Abstract
The present study has concentrated on an investigation into the mechanism of release of LH from an in vitro pituitary cell perifusion system. Using pituitary cells prepared mainly from prepubertal female rats, various LH secretogogues were investigated, as well as the influence of the addition of physiological concentrations of melatonin on the resultant LH release. The presence of melatonin receptors in this and other tissues was also investigated using radioligand binding assays. LHRH induced LH secretion increased to peak values but began to fall before stimulation was discontinued. This secretory pattern was repeated for other secretagogues such as increased intracellular calcium, altered K+/Na+ ionic ratio, and to some extent, dibutryl cAMP. When LHRH was pulsed with various wash intervals the degree of refractoriness compared with the response to the previous dose varied with the dose given and the wash interval. Shorter wash intervals, or higher doses of LHRH, increased this refractoriness. Melatonin, in physiological doses, inhibited LHRH induced LH secretion when given 1.5 hours prior to, and during the stimulation period. Inclusion of melatonin during the stimulation period alone reduced this inhibitory effect. Four analogues of melatonin were far less potent in inhibiting LH release. Furthermore, melatonin reduced the calcium induced release of LH, but did not reduce the LH released by altered K+/Na+ ratio in the perifusion buffer. Therefore, the action of melatonin is probably exerted intracellularly, at the site of, or subsequent to, the action of calcium. In agreement with this finding, results from binding studies using radiolabelled melatonin, show evidence for the existence of a small population of melatonin receptors in the pituitary cytoplasm of prepubertal female rats, and not in the membrane fraction. No binding was found in adult female rats.