Effect of linear alkylbenzene sulfonate (LAS) on ion transport and intracellular calcium in kidney distal epithelial cells (A6)

Abstract

Linear alkylbenzene sulfonate (LAS) is found in near-shore environments receiving wastewater from urban treatment plants in a concentration reported to have physiological and toxic effect on aquatic organisms. The aim of this study was to investigate the effect LAS on ion transport and homeostasis in epithelia cells. A6 cells form a polarised epithelium when grown on permeable supports, actively absorb sodium and secrete chloride. Only the addition of LAS (100 μ m) to the apical solution of A6 epithelia resulted in an increase in the active ion transport measured as short circuit current (SCC) and transepithelial conductance (G t). This increase could not be affected by the sodium channel inhibitor amiloride (100 μ m), indicating that LAS stimulated the chloride secretion. Change in the intracellular calcium concentration (Ca 2+) i was measured in fura-2 loaded A6 cells, since it known that increase in (Ca 2+) i stimulate chloride secretion. LAS induced a concentration-dependent increase in (Ca 2+) i from 5 to 200 μ m, where the half-maximal stimulating concentration on 100 m m resulted in an increase in (Ca 2+) i from 108±15 to 570±26 n m ( n=4; P<0.01). The increase in (Ca 2+) i could be blocked by the calcium chelator ethylenebis(5-oxyethylenenitrilo)tetraacetic acid (EGTA), showing that the effect of LAS was due to influx of extracellular calcium. Furthermore, it was shown that the calcium channel inhibitor verapamil (0.2 m m) abolished the LAS induced increase in (Ca 2+) i and Gt when applied to the apical solution. However, verapamil has no inhibitory effect on these parameters when the non-ionic detergent Triton X-100 (100 μ m) was added to A6 cells. These results indicate that LAS induced a specific activation of calcium channels in the apical membrane of A6 epithelia, leading to increase in (Ca 2+) i and thereby increased chloride secretion as a result of stimulation of calcium-dependent chloride channels in the apical membrane. The change in ion homeostasis is thought to be the fundamental reason to the physiological and toxic effects induced by LAS in marine organism.