Studies on the Release of Follicle-Stimulating Hormonein Vitrofrom Rat Pituitary Glands Stimulated by Hypothalamic Follicle-Stimulating Hormone-Releasing Factor

Abstract

A specific and sensitive method for the assay of FSH-releasing factor (FRF) in vitro was developed. Pituitary halves of Wistar rats, ovariectomized and pretreated with estradiol benzoate and progesterone, were incubated for 2 hr. at 37[degree]C in a Krebs Ringer bicarbonate-glucose medium, in the presence or absence of FRF. At the end of the incubation period, the pituitary tissue and the supernatant were assayed for FSH using the Steelman-Pohley method. Control studies established that there was no evidence of inactivation of FSH under the conditions used for incubation and that hypothalamic extracts, injected directly into Steelman-Pohley rats, did not stimulate the ovaries, nor potentiate their response to FSH. Different substances such as pitressin, [alpha]- and [beta] -MSH, serotonin, melatonin and cortical extract were shown not to act on the release of FSH from the pituitary. A graded response in release of FSH by pituitary tissue stimulated with graded doses of a hypothalamic extract or purified FRF was observed. On the other hand, it was demonstrated that the FSH-releasing process was related to the constant presence of FRF in the tissue. It stopped if there was a lack of the releasing factor but could be reinitiated by adding FRF. The pituitary tissue was responsive to stimulation by FRF, in conditions used in this work, for at least a 4-hr. period. If during this time the incubation medium was renewed and FRF was repeatedly added the amount of FSH released per hr. of incubation was almost constant. At the end of the incubation period the total amount of FSH present in incubation media and tissues treated with FRF was about 40% higher than in the control. This seems to suggest that FSH was synthesized during the incubation period in pituitary tissue treated with FRF. Incubations with puromycin and actinomycin D showed that the stimulating action of FRF on FSH release was significantly but-only partially inhibited by these antibiotics. As inhibition of this action by puromycin (39%) and actinomycin D (21%) was not equivalent to that of total synthesis of proteins (95%) and RNA (84%), respectively, this may suggest that FRF does not act directly and solely on stimulation either of biosynthesis of the FSH polypeptide chain or of synthesis of the messenger RNA specific for FSH. It seems, on the contrary, to act on the stimulation of the release of this hormone (as shown in experiments with antibiotics). To explain the synthesis of FSH during the incubation period in pituitary tissue treated with FRF, a 2-stage reaction may be envisaged: FRF inducing the release of FSH, which is in the 2nd stage resynthesized in the tissue. Another possibility would be a dual action of FRF on release and synthesis of FSH.