1. The regulation of Maxi Cl- channels by 17β-oestradiol and non-steroidal triphenylethylene antioestrogens represents a rapid, non-classical effect of these compounds. In the present study we have investigated the signalling pathways used for the regulation of Maxi Cl- channel activity by oestrogens and antioestrogens in C1300 neuroblastoma cells. 2. Whole-cell Maxi Cl- currents were readily and reversibly activated by tamoxifen, toremifene and the membrane-impermeant ethyl-bromide tamoxifen, only when applied to the extracellular medium. 3. Pre-treatment of C1300 cells with oestrogen or cAMP prevented the antioestrogen-induced activation of Maxi Cl- channels. The inhibitory effect of 17β-oestradiol and cAMP was abolished by the kinase inhibitor staurosporine. 4. Current activation was unaffected by the removal of intracellular Ca2+ and Mg2+, but was completely abolished in the presence of okadaic acid. These results are consistent with the participation of an okadaic acid-sensitive serine/threonine protein phosphatase in the activation of Maxi Cl- channels. However, neither oestrogen or antioestrogen treatment modified the total activity of the two major serine/threonine phosphatases, PP1 and PP2A, in C1300 cells. 5. Although the role of these Maxi Cl- channels remains unknown, our findings suggest strongly that their modulation by oestrogens and antioestrogens is linked to intracellular signalling pathways.