A role for tyrosine phosphorylation in the regulation and sensitization of adenylate cyclase by melatonin

Mimicking short photoperiod melatonin signals (16 h exposure) on primary cell cultures of melatonin target cells of the ovine pars tuberalis (PT) results in an enhanced cAMP response to forskolin stimulation relative to untreated cells, a phenomenon termed sensitization. The sensitized response of PT cells may be an important aspect of the interpretation of the melatonin signal to initiate appropriate seasonal physiological responses. The aim of this study is to add to our understanding of the molecular mechanisms involved in the sensitization of PT cells by melatonin, We demonstrate that sensitization of PT cells by melatonin is mediated via a G(i)-coupled melatonin receptor. The sensitized cAMP response is not only obtained with the pharmacological tool forskolin, but also with cholera toxin, an activator of G(s alpha). Changes in the level of G(s alpha) or G(i alpha) G-protein subunits are ruled out as part of the sensitization mechanism. However, changes in tyrosine phosphorylation may be involved as tyrosine kinase inhibitors sensitize ovine PT cells and tyrosine phosphatase inhibitors significantly blunt adenylate cyclase activity, including the sensitized response to melatonin. The adenylate cyclase isoforms mediating the sensitized response may be broad as 7 of the 9 isoforms of adenylate cyclase are expressed in the PT.