CONTROL OF THE INTRACELLULAR CA2+-CONCENTRATION AND THE INOSITOL PHOSPHATE ACCUMULATION IN DOG THYROCYTE PRIMARY CULTURE - EVIDENCE FOR DIFFERENT KINETICS OF CA2+-PHOSPHATIDYLINOSITOL CASCADE ACTIVATION AND FOR INVOLVEMENT IN THE REGULATION OF H2O2 PRODUCTION

Carbachol, through a muscarinic receptor, thyrotropin-releasing hormone (TRH), prostaglandin F2-alpha (PGF2-alpha), bradykinin, and adenosine triphosphate (ATP) increased the apparent [Ca2+]i (intracellular free Ca2+ concentration) of dog thyrocytes in primary culture. The [Ca2+]i measured by the Quin-2 technique rose immediately after the addition of the agonists and reached a maximal value after less than 30 seconds. Afterwards, the [Ca2+]i declined to a plateau higher than the basal level when the cells were triggered with carbachol. By contrast, in most experiments with PGF2-alpha and in the case of bradykinin, TRH, and ATP, the [Ca2+]i returned to the basal value. If the extracellular Ca2+ was chelated by excess of EGTA, the addition of all agents caused a sharp reduced transient rise in the [Ca2+]i followed by a decline of the [Ca2+]i often below the basal level (especially in the case of carbachol). It is suggested that the first transient phase of these responses is due at least in part to the mobilisation of Ca2+ from intracellular stores whereas the second sustained phase of the response to carbachol mainly originates from an increased Ca2+ influx into the thyrocytes. Carbachol, bradykinin, TRH, PGF2-alpha, and ATP also increased generation of inositol phosphates in dog thyrocytes. This effect was sustained when the cells were triggered with carbachol and was more transient with bradykinin, TRH, PGF2-alpha, or ATP. All these agents and the phorbdester TPA as well as forskolin enhanced to various extent the thyrocyte H2O2 generation. This enhancement was severely reduced in the absence of extracellular Ca2+ and was mimicked by Ca2+ ionophores in the presence of extracellular Ca2+ especially in synergy with protein kinase C activators. These data suggest that the dog thyrocyte H2O2 generation, the limiting step of the thyroid hormone synthesis, is modulated by carbachol, TRH, PGF2-alpha, bradykinin, and ATP through their action on the Ca2+-phosphatidylinositol cascade.