Effect of Typha capensis (Rohrb.)N.E.Br. rhizome extract F1 fraction on cell viability, apoptosis induction and testosterone production in TM3-Leydig cells.

Typha capensis (Rohrb.)N.E.Br. (bulrush) is used by traditional healers in Southern Africa to treat male reproductive problems. This study aimed at investigating the effects of T. capensis on TM3-Leydig cells. T. capensis rhizome crude extract obtained from autumn, winter, spring and summer harvest was fractionated using HPLC into four fractions, and TM3-Leydig cells were incubated with different concentrations of the F1 fraction (0.01, 0.02, 0.1, 1, 10 and 100 µg/ml) for 24, 48 and 96 hr respectively. The following parameters were evaluated: cell morphology, viability (MTT assay), testosterone production (testosterone ELISA test), apoptosis (Annexin V-Cy3 binding) and DNA fragmentation (TUNEL assay). Results revealed that the summer harvest obtained the highest amount of extract. The F1 fraction of all harvests was the most effective. This fraction significantly enhanced testosterone production in TM3 cells in a dose-dependent manner with maximum effect at 0.1 µg/ml. At higher concentrations, lower testosterone production was observed. Cell viability including apoptosis was not affected at concentrations used by the traditional healers to treat patients. This study shows that T. capensis enhanced testosterone production and might be useful to treat male infertility and ageing male problems.

Effects of selenium on the proliferation, apoptosis and testosterone production of sheep Leydig cells in vitro.

The objective of this study was to investigate the effects of selenium (Se) on in vitro proliferation, apoptosis and testosterone production of sheep Leydig cells and its underlying mechanism. Leydig cells were collected from 8-month-old sheep and divided into four treatment groups (0, 2.0, 4.0 and 8.0 µmol/L Se). After treatment with Se for 48 h, the MTT and flow cytometric assay were used to detect cell proliferation and apoptosis. Testosterone level in the culture medium was determined by ELISA. The mRNA expression and protein abundance of cell cycle, apoptosis and testosterone synthesis-related genes were detected using real-time PCR and western blot analysis. The results showed that the highest percentage of live and apoptotic cells was obtained in the 2.0 and 8.0 µmol/L group, respectively. In the Se treatment groups, the proliferation rate of Leydig cells and the expression of cell cycle-related genes were decreased with the increasing Se supplementation in the culture medium. The percentage of apoptotic cells was increased with the increasing Se level, which was consistent with the expression of pro-apoptosis genes. The highest GSH-Px activity and lowest ROS content were also observed in the 2.0 µmol/L group. Appropriate Se level (2.0 µmol/L) can significantly increase the expression of p-ERK1/2, StAR and 3ß-HSD, and improve the testosterone synthesis. Compared with the control group, PD0325901 could significantly inhibit the production of testosterone and the protein abundance of p-ERK1/2, StAR and 3ß-HSD. Se treatment can mitigate the inhibition effect of PD0325901 and the testosterone secretion between the 2.0 µmol/L and control group was not significantly different. These results demonstrate that Se can affect the proliferation and apoptosis of Leydig cells by regulating cellular oxidative stress and the expressions of cell cycle and apoptosis-related genes. Se can also enhance the testosterone production of Leydig cells by activating the ERK signaling pathway and the expression of its downstream genes (StAR and 3ß-HSD), which could be closely related to the regulating roles of Se in male fertility and spermatogenesis.

Dehydroepiandrosterone inhibits cell proliferation and improves viability by regulating S phase and mitochondrial permeability in primary rat Leydig cells.

Dehydroepiandrosterone (DHEA) is widely used as a nutritional supplement and exhibits putative anti­aging properties. However, the molecular basis of the actions of DHEA, particularly on the biological characteristics of target cells, remain unclear. The aim of the current study was to investigate the effects of DHEA on cell viability, cell proliferation, cell cycle and mitochondrial function in primary rat Leydig cells. Adult Leydig cells were purified by Percoll gradient centrifugation, and cell proliferation was detected using a Click-iT® EdU Assay kit and cell cycle assessment performed using flow cytometry. Mitochondrial membrane potential was detected using JC-1 staining assay. The results of the current study demonstrate that DHEA decreased cell proliferation in a dose­dependent manner, whereas it improved cell viability in a time­dependent and dose­dependent manner. Flow cytometry analysis demonstrated that DHEA treatment increased the S phase cell population and decreased the G2/M cell population. Cyclin A and CDK2 mRNA levels were decreased in primary rat Leydig cells following DHEA treatment. DHEA treatment decreased the transmembrane electrical gradient in primary Leydig cells, whereas treatment significantly increased succinate dehydrogenase activity. These results indicated that DHEA inhibits primary rat Leydig cell proliferation by decreasing cyclin mRNA level, whereas it improves cells viability by modulating the permeability of the mitochondrial membrane and succinate dehydrogenase activity. These findings may demonstrate an important molecular mechanism by which DHEA activity is mediated.

The effect of estrogen supplementation on cell proliferation and expression of c-kit positive cells in the rat prostate.

BACKGROUND: Benign prostatic hyperplasia (BPH) is associated with the proliferation of prostate tissue and an increase in smooth muscle tone. However, the way in which the hormonal environment affects cell proliferation and prostatic interstitial cells (PIC) responsible for the maintenance of the smooth muscle tone is not clear. The present study investigated the effect of estrogen supplementation on cell proliferation, androgen/estrogen ratio, and expression and/or distribution of PIC. METHODS: Male Sprague-Dawley rats were anesthetized with isoflurane/oxygen breathing mixture and subcutaneously implanted with silastic capsules. These capsules were either empty or filled with a 10 or 20 mg of crystalline estrogen. RESULTS: Estrogen exerted a potent effect on ventral prostate weight, which was manifested as a significant decrease between controls and the E(10)- and E(20)-treated rats. Active cell proliferation detected as Ki67-positive nuclei was observed in the stromal and epithelial cells of the ventral prostatic lobes from estrogen-treated rats and controls. Estrogen supplementation caused a significant and dose-dependent increase in prostatic estradiol and 5alpha-dihydrotestosterone (DHT) concentration but the ratios of either DHT/E(2) or E(2)/DHT were not significantly affected. PIC were observed in the region between the fibromuscular stroma and the glandular epithelium in all three experimental groups. E(20)-treated rats showed a higher expression of PIC than controls and E(10)-treated rats. CONCLUSIONS: The present study provides novel information regarding the synergistic role of estrogens and androgens in the prostate: estrogen may prevent prostatic hyperplasia via mechanisms other than affecting cell proliferation or DHT/estrogen ratio.

The effects of palm kernel cake based diet on spermatogenesis in Malin x Santa-Ines rams.

Testes from nine male Malin x Santa-Ines rams with an average body weight of 43.1+/-3.53 kg, were used to study the effects of palm kernel cake (PKC) based diet on spermatogenic cells and to assess copper (Cu) levels in liver, testis and plasma in sheep. Animals were divided into three groups and randomly assigned three dietary treatments using restricted randomization of body weight in completely randomized design. The dietary treatments were 60% palm kernel cake plus 40% oil palm frond (PKC), 60% palm kernel cake plus 40% oil palm frond supplemented with 23 mg/kg dry matter of molybdenum as ammonium molybdate [(NH(4))(6)Mo(7)O(24).4H(2)O] and 600 mg/kg dry matter of sulphate as sodium sulphate [Na(2)SO(4)] (PKC-MS) and 60% concentrate of corn-soybean mix+40% oil palm frond (Control), the concentrate was mixed in a ratio of 79% corn, 20% soybean meal and 1% standard mineral mix. The results obtained showed that the number of spermatogonia, spermatocytes, spermatids and Leydig cells were not significantly different among the three treatment groups. However, spermatozoa, Sertoli cells and degenerated cells showed significant changes, which, may be probably due to the Cu content in PKC. Liver and testis Cu levels in the rams under PKC diet was found to be significantly higher (P<0.05) than rams in Control and PKC-MS diets. Plasma Cu levels showed a significant increase (P<0.05) at the end of the experiment as compared to at the beginning of the experiment for PKC and Control. In conclusion, spermatogenesis is normal in rams fed the diet without PKC and PKC supplemented with Mo and S. However spermatogenesis was altered in the PKC based diet probably due to the toxic effects of Cu and the significant changes in organs and plasma. Thus, Mo and S play a major role in reducing the accumulation of Cu in organs.

Prenatal induced chronic dietary hypothyroidism delays but does not block adult-type Leydig cell development.

Transient hypothyroidism induced by propyl-2-thiouracyl blocks postpartum Leydig cell development. In the present study, the effects of chronic hypothyroidism on the formation of this adult-type Leydig cell population were investigated, using a more physiological approach. Before mating, dams were put on a diet consisting of an iodide-poor feed supplemented with a low dose of perchlorate and, with their offspring, were kept on this diet until death. In the pups at day 12 postpartum, plasma thyroid-stimulating hormone levels were increased by 20-fold, whereas thyroxine and free tri-iodothyronine levels were severely depressed, confirming a hypothyroid condition. Adult-type progenitor Leydig cell formation and proliferation were reduced by 40-60% on days 16 and 28 postpartum. This was followed by increased Leydig cell proliferation at later ages, suggesting a possible slower developmental onset of the adult-type Leydig cell population under hypothyroid conditions. Testosterone levels were increased 2- to 10-fold in the hypothyroid animals between days 21 and 42 postpartum compared with the age-matched controls. Combined with the decreased presence of 5alpha-reductase, this implicates a lower production capacity of 5alpha-reduced androgens. In 84-day-old rats, after correction for body weight-to-testis weight ratio, plasma insulin-like factor-3 levels were 35% lower in the hypothyroid animals, suggestive of a reduced Leydig cell population. This is confirmed by a 37% reduction in the Sertoli cell-to-Leydig cell ratio in hypothyroid rats. In conclusion, we show that dietary-induced hypothyroidism delays but, unlike propyl-2-thiouracyl, does not block the development of the adult-type Leydig cell population.

Reversible changes in the antifertility induced by Aegle marmelos in male albino rats.

To study the effects of Aegle marmelos on the testicular reproductive system, a 50% ethanolic extract of Aegle marmelos leaves (AMLEt) was fed orally to male albino rats at the dose levels of 200 and 300 mg/kg body wt./day for 60 days. Recovery was assessed for an additional 120 days. Oral administration of AMLEt did not cause body weight loss. The motility and sperm concentration were significantly reduced along with complete inhibition of fertility at a dose of 300 mg/kg. The level of serum testosterone also declined and spermatogenesis was impaired. The number of normal tubules and the height of epithelial cells of the caput and cauda were reduced significantly. The cross sectional surface area of Sertoli cells and mature Leydig cells was reduced along with a dose dependent reduction of preleptotene and pachytene spermatocytes. Thus the antifertility effects of Aegle marmelos seemed to be mediated by disturbances in structure and function in testicular somatic cells including Leydig and Sertoli cells resulting in an alteration in physio-morphological events of spermatogenesis. However, complete recovery was observed after a 120 day withdrawal.

The inhibitory effects on adult male reproductive functions of crude garlic (Allium sativum) feeding.

AIM: to investigate the effects of crude garlic on adult male rat reproductive functions. METHODS: Thirty male rats were divided into five groups: group 1 (untreated) and groups 2, 3, 4 and 5 were fed for 30 days with 5%, 10%, 15% and 30% crude garlic, respectively. Testes and accessory organs were weighed and some markers were assessed. Light and electron microscopy observations were also performed. RESULTS: A significant decrease was observed in the body weight of groups 4 (14%; P < 0.01) and 5 (20%; P < 0.01); of the prostate weight in group 5 (29.1%; P < 0.05) and of seminal vesicle weight in groups 3 (14.4%; P < 0.01), 4 (18.3%; P < 0.01) and 5 (27.3%; P < 0.01). In contrast, testis and epididymis weights were unchanged. In epididymis tissue, the alpha glucosidase activity and the spermatozoa density were unchanged. The treatment resulted in a significant decrease in testosterone serum levels in groups 3 (77.3%; P < 0.01), 4 (77.3%; P < 0.01) and 5 (90.9%; P < 0.01), associated with a significant increase in LH serum levels (P < 0.01). Testicular histology showed a dose-dependent increase in the percentage of empty seminiferous tubules. Moreover, testicular function was affected; a significant decrease in phosphatase acid activity (P < 0.01) and testosterone (P < 0.05) contents were observed. CONCLUSION: Crude garlic consumption during 1 month reduced testosterone secretion and altered spermatogenesis at 10%, 15% and 30% doses.

Androgenic regulation of novel genes in the epididymis.

The epididymis is critically dependent on the presence of the testis. Although several hormones, such as retinoids and progestins, and factors secreted directly into the epididymal lumen, such as androgen binding protein and fibroblast growth factor, might play regulatory roles in epididymal function, testosterone (T) and its metabolites, dihydrotestosterone (DHT) and estradiol (E2), are accepted as the primary regulators of epididymal structure and functions, with the former playing the greater role. To ascertain the molecular action of androgens on the epididymis, three complementary approaches were pursued to monitor changes in gene expression in response to different hormonal milieux. The first was to establish changes in gene expression along the epididymis as androgenic support is withdrawn. The second was to determine the sequence of responses that occur in an androgen deprived tissue upon re-administration of the two metabolites of T, DHT and E2. The third was to study the effects of androgen withdrawal and re-administration on gene expression in immortalized murine caput epididymidal principal cells. Specific responses were observed under each of these conditions, with an expected major difference in the panoply of genes expressed upon hormone withdrawal and re-administration; however, some key common features were the common roles of genes in insulin like growth factor/epidermal growth factor and the relatively minor and specific effects of E2 as compared to DHT. Together, these results provide novel insights into the mechanisms of androgen regulation in epididymal principal cells.

Regulation of steroidogenesis by Cordyceps sinensis mycelium extracted fractions with (hCG) treatment in mouse Leydig cells.

The in vitro effect of extracted fractions of Cordyceps sinensis (CS) mycelium on hCG-treated testosterone production from purified normal mouse Leydig cells was examined. Different fractions extracted from CS (F1-water soluble polysaccharide, F2- water soluble protein and F3- poorly water soluble polysaccharide, and protein) were added to Leydig cells with hCG, and the production of testosterone was determined by radioimmunoassay (RIA). Testosterone productions stimulated by hCG in mouse Leydig cells were suppressed by F2 at 10 mg/ml and F3 at doses from 3 to 10 mg/ml, respectively. F2 and F3 at 10 mg/ml did inhibit dbcAMP-stimulated testosterone productions which indicated that F2 and F3 might affect steroidogenesis at the site after the formation of cyclic AMP. Finally, cycloheximide inhibited F2- and F3-treated mouse Leydig cell testosterone production.

Insights into GABA receptor signalling in TM3 Leydig cells.

Gamma-aminobutyric acid (GABA) is an emerging signalling molecule in endocrine organs, since it is produced by endocrine cells and acts via GABA(A) receptors in a paracrine/autocrine fashion. Testicular Leydig cells are producers and targets for GABA. These cells express GABA(A) receptor subunits and in the murine Leydig cell line TM3 pharmacological activation leads to increased proliferation. The signalling pathway of GABA in these cells is not known in this study. We therefore attempted to elucidate details of GABA(A) signalling in TM3 and adult mouse Leydig cells using several experimental approaches. TM3 cells not only express GABA(A )receptor subunits, but also bind the GABA agonist [(3)H]muscimol with a binding affinity in the range reported for other endocrine cells (K(d) = 2.740 +/- 0.721 nM). However, they exhibit a low B(max) value of 28.08 fmol/mg protein. Typical GABA(A) receptor-associated events, including Cl(-) currents, changes in resting membrane potential, intracellular Ca(2+) or cAMP, were not measurable with the methods employed in TM3 cells, or, as studied in part, in primary mouse Leydig cells. GABA or GABA(A) agonist isoguvacine treatment resulted in increased or decreased levels of several mRNAs, including transcription factors (c-fos, hsf-1, egr-1) and cell cycle-associated genes (Cdk2, cyclin D1). In an attempt to verify the cDNA array results and because egr-1 was recently implied in Leydig cell development, we further studied this factor. RT-PCR and Western blotting confirmed a time-dependent regulation of egr-1 in TM3. In the postnatal testis egr-1 was seen in cytoplasmic and nuclear locations of developing Leydig cells, which bear GABA(A) receptors and correspond well to TM3 cells. Thus, GABA acts via an atypical novel signalling pathway in TM3 cells. Further details of this pathway remain to be elucidated.

Goliath, a ring-H2 mitochondrial protein, regulated by luteinizing hormone/human chorionic gonadotropin in rat leydig cells.

We have cloned the rat homologue of the ring-H2 protein Goliath involved in Drosophila development. The rat Goliath mRNA (1.85 kb) was translated as a major ubiquitous protein species of 28-kDa and three larger isoforms (50, 46, and 36 kDa) expressed mainly in liver, lung, stomach, heart, and thymus and barely detectable in other tissues (kidney, skeletal muscle, brain, testis, intestine, and spleen). By immunohistochemistry on rat testis sections, we localized the protein in interstitial tissue and seminiferous tubules. In tubules, Goliath was expressed mainly in postmeiotic germ cells and to a much lesser extent in Sertoli cells. In the interstitium, Goliath was exclusively present in Leydig cells. Using a series of immunolabeling, cellular fractionation, and electron microscopy experiments, we established that Goliath is present in mitochondria of the R2C Leydig cell line. Using short-term hypophysectomized animals, we showed that Goliath is regulated by LH/hCG in Leydig cells but not in germ cells. This regulation in Leydig cells concerned only the 50-kDa isoform. This report is the first description of a differential regulation of the Goliath protein between germ cells and Leydig cells.

Anatomical and functional evidence for a neural hypothalamic-testicular pathway that is independent of the pituitary.

Testosterone (T) secretion is classically considered to be under the primary control of pituitary LH, itself regulated by the hypothalamic peptide LH-releasing hormone. Secretagogues present in the general circulation and/or manufactured in the testis can also alter Leydig cell activity independently of the pituitary. Finally, spanchnic innervation regulates testicular LH receptors and blood flow. In the present work, we provide evidence that, in addition, there may be a neural brain-testicular circuit that regulates T release function independently of LH release. We had recently reported that the intracerebroventricular injection of IL-1beta, corticotropin-releasing factor, or beta-adrenergic agonists significantly interfered with the T response to human chorionic gonadotropin through mechanisms that did not involve LH. Here, we show that the injection of the transganglionic retrograde tracer pseudorabies virus into the testes caused viral staining in the spinal cord, the brain stem, and the hypothalamus. This observation indicates the presence of a neural pathway between the central nervous system and the testis. We then demonstrated that spinal cord injury significantly interfered with this staining, thus supporting the hypothesis that the proposed circuit travels through the cord. Finally, we showed that spinal cord injury completely abolished the ability of intracerebroventricularly injected IL-1beta or corticotropin-releasing factor to blunt the T response to human chorionic gonadotropin, which suggests that these two secretagogues act within the brain to stimulate a neural pathway that interferes with Leydig cell function independently of the pituitary. The hitherto unsuspected brain-testicular circuit that these experiments have uncovered may play a role in pathologies, so far unexplained, that are characterized by decreased T levels despite normal LH production.

Functional compensation by Egr4 in Egr1-dependent luteinizing hormone regulation and Leydig cell steroidogenesis.

The Egr family of zinc finger transcription factors, whose members are encoded by Egr1 (NGFI-A), Egr2 (Krox20), Egr3, and Egr4 (NGFI-C) regulate critical genetic programs involved in cellular growth, differentiation, and function. Egr1 regulates luteinizing hormone beta subunit (LHbeta) gene expression in the pituitary gland. Due to decreased levels of LHbeta, female Egr1-deficient mice are anovulatory, have low levels of progesterone, and are infertile. By contrast, male mutant mice show no identifiable defects in spermatogenesis, testosterone synthesis, or fertility. Here, we have shown that serum LH levels in male Egr1-deficient mice are adequate for maintenance of Leydig cell steroidogenesis and fertility because of partial functional redundancy with the closely related transcription factor Egr4. Egr4-Egr1 double mutant male mice had low steady-state levels of serum LH, physiologically low serum levels of testosterone, and atrophy of androgen-dependent organs that were not present in either Egr1- or Egr4-deficient males. In double mutant male mice, atrophic androgen-dependent organs and Leydig cell steroidogenesis were fully restored by administration of exogenous testosterone or human chorionic gonadotropin (an LH receptor agonist), respectively. Moreover, a normal distribution of gonadotropin-releasing hormone-containing neurons and normal innervation of the median eminence in the hypothalamus, as well as decreased levels of LH gene expression in Egr4-Egr1-relative to Egr1-deficient male mice, indicates a defect of LH regulation in pituitary gonadotropes. These results elucidate a novel level of redundancy between Egr4 and Egr1 in regulating LH production in male mice.

Evidence for the involvement of non-androgenic testicular factors in the regulation of hypothalamic somatostatin and GHRH mRNA levels.

The growth hormone (GH) secretory pattern is dependent on sex and developmental stage. It is generally accepted that in the male rat this pattern is markedly influenced by androgens secreted by the Leydig cells. Recent findings, however, point to the existence of other non-androgenic testicular factors produced by the Sertoli cells and which regulate in vivo the GH responses to growth hormone releasing hormone (GHRH). The aim of this work was to investigate the role played by non-androgenic testicular factors on hypothalamic somatostatin (SST) and GHRH mRNA levels. Seventy-day-old male Sprague-Dawley rats were used throughout the work. They were divided into five groups: (1) control rats; (2) gonadectomized rats; (3) gonadectomized rats supplemented with exogenous administration of dihydrotestosterone (DHT); (4) ethylene dimethane sulphonate (EDS)-treated rats; (5) EDS-treated rats supplemented with exogenous administration of DHT. EDS is a cytotoxic agent that specifically destroys the Leydig cells. The rats were killed after 15 days of treatment. Hypothalamic SST mRNA levels were determined by Northern blot and by in situ hybridization, and GHRH mRNA levels assessed by Northern blot. We found that selective removal of Leydig cells with EDS greatly reduced the SST mRNA content in the periventricular nucleus of the hypothalamus. These levels were significantly lower than those found in gonadectomized rats. Furthermore, replacement treatment with dihydrotesterone (DHT) did not completely restore SST mRNA levels in EDS-treated rats, contrasting with the complete recovery of SST mRNA levels in gonadectomized rats. On the other hand, gonadectomy and EDS treatment produced a significant reduction in GHRH mRNA levels. DHT administration reversed the action of gonadectomy, but did not restore GHRH mRNA content in EDS-treated rats. These data suggest that, in addition to testosterone, as yet unidentified non-androgenic testicular factors can significantly influence SST and GHRH mRNA levels. This may indicate that non-androgenic testicular factors acting at hypothalamic level may be important in the neuroregulation of GH secretion and in the maintenance of sexual dimorphism in GH secretory pattern.

Effect of vitamin D repletion on testicular function in vitamin D-deficient rats.

Freshly weaned 30-day-old male Wistar rats were fed a vitamin D-deficient diet adequate in calcium and phosphorus for 3 months. On the 120th day of age three different doses of vitamin D were injected intramuscularly into three groups of rats and maintained for another month with water and a normal diet ad libitum. One group was continued with a vitamin D-deficient diet up to the 150th day. One group of animals was killed by decapitation on the 120th day and testicular functions like sperm count in testis and epididymis, testicular glutamyl transpeptidase activity and Leydig cell count as well as body weight were noted. On the 150th day animals of all groups were killed and testicular function was studied. Body weight and testicular function decreased significantly on the 120th and 150th day of age in vitamin D-deficient rats as compared to age-matched normal control rats. Injection of lower doses of vitamin D on the 120th day of age improved testicular function after 1 month whereas administration of a high dose of vitamin D caused a deterioration of the testicular function. The result suggests that retardation of spermatogenesis due to disturbances in Sertoli and Leydig cell function in vitamin D deficiency is reversible and can be corrected by supplementing an optimal dose of vitamin D.

Phospholipid methylation decreases in human chorionic gonadotrophin-induced desensitized rat Leydig cells.

Phospholipid methylation in Leydig cells from desensitized rats was studied. The incorporation of L-[methyl-3H]methionine into phospholipids in intact Leydig cells decreased when animals were injected with a single high dose of human chorionic gonadotrophin (hCG). This effect was detected on the first day after hCG injection and remained up to 12 days after treatment. The inhibition was not due to a reduced uptake of L-[methyl-3H]methionine. A decreased phospholipid methylation with unaltered phospholipid methyltransferase activity was observed on days 1, 6 and 12 after the hCG. On day 3 after hCG injection, phospholipid methyltransferase activity and phospholipid methylation in intact Leydig cells were both inhibited by 40%. Also, a minimal amount of LH free receptors and the lowest number of total receptors was observed at this time. Thus, a relationship between the reduced enzymatic activity and the maximal decrease in LH surface receptors is suggested. In addition, the decreased incorporation of L-[methyl-3H]methionine into phospholipids on days 1, 6 and 12 after hCG injection, could be associated with other cellular changes related to the desensitization process.

Fetal Leydig cells: cellular origin, morphology, life span, and special functional features.

The Leydig cells, responsible for testicular androgen production, have two growth phases during the life-span of mammals. The fetal population appears during fetal life and is responsible for the androgen-induced differentiation of the male genitalia. The fetal Leydig cells disappear after birth, and the other population, the adult Leydig cells, appears during puberty and persists for the whole adult life. The fetal Leydig cells, evidently due to the intrauterine endocrine milieu and their special functional requirements in genital differentiation, differ both morphologically and functionally from the adult population. The purpose of this review is to elucidate the special features of the mammalian fetal Leydig cell population, which presents an intriguing experimental model for studies of function and regulation of steroidogenic cells.

Dose-response effects of gonadotropin-releasing hormone on plasma concentrations of gonadotropins and testosterone in fertile and subfertile stallions.

Five fertile and five subfertile stallions were treated with a single intravenous injection of saline the first week followed by a single intravenous injection of varying doses of gonadotropin-releasing hormone (5, 10, 25, 100, 500 micrograms) given in a randomized fashion over the next 5 weeks during the nonbreeding season. Blood samples were collected periodically before and after treatment for analysis of luteinizing hormone, follicle stimulating hormone, and testosterone content by radioimmunoassay. Before treatment, semen samples were collected every other day for 3 weeks for analysis of volume, concentration, motility, pH, and morphology. Basal plasma levels of luteinizing hormone were higher (P < 0.05) in the subfertile group, follicle stimulating hormone levels tended to be higher (P < 0.10) in the subfertile group, and testosterone levels were similar in the two groups. A significant linear-log dose-response relationship was observed for plasma luteinizing hormone (P < 0.05) and follicle stimulating hormone (P < 0.05) to exogenous gonadotropin-releasing hormone in both the fertile and subfertile group. A linear-log dose-response relationship was also observed for plasma testosterone (P < 0.05) in the fertile group. The magnitude of the luteinizing hormone and follicle stimulating hormone response to gonadotropin-releasing hormone across doses was similar in both groups of stallions. A significant testosterone response to gonadotropin-releasing hormone in the subfertile group of stallions was not observed (P > 0.05). Mean testosterone concentrations after treatment in terms of net increase and percent of baseline were significantly lower (P < 0.05) in the subfertile group compared to the fertile group.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of megadoses of pyridoxine on spermatogenesis and male reproductive organs in rats.

Although it has been indicated that many neurotoxicants also cause reproductive toxicity, the reproductive toxicity of megadoses of pyridoxine, which is a neurotoxicant, has not been studied. In this paper, we studied the effects of megadoses of pyridoxine on male reproductive organs. Pyridoxine hydrochloride, 125 mg/kg, 250 mg/kg, 500 mg/kg or 1000 mg/kg, daily, was intraperitoneally injected into Wistar male rats 5 days a week for 2 or 6 weeks, and its effects on the male reproductive organs were investigated. After 2 weeks of administration, absolute weights of the testis in the 500 and 1000 mg/kg epididymis in all the exposed groups and prostate gland in the 1000 mg/kg group decreased, and mature spermatid counts in the testis decreased in the 1000 mg/kg group. After 6 weeks administration, the absolute and relative weights of the testis, epididymis, prostate gland and seminal vesicle decreased in the 500 mg/kg and 1000 mg/kg groups, and mature spermatid counts in the testis and sperm counts in the epididymis decreased in these groups. Among the marker enzymes of the testicular cells, LDH-X activity decreased, and beta-glucuronidase activity, cytochrome P-450 content and cytochrome b5 content increased in the 1000 mg/kg group. Plasma testosterone concentration did not significantly alter in all the exposed groups. From these results, it was concluded that megadoses of pyridoxine affected the spermatogenesis and decreased reproductive organ weights in the rat.