect. When the Galphah7 antibody was applied, the GTPgammaS-stimulated channel activity was significantly inhibited to control level, suggesting that Galphah is involved in activation of the Maxi-K+ channel in smooth muscle cells.
Subject(s)
GTP Phosphohydrolases/metabolism , GTP-Binding Proteins , Muscle, Smooth, Vascular/metabolism , Potassium Channels, Calcium-Activated , Potassium Channels/metabolism , Transglutaminases/metabolism , Animals , Guanosine 5'-O-(3-Thiotriphosphate)/pharmacology , Large-Conductance Calcium-Activated Potassium Channels , Protein Glutamine gamma Glutamyltransferase 2 , RabbitsABSTRACT
Large conductance Ca(2+)-activated K+ channel was identified and studied in excised inside-out membrane patches of freshly dispersed smooth muscle cells from rabbit gastric antrum. The current-voltage relationship of the single channel was linear from -80 to +80 mV of pipette voltage in which single channel conductance was 249 +/- 17.8 pS (n = 19) in symmetrical concentration of K+ (145 mM) across the patch. Activity of the channel (NPo) depended not only on cytoplasmic calcium concentration but also on membrane potential. MgATP increased NPo in a dose-dependent manner and Mg2+ was prerequisite for the effect. Okadaic acid (100 nM), inhibitor of protein phosphatases, increased NPo further in the presence of MgATP. Therefore, it would be concluded that activity of the calcium-activated K+ channel in gastric smooth muscle cells was controlled by phosphorylation state of the channel protein and the state is further modulated by membrane-delimited protein kinase and protein phosphatase activities.
