It is well established that TGF- stimulates human lung fibroblasts (HLF) to differentiate into myofibroblasts. We characterized lysophosphatidic acid (LPA)-activated Cl^- channels (ICl_LPA) in cultured human lung fibroblasts and myofibroblasts and investigated the influence of ICl_LPA on fibroblast to myofibroblast differentiation. ICl_LPA was recorded using the amphotericin perforated-patch technique. ICl_LPA was activated using LPA or sphingosine-1-phosphate (S1P). Phenotype was determined by Western blotting and immunohistochemistry using an anti--SMA antibody. RT-PCR was performed to determine which phospholipid growth factor receptors are present in HLF. We found that HLF cultured in TGF- (myofibroblasts) had significantly elevated alpha-smooth muscle actin (-SMA) levels and ICl_LPA current density compared to control fibroblasts. ICl_LPA activation was blocked by 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) ,5- nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) and LPA receptor-specific antagonist dioctyl-glycerol pyrophosphate (DGPP;1uM). DIDS and NPPB, in a dosedependent manner, significantly reduced -SMA levels in HLF stimulated with TGF-. These results demonstrate the receptor mediated activation of ICl_LPA by LPA and S1P in cultured human lung myofibroblasts, with only minimal ICl_LPA activity in fibroblasts. This chloride channel activity appears to play a critical role in the differentiation of human lung fibroblasts to myofibroblasts.