NONGENOMIC EFFECTS OF THYROID HORMONE ON THE INTRACELLULAR PH OF L-6 MYOBLASTS: SIGNAL TRANSDUCTION

Thyroid hormones activate by a nongenomic mechanism the Na/H exchanger of L-6 cells from rat skeletal muscle, giving rise to an increase of intracellular pH, both at the steady state and under acid load with ammonium chloride (S. Incerpi et al., Endocrinology 140:683-689, 1999). Intracellular pH was measured by the fluorescent indicator 2’,7’bis(carboxyethyl)-5(6)-carboxyfluorescein. The effect was specific as shown by the ineffectiveness of the most common thyroid hormone analogs and by the use of 5-(ethyl-N-isopropyl)amiloride and 4,4’-diisothiocyano-stilbene-2,2’disulfonic acid. The signal transduction appears to be mediated by both intracellular calcium mobilization and protein kinase activity. (S. Incerpi, Curr. Op. Endocrinol. Diab. 9:381-386, 2002). The involvement of intracellular calcium was measured by loading the L-6 myoblasts with the fluorescent probe Fura-2. Thyroid hormone gave rise to a peak after about 90 seconds for L-T3 and 5 min for L-T4, and the thyroid hormone analogs were ineffective. The action of the most common intracellular calcium modulators, such as caffeine and ryanodine was investigated. The effect of thyroid hormone on the intracellular pH appears to be mediated also by protein kinase C, MAP Kinase and Akt activity as assessed by the use of inhibitors and immunoblotting experiments.