The intracellular taurine release evoked by hypotonic shock is accomplished by volume-activated Cl- channels whose activity has been related to the expression of the multidrug resistance protein (MDR-1). We studied taurine transport in 9HTEo- cells and in the derived cell line 9HTEo-/Dx expressing MDR-1. [3H]taurine release from preloaded cells increased upon reduction of extracellular osmolality. This process was not inhibited by preincubation with phorbol 12-myristate 13-acetate but was reduced by inhibitors of volume-sensitive Cl- channels such as 1,9-dideoxiforskolin, La3+, and arachidonate. Verapamil, a substrate of MDR-1, increased the osmotically evoked taurine efflux. Replacement of extracellular Cl- with I- or gluconate or of extracellular Na+ with Li+ significantly reduced the taurine efflux, whereas substitution of N-methyl-D-glucamine for Na+ increased it. Application of ATP and 2-chloroadenosine stimulated the efflux in isotonic medium. No differences were seen between 9HTEo- and 9HTEo-/Dx cells with respect to hypotonically induced taurine efflux and the response to phorbol ester, channel blockers, ion replacement, and purinergic agents. Our results reveal novel properties of the osmotically induced taurine release and demonstrate its independence from MDR-1 gene expression.