Involvement of stretch-activated Cl- channels in ramification of murine microglia

A stretch-activated Cl- current (ICl) was investigated in cultured murine microglia using the whole-cell configuration of the patch-clamp technique. After application of membrane stretch, a Cl- current appeared within seconds, and its amplitude increased further within 3-8 min. ICl underwent rundown, which was prevented by addition of 4 mM ATP to the intracellular perfusing solution. The stretch-activated Cl- current exhibited outward rectification and did not show any voltage-dependent gating. Lowering the concentration of extracellular Cl- from 142 to 12 mM by equimolar substitution of Cl- with gluconate shifted the reversal potential of ICl by 41.6 +/- 1.8 mV in the depolarizing direction. 4, 4'-Diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) and 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS) blocked ICl in a voltage- and time-dependent manner. At a test potential of +40 mV, a half-maximal blockade at 16.1 microM DIDS and at 71.0 microM SITS was determined for ICl. At a concentration of 200 microM, 5-nitro-2-(3-phenylpropylamino)benzoic acid or flufenamic acid blocked ICl by 88% and 75%, respectively. Each of these four Cl- channel blockers reversibly inhibited the ramification process of microglia, whereas blockers of voltage-gated Na+ and K+ channels did not affect the transformation of microglia from their ameboid into the ramified phenotype. It is suggested that in microglia functional stretch-activated Cl- channels are required for the induction of ramification but not for maintaining the ramified shape.