K. Purvis

Prolactin and Leydig Cell Responsiveness to LH/hCG in the Rat

The effects of prolactin and 2-bromo-alpha-ergocryptine (CB-154) on Leydig cell function in intact and hypophysectomized male rats were studied. The conclusions can be summarized as follows: prolactin (1) has a direct stimulatory effect on the number of LH receptors on rat Leydig cells, (2) has no effect on the characteristics of the dose-response curve of isolated Leydig cells (hCG stimulated androgen production) in vitro even after treatment with pharmacological doses in vivo, and (3) acts synergistically with LH to stimulate the quantity of androgen produced by the Leydig cells in response to hCG in vitro and to increase the sensitivity of the hCG-dose-response curve. Treatment of intact rats with CB-154 reduced the quantity of androgen produced by the Leydig cells in vitro after exposure to hCG and decreased LH binding to the same cells by 50%. These results suggest that under normal conditions, endogenous prolactin plays a key role in maintaining the functional integrity of rat Leydig cells.

Endocrine Correlates of Meiosis in the Male Rat

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.

Temporal Relationship Between hCG Induced Desensitization of LH/hCG Responsive Adenylyl Cyclase and Down regulation of LH/hCG Receptors in the Rat Testis

A single s.c. injection of 75 IU of human chorionic gonadotropin (hCG) into adult male rats caused a transient desensitization of the LH/hCG responsive adenylyl cyclase (AC) in membrane particles from rat testis. Two hours after the injection of hCG, LH/hCG responsive AC was reduced by 40%, whereas LH binding was still normal. After 24 hr LH/HCG responsive AC was lost, whereas LH binding only showed a marginal decrease. During the next 2-3 days there was a gradual loss of LH/HCG receptors, which reached a nadir at day 4 (14% of control levels). At day y and 9 following the hCG injection LH/hCG receptors and LH/hCG responsive AC gradually returned towards normal. Fluoride (F-) stimulated AC activity showed only minor changes throughout the period investigated. The initial loss of AC responsiveness to hCG was associated with maximal levels of circulating hCG. However, desensitization persisted for several days after hCG was cleared form the circulation. AC desensitization preceded LH receptor downregulation and may support the notion that uncoupling of the catalytic subunit of the AC from Leydig cell LH/hCG receptors is a requirement for subsequent receptor internalization.

LH Receptors and Leydig Cell Responsiveness to hCG in vitro

The in vitro responsiveness of isolated Leydig cells from 30-day-old rats to hCG was assessed in terms of androgen and cAMP production, under conditions where the quantity of LH/hCG receptors on the Leydig cell membranes was reduced to varying degrees. Reduction in the receptor population was achieved by a single, in vivo injection of 75 IU hCG 3 and 5 days prior to incubation. When receptor binding was reduced by approximately 30%, neither the sensitivity (dose of hCG needed to give a half maximum response) nor the magnitude of the Leydig cell response to hCG in terms of androgen production was affected. However, a reduction in [125I]-hLH receptor binding by 80% was associated with a decrease in the in vitro sensitivity of Leydig cells to hCG, resulting in a displacement of dose-response curve to the right. Maximal quantity of androgen produced was not reduced. In the case of cAMP production, the same reduction in LH binding was associated with a decrease in both sensitivity and magnitude of the maximal ...

Testicular Cyclic Nucleotide Phosphodiesterase in the Rat. Kinetic Properties and Changes with Age

The assay for cyclic nucleotide phosphodiesterase has been applied, with certain modifications, to the measurement of the soluble forms of these enzymes in the rat testis. The homogenization and incubation conditions were adjusted to achieve linear product formation as a function of time and protein concentrations and the resulting products were isolated by ion exchange chromatography using 5 mM HCl as the eluting agent. Phosphodiesterase activities were present in the testicular cytosol (105,000 g supernatant) of adult rats which were capable of hydrolyzing cAMP with a high (Km2 microM) and low Km20 microM) affinity and GMP with a relatively high affinity (Km3 microM). The low affinity cAMP enzyme activity could be stimulated with divalent ions such as calcium, magnesium, and manganese. At 18 days of age, all three enzyme activities were present in the testis, although both the high and low affinity cAMP phosphodiesterase displayed maximal rates (Vmax) that were only one third of the adult testis (when expressed per mg protein).