Maturation of hypothalamic-pituitary-gonadal function in normal human fetuses: circulating levels of gonadotropins, their common alpha-subunit and free testosterone, and discrepancy between immunological and biological activities of circulating follicle-stimulating hormone.

The recent availability of both cordocentesis, a low risk and effective technique for fetal blood sampling, and ultrasensitive/highly specific two-site immunofluorometric assays (IFMA) for pituitary and chorionic glycoprotein hormone (I-LH, I-FSH, and I-CG) measurement prompted us to study the maturation of hypothalamic-pituitary-gonadal function in 114 normal human fetuses (49 females and 65 males) from 17-40 weeks gestation. The subjects were selected from 216 consecutive cordocenteses carried out for rapid karyotyping and diagnosis of fetal infection or hematological disorders. In addition, FSH bioactivity (B-FSH) was measured by rat Sertoli cell aromatase induction assay, glycoprotein hormone alpha-subunit (alpha-SU) by RIA, and circulating free testosterone (fT) by direct analog technique. No significant cross-reactions were recorded in the different measurement methods. In particular, alpha-SU did not interfere in any IFMA, and CG cross-reactivity in LH IFMA was 0.5%. Circulating I-LH, I-FSH, and B-FSH levels at 17-24 weeks gestation were significantly higher in female than in male fetuses (I-LH, 48 +/- 4 vs. 6.3 +/- 0.7 U/L; I-FSH, 35 +/- 2 vs. 0.7 +/- 0.1 U/L; B-FSH, 131 +/- 17 vs. 43.4 +/- 5.4 U/L). During the last weeks of gestation, a significant decrease in I-LH and I-FSH levels was seen in both female and male fetuses (I-LH, 0.24 +/- 0.05 and 1.0 +/- 0.3 U/L; I-FSH, 0.45 +/- 0.1 and 0.5 +/- 0.1 U/L), while serum B-FSH remained elevated, but the previously recorded difference between sexes disappeared (54.3 +/- 7.2 and 58.7 +/- 7.3 U/L). Circulating I-CG and alpha-SU levels at midgestation were elevated in both female and male fetuses (I-CG, 117 +/- 29 and 191 +/- 44 U/L; alpha-SU, 143 +/- 16 and 105 +/- 9 micrograms/L, respectively) and decreased thereafter (I-CG, 42 +/- 9 and 26 +/- 6 U/L; alpha-SU, 60 +/- 15 and 37 +/- 6 micrograms/L). Serum fT levels at midgestation were significantly lower in females than in males (4.3 +/- 0.9 vs. 10.0 +/- 0.8 pmol/L) and increased until term, when the difference between sexes disappeared (16.2 +/- 1.8 vs. 17.6 +/- 1.6 pmol/L).(ABSTRACT TRUNCATED AT 400 WORDS)

Immunoreactivity in pituitary gonadotropes and hypothalamic vasopressinergic neurons for the monoclonal antibody INO.

The monoclonal antibody INO (inhibitor of neurite outgrowth) has been shown to bind to a complex of laminin and a heparan sulfate proteoglycan and to block the action of this complex in promoting neurite outgrowth. We now report that the same antibody binds to cytoplasmic constituents in rat adenohypophyseal gonadotropes, as well as to vasopressinergic neurons in the hypothalamus and their terminals in the neurohypophysis. INO immunoreactivity in fixed sections of pituitary does not colocalize with the immunoreactive laminin in blood vessels and glandular basement membranes, although when unfixed tissue is washed in buffer prior to fixation, the INO immunoreactivity appears in these laminin-rich structures. These observations suggest similarities between the INO hypophyseal antigen and the neurite-promoting proteoglycan complex characterized in conditioned media. Presence of this complex in specific neurosecretory cell types suggests that it is involved with specific secretory products with function yet to be determined.

The effect of ovarian steroid feedback upon radioimmunoreactive luteinizing hormone releasing hormone in the hypothalamus.

A radioimmunoassay (RIA) method for luteinizing hormone (LH) releasing hormone (RH) utilizing rabbit antiserum against synthetic (Glu1)-LH-RH coupled with human serum albumin at the N-terminus, is described. This assay system for LH-RH also cross-reacted with several LH-RH analogues or fragments, but not with pituitary trophic hormones. The assay was performed on the hypothalamic extracts of adult ovariectomized rats and female immature rats which had been treated with estradiol. The FSH and LH levels in the pituitary gland and serum of the same animals were determined by RIA. The radioimmunoreactive LH-RH content of the stalk median eminence markedly increased seven days after ovariectomy. The serum levels and the pituitary contents of FSH and LH of the same rats were also significantly augmented. In immature rats, the hypothalamic content of LH-RH, as measured by RIA, was significantly increased one hour after the injection of estradiol. The FSH and LH levels in the pituitary showed a significant rise after 7 hours.

PIP: To determine hypothalamic luteinizing hormone-releasing hormone (LH-RH) content and possible steroid feedback effects from the ovary, female rats were ovariectomized and killed 7 days later; another group was injected sc with 50 mcg estradiol benzoate and killed 1 hour or 7 hours later. The LH-RH content of the stalk median eminence of the hypothalamus as well as the levels of follicle stimulating hormone (FSH) and LH in the pituitary were measured by radioimmunoassay. In the castrated group there were significant increases in the LH-RH content of the median eminence (p less than .001) and in the LH and FSH levels in the serum and pituitary glands. Rats given the estradiol injection showed a significant increase in hypothalamic LH-RH 1 hour postinjection and in pituitary LH 7 hours postinjection.

Immunochemical staining of the rat adenohypophysis in organ culture.

Rat anterior pituitaries were cytologically studied following cultivation in organ culture, with and without the addition of hypothalamic and cortical extracts. Although five distinct cell types could be identified with classical stains in the uncultivated glands, the peroxidase-labeled antibody technique (using antibodies against STH, LTH, FSH, LH and TSH) showed that not all of the immune-specific cell types were being identified with the classical stains. This discrepancy was magnified following culture as chromophilic cells seen with classic stains decreased in number with an increase in culture time. The peroxidase technique, however, revealed that all cells remained constant in type and number regardless of time in culture. While the addition of either hypothalamic or cortical extract to the culture medium produced cytological alterations demonstrated by the classical dyes, the antibody technique showed no such alterations. Such a comparison of staining techniques emphasizes the hazards of relying solely on histological procedures to reveal the hormonal activity of the pituitary gland.