Summary. The responsiveness of decapsulated testes and isolated Leydig cell preparations from rats (30–80 days of age) to a constant dose of 3 ng hCG/2 ml was assessed by comparison of the production of testosterone and 'total 17β-hydroxy androgen' (17β-HA). When testosterone secretion was used as the index of response, there was a marked increase in the production with age by decapsulated testes and also by equal numbers of Leydig cells. When 17β-HA was taken as the response parameter this increase was only marginal for the decapsulated testes and there was an age-dependent decrease when expressed per 106 cells. These differences probably reflect changes in the metabolism of testosterone to 5α-reduced products with increasing age because 80% of androgen secreted at 30 days is 3α-androstanediol and 86% is secreted as testosterone at 80 days. We conclude that for studies on hCG responsiveness and the steroidogenic capacity of immature rat Leydig cells (a) testosterone is an inappropriate response parameter and (b) this response undergoes a decrease rather than an increase during prepubertal development.
K. Purvis, O. P. F. Clausen and V. Hansson
K. Purvis, O. P. F. Clausen and V. Hansson
Summary. Treatment of immature, hypophysectomized male rats with 50 μg ovine FSH (NIH-FSH-S12) twice a day for 5 days stimulated the maximum quantity of 17β-hydroxyandrogen produced by isolated Leydig cells in response to hCG. Pretreatment of the FSH preparation with an LH antiserum in one study markedly reduced and in another study completely abolished this stimulatory effect of FSH, but only slightly impaired the capacity of the hormone to stimulate the Sertoli cell in vivo (epididymal androgen-binding protein). Administration of another highly potent FSH preparation (LER-1881) had no discernible effects on the dose–response characteristics of the Leydig cells but was superior to the NIH-FSH-S12 in its capacity for stimulating the Sertoli cell. When all hormone preparations were tested for their ability to stimulate steroid secretion from normal Leydig cells in vitro, a close correlation was obtained between their Leydig cell-stimulating activity (a measure of LH contamination) and their capacity to alter Leydig cell responsiveness after in-vivo treatment. FSH treatment had no effects on specific LH binding per 106 Leydig cells. It is concluded that the stimulatory influence of FSH on rat Leydig cells may to some extent be a result of the LH contaminating the hormone preparation.
O. P. F. Clausen, K. Purvis and V. Hansson
Summary. The technique of microflow fluorometry (MFF) was used to identify the proportion of haploid cells (from the tubules) in interstitial cell suspensions. The MFF estimates of the degree of contamination by tubular elements correlated well with the numbers of cells with Leydig cell morphology and those staining positively for 3β-hydroxysteroid dehydrogenase.
K. Purvis, O. P. F. Clausen and V. Hansson
Summary. The response of rat Leydig cells to hCG in vitro was examined using decapsulated testes and enriched Leydig cell suspensions of a known purity. The secretion of testosterone and 5α-androstan-3α,17β-diol was used as the response and medium extracts were assayed before and after celite chromatography. At least 4 important phases in the development of steroidogenic function of the Leydig cells were detected. (1) A period after birth up to 10 days of age when the Leydig cell preferentially secretes testosterone. The cells may constitute the remnants of a fetal Leydig cell population. (2) A period which begins between 10 and 15 days of age and continues until 35–40 days of age, when the major products of the Leydig cell are 5α-reduced metabolites of testosterone, especially 5α-androstan-3α,17β-diol. (3) A phase at 20–30 days of age when responsiveness of the Leydig cell to hCG in vitro undergoes a distinct increase (the quantity of androgen produced with maximum hCG stimulation) and attains adult levels, although the sensitivity of the Leydig cells to hCG (the dose of hCG eliciting a half-maximal response, ED50) is gradually decreased after Day 20. (4) A period from 35–40 days of age into adulthood when testosterone increasingly becomes the major secretory product of the Leydig cell.
Hypophysectomy for 8 days decreased the quantity of androgens produced per 106 Leydig cells in response to hCG to an extent dependent on age. At 28 days of age steroidogenic function was impaired to a much greater extent (reduced by some 80–90%) than at 38 or 58 days (reduced by 40–50%). At all ages the sensitivity of the Leydig cells to hCG was increased after hypophysectomy.
A. J. Smith, O. P. F. Clausen, B. Kirkhus, T. Jahnsen, O. M. Møller and V. Hansson
Summary. The testes of the blue fox (Alopex lagopus) showed marked seasonal variations in size. Testicular weight and volume increased rapidly during January and February to reach maximal values by the beginning of the breeding season (~15 March). During May and June the weights and volumes of the testes declined gradually to the quiescent state which lasted from July until October. Quantitation by DNA flow cytometry of the seasonal changes in the relative numbers of haploid (1C), diploid (2C) and tetraploid (4C) cell numbers in the testis showed that the increase in testis size from December to February was associated with a rapid expansion of the haploid cell compartment as spermatogenesis resumed. In addition, an increase in number of more mature cell types within the haploid cell population was observed over a 2-month period before the breeding season. The decline in testicular size from the middle of April until October was associated with a reduction in both the absolute and relative sizes of the haploid and tetraploid cell populations and a concomitant increase in the relative numbers of diploid cells. Measurements of the activity of the soluble Mn2 + -dependent adenylate cyclase revealed seasonal variations that closely paralleled those of the haploid cell population, indicating that, as in other species, the enzyme may be associated with maturing germ cells.
A. J. Smith, M. Mondain-Monval, P. Simon, K. Andersen Berg, O. P. F. Clausen, P. O. Hofmo and R. Scholler
Summary. Bromocriptine administration in the form of slow-release injections to male blue foxes during March–May abolished the normal spring rise in plasma prolactin concentrations seen in May and June. The spring moult was prevented and the treated animals retained a winter coat of varied quality and maturity until the end of the study in August.
Plasma testosterone concentrations fell normally from March until August. Testicular regression was, however, delayed, although there were individual variations in response. Estimation by DNA flow cytometry in early July of the relative numbers of haploid, diploid and tetraploid cells in the testis showed that, in the treated animals, 74–80% of the cells were haploid (maturing germinal cells), 4–6% tetraploid (mainly primary spermatocytes) and the rest diploid cells (somatic cells and the remaining germinal cell types). In the control males, however, no haploid cells were detected and the majority of cells were diploid (93–99%). At castration in August, histological examination revealed various stages of testicular regression in the treated and control animals.
N. B. Oldereid, H. Rui, O. P. F. Clausen and K. Purvis
Summary. The sperm qualities of 350 men under fertility investigation were compared in relation to their smoking habits. The sperm variables included number, motility, morphology and vitality. Sperm motility was assessed objectively by laser–Doppler spectroscopy. In a randomly selected group, sperm samples were subjected to flow cytometry to assess the levels of DNA condensation. No significant differences (Kruskal–Wallis' test) in any aspect of sperm quality including DNA distribution could be demonstrated between non-smokers, moderate smokers (1–14 cigarettes/day) and heavy smokers (15–40 cigarettes/day). This was true when the data were pooled and when oligozoospermic/hypozoospermic ejaculates (1–39 × 106/ml) and asthenozoospermic ejaculates (<25% of sperm cells with progressive movement) were analysed separately. The distribution of non-smokers, moderate and heavy smokers was the same in groups of men with normal sperm quality as those with impaired quality. The present study does not provide support for the contention that smoking has deleterious effects on sperm quality, at least using conventional parameters.
Keywords: cigarette smoking; sperm quality; human infertility; flow cytometry; laser–Doppler spectroscopy
A. J. Smith, M. Mondain-Monval, K. Andersen Berg, J. O. Gordeladze, O. P. F. Clausen, P. Simon and R. Scholler
Summary. Testicular weight in young male blue foxes increased steadily from 12 weeks of age (0·4–0·7 g) to reach peak values at the time of the mating season in March–April (5·2–6·6 g), before declining rapidly during May to low values in August at 63 weeks of age (1·3–1 ·6 g). Primary spermatocytes were found in the spermatogenic epithelium at 20 weeks of age and by late December (29 weeks of age) elongated spermatids were seen. There was a good correlation between the seasonal variations in the presence of germ cell types assessed by quantitative analysis of testicular histology and the variations in numbers of haploid, diploid and tetraploid cells measured by DNA flow cytometry: no haploid cells were found before the end of November and peak numbers were observed in March.
Plasma FSH concentrations were increased from December onwards (with the exception of April). There were no clearcut seasonal variations in plasma LH concentrations although values were consistently lower in April. Testosterone concentrations were low for most of the year but increased from the end of January to the middle of April. There was no detectable seasonal variation in LH release in response to LHRH injection, and no typical pattern in plasma FSH concentrations during the first 100 min after injection. Plasma testosterone concentrations after LHRH injection rose gradually during testicular development.
There were large seasonal variations in soluble Mn2+-dependent adenylate cyclase activity in the testis, that paralleled the changes in testicular weight and haploid cell content. Values were low until December and reached a peak at the time of the mating season before falling to basal levels again by June.
The results suggest that immature male blue foxes reach full testicular development (indistinguishable from that of older animals) by the first mating season after birth, at an age of about 40 weeks.
Nicolet H. P. M. Jutte, R. Jansen, J. A. Grootegoed, F. F. G. Rommerts, O. P. F. Clausen and H. J. van der Molen
Summary. During incubation of fragments of seminiferous tubules in the absence of glucose, pachytene spermatocytes and round spermatids died within 24 h, while Sertoli cells were still viable. The germ cells survived for at least 72 h in seminiferous tubule fragments which were incubated in the presence of glucose. Lactate rather than glucose is essential for [3H]uridine incorporation and survival of isolated pachytene spermatocytes. However, if the spermatocytes were incubated in the presence of Sertoli cells, glucose maintained the incorporation of [3H]uridine into the germ cells. Sertoli cells secreted lactate in the presence of glucose and the lactate secretion was stimulated 2–4-fold by FSH. It is concluded that the activity and survival of pachytene spermatocytes in vitro can be regulated by the supply of lactate from Sertoli cells.
A. J. Smith, M. Mondain-Monval, K. Andersen Berg, P. Simon, M. Forsberg, O. P. F. Clausen, T. Hansen, O. M. Møller and R. Scholler
Summary. Melatonin administration to male blue foxes from August for 1 year resulted in profound changes in the testicular and furring cycles. The control animals underwent 5-fold seasonal changes in testicular volume, with maximal values in March and lowest volumes in August. In contrast, melatonin treatment allowed normal redevelopment of the testes and growth of the winter coat during the autumn but prevented testicular regression and the moult to a summer coat the following spring. At castration in August, 88% of the tubular sections in the testes of the controls contained spermatogonia as the only germinal cell type, whereas in the treated animals 56–79% of sections contained spermatids or even spermatozoa. Semen collection from a treated male in early August produced spermatozoa with normal density and motility.
Measurement of plasma prolactin concentrations revealed that the spring rise in plasma prolactin values (from basal levels of 1·6–5·4 ng/ml to peak values of 4·1–18·3 ng/ml) was prevented; values in the treated animals ranged during the year from 1·8 to 6·3 ng/ml. Individual variations in plasma LH concentrations masked any seasonal variations in LH release in response to LHRH stimulation, but the testosterone response to LH release after LHRH stimulation was significantly higher after the mating season in the treated animals, indicating that testicular testosterone production was maintained longer than in the controls.
The treated animals retained a winter coat, of varied quality and maturity, until the end of the study in August.