The thiazolidinedione (TZD) drugs rosiglitazone (Ro) and pioglitazone (Po) are PPAR agonists in widespread clinical use as insulin-sensitizing agents in type 2 diabetes. Based on recent evidence implicating PPAR as a positive modulator of intestinal epithelial differentiation, we hypothesized that TZD drugs might attenuate intestinal secretory function. To evaluate this possibility, we examined the effects of Ro and Po on electrogenic Cl^- secretion (Isc) in mouse intestinal segments and in cultured human intestinal epithelial cells (HT29-Cl.19A). As hypothesized, oral administration of Ro (20 mg/kg/day) to mice for 8-28 days markedly reduced intestinal Isc responses to cAMP (forskolin) and Ca²⁺ (carbachol) dependent stimuli. In these Ro-treated mice, cholera toxin-induced intestinal fluid accumulation was reduced 65%. With continued Ro treatment, the Isc response to carbachol recovered significantly, whereas that to forskolin remained attenuated. Treatment of HT29 cells for 5 days with 10 µM Ro or Po in vitro brought about a similar hyposecretory state. In HT29 cells, the loss of cAMP-dependent Cl^- secretion was attributable to a reduced expression of CFTR Cl^- channel, KCNQ1 K⁺ channel, and Na-K-2Cl cotransporter-1 proteins. The transient loss of Ca²⁺-dependent Cl- secretion involved an impairment of basolateral Ca²⁺-stimulated K⁺ channel activity without a detectable loss of K_Ca3.1 channel protein. Our results establish TZD drugs as important modulators of intestinal Cl^- secretory function.