Effects of prolactin on the morphology and function of rat Leydig cells: short-term versus long-term administration

Summary The bolus administration of prolactin (PRL) to adult rats did not cause any apparent change in the basal and luteinizing hormone (LH)-stimulated blood levels of testosterone (as estimated by radioimmune assay). Prolonged PRL infusion did not affect either basal testosterone plasma concentration or the morphology of Leydig cells (as evaluated by electron microscopy and stereology). Conversely, prolonged PRL treatment notably increased the gonadotrophic effects of chronic LH administration; this mainly consisted of a rise in the blood concentration of testosterone and a conspicuous hypertrophy of Leydig cells. The LH-induced increase in the volume of Leydig cells was the result of an increase in the volumes of all the organelles involved in steroid synthesis (i.e., smooth endoplasmic reticulum, peroxisomes and mitochondria). However, the trophic effects of PRL infusion exclusively concerned smooth endoplasmic reticulum and peroxisomes. In the light of these findings, the hypothesis is advanced that the mechanism underlying the gonadotrophic action of PRL involves an enhancement of the endogenous cholesterol synthesis, which could provide an abundance of precursors for testosterone synthesis, the post-cholesterol steps of which, in turn, would be exclusively controlled by LH.     

Comparison of components of the testis interstitium with testosterone secretion in hamster, rat, and guinea pig testes perfused in vitro

Components of the testis and cytoplasmic organelles in Leydig cells were quantified with morphometric techniques in hamster, rat, and guinea pig. Testosterone secretory capacity per gram of testis and per Leydig cell in response to luteinizing hormone (LH) (100 ng/ml) stimulation was determined in these three species from testes perfused in vitro. Numerous correlations were measured among structures, and between structures and testosterone secretion, to provide structural evidence of intratesticular control of Leydig cell function. Testosterone secretion per gm testis and per Leydig cell was significantly different in the three species: highest in the guinea pig, intermediate in the rat, and lowest in the hamster. The volume of seminiferous tubules per gm testis was negatively correlated, and the volumes of interstitium, Leydig cells, and lymphatic space per gm testis were positively correlated with testosterone secretion. No correlations were observed between volumes of blood vessels, elongated spindleshaped cells, or macrophages per gm testes and testosterone secretion. The average volume of a Leydig cell and the volume and surface area of smooth endoplasmic reticulum (SER) and peroxisomes per Leydig cell were positively correlated, and the volume of lysosomes and surface area of inner mitochondrial membrane per Leydig cell were negatively correlated with testosterone secretion. No correlations were observed between volume and surface area of rough endoplasmic reticulum (RER), Golgi apparatus, and lipid, and volume of ribosomes, cytoplasmic matrix, and the nucleus with testosterone secretion per Leydig cell. These results suggest that Leydig cell size is more important than number of Leydig cells in explaining the difference in testosterone-secreting capacity among the three species, and that this increase in average volume of a Leydig cell is associated specifically with increased volume and surface area of SER and peroxisomes. An important unresolved question is what is the role of peroxisomes in Leydig cell steroidogenesis.     

Taenia taeniaeformis: inhibition of rat testosterone production by excretory-secretory product of the cultured metacestode

In 3- to 5-month-old male Sprague-Dawley rats infected with the hepatic metacestode, Taenia taeniaeformis, the serum testosterone level was significantly lower than in comparable uninfected controls. By transmission electron microscopy, testicular Leydig cells of infected rats had less smooth endoplasmic reticulum than control Leydig cells. Cultured metacestodes isolated from the hepatic cysts secreted or excreted substances into the incubation medium. The effect of the excretory-secretory product on testosterone concentration in the sera and testes of 15-day-old rats was examined. Subcutaneous injection of 50-200 micrograms of excretory-secretory product/0.1 ml saline/rat for 2 days significantly reduced human chorionic gonadotropin-stimulated serum and testicular testosterone concentrations. Furthermore, the effect of the excretory-secretory product on isolated rat Leydig cell testosterone production was examined. Rat Leydig cells produced testosterone in vitro and, in the presence of 50 IU human chorionic gonadotropin/ml incubation medium, they responded with approximately 100% increase in testosterone production. Addition of 2-10 micrograms excretory-secretory product protein/ml of culture medium significantly reduced the testosterone production by rat Leydig cells in vitro. These results indicate that excretory-secretory product of cultured T. taeniaeformis metacestodes has a direct inhibitory effect on Leydig cell testosterone production under stimulation with human chorionic gonadotropin.     

Ethanol-induced inhibition of testosterone biosynthesis in rat Leydig cells: central role of mitochondrial NADH redox state

The mechanisms by which ethanol (EtOH) inhibits the human chorionic gonadotropin (hCG)-stimulated testosterone synthesis was studied in isolated rat Leydig cells in vitro. EtOH inhibited steroidogenesis, but this inhibition was reversed by L-glutamate (Glu) and an uncoupler of the oxidative phosphorylation, 2,4-dinitrophenol (DNP). The mechanism of EtOH-induced inhibition was studied by measuring steroidogenic precursors and comparing them with the cytosolic and mitochondrial NADH redox states during uncoupling or in the presence of Glu. DNP had a dual effect. Low concentrations abolished the EtOH-induced inhibition of progesterone to testosterone formation suggesting that the inhibitory step was at or before progesterone formation. A large concentration led to an overall decrease in steroidogenesis indicating toxic effects on steroidogenesis. The mitochondrial NADH/NAD+ ratio, measured as the 3-hydroxybutyrate/acetoacetate ratio, decreased simultaneously when steroidogenesis was stimulated, either during uncoupling or in the presence of Glu, whereas cytosolic NADH/NAD+ ratio, measured as lactate/pyruvate ratio showed no response. These results demonstrate that the rise in the mitochondrial NADH/NAD+ ratio rather than in the cytosolic one is connected with the inhibition of testosterone synthesis by EtOH in isolated Leydig cells. The EtOH-induced high mitochondrial NADH/NAD+ ratio may deplete mitochondrial oxalacetate concentrations. This can decrease the activity of several transport shuttles and interrupt the flow of mitochondrial citrate into the smooth endoplasmic reticulum, which then reflects to decreased rate of steroidogenesis in the presence of ethanol.     

Changes in Leydig cell ultrastructure and function during pubertal development in the boar

Changes in the ultrastructure of Leydig cells during pubertal development in the boar (40 to 250 days of age) were assessed using quantitative morphometric procedures, and the results were compared to the in vitro steroid-producing capacity and gonadotropin sensitivity of testicular tissue obtained from the same boars. Volume of individual Leydig cells declined through 100 days of age, increased rapidly to a peak at 130-160 days (i.e., puberty), and then declined to intermediate levels by 220-250 days of age. The pattern of change in the number of intracellular organelles per Leydig cell was very similar to the change that occurred in Leydig cell volume. Changes in the total intracellular volume occupied by each type of organelle were highly correlated with changes in Leydig cell volume (r = 0.40-0.99, p less than 0.01), and this was particularly true for the nucleus (r = 0.63), mitochondria (r = 0.88), smooth endoplasmic reticulum (SER; r = 0.97), and total cytoplasm (r = 0.99) of the boar Leydig cell. In vitro production of testosterone and estradiol, expressed per Leydig cell, also peaked at 130-160 days, and was highly correlated to average Leydig cell volume, volume of SER, and number and total volume of mitochondria (r = 0.63-0.84; p less than 0.01). Observations in the present study indicated that onset of puberty in boars coincides with a dramatic increase in average Leydig cell size and SER volume per Leydig cell, accompanied by an increase in number of other intracellular organelles, including mitochondria, lysosomes, and lipid droplets, and a peak in the steroid-producing capacity per Leydig cell. A decline in Leydig cell size, intracellular organelles, and sensitivity to gonadotropin stimulation occurred postpubertally.     

Morphometric analysis of Leydig cells in the normal rat testis

Leydig cells are thought to be the source of most, if not all, the testosterone produced by the testis. The goal of this study was to obtain quantitative information about rat Leydig cells and their organelles that might be correlated with pertinent physiological and biochemical data available either now or in the future. Morphometric analysis of Leydig cells in mature normal rats was carried out on tissue fixed by perfusion with buffered glutaraldehyde, and embedded in glycol methacrylate for light microscopy and in Epon for electron microscopy. In a whole testis, 82.4% of the volume was occupied by seminiferous tubules, 15.7% by the interstitial tissue, and 1.9% by the capsule. Leydig cells constituted 2.7% of testicular volume. Each cubic centimeter (contained approximatelyy 1 g) of rat testis contained about 22 million Leydig cells. An average Leydig cell had a volume of 1,210 micron3 and its plasma membrane had a surface area of 1,520 micron2. The smooth endoplasmic reticulum (SER), the most prominent organelle in Leydig cells and a major site of steroidogenic enzymes, had a surface area of approximately 10,500 micron2/cell, which is 6.9 times that of the plasma membrane and is 60% of the total membrane area of the cell. The total surface area of Leydig SER per cubic centimeter of testis tissue is approximately 2,300 cm2 or 0.23 m2. There were 3.0 mg of Leydig mitochondria in 1 g of testis tissue. The average Leydig cell contained approximately 622 mitochondria, measuring on the average 0.35 micron in diameter and 2.40 micron in length. The mitochondrial inner membrane (including cristae), another important site of steroidogenic enzymes, had a surface area of 2,920 micron2/cell, which is 1.9 times that of the plasma membrane. There were 644 cm2 of inner mitochondrial membrane/cm3 of testis tissue. These morphometric results can be correlated with published data on the rate of testosterone secretion to show that an average Leydig cell secretes approximately 0.44 pg of testosterone/d or 10,600 molecules of testosterone/s. The rate of testosterone production by each square centimeter of SER is 4.2 ng/d or 101 million molecules/s: the corresponding rate for each square centimeter of mitochondrial inner membrane is 15 ng testosterone/d or 362 million molecules/s.     

In vitro stimulation and inhibition of steroid hormone release from postovulatory follicles of the tilapia,Oreochromis mossambicus

Summary Postovulatory follicles of the tilapia, Oreochromis mossambicus, were incubated with graded doses of salmon gonadotropin to identify the steroid hormones released by this tissue. In addition, the effects of either cytochalasin B or colchicine on steroid hormone release were studied. After the incubation, the tissue was examined by electron microscopy. Postovulatory follicles released testosterone and estradiol-17B in a dose-dependent manner with gonadotropin. There was no detectable release of progesterone or 17a-OH-progesterone. When stimulated with high doses of gonadotropin, the steroidogenic cells showed an increase in smooth endoplasmic reticulum, Golgi complexes, and lipid droplets. Also, microfilaments became arranged in orderly bundles and were found close to the numerous secretory vesicles and lipid droplets. Upon incubation with gonadotropin and either colchicine or cytochalasin B, the cells still appeared steroidogenic, but the filaments were not organized nor associated with vesicles or lipid droplets. Release of steroid hormone decreased significantly. Also in these tissues, vesicles were no longer numerous in the apical region of the granulosa cells, but were located primarily near smooth endoplasmic reticulum and Golgi complexes. This suggests that disruption of the cytoskeleton results in reduced steroid hormone synthesis or release.     

乌梢蛇精巢间质细胞微细结构与血清 睾酮浓度的相关性

用光镜及透射电镜观察了乌梢蛇(Zaocys dhumnades)精巢间质细胞的显微与超微结构,并利用放射免疫测定法测定了血清中睾酮浓度.结果表明,在一个年生殖周期中,乌梢蛇间质组织所占区域相对大小、间质细胞数量和显微结构均存在较明显的变化;5月份的间质细胞具有发达的管状嵴线粒体、丰富的滑面内质网、大量的脂滴等合成和分泌...     

Testicular function and Leydig cell ultrastructure in long-term bilaterally cryptorchid rams

This study was conducted to investigate the effects of bilateral cryptorchidism induced in adult rams on testicular function and Leydig cell ultrastructure. The results indicated that long-term bilateral cryptorchidism resulted in decreased testicular size, degeneration of seminiferous tubules, elevated serum LH levels, maintenance of normal testosterone concentrations in peripheral and spermatic vein serum, impairment of the magnitude and duration of androgen response to exogenous luteinizing hormone (LH), a 13-fold reduction in total number of Leydig cells/paired testes, and a 3-fold hypertrophy in the average size of remaining Leydig cells. Based on quantitative morphometry, the hypertrophied Leydig cells exhibited significant increases in the volume of intracellular organelles, including the cell nucleus, mitochondria, smooth and rough endoplasmic reticulum, lysosome-like bodies and lipid vesicles. Quantitatively, the hypertrophy alone was not enough to offset the loss in number of Leydig cells and was insufficient to explain the maintenance of normal levels of testosterone in jugular and spermatic venous blood. The additional mechanisms responsible for production of normal serum testosterone levels in the cryptorchid ram remain to be elucidated.     

Effects of Cordyceps sinensis on testosterone production in normal mouse Leydig cells.

The stimulatory effect of Cordyceps sinensis (CS) on MA-10 mouse Leydig tumor cell steroidogenesis was previously demonstrated in our laboratory. In the present studies, we further determined the effect of CS on steroidogenesis in purified normal mouse Leydig cells. Different concentrations of CS (0.1-10 mg/ml) were added to Leydig cells without or with human chorionic gonadotropin (hCG) (50 ng/ml), and the steroid production was determined by radioimmunoassay (RIA). The results illustrated that CS stimulated normal mouse Leydig cell steroidogenesis in a dose-dependent relationship. CS at 3 mg/ml significantly stimulated testosterone production (p<0.05). Concerning the temporal relationship, CS at 3 mg/ml stimulated maximal testosterone production between 2 to 3 hr. Interestingly, hCG-stimulated testosterone productions were suppressed by CS in a dose-dependent relationship. CS also reduced dbcAMP-stimulated testosterone productions, which indicated that CS affected signal transduction pathway of steroidogenesis after the formation of cyclic AMP. Moreover, cycloheximide inhibited CS-treated mouse Leydig cell testosterone production, suggesting that new protein synthesis was required for CS-stimulated steroidogenesis.     

Seasonal changes of the Leydig cells of viscacha (Lagostomus maximus maximus). A light and electron microscopy study

The Leydig cells of viscacha (seasonal rodent) show cytoplasmic hypertrophy and regional distribution during the breeding period (summer-autumn). The dominant organelles are smooth endoplasmic reticulum (SER) and mitochondria. A moderately well-developed Golgi, abundant lipid inclusions, dense bodies like lysosomes in different stages, and centrioles are observed. Extensive or focal desmosome and gap-like junctions between neighbouring Leydig cells are present. These cells exhibit an evident hypotrophy and an increase in the number of dense bodies during the gonadal regression in winter (July and August). Cells in different stages of involution are observed in this period. Their nuclei are irregular and heterochromatic. The cytoplasm contains few mitochondria. The vesicular SER is scarse. Irregular and large intercellular spaces with microvilli and amorphous material are present. The junctional complexes are absent. The nuclear and cytoplasmic volume and development of SER and mitochondria increase during the recovery period (spring). The lipid inclusions decrease. Dilatations of the intercellular space with microvilli and limited by focal desmosome-like junctions are observed. In conclusion, the Leydig cells of Lagostomus maximus maximus show deep changes alongside the reproductive cycle. The photoperiod variations, through pineal hypothalamus pituitary axis and the hormone melatonin, are probably responsible for them. Moreover, the fall of serum and tubular testosterone would be one of the factors responsible for gonadal regression.     

Ultrastructure and morphometry of testicular Leydig cells and the interstitial components correlated with testosterone in aging rats

The ultrastructure of testicular interstitium in young and aged adult rats was analysed using morphometric methods, and the plasma testosterone concentration was measured. With increasing age there was an augumentation in the volume of collagen fibrils in the intercellular matrix and in blood vessels. During the aging process (approximately two years) the average volume of the Leydig cell decreased from 1364 m³ to 637 m³, but the number of Leydig cells in paired testes increased from 53x10⁶ to 113x10⁶. The absolute volume of smooth surfaced endoplasmic reticulum (SER) per Leydig cell amounted in aged rats to 78% of that in young adult rats. The total amount of SER in paired testes increased by 62% with aging. The present analysis suggests that the ability of SER to maintain peripheral testosterone concentration decreases with age. In young adult rats the absolute volume of peroxisomes per Leydig cell correlated significantly with the concentration of testosterone in blood and also with the absolute volume of SER per Leydig cell. These results combined with ultrastructural observations of close apposition of peroxisomes and SER suggest that peroxisomes have a role in testosterone secretion by Leydig cells.Visiting scientist to Laboratory of Electron Microscopy (Director: Prof. L.J. Pelliniemi)     

Release of arachidonic acid and the effects of corticosteroids on steroidogenesis in rat testis Leydig cells.

The release of arachidonic acid by luteinizing hormone (LH) and the effects of inhibiting phospholipase A2 (PLA2) in vivo and in vitro on LH stimulated steroidogenesis in rat testis Leydig cells has been investigated. It was found that arachidonic acid is rapidly incorporated into phospholipids and is released within 1 min after addition of LH. The effects of treating adult rats with dexamethasone and human chorionic gonadotropin (hCG) in vivo on steroidogenesis and prostaglandin synthesis in Leydig cells isolated 6 h later were determined. It was found that hCG caused a marked increase in prostaglandin F2 alpha formation which was inhibited by treatment with dexamethasone. LH-stimulated testosterone production was inhibited in the hCG treated rats and dexamethasone caused a further decrease. Treatment with dexamethasone alone also caused a decrease in the response to LH. HCG, but not dexamethasone, had similar inhibitory effects on LH-stimulated cyclic AMP production. Similarly, the PLA2 inhibitors quinacrine, dexamethasone and corticosterone, added to the Leydig cells in vitro, inhibited LH-stimulated testosterone production but not cyclic AMP production. 11-Dehydrocorticosterone also inhibited LH-stimulated testosterone production, but higher concentrations were required to give 50% inhibition compared to corticosterone (50 and 25 microM, respectively). Ring A-reduced metabolites of corticosterone and progesterone were also found to inhibit LH-stimulated steroidogenesis. The results obtained in this and previous studies are consistent with the activation of PLA2, (either directly by LH and/or via cyclic AMP), which results in the release of arachidonic acid and the formation of leukotrienes, which stimulate steroidogenesis in the Leydig cell. This study also indicates that corticosteroids and their metabolites may exert inhibitory effects at other sites in the steroidogenic pathways, in addition to PLA2.     

Gonadotropin binding and stimulation of cyclic adenosine 3':5'-monophosphate and testosterone production in isolated Leydig cells.

Gonadotropin binding and stimulation of cyclic adenosine 3':5'-monophosphate (cyclic AMP) formation and testosterone synthesis were studied in collagenase-dispersed interstitial cells from the adult rat testis. Binding of 125I-human chorionic gonadotropin (hCG) by isolated Leydig cells was of high affinity (Ka = 10(10) M-1) and low capacity, equivalent to approximately 6000 sites/cell. The binding data were consistent with the presence of a single order of receptors, with no interaction between binding sites. Stimulation of testosterone synthesis by increasing concentrations of hCG was completely dissociated from changes in cyclic AMP formation, and maximum activation of steroidogenesis was induced by hCG concentrations which had no effect upon cyclic AMP production. Kinetic analysis of gonadotropin-induced responses in dispersed Leydig cells also showed a marked dissociation between steroidogenesis and cyclic nucleotide formation. Low concentrations of hCG caused maximum stimulation of testosterone production which was not accompanied by a rise in cyclic AMP formation at any time after addition of gonadotropin. Higher concentrations of hCG caused marked elevations of cyclic AMP at progressively earlier time intervals, but did not alter the 20 to 30 min lag period required for induction of testosterone synthesis. These observations indicated that occupancy of gonadotropin receptors occurs over a much wider range of hCG concentration than that required for maximum steroidogenesis.     

Ultrastructural and biochemical characteristics of leydig cells from newborn androgen-insensitive rats

Summary Leydig cells of the testis of newborn pseudohermaphrodite (tfm) rats have an ultrastructure similar to that of the normal, containing well developed organelles and inclusions. The cytoplasm is filled with smooth endoplasmic reticulum forming a network of interconnected tubules. Lipid droplets are surrounded by cisternae of smooth endoplasmic reticulum and are in close association with pleomorphic mitochondria. Many of the latter are cup-shaped and have tubular cristae and intramitochondrial dense bodies.Essentially, these are characteristics of normal Leydig cells. Accordingly, the production of testosterone by testes from newborn tfm rats is the same as that by testes from normal newborns and adults. However, it is significantly higher than that by testes of tfm adults. Also, the plasma testosterone levels of newborn tfm rats are the same as in the normal newborn, but lower than in normal adults and much lower than in adult tfm animals.Thus, since in the tfm rat the morphology of Leydig cells, androgen production, and maintenance of plasma levels of testosterone are normal in the newborn, but become abnormal with advancing age, it appears that defective androgen action rather than insufficient androgen production is the cause of male pseudohermaphroditism.     

Desensitization of mouse Leydig cells in vivo: evidence for the depletion of cellular cholesterol

The study presents a characterization of the refractory state in purified mouse Leydig cells desensitized by a single injection of human chorionic gonadotropin (hCG) in vivo. The treatment of mice with 1 microgram hCG i.p. for 48 h followed by Leydig cell isolation and purification resulted in a decrease in the maxima of hCG-induced cAMP accumulation and testosterone production by approximately 70% and approximately 55%, respectively, when compared to cells of control mice. Despite a 55% reduction in 125I-hCG binding sites, the sensitivity of stimulation was not changed. The refractoriness in testosterone production in vitro was also present when the Leydig cells were stimulated with cholera toxin or dibutyryl cAMP; however, it was not observed when testosterone production was induced by the addition of pregnenolone or 20 alpha- and 22(R)-hydroxycholesterol. Mouse lipoproteins, high-density lipoprotein (HDL) and low-density lipoprotein (LDL) in natural composition, were also able to overcome the steroidogenic block (although not always completely). On the basis of the cholesterol content of the lipoproteins, the two classes were similarly effective. They increased maximal hCG-induced testosterone production not only in desensitized cells, but also in control cells (by 80-100%), whereas their effect on basal testosterone production was negligible. In desensitized cells from hCG-treated mice (2 micrograms i.p., 48 h) cellular unesterified and esterified cholesterol were decreased by 21% and 81%, respectively, when compared to control cells. This loss occurred in the face of unchanged plasma cholesterol levels. In conclusion, our data indicate that the impaired steroidogenesis in mouse Leydig cells desensitized in vivo by a single injection of hCG is the result of a depletion in cellular cholesterol, rather than of an impaired conversion of cholesterol to testosterone.     

The effects of cytochalasin B on the testosterone synthesis by interstitial cells of rat testis

The present study examined the effects of cytochalasin B on various steps in the luteinizing hormone (LH)-stimulated increase in testosterone synthesis by collagenase-dispersed interstitial cells of adult rat testis. Cytochalasin B at a concentration range of 0.1–50 μM inhibited the LH-stimulated increase in testosterone synthesis in a dose-dependent manner. Both intracellular and medium (released) testosterone levels were reduced, thus indicating that the decrease was not due to the accumulation of testosterone inside the cell as a result of cytochalasin B treatment. Cytochalasin B also inhibited the 8-bromocyclic AMP and pregnenolone-stimulated testosterone synthesis in a similar dose-dependent manner. Cytochalasin B at the two higher doses (10 and 50 μM) also inhibited the LH-stimulated generation of cyclic AMP by interstitial cells. However, this drug had no effect on basal testosterone synthesis except at the highest concentration added.Previous studies on adrenocorticotropic hormone (ACTH)- and LH-stimulated increase in glucocorticoid and testosterone synthesis in adrenal and Leydig cells, respectively, demonstrated that cytochalasin B or anti-actin inhibited the transport of cholesterol into mitochondria. The present studies suggest that cytochalasin B inhibits at least two additional steps in the LH-stimulated increase in testosterone synthesis: (1) the generation of cyclic AMP at the level of the plasma membrane, and (2) the conversion of pregnenolone to the testosterone at the level of the smooth endoplasmic reticulum. It remains to be established whether these are direct effects of cytochalasin B, or whether they are mediated by disruption of microfilaments by cytochalasin B.     

Rapid mechanisms of glucocorticoid signaling in the Leydig cell

Stress-mediated elevations in circulating glucocorticoid levels lead to corresponding rapid declines in testosterone production by Leydig cells in the testis. In previous studies we have established that glucocorticoids act on Leydig cells directly, through the classic glucocorticoid receptor (GR), and that access to the GR is controlled prior to the GR by a metabolizing pathway mediated by the type 1 isoform of 11beta-hydroxysteroid dehydrogenase (11betaHSD1). This enzyme is bidirectional (with both oxidase and reductase activities) and in the rat testis is exclusively localized in Leydig cells where it is abundantly expressed and may catalyze the oxidative inactivation of glucocorticoids. The predominant reductase direction of 11betaHSD1 activity in liver cells is determined by an enzyme, hexose-6-phosphate dehydrogenase (H6PDH), on the luminal side of the smooth endoplasmic reticulum (SER). Generation of the pyridine nucleotide cofactor NADPH by H6PDH stimulates the reductase direction of 11betaHSD1 resulting in increased levels of active glucocorticoids in liver cells. Unlike liver cells, steroidogenic enzymes including 17beta-hydroxysteroid dehydrogenase 3 (17betaHSD3) forms the coupling with 11betaHSD1. Thus the physiological concentrations of androstenedione serve as a substrate for 17betaHSD3 utilizing NADPH to generate NADP+, which drives 11betaHSD1 in Leydig cells primarily as an oxidase; thus eliminating the adverse effects of glucocorticoids on testosterone production. At the same time 11betaHSD1 generates NADPH which promotes testosterone biosynthesis by stimulating 17betaHSD3 in a cooperative cycle. This enzymatic coupling constitutes a rapid mechanism for modulating glucocorticoid control of testosterone biosynthesis. Under stress conditions, glucocorticoids also have rapid actions to suppress cAMP formation thus to lower testosterone production.     

Acute response of testicular interstitial tissue in rats to the cytotoxic drug ethane dimethanesulphonate

Summary The cytotoxic effects of ethane dimethanesulphonate upon rat Leydig cells were examined ultrastructurally up to 3 days after treatment and related to changes in serum levels of gonadotrophins and testosterone. Six hours after administration of ethane dimethanesulphonate the usual tubulo-vesicular morphology of Leydig-cell smooth endoplasmic reticulum was converted to small vesicles and the Golgi apparatus showed focal hypertrophy into anastomosing tubules. These changes became more marked by 12 h with many Leydig cells exhibiting karyopyknosis and hyperchromatism. Necrotic Leydig cells were often engulfed by macrophages, the latter containing pyknotic fragments of Leydig cells within their cytoplasm. One day after administration, advanced necrosis of Leydig cells occurred, many of which were phagocytosed by macrophages, and on day 3, destruction of Leydig cells was complete resulting in their elimination from the interstitial tissue, which contained only loose connective tissue and macrophages. Structural alterations to the Leydig cells from 6–24 h was reflected by a significant reduction in serum testosterone levels which further declined to the limits of detection accompanying the abolition of Leydig cells on day 3. These changes were paralleled by a significant elevation of serum LH and FSH levels suggesting diminished feedback regulation of pituitary gonadotrophin secretion. The results indicate that ethane dimethanesulphonate is a rapidly acting Leydig cell toxin which may be a useful experimental tool in further studies of spermatogenic function mediated via Sertoli cell-Leydig cell interaction.