Involvement of K +channels and calcium-dependent pathways in the action of T 3 on amino acid accumulation and membrane potential in Sertoli cells of immature rat testis

Abstract

The purpose of this study was to investigate the involvement of calcium in K + currents and its effects on amino acid accumulation and on the membrane potential regulated by tri-iodo-L-thyronine (T 3) in Sertoli cells. Immature rat testes were pre-incubated for 30 min in Krebs-Ringer bicarbonate buffer and incubated for 60 min in the presence of [ 14C]methylaminoisobutyric acid with and without T 3 or T 4 (dose-response curve). Specific channel blockers or chelating agents were added at different concentrations during pre-incubation and incubation periods to study the basal amino acid accumulation and a selected concentration of each drug was chosen to analyze the influence on the stimulatory hormone action. All amino acid accumulation experiments were carried out in a Dubnoff metabolic incubator at 32 °C, pH 7.4 and gassed with O 2:CO 2 (95:5; v/v). Seminiferous tubules from immature Sertoli cell-enriched testes were used for the electrophysiology experiments. Intracellular recording of the Sertoli cells was carried out in a chamber perfused with KRb with/without T 3, T 4 or blockers and the membrane potential was monitored. We found that T 3 and T 4 stimulated α-[1- 14C] methylaminoisobutyric acid accumulation in immature rat testes and induced a membrane hyperpolarization in Sertoli cells. The action of T 3 on amino acid accumulation and on the hyperpolarizing effect was inhibited by the K +-ATP channel blocker tolbutamide as well as the voltage-dependent Ca 2+ channel blocker verapamil. These results clearly demonstrate for the first time the existence of an ionic mechanism related to Ca 2+ and K + fluxes in the rapid, nongenomic action of T 3.