Purchase this article with an account.
SP Srinivas, C Maertens, B Nilius; Fluoxetine Inhibits Volume Regulated Anion Channels (VRAC) In Cultured Bovine Trabecular Meshwork Cells (BTMC) . Invest. Ophthalmol. Vis. Sci. 2002;43(13):4029.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Purpose: Acute osmotic swelling activates VRAC of unknown molecular identity in many cell types. VRAC are necessary, if not sufficient, for regulatory volume decrease. A small VRAC activity may prevail under isosmotic conditions and skew the membrane potential to a depolarized state. This study has examined VRAC in cultured BTMC. Methods: BTMC were grown on glass coverslips coated with 0.2% gelatin. Expression of mRNAs for CLC family Cl- channels and CFTR was examined by RT-PCR using total RNA from BTMC. Volume-sensitive Cl- currents (ICl (swell)) were induced by exposing cells to 25% hyposmotic Ringers. These experiments were carried out at room temperature using whole cell configuration of the patch clamp technique. A step protocol (1 sec voltage steps from -100 to +100 mV; 20 mV increments) was employed to assess current-voltage relations. A voltage ramp protocol was employed to obtain time course of ICl (swell) during hyposmotic shocks. Results: RT-PCR analyses gave positive bands for the expression of CLC-2 and CLC-5 isoforms. Expression patterns were confirmed by sequencing the PCR products. CLC-3 and CFTR were not expressed. CLC-5 expression alone was enhanced by exposure to dexamethasone (5 days at 500 nM). ICl (swell) was induced in response to hyposmotic shock (225 mosM) with characteristics of outward rectification (n=3). NPPB inhibited ICl (swell) by ≷90 % at 100 µM (n=3; calculated at +100 mV). Similarly, fluoxetine inhibited ICl (swell) by 100 % at 50 µM (n=6; also calculated at +100 mV). Both drugs inhibited ICl (swell) rapidly and in a reversible manner as reported in other cell types. Conclusion: BTMC express VRAC with characteristics similar to those expressed by vascular endothelial cells (Nilius et al., Gen Pharmacol, 27: 67-77, 1996). Since BTMC express L-type Ca2+channels, VRAC activity could influence their conductance through depolarization of Em. CLC-5 expression in BTMC indicates that it is not kidney specific.
This PDF is available to Subscribers Only