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The testis homogenate from Red bull (3-17 months of age) show specific binding of hLH and hFSH with Kd of 1.3 +/- 0.1 X 10(-10)M and 1.5 +/- 0.1 X 10(-9)M, respectively. Prepubertal bulls (3-5 3/4 months) exhibit the highest binding capacity of both LH and FSH, revealing the fact that LH and FSH receptors are present in high concentrations prior to the first rise in plasma T. The testicular T concentration was significantly higher in the four prepubertal bulls than in the four older animals in spite of the fact that circulating T is lower before puberty. Thus, local androgen effects within the testis may occur before peripheral androgenization. The prepubertal bull Leydig cell also displayed steroidogenic capacity. The ratio of biologically active (testosterone, dihydrotestosterone) to inactive androgens (androstenedione) increased as the animals approached puberty. Such qualitative changes in steroid production may be one of the factors responsible for peripheral androgenization.

Using ABP as an index of Sertoli cell secretory function, several important features of the Sertoli cell have emerged: 1. The stimulation of ABP production by FSH clearly points to the Sertoli cell as a target cell for FSH (3,4,9-16,21,24). 2. The dramatic effects of androgens on ABP production both in immature and mature rats also suggest that the Sertoli cell is a target cell for androgen (3,12,14,16,25). 3. The striking reduction in ABP production in the cryptorchid testis raises the question whether impairment of Sertoli cell function is the primary reason for the loss of germ cells that occurs in this condition (20). 4. Drugs like nitrofurazone or ethionine, or X-irradiation only slightly affect the secretory function of the Sertoli cells (ABP production), indicating that these treatments most probably have direct effects on the germ cells as well. Thus, measurement of ABP production rate is a very important tool in order to evaluate how hormones, drugs and physical injuries might affect the secretory function of the Sertoli cell. This test system might be of great use in order to study the physiology and hormonal regulation of the Sertoli cells. It might also be valuable in pharmacological and toxicological studies.

Seminiferous tubules contain a cytoplasmic androgen receptor similar to the receptors in the epididymis and ventral prostate. The presence of a cytoplasmic receptor indicates that androgens maintain spermatogenesis by a direct action on certain types of cells within the seminiferous tubule. The Sertoli cell appears to be one of the cell types containing androgen receptors and the receptor might also be present in spermatogonia, primary spermatocytes, or peritubular cells. The Sertoli cell is stimulated by FSH to produce an androgen-binding protein which may serve to increase the accumulation of androgen in the seminiferous epithelium and make it available for binding by intracellular androgen receptors. This may be a way in which FSH enhances the action of androgen on spermatogenesis. Androgens act on the Sertoli cell to increase its response to FSH. This action of androgens on the Sertoli cell results in increased production of androgen-binding protein and may enhance the production of other substances which exert trophic effects on spermatogenesis.

Testicular androgen binding protein (ABP) was purified from the epididymis of 1500 adult rabbits by the sequential use of ammonium sulphate precipitation, ion exchange chromatography on DEAE cellulose, gel filtration on Sephadex G-200, hydroxyl-apatite chromatography and preparative polyacrylamide gel electrophoresis. This procedure yielded a 1000-fold increase in specific activity compared to that of the 1500,000 x g supernatant, and the recovery of active ABP was about 3-5%. ABP is acid glycoprotein with a molecular weight of 65-68,000 daltons. Antisera to rabbit ABP raised in quinea pigs inhibit 3H-DHT binding to ABP as measured by SS-PAGE. When diluted rabbit serum containing TeBG is treated with the same dilutions of these antisera, identical binding inhibition curves are found. Thus, ABP and TeBG in rabbits appear to possess identical immunological determinants.

The Sertoli cell appears to be a target cell for both FSH and androgen. FSH stimulates the Sertoli cell to produce an androgen-binding protein (ABP) which may serve to increase the accumulation of androgen in the seminiferous tubular epithelium and make it available for binding by intracellular androgen receptors. This might be one way by which FSH enhances the action of androgen on spermatogenesis. Androgen acts on the Sertoli cell to increase its response to FSH resulting in increased production of ABP and of other substances which exert trophic effects on spermatogenesis.

The levels of testosterone, dihydrotestosterone (DHT), 5alpha-androstan-3alpha,17beta-diol and androgen-binding protein (ABP) were measured in various segments of the epididymis from adult rats which had been unilaterally orchidectomized for 4 weeks. On the 'intact' side, ABP concentrations were highest in the caput region. The segmental distribution of DHT closely followed that of ABP with the highest concentration in the caput (40 ng/g tissue) and lowest in the cauda (10 ng/g tissue) epididymidis. There was a high degree of correlation (r = 0.98) between the concentration of DHT in the epididymis and ABP levels. 'Castration' completely abolished the DHT gradient. The levels of testosterone and androstanediol were lower than those of DHT; most was present in the corpus epididymidis. The relative differences were reduced after 'castration'. It is concluded that ABP in the rat epididymis is the primary factor for determining the concentration of DHT in the epididymal fluid.

Changes in the proportion of cells within various DNA classes of dispersed testicular cells from the developing rat were monitored by microflow fluorometry and correlated with changes in the function of the pituitary (FSH), of the Leydig cells (androgens) and Sertoli cells (androgen-binding proteins, ABP). Peaks of androgens and of FSH appeared simultaneously and coincided with an accumulation of tetraploid cells and with the first appearance of haploid cells in the testis and ABP in the epididymis. Estrogen treatment (5 micrograms/day) of developing rats from day 7 completely prevented the appearance of haploid cells in the testis as well as ABP in the epididymis. In these animals the wave of tetraploid cells started and progressed normally, indicating that transformation and progression of germinal cells to the stage of the primary spermatocytes were taking place. A combined treatment with FSH and dihydrotestosterone propionate (DHTP) resulted in a premature start of Sertoli cell secretion of ABP into the epididymis, but in a normal appearance of haploid and tetraploid cells. The time correlation between peaks in FSH/androgens, the start of Sertoli cell secretion, and the occurrence of haploid cells in the testis stresses the importance of these two hormones for normal Sertoli cell function and the importance of a functional Sertoli cell for the completion of meiosis.

The seasonal changes in testicular weight in the blue fox were associated with considerable variations in plasma concentrations of LH, prolactin, androstenedione and testosterone and in FSH-binding capacity of the testis. An increase in LH secretion and a 5-fold increase in FSH-binding capacity were observed during December and January, as testis weight increased rapidly. LH levels fell during March when testicular weight was maximal. Plasma androgen concentrations reached their peak values in the second half of March (androstenedione: 0.9 +/- 0.1 ng/ml: testosterone: 3.6 +/- 0.6 ng/ml). A small temporary increase in LH was seen in May and June after the breeding season as testicular weight declined rapidly before levels returned to the basal state (0.5-7 ng/ml) that lasted until December. There were clear seasonal variations in the androgenic response of the testis to LH challenge. Plasma prolactin concentrations (2-3 ng/ml) were basal from August until the end of March when levels rose steadily to reach peak values (up to 13 ng/ml) in May and June just before maximum daylength and temperature. The circannual variations in plasma prolactin after castration were indistinguishable from those in intact animals, but LH concentrations were higher than normal for at least 1 year after castration.

The specific androgen receptors for testosterone (T) and 5alpha-dihydrotestosterone (DHT) in the cytosol fraction of the anterior pituitary of rats have been further characterized using electrophoresis and isoelectric focusing in polyacrylamide gels. After labeling of the cytosol fraction in vivo and in vitro, we were able to demonstrate androgen-protein complexes moving with an electrophoretic mobility (Rf) of 0.5 in 3.25% acrylamide gels containing 0.5% agarose and 10% glycerol. This method was used for quantitative measurements of pituitary androgen receptors, allowing multiple samples to be run simultaneously with little or no non-specific binding. There was no measurable dissociation of the androgen-receptor complexes during electrophoresis. When radioactive testosterone (1 nM) was added to pituitary cytosol fractions in vitro, there was an increase in the binding up to 4 hours of incubation at 0 C and little or no increase between 4 and 24 hours. All the binding studies therefore were done by incubation overnight at 0 C. When cytosol fractions were incubated with increasing concentrations of radioactive testosterone, a typical saturation curve was found. Scatchard plot analysis showed a binding capacity of 12.0 femtomoles/mg protein and the equilibrium constant of dissociation was estimated to be 3.4 +/- 0.7 (SD) X 10(-10)M. Like other androgen-receptor complexes, the testosterone-receptor complex in the anterior pituitary gland had an extremely slow rate of dissociation at 0 C (t1/2 greater than 4 days). The steroid specificity of the cytoplasmic androgen receptors was tested in vitro by the competing efficiency of different unlabeled steroids for [3H] testosterone binding. T and DHT caused the same inhibition of [3H]T to the receptors. However, since metabolism, of DHT to 5alpha-androstane-3alpha,17beta-diol occurred even at 0 C, the affinity of DHT for the receptor is probably somewhat underestimated. Cyproterone acetate had approximately half the affinity for the receptor compared with T, whereas lower affinities were found for progesterone and 17beta-estradiol. Cortisol did not appear to have any affinity for the receptors. Isoelectric focusing in polyacrylamide gels showed a peak of bound radioactivity with an isoelectric point of 5.8. Thus, the characteristics of the cytoplasmic androgen receptors of the anterior pituitary gland are very similar to those of the androgen receptors described in the ventral prostate, epididymis, and testis.