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Rabbit antisera to rat pituitary follicle-stimulating hormone (FSH) and to rat luteinizing hormone (LH) were used, in an immunocytochemical probe, to determine the ontogeny and distribution of immunoreactive, endogenous, intraovarian FSH and LH in immature rats. Ovaries from rats 4, 8, 12, and 21 days of age were studied. Both gonadotrophins were first immunodetectable on Day 8. In reactive primordial follicles, LH was restricted to the cytoplasm and nuclei of the surrounding follicle cells. In those follicles possessing both squamous and cuboidal follicle cells, i.e., transitional between primordial and primary, LH was found in both the cytoplasm and nuclei of both follicle cell types. In primary follicles, LH was no longer present in granulosa cells but was concentrated in germ cell cytoplasm. In contrast, in primordial follicles, FSH was restricted to the germ cell but was present in both the oocyte cytoplasm and germinal vesicle. In transitional and primary follicles, FSH remained within the oocyte cytoplasm and germinal vesicle but also became detectable within the cytoplasm and nuclei of granulosa cells. These findings raise some important new questions regarding the role(s) of the gonadotrophins in early follicular development.

In both intact and castrated male and female rats, administration of the A-ring reduced androgen, dihydrotestosterone (DHT), consistently failed to stimulate prolactin (PRL) secretion although it inhibited LH release and, in males, stimulated ventral prostate growth. In intact females, but not in the other types of rat, DHT actually suppressed PRL release. These findings do not support generalizations, based entirely on findings with testosterone, that both "androgens" and estrogens exert stimulatory actions on PRL secretion. The distinct stimulatory effects of testosterone and its esters on PRL secretion seem attributable, not to their androgenic actions per se, but to the ability of testosterone to form estrogenic metabolites. This ability does not appear to be shared by the "pure" androgen, DHT.

In studies designed to immunocytochemically map the distribution of endogenous pituitary FSH and LH in preovulatory follicles in the rat, we discovered that both gonadotropins were detectable in both primordial and primary follicles. These findings were explored and the results are reported here.

Publisher Summary This chapter presents a line of evidence that for at least one anterior pituitary hormone, feedback control is not only quantitative but also qualitative. This means that the target gland—in this case, the gonad in its “back-talk” to the pituitary or brain-pituitary system—can dictate not only the amount of tropic hormone to be secreted but also the kind of tropic hormone. The chapter presents an evidence that in a rat, there is a set of changes in the biological and chemical characteristics of the FSH secreted by the pituitary that seems to bear no fixed relationship to either the degree or direction of any concomitant quantitative changes in FSH and LH secretory rates. These qualitative changes can be induced rapidly by alterations in the gonadal steroid environment to which the pituitary or brain-pituitary system is exposed. Furthermore, they can occur within a physiological range of circulating sex steroid levels. In the absence of gonadal steroids, the pituitary of the rat synthesizes, stores, and releases a type of FSH that is called “neuter-FSH.” In response to androgen replacement, both stored and circulating forms of FSH change in three ways. The pituitary begins to secrete a different hormone, “andro-FSH,” for which the index of discrimination, apparent molecular size, and capacity to survive in the circulation all are greater than they are for “neuter-FSH.” Ovarian steroid(s) appear to have an opposite effect. The index of discrimination for the FSH in female rat pituitaries is low, and the apparent molecular size of this FSH is even less than that of “neuter-FSH.” It seems reasonable to speculate that the female rat releases a relatively short-lived “gyno-FSH” into the circulation. The chapter also discusses the discovery of the types of FSH heterogeneity.

The question of whether endogenous prolactin might be detectable in normally lactating rat mammary glands (LMG) was examined by light microscopic immunohistochemistry. In an initial study, sections of LMG were incubated sequentially with I) rabbit anti-rat prolactin (APRL); II) anti-rabbit gamma globulin: peroxidase conjugate (ARgammaG-per), prepared from goat anti-rabbit gamma globulin and horseradish peroxidase, and III) 3,3'-diaminobenzidine mixed with H2O2(DAB). Use of this three-step procedure led to staining of stromal, parenchymal and intraluminal elements. However, control procedures which ommitted step I produced similar staining, caused by direct binding of the ARgammaG-per to endogenous tissue components (possibly rat immunoglobulins). This precluded detection of indirect, APRL-linked, binding of ARgammaG-per. In subsequent experiments, direct binding of ARgammaG-per was prevented by preincubating LMG sections with goat anti-monkey gamma globulin (AMgammaG), following which, the three-step reaction sequence (APRL, ARgammaG-per and DAB) led to APRL-dependent staining, both within the alveolar lumina and in the apices of epithelial cells. This staining, presumably of endogenous prolactin, appeared to be specific, since it did not occur when absorbed APRL (APRL admixed with highly purified rat prolactin) was substituted for APRL in step I of the reaction sequence. In contrast, when an incubation with purified rat prolactin was interposed between the AMgammaG pretreatment and the three-step reaction sequence, staining of epithelial elements was intensified, indicating that these cells had contained unoccupied prolactin-binding sites. The apparent presence of both prolactin-binding sites and endogenous immunoreactive prolactin in LMG alveolar cells strongly suggests that this protein hormone can enter these target cells under physiological conditions.

Is the entry hypothesis compatible with all the existing data about "the" second messenger for prolactin listed in Section II? All of these messengers, in some way either participate in, or modify, prolactin's actions or, in an end point-dependent manner, may actually mimic prolactin. There remains considerable uncertainty as to whether these findings reflect phenomena, some independent of and others quite dependent upon entry, on the one hand, or merely portions of a relatively large number of molecular cascades, some (but not necessarily all) begun initially at the plasmalemma and many (if not all) orchestrated toward completion by intracellular prolactin or agonist-receptor complex.