In porcine thyroid cells in primary culture, thyrotropin (TSH) provokes important morphological changes associated with specific thyroidal function recovery. TSH influences cell-cell interactions and follicular morphogenesis. In the present report, we identify an ectotyrosine phosphatase activity controlled by chronic treatment with TSH. Specific substrates and various inhibitors allowed us to characterize this activity. This tyrosine phosphatase is located at the outer surface of thyroid cells as demonstrated by phosphotyrosylhistone hydrolysis. Control of cell viability and trypsin treatment confirm this extracellular localization. These data show that TSH activates ectotyrosine phosphatase activity, suggesting a role for surface protein phosphorylation in regulating specific functions of porcine thyroid cells in suspension.