Monitoring of spermatogenesis in man--measurement of Sertoli cell- or germ cell-secreted proteins in semen or blood.
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Blood Proteins↗
Biomedical subjects
R M Sharpe
Publications and source records attributed to R M Sharpe.
Evaluation of the role of germ cells in regulating the route of secretion of immunoactive inhibin from the rat testis.
During normal sexual maturation of the male rat there is a progressive change in the route of secretion of inhibin by the Sertoli cell, from a predominantly basal route of secretion in prepuberty to a predominantly apical route of secretion in adulthood. This change may be monitored by comparing the levels of inhibin in testicular (TV), spermatic and peripheral (PV) venous blood and the levels in testicular interstitial fluid (IF). This study has assessed the role of germ cells in effecting this change by assessing (a) the effect of total germ cell depletion by X-irradiation of the males in utero, and (b) the effect of selective germ cell depletion in adulthood using the testicular toxicant, methoxyacetic acid (MAA). Female rats were X-irradiated on day 20 of gestation to produce male offspring whose testes were germ-cell deficient. Blood and IF samples were collected from groups of these offspring and age-matched controls at 35 and 100 days of age. In blood and IF samples, inhibin concentrations were significantly higher at 35 days of age than at 100 days. The absence of germ cells in X-irradiated animals did not affect the age-related fall in inhibin levels, nor the change in the predominant route of secretion of inhibin from the testis into blood. Testosterone was almost undetectable in 35-day-old controls, but was raised significantly by 100 days of age. In X-irradiated animals, testosterone levels were increased significantly at 35 days of age, and the levels in most samples were increased even more substantially by 100 days of age. However, PV levels of testosterone in 100-day-old X-irradiated animals were significantly lower than in controls. LH and FSH levels were raised in X-irradiated animals compared with their age-matched controls, but FSH levels in X-irradiated animals still fell with age, as in the controls. The role of specific germ cell types in regulating the route of secretion of inhibin from the normal adult testis was studied after depletion (80-100%) of pachytene and later spermatocytes by a single oral administration of MAA (650 mg/kg) to adult rats. At 3 days after MAA treatment, coincident with the loss of pachytene spermatocytes, plasma inhibin levels were increased significantly in blood and IF samples, and this was associated with a dramatic change in the route of secretion of inhibin from the testis, with increased secretion of this peptide via the base of the Sertoli cell into IF and TV blood.(ABSTRACT TRUNCATED AT 400 WORDS)
Animals↗
Evaluation of changes in the secretion of immunoactive inhibin by adult rat seminiferous tubules in vitro as an indicator of early toxicant action on spermatogenesis.
A method for culturing isolated seminiferous tubules (ST) from adult rats for 1-3 days has been developed and optimized rigorously on the basis of the secretion of immunoactive inhibin under basal conditions and after maximal stimulation with rat FSH or dibutyryl cyclic AMP. The effect on these cultures of three known testicular toxicants was assessed. Of these, two are thought to act on the Sertoli cell, meta-dinitrobenzene (mDNB) and nitrobenzene (NB), while the third, methoxy acetic acid (MAA), is thought to act on pachytene spermatocytes. In addition, the effect of a possible testicular toxicant, 3-mononitrotoluene (3-MNT), was investigated. These data were compared with those obtained using cultures of immature rat Sertoli cells (SC) or SC + germ cells and with data on the effect of equivalent doses of the compounds on the secretion of immunoactive inhibin in vivo. In studies designed to optimize conditions for the secretion of immunoactive inhibin by ST in culture, significant effects were found of the type of culture medium used, the duration of culture, the total and individual length of tubules used, etc. All subsequent studies with toxicants utilized optimal conditions. Addition of either mDNB or NB to ST cultures at 10(-5) or 10(-3) M, or MAA at 10(-4) M, stimulated basal secretion of immunoactive inhibin by two- to fourfold on Days 1, 2, or 3 of culture while FSH or dibutyryl cyclic AMP-stimulated secretion of immunoactive inhibin was either unaffected or was enhanced to a small extent. At the same doses, mDNB or NB also enhanced secretion of immunoactive inhibin by SC cultures. although these effects were more variable and of smaller magnitude than the effects on ST cultures. In contrast, addition of up to 10(-3) MAA to cocultures of SC + germ cells had no effect on the secretion of immunoactive inhibin. Exposure of rats in vivo to levels of mDNB, NB, or MAA similar to those which stimulated secretion of immunoactive inhibin in vitro resulted in a two- to fourfold increase in the levels of immunoactive inhibin in testicular interstitial fluid (IF) at 1 and 3 days post-treatment, and this was associated with early impairment of spermatogenesis (as judged by testis weight). In contrast to these effects, addition of 3-MNT to ST or SC cultures had no effect except at 10(-3) M, when the secretion of immunoactive inhibin was increased marginally.(ABSTRACT TRUNCATED AT 400 WORDS)
Acetates↗
Evidence that secretion of immunoactive inhibin by seminiferous tubules from the adult rat testis is regulated by specific germ cell types: correlation between in vivo and in vitro studies.
This study has assessed whether depletion of specific germ cell types is able to alter the secretion of immunoactive inhibin by adult Sertoli cells in vivo and in vitro. Pachytene and later spermatocytes were depleted (80-100%) by a single administration of methoxy acetic acid (MAA; 650 mg/kg) to adult rats. At intervals between 1 and 42 days posttreatment, rats were killed, and the blood levels of FSH, LH, and testosterone were determined together with the levels of immunoactive inhibin in plasma and testicular interstitial fluid (IF). At the same time intervals, seminiferous tubules (ST; 5 x 2 cm) were isolated from control and MAA-treated rats and cultured for 24-72 h in the presence or absence of rat FSH, (Bu)2cAMP, or MAA under rigorously optimized conditions. The hormonal changes observed were related to the presence/absence of specific germ cell types, as determined by assessment of testicular morphology in perfusion-fixed testes from similarly treated rats. One to 3 days after MAA treatment, coincident with the depletion of pachytene spermatocytes, blood levels of FSH were increased significantly compared with controls; FSH returned to control levels at 7-14 days (when early spermatids were depleted), but were increased again at 21-35 days (when late spermatids were depleted). In contrast, while the plasma levels of immunoactive inhibin were increased 2-fold 3 days posttreatment, they were comparable to controls at 7-14 days, but were decreased substantially at 21-28 days. The levels of immunoactive inhibin in testicular IF were more than doubled 1 and 3 days posttreatment, but were comparable to control levels at all other times. Blood levels of LH showed a similar pattern of change to FSH, although only at 21-28 days after MAA treatment was there a significant increase, while blood levels of testosterone were comparable at all times in control and MAA-treated rats. To confirm that the changes observed in vivo after MAA treatment were indicative of changes in Sertoli rather than Leydig cell secretion of immonoactive inhibin, its secretion by isolated ST was assessed, and a pattern of change similar to that in plasma was observed. Thus, when cultured for 24 h under basal conditions, ST from rats 1-3 days after MAA treatment showed a 2- to 3-fold increase in secretion of immunoactive inhibin, which returned to control levels at 7-14 days before being reduced substantially at 21-28 days and then recovering to control levels; similar changes were observed for FSH- and (Bu)2cAMP-stimulated secretion of immunoactive inhibin.(ABSTRACT TRUNCATED AT 400 WORDS)
Acetates↗
Evidence for the control of testicular interstitial fluid volume in the rat by specific germ cell types.
Following on from our recent evidence that Sertoli cells may regulate testicular interstitial fluid (IF) volume, this study has assessed whether depletion of specific germ cell types in vivo is associated with changes in recovered IF volume. Germ cell depletion was induced by either a single oral administration of 650 mg methoxyacetic acid (MAA)/kg or exposure of the testes to local heating (43 degrees C for 30 min). Treatment with MAA induced depletion or loss of most pachytene and later spermatocytes at 1-3 days and, because of maturation depletion, this resulted in the specific depletion of later germ cell types at 7-35 days. Testicular IF volume was unchanged at 1-7 days after MAA treatment but was increased significantly (P less than 0.01) at 14 days and was nearly doubled (P less than 0.001) at 21 days, before returning to control levels at 28-42 days. Serum LH (and FSH) levels were generally higher in MAA-treated rats, especially at 21 and 28 days, but there was no obvious correlation between LH levels and IF volume changes. Similarly, there was no relationship between IF volume changes and testicular weight or IF levels of testosterone. The increase in IF volume at 14-21 days after MAA treatment coincided with specific depletion of the later elongate spermatids (steps 14-19) and, when these cells reappeared in the testis, IF volume normalized. This possible causal association was studied further in rats exposed to local testicular heating which, within 3 days, caused major depletion of pachytene spermatocytes and early (step 1-8) spermatids.(ABSTRACT TRUNCATED AT 250 WORDS)
Acetates↗
[Germ cells and post-natal development of testicular function: in vitro studies].
Studies in recent years have clearly established that, in addition to the well known endocrine regulation by gonadotrophin hormones, spermatogenesis is under the modulatory control of a complex set of paracrine regulators. Whereas the role of Leydig cells (testosterone) and of Sertoli cells (nurce cells of germ cells) in spermatogenesis has focused most of the attention, until recently little was known about the contribution of germ cells in the spermatogenetic process. This was the aim of the present experiments. We have used, in vitro, 3 complementary approaches; 1) we measured the influence of the removal of germ cells contaminating Sertoli cell cultures by a hypotonic treatment; 2) in coculture, we examined to what extend isolated germ cells could affect Sertoli cell function; 3) we investigated the effects of germ cell conditioned media on Sertoli cell cultures. Our results indicate that germ cells are able to modulate Sertoli cell function in vitro. This germ cell influence varies according to: 1) the germ cell fraction tested (pachytene spermatocytes, early spermatids or cytoplast from elongated spermatids/residual bodies); 2) the parameter of Sertoli cell function studied (inhibition of oestradiol; stimulation of androgen-binding protein, transferrin...); 3) the age of the Sertoli cell donors; 4) the hormonal environment (+/- FSH). Furthermore we wave demonstrated that germ cell effects were partly at least mediated via proteinaceous factor(s) detected in germ cell spent media. Taking into account previous in vivo studies and these in vitro results, we have hypothesized that germ cells, in conjunction with hormones (LH, FSH, testosterone) play an important role in the ontogenesis of Sertoli cells and therefore in spermatogenesis.
Androgen-Binding Protein↗
Regulation of Sertoli cell inhibin production and of inhibin alpha-subunit mRNA levels by specific germ cell types.
To elucidate the endocrine and paracrine regulation of testicular inhibin production, the effects of follicle-stimulating hormone (FSH), (Bu)2cAMP, germ cells (either crude or enriched preparations) and germ cell-conditioned media on inhibin production (immuno- and bio-activities) and the levels of alpha- and beta B-subunit mRNAs were assessed in cultured Sertoli cells isolated from 20-day-old rats. FSH and (Bu)2-cAMP stimulated both secreted and intracellular inhibin levels in a dose-dependent manner. Using cDNA probes corresponding to the alpha-subunit and the beta B-subunit of rat inhibin it was also shown that both FSH and (Bu)2cAMP markedly increased the level of alpha-subunit mRNA but had no effect on the beta B-subunit mRNA. Addition of a crude mixture of germ cells to Sertoli cell monolayers was found to enhance inhibin secretion. Of the different germ cell fractions tested in co-culture, early spermatids reproducibly stimulated both basal and (Bu)2cAMP-induced production of inhibin whereas pachytene spermatocytes only increased the latter; cytoplasts from elongated spermatids (CES) had no effect. Co-culture of Sertoli cells with liver epithelial cells (LEC) significantly enhanced (Bu)2cAMP-induced inhibin levels. Media conditioned by early spermatids consistently and dramatically stimulated the secretion of both bioactive and immunoactive inhibin by Sertoli cells while spent media from pachytene spermatocytes displayed less activity. CES-conditioned media had only minor stimulatory effects, which may have resulted from the contamination of this fraction by spermatids. Media conditioned by LEC had no effect on inhibin production, confirming that the activity of this cell line is not mediated via a diffusible factor. Early spermatids were found to increase levels of the alpha-subunit mRNA. The current study provides evidence for the involvement of germ cells, in particular of early spermatids, in the local testicular regulation of inhibin gene expression and production in the rat. This may be of crucial importance for the ontogeny of this parameter of Sertoli cell function, and has important implications with regard to the postulated endocrine and paracrine roles of inhibin.
Animals↗
Cell-cell interactions in the control of spermatogenesis as studied using Leydig cell destruction and testosterone replacement.
This review centers around studies which have used ethane dimethane sulphonate (EDS) selectively to destroy all of the Leydig cells in the adult rat testis. With additional manipulations such as testosterone replacement and/or experimental induction of severe seminiferous tubule damage in EDS-injected rats, the following questions have been addressed: 1) What are the roles and relative importance of testosterone and other non-androgenic Leydig cell products in normal spermatogenesis and testicular function in general? 2) What are the factors controlling Leydig cell proliferation and maturation? 3) Is it the Leydig cells or the seminiferous tubules (or both) which control the testicular vasculature? The findings emphasize that in the normal adult rat testis there is a complex interaction between the Leydig cells, the Sertoli (and/or peritubular) cells, the germ cells, and the vasculature, and that testosterone, but not other Leydig cell products, plays a central role in many of these interactions. The Leydig cells drive spermatogenesis via the secretion of testosterone which acts on the Sertoli and/or peritubular cells to create an environment which enables normal progression of germ cells through stage VII of the spermatogenic cycle. In addition, testosterone is involved in the control of the vasculature, and hence the formation of testicular interstitial fluid, presumably again via effects on the Sertoli and/or peritubular cells. When Leydig cells regenerate and mature after their destruction by EDS, it can be shown that both the rate and the location of regenerating Leydig cells is determined by an interplay between endocrine (LH and perhaps FSH) and paracrine factors; the latter emanate from the seminiferous tubules and are determined by the germ cell complement. Taken together with other data on the paracrine control of Leydig cell testosterone secretion by the seminiferous tubules, these findings demonstrate that the functions of all of the cell types in the testis are interwoven in a highly organized manner. This has considerable implications with regard to the concentration of research effort on in vitro studies of the testis, and is discussed together with the need for a multidisciplinary approach if the complex control of spermatogenesis is ever to be properly understood.
Animals↗
Changes in Sertoli cell function in vitro induced by nitrobenzene.
Nitrobenzene (NB) has been identified as a testicular toxicant in vivo, but its site of action remains unknown. In the present study, the effect of NB on the Sertoli cell was assessed in vitro using Sertoli cell and Sertoli-germ cell cocultures. The parameters measured were the exfoliation of germ cells; the secretion of lactate, pyruvate, and inhibin; and gross cellular morphology. The effect of metadinitrobenzene (mDNB), a related compound which is a known Sertoli cell toxicant, was assessed for comparison. Gross morphological changes including vacuolation of Sertoli cells were observed following treatment of cultures with 10(-3) M NB. Exposure of cocultures to NB also resulted in dose-dependent exfoliation of predominantly viable germ cells. NB (greater than 5 X 10(-4) M) and mDNB at the single dose level used (10(-4) M) stimulated the secretion of lactate and pyruvate significantly by Sertoli cells, an effect that was more marked in the absence of germ cells. Comparable changes were observed in follicle stimulating hormone (FSH)-stimulated cultures. Inhibin secretion by Sertoli cells was also altered by exposure to NB but in a biphasic manner, with low (10(-8) to 10(-6) M) and high (10(-4) to 10(-3) M) doses enhancing inhibin secretion while intermediate (10(-5) M) doses had no effect. These effects were evident in both culture systems but inhibin secretion by Sertoli-germ cell cocultures was always greater than that by Sertoli cell cultures. However, these effects of NB on inhibin secretion were not evident in FSH-stimulated cultures. In contrast to the effects of NB, mDNB had no effect on basal secretion of inhibin but blocked the stimulatory effect of FSH. It is concluded that NB, like mDNB, is probably a Sertoli cell toxicant in view of its similar disruptive effects on various parameters of Sertoli cell function. However, NB is far less toxic than mDNB at equivalent concentrations in vitro. The present study is the first to evaluate the potential of inhibin secretion by Sertoli cells in culture as an additional marker of toxicant action, and concludes that it merits further study in this context.
Animals↗
Intratesticular control of steroidogenesis.
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Animals↗
The levels and possible involvement of leukotriene B4 and prostaglandin F2 alpha in the control of interstitial fluid volume in the rat testis.
The possible involvement of two arachidonic acid metabolites, prostaglandin F2 alpha (PGF2 alpha) and leukotriene B4 (LTB4), in the stimulatory effect of human chorionic gonadotrophin (hCG) on the volume of interstitial fluid (IF) in the rat testis has been investigated. Administration of hCG caused a time- and dose-dependent increase in the IF levels of PGF2 alpha while LTB4 showed no clear dose-dependence, but did decrease significantly at 2-8 h after injection of 100 IU hCG. Administration of ethane dimethane sulphonate (EDS), which specifically destroys Leydig cells, decreased the volume of IF but the IF levels of LTB4 and PGF2 alpha were unchanged. This indicates that although the absence of Leydig cells results in significant changes in IF volume, LTB4 and PGF2 alpha are probably not involved in these changes. Furthermore, these findings suggest that Leydig cells are not the only contributors to LTB4 and prostaglandins E2 and F2 alpha in testicular IF. In rats injected peripherally with hCG, intratesticular administration of the cycloxygenase inhibitor, indomethacin, decreased PGF2 alpha levels in IF after 2 and 4 h but not at later times. However, no inhibitory effect of indomethacin on the hCG-stimulated increase in IF volume was detected; indeed, at 2 h after injection of indomethacin + hCG there was a significant increase in IF volume in the indomethacin-injected testis. It is concluded that, although prostaglandins and LTB4 are secreted into IF in the rat testis, these arachidonic acid metabolites are apparently not the primary mediators of the increase in IF volume that follows hCG treatment.
Analysis of Variance↗
The effects of sexual maturation and altered steroid synthesis on the production and route of secretion of inhibin-alpha from the rat testis.
This study has determined the route of secretion of inhibin-alpha into blood by the rat testis during sexual maturation, and in adult animals in which Leydig cell steroidogenesis was stimulated with human CG (hCG) or suppressed with aminoglutethimide. In each rat, inhibin-alpha levels were measured in samples of testicular (TV), spermatic (SV), and peripheral (PV) venous blood plasma, and in testicular interstitial fluid (IF). The IF and TV plasma reflect inhibin-alpha secretion via the base of the Sertoli cell while that secreted via the apex of the Sertoli cell (which is resorbed from the rete testis) was determined from the difference between SV and TV levels of inhibin-alpha. During sexual maturation, inhibin-alpha levels in IF and all plasma samples declined from maximal values at 28 days of age to minimal values at 100 days of age, in contrast to testosterone levels which showed the reverse pattern. There was a major change with age in the route of secretion of inhibin-alpha from the testis into blood. In immature (28-35 days) rats, most inhibin-alpha (58-65%) leaving the testis in blood was derived from that secreted via the base of the Sertoli cell with a relatively small contribution (35-42%) from apically-secreted inhibin-alpha. However, the latter made a progressively increasing contribution between 45 and 100 days of age (adults) and in adult rats the vast majority of inhibin-alpha (95%) leaving the testis in blood was derived from apically-secreted inhibin-alpha. This change was due primarily to a progressive reduction with age in the secretion of inhibin-alpha via the base of the Sertoli cell, a change which was confirmed by inhibin bioassay. Stimulation of steroidogenesis in the adult testis with hCG significantly increased inhibin-alpha and testosterone levels in IF and all plasma samples. The concomitant administration of hCG and aminoglutethimide (to block steroidogenesis) prevented the hCG-induced increase in testosterone levels, but still led to significant increases in inhibin-alpha secretion which were comparable to those seen with the use of hCG alone. The administration of aminoglutethimide (AMG) on its own did not alter the inhibin-alpha secretion profile from that seen in controls, but it did significantly reduce the levels of testosterone in all fluids. In rats treated with hCG +/- AMG there was a small change in the route of secretion of inhibin-alpha into blood, with an increased contribution (24-37%) from inhibin-alpha secreted via the base of the Sertoli cell, when compared with controls (7-16%).(ABSTRACT TRUNCATED AT 400 WORDS)
Aminoglutethimide↗
Focal disruption of spermatogenesis in the testis of adult rats after a single administration of human chorionic gonadotrophin.
The effect of a single injection of 100 i.u. human chorionic gonadotrophin (hCG) upon the morphology of the rat testis was studied by light and electron-microscopy from 12-48 h after treatment. Twelve hours after injection of hCG, emigration of leukocytes occurred across the intertubular blood vessels and, both at this time and at 24 h, infiltrations of leukocytes were observed within the extracellular tissue spaces. Furthermore, 12 h after hCG, the seminiferous epithelium showed focal disruption of spermatogenesis involving germ cell degeneration and pyknosis. Focal damage to the seminiferous epithelium persisted at 24 h and 48 h after injection of hCG, the affected seminiferous tubules showing failure of spermiation, accumulation of extracellular vacuoles and degeneration or partial loss of spermatogonia and primary spermatocytes. The observations show that, after stimulation of the Leydig cells with hCG, the intertubular tissue exhibits an inflammatory-type response and this is associated with focal tissue destruction in the seminiferous tubules. It is concluded that a high dose of hCG exerts anti-spermatogenic effects upon the testis and raises the possibility of unexpected interference with tests of normal Leydig cell function in both laboratory and clinical investigations.
Animals↗
Assessment of the contribution of Leydig cells to the secretion of inhibin by the rat testis.
Cultured Leydig cells secreted 1.3-4.3 ng 1-26 alpha-inhibin/10(6) cells/24 h, and although this was unaffected by human chorionic gonadotrophin (hCG), these cells could contribute to the intratesticular and blood levels of inhibin. The present study evaluated this contribution in rats in which the Leydig cells were destroyed by injection of ethane dimethane sulphonate (EDS). In these animals, inhibin levels increased in testicular interstitial fluid (IF), and in testicular (TV) and spermatic (SV) venous blood. In EDS-treated rats supplemented for 21 days with 1 or 25 mg testosterone esters to maintain full spermatogenesis and/or suppress the elevated follicle-stimulating hormone (FSH) levels and prevent Leydig cell regeneration, significant changes occurred in the levels of inhibin in IF, in TV and SV plasma and in the route of secretion of inhibin from the testis (i.e. via IF or seminiferous tubule fluid). However, none of these changes was related to the presence or absence of Leydig cells. It is concluded that Leydig cells make little contribution to the intratesticular and blood levels of inhibin in the adult rat.
Animals↗
Evaluation of the effect of selective germ cell depletion on subsequent spermatogenesis and fertility in the rat.
Rats were treated with a single high dose of methoxy acetic acid (MAA; 650 mg/kg) specifically to deplete seminiferous tubules of pachytene and later spermatocytes. The impact of this selective depletion on subsequent spermatogenesis, sperm output and fertility was then evaluated at intervals ranging from 3 days to 10 weeks. Cauda epididymal sperm number was reduced progressively beyond 2 weeks post-treatment and reached a nadir at 5-6 weeks (28-34% of control values) before recovering progressively back to control levels at 10 weeks. Sperm motility was reduced significantly at 4-7 weeks post-treatment with a nadir at 6 weeks (35% of control values). Thus, at 5-6 weeks after MAA treatment, motile sperm output was reduced by 82-88%. Despite these changes, there was little evidence for infertility in the majority of treated males during a serial mating trial. Evaluation of seminiferous tubule morphology combined with germ cell counts at stage VII of the spermatogenic cycle confirmed that, initially, MAA induced the specific loss of pachytene and later spermatocytes at all stages other than early to mid stage VII. Maturation depletion of germ cells at later intervals was consistent with the initial effects of MAA, although at 21 days post-treatment a number of unpredicted (? secondary) changes in spermatogenesis were observed. These were (a) a reduction in number of pachytene spermatocytes at late stage VII/early stage VIII, (b) retention of sperm at stages IX-XIV, and (c) increased degeneration of pachytene spermatocytes and round spermatids at stage VII and of secondary spermatocytes at stages XIV-I. Whilst none of these changes was severe, together they probably accounted for the unexpectedly prolonged drop in sperm output. It is concluded that whilst deleterious changes in spermatogenesis may occur secondarily following MAA treatment, for the most part spermatogenesis proceeds normally and fertility is largely maintained despite a massive but transient decrease in sperm output.
Acetates↗
Evaluation of the relative importance of endocrine and paracrine factors in control of the levels of inhibin in testicular interstitial fluid.
This study aimed to identify the role of endocrine (FSH, LH, testosterone) or paracrine (Leydig or germ cell) factors in control of the secretion of inhibin into testicular interstitial fluid (IF). This was done by measuring inhibin and testosterone levels in IF, and serum gonadotrophin and testosterone levels in adult rats following the destruction of Leydig cells with ethane dimethane sulphonate (EDS), alone or in combination with testosterone ester (TE) supplementation at various doses initiated at various times after EDS treatment. The effect of germ cell loss (induced by local testicular heating) on its own or in combination with the above treatments was also assessed. Treatment with EDS led to major increases in the levels of inhibin in IF and of FSH and LH in serum whilst testosterone levels in IF and serum fell to undetectable levels. Supplementation with TE (1-25 mg) for 21 days from the time of EDS treatment failed to prevent the initial (+3 days) increase in IF levels of inhibin but thereafter suppressed inhibin to control levels or lower and grossly suppressed FSH and LH levels, irrespective of whether the dose of TE administered did (25 or 5 mg) or did not (1 mg) prevent major seminiferous tubule damage. Partial regeneration of Leydig cells and normalization of testosterone levels occurred in rats 21 days after treatment with EDS alone but this failed to normalize inhibin and gonadotrophin levels. When supplementation with TE (25 mg) was initiated at 3, 6 or 9 days after EDS treatment, IF levels of inhibin were normalized within 3 days and maintained thereafter in parallel with suppression of serum FSH and LH to below control levels. Seminiferous tubule damage induced by local testicular heating (43 degrees C for 30 min) led to increased IF levels of inhibin 3 and 14 days later, in parallel with increased serum levels of FSH (but not LH). Suppression of FSH to subnormal levels in heat-exposed rats by TE treatment (25 mg) restored IF inhibin to control levels or below, a change which still occurred when Leydig cells were destroyed by EDS treatment. It is concluded that secretion of inhibin via the base of the Sertoli cell into testicular IF is controlled primarily by FSH, although local factors may play a minor role. These findings have important implications regarding the possible paracrine role(s) of inhibin in IF during puberty and in the normal adult testis.
Animals↗
Interstitial fluid volume in the rat testis: androgen-dependent regulation by the seminiferous tubules?
Regulation of testicular interstitial fluid (IF) volume has been investigated in adult male rats in which the Leydig cells were selectively destroyed with a single i.p. injection of ethane dimethane sulphonate (EDS). Following this treatment, some animals also received testosterone supplementation by s.c. injection every 3 days, beginning either from the time of EDS injection, or 3-12 days afterwards. The volume of IF obtained by drip collection was determined, and testosterone and gonadotrophin concentrations measured in blood and in IF. Testosterone levels in IF and serum became undetectable by 3 days after EDS treatment. IF volume was reduced by 50% (P less than 0.01) to reach a minimum level between 6 and 9 days after treatment. However, this decline was prevented in the absence of Leydig cells by supplementation with testosterone from the time of EDS injection, a treatment which also kept gonadotrophins at minimum or undetectable levels. Furthermore, the reduced IF volume seen up to 9 days after treatment with EDS alone could be restored to control levels within 3 days by a single injection of testosterone. The results obtained demonstrate that androgens, but not Leydig cells or gonadotrophins, are required for the maintenance of interstitial fluid volume in the adult rat testis. It is suggested that the seminiferous tubules may mediate this response, through an androgen-dependent mechanism.
Animals↗
The route of secretion of inhibin from the rat testis.
Levels of immunoactive and bioactive inhibin were measured in venous blood collected at a point just before (testicular venous) and after (spermatic venous) its passage through the mediastinal venous plexus over the anterior pole of the rete testis, and compared with levels in peripheral venous blood and testicular interstitial fluid (IF). In 15 control rats, levels of inhibin were highest in IF (8900 +/- 432 ng/l; mean +/- SEM) and lowest in peripheral (290 +/- 32 ng/l) and testicular (288 +/- 34 ng/l) venous blood, whilst levels in spermatic venous blood (633 +/- 99 ng/l) were always higher (P less than 0.002) than the levels in testicular venous blood. The latter difference was either reduced or abolished after disruption of spermatogenesis by local heating of the testes 8, 14, or 21 days previously, and by ligation of the efferent ducts for 6 h or more, but was not affected by acute removal of the epididymis. It is concluded that inhibin secreted into seminiferous tubule fluid may be reabsorbed from the rete testis and this may be the major route by which it reaches the peripheral bloodstream in rats with normal spermatogenesis.
Animals↗