Intracellular and plasma membrane-initiated pathways involved in the [Ca2+]i elevations induced by iodothyronines (T3 and T2) in pituitary GH3 cells

Del Viscovo A, Secondo A, Esposito A, Goglia F, Moreno M, Canzoniero LM. Intracellular and plasma membrane-initiated pathways involved in the [Ca2+](i) elevations induced by iodothyronines (T3 and T2) in pituitary GH(3) cells. Am J Physiol Endocrinol Metab 302: E1419-E1430, 2012. First published March 13, 2012; doi: 10.1152/ajpendo.00389.2011.-The role of 3,5,3'-triiodo-L-thyronine (T3) and its metabolite 3,5-diiodo-L-thyronine (T2) in modulating the intracellular Ca2+ concentration ([Ca2+](i)) and endogenous nitric oxide (NO) synthesis was evaluated in pituitary GH(3) cells in the absence or presence of extracellular Ca2+. When applied in Ca2+-free solution, T2 and T3 increased [Ca2+](i), in a dose-dependent way, and NO levels. Inhibition of neuronal NO synthase by N-G-nitro-L-arginine methyl ester and L-N-5-(1-iminoethyl)ornithine hydrochloride significantly reduced the [Ca2+](i) increase induced by T2 and T3. However, while depletion of inositol trisphosphate-dependent Ca2+ stores did not interfere with the T2- and T3-induced [Ca2+](i) increases, the inhibition of phosphatidylinositol 3-kinase by LY-294002 and the dominant negative form of Akt mutated at the ATP binding site prevented these effects. Furthermore, the mitochondrial protonophore carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone prevented the increases in both [Ca2+](i) and NO elicited by T2 or T3. Interestingly, rotenone blocked the early [Ca2+](i) increases elicited by T2 and T3, while antimycin prevented only that elicited by T3. Inhibition of mitochondrial Na+/Ca2+ exchanger by CGP37157 significantly reduced the [Ca2+](i) increases induced by T2 and T3. In the presence of extracellular calcium (1.2 mM), under carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone, T2 and T3 increased both [Ca2+](i) and intracellular Na+ concentration; nimodipine reduced the [Ca2+](i) increases elicited by T2 and T3, but inhibition of NO synthase and blockade of the Na+/H+ pump by 5-(N-ethyl-N-isopropyl) amiloride prevented only that elicited by T3; and CB-DMB, bisindolylmaleimide, and LY-294002 (inhibitors of the Na+/Ca2+ exchanger, PKC, and phosphatidylinositol 3-kinase, respectively) failed to modify the T2- and T3-induced effects. Collectively, the present results suggest that T2 and T3 exert short-term nongenomic effects on intracellular calcium and NO by modulating plasma membrane and mitochondrial pathways that differ between these iodothyronines.