Human thyroid follicular cells in culture synthesise plasminogen activators, both urokinase (uPA) and tissue-type (tPA). The PAs secreted into the culture medium were active (2.550 U/ml) and able to mediate the conversion of plasminogen to plasmin. Secreted PA activity (PAA) was markedly increased by epidermal growth factor (EGF) and protein kinase C (PKC) activation with TPA. There was a corresponding increase in uPA and tPA mRNA levels and in uPA and tPA protein secreted by the cells. EGF and TPA both inhibited thyroid cell function (TPA>EGF). This was measured by the uptake and organification of iodide and was dependent on the presence of insulin and TSH. At TSH concentrations optimal for iodide uptake, secreted PAA was at its nadir. These changes in PAA by TSH were reflected by changes in uPA and tPA mRNA levels measured by qRT-PCR, and in protein expression. Insulin and TSH alone and in combination decreased PAA and the mRNAs for uPA and tPA. Forskolin, an activator of adenylate cyclase, mimicked the effects of TSH. We investigated the signalling pathways controlling the enhanced secretion of uPA and tPA by EGF and TPA and found that they differed. Time course studies of PAA and mRNA levels revealed early effects by TPA (≥4 h) and late effects by EGF (≥24 h). All of the effects of TPA could be reversed by inhibition of protein kinase C beta with 100 nM LY396196 whereas the effects of EGF were largely unaffected. EGF effects were however inhibited by inhibition of p42/44 MAPK with 0.02 mM PD98059. TPA preferentially increased uPA whereas the effects of EGF were greater on tPA. We investigated the autocrine function of PAs by inhibiting PAA with serine protease inhibitors. Although these increased thyroid iodide uptake modestly, they did not prevent the inhibitory effects of EGF or TPA suggesting no direct causal link between these effects. We conclude that normal follicular cells can secrete active PAA and that the ability to take up iodide is inversely correlated with PAA secretion. Growth factors acting through PKC and p42/44 MAPK can increase PAA secretion from thyroid cells which may aid in tissue remodelling required for growth.