Triiodo- L-thyronine enhances TRH-induced TSH release from perifused rat pituitaries and intracellular Ca 2+ levels from dispersed pituitary cells

Abstract

There is now increasing evidence that Ca 2+ serves as the first messenger for the prompt and non-genomic effects of 3,5,3′ triiodo- L-thyronine (T 3) in several tissues. We have previously shown that the first phase of thyroid stimulating hormone (TSH) release in response to thyrotropin-releasing hormone (TRH) can be potentiated by messenngers of hypothalamic origin, by a Ca 2+-dependent phenomenon involving the activation of dihydropyridine-sensitive Ca 2+ channels. By perifusing rat pituitary fragments, we have investigated whether T 3 would modify TSH release when the hormone is applied for a short time (i.e. 30 min) before a 6 min pulse of physiological concentrations of TRH, thus excluding the genomic effect of T 3. We show that: (1) increasing concentrations of T 3 (100 nM−10 μM) in the perifused medium potentiates the TRH-induced TSH release in a dose-dependent manner; (2) the T 3 potentiation is not reproduced by diiodothyronine and T 3 does not potentiate the increase of TSH release induced by a depolarizing concentration of KCl; (3) the protein synthesis inhibitor cycloheximide, does not significantly modify the effect of T 3; (4) addition of Co 2+, nifedipine, verapamil, or ω-conotoxin in the medium, at a concentration which does not modify the TSH response to TRH, reverses the T 3 potentiation of that response. We also tested whether T 3 would change intracellular concentrations of Ca 2+, by measuring [Ca 2+] i with fura-2 imaging on primary cultures of dispersed pituitary cells, either in basal conditions or after stimulation by TRH or/and T 3. Both substances induced a fast increase of [Ca 2+] i, with a peak at 15 s, followed by a subsequent progressive decay with TRH and a rapid return with T 3. Our data suggest that T 3 enhances TRH-induced TSH release by a protein synthesis-independent and Ca 2+-dependent phenomenon, probably due to an increase in Ca 2+ entry through the activation of dihydropyridine- and ω-conotoxin-sensitive Ca 2+ channels. They also show that T 3 may acutely enhance [Ca 2+] i in pituitary cells. These findings support the idea of the occurence of a prompt and stimulatory role of T 3 at the plasma membrane level in normal rat pituitary gland.