Characterization of Volume-activated Chloride Currents in Endothelial Cells from Bovine Pulmonary Artery

Abstract.

We have measured the kinetic and pharmacological properties of volume-activated Cl⁻ currents (I _Cl,vol) in endothelial cells, and tried to correlate them with those of the already described volume-activated current I _Cln. Both conductances show a similar permeability sequence for monovalent anions, and they are blocked by extracellular ATP. In the present report, we demonstrate by Western blot and RT-PCR that cultured endothelial cells from bovine pulmonary artery (CPAE) contain pI _Cln. The expression of this protein has been shown to be closely associated with the I _Cln current.

I _Cl,vol showed however, in contrast with I _Cln, no striking inactivation at positive potentials. This property is also at variance with that of the volume-activated current related to MDR-1. Activation of I _Cl,vol at potentials more negative than −80 mV was not time dependent, which excludes a major contribution of a ClC-2 related current. The antiviral nucleoside analogue AZT (3′-azido-3′-deoxythymidine) inhibited I _Cl,vol by 21 ± 2.7% (n = 10), at a concentration of 100 μm. Another antiviral drug, acyclovir (ACV, 9-[(2-hydroxyethoxy)methyl]guanine) blocked I _Cl,vol by 27 ± 6.2% at 100 μm (n = 11). Both blocking effects are much smaller than those reported for I _Cln.

The phenol derivative gossypol, which blocks I _Cln-related currents, efficiently inhibited I _Cl,vol in CPAE cells (67 ± 2.1% at 1 μm, n = 7, K _I = 0.4 μm).

The presence of pI _Cln in CPAE cells and the similar qualitative pharmacological profile of I _Cl,vol and I _Cln support the hypothesis that pI _Cln is a good molecular candidate for I _Cl,vol in endothelial cells. The discrepant kinetic properties may indicate that these time-dependent currents at high positive or negative potentials are not intrinsic properties of the channels, but are caused by time-dependent depletion/accumulation phenomena due to the large amplitudes of these currents.