Suppression of the intratesticular testosterone is associated with quantitative changes in spermatogonial populations in intact adult rats

To investigate the interactions between testosterone and FSH on quantitative aspects of spermatogenesis, intact adult rats were implanted sc with 0.5- to 50-cm long testosterone-filled silastic capsules (TC) for 8 weeks. Serum testosterone levels were suppressed slightly in rats bearing 1-cm TC implants but were elevated 6- to 30-fold when 5 cm or longer TC implants were used. These changes in serum testosterone were associated with a biphasic, dose-dependent response in testicular testosterone concentrations, reaching a minimum of 10-15% of the control values in rats bearing the 1- or 5-cm TC implants, but rebounding to 80% when 50-cm long TC implants were used. Meanwhile, serum FSH was comparably suppressed by 40-70%. Complete spermatogenesis was observed in all experimental animals with the exception of three rats with 1-cm TC implants in which elongated spermatids were absent or reduced in number. Enumeration of various cell types in the basal compartment of whole mounted seminiferous tubules revealed a 20% increase in type A1 spermatogonia in rats with the 1-cm TC implants. On the other hand, a 15-20% reduction in type B spermatogonia and preleptotene spermatocytes was noted in rats receiving 5-cm or longer TC implants. These results demonstrate that complete spermatogenesis can be maintained in intact animals in the presence of 10-15% of the normal testicular testosterone concentration. Failure to maintain normal testis weight, testicular testosterone concentration, or germ cell number at a 30-fold increase in serum testosterone demonstrates that high levels of serum testosterone may not be beneficial for quantitative spermatogenesis. Furthermore, both quantitative and qualitative differences were observed in different phases of spermatogenesis in the presence of comparable serum FSH and testicular testosterone concentrations. These results suggest that factors other than these two hormones are also involved in the regulation of spermatogenesis.     

Synergistic effects of follicle-stimulating hormone and testosterone on the maintenance of spermiogenesis in hypophysectomized rats: relationship with the androgen-binding protein status

The present study examined the relationship between the functional status of Sertoli cells and the maintenance and restoration of spermatogenesis in immature hypophysectomized (HPX) rats given various doses of exogenous testosterone with or without daily injections of FSH for 90 days. Subcutaneous implantation of a 2- to 10-cm testosterone capsule (TC) increased serum testosterone levels of HPX rats 2-10 times above the normal control levels, but did not significantly increase the testicular testosterone level. Daily injections of FSH significantly increased the accumulation of testosterone in testes of TC-implanted HPX rats. Maintenance of early spermiogenesis was observed in all TC-implanted animals. Although elongated spermatids were present, step 18-19 spermatids at the luminal edge of stages VII-VIII epithelium were only observed in rats bearing 10-cm TC implants. Daily injection of FSH resulted in the completion of spermiogenesis in all TC-implanted animals, and the number of step 18-19 spermatids was dependent on the length of TC implants used. These results demonstrate the importance of the synergism of FSH and testosterone in the final steps of spermiogenesis. The androgen-binding protein (ABP) content per testis of the HPX rats was stimulated by TC implants. However, a significant increase in epididymal ABP was only noted in rats bearing 10-cm TC implants. Injection of FSH resulted in a significant increase in the testicular ABP content in rats bearing 2- or 5-cm TC, but not in those with 10-cm TC implants. In addition, the epididymal ABP content was significantly stimulated by FSH in all TC-implanted animals. The ABP status in the testis and its transport toward the epididymis are closely related to the extent of maintenance of spermiogenesis. It is speculated that the production of ABP by Sertoli cells and the biochemical properties of ABP molecules may have some role in the control of the final steps of spermiogenesis.     

Hormonal control of spermatogenesis.

FSH and testosterone (T) secretion are essential for the successful completion of spermatogenesis. Because there are no receptors for FSH or testosterone on germ cells, there are intermediate steps in this action, the nature of which are unknown. However, as the Sertoli cell contains receptors for both FSH and T, it is likely that these hormones exert their influence on germ cells by modulating Sertoli cell function. Both FSH and T exert synergistic actions on germ cells, but T has a specific action on the later stages of spermatid maturation. FSH, by its ability to stimulate Sertoli cell mitosis during testicular development, can influence the spermatogenic capacity of the adult testis.     

Sertoli and germ cell development in hypogonadal (hpg) mice expressing transgenic follicle-stimulating hormone alone or in combination with testosterone

We recently created a novel transgenic (tg) model to examine the specific gonadal actions of FSH, distinct from LH effects, by expressing tg-FSH in gonadotropin-deficient hypogonadal (hpg) mice. Using this unique in vivo paradigm, we now describe the postnatal cellular development in seminiferous tubules selectively stimulated by tg-FSH alone or combined with testosterone (T). In the alphabeta.6 line, tg-FSH stimulated the maturation and proliferation ( approximately 2-fold) of Sertoli cells in hpg testes. Total Sertoli cell numbers were also significantly increased (1.5-fold) independently of FSH effects by T treatment alone. Selective FSH activity in alphabeta.6 hpg testes increased total spermatogonia numbers 3-fold, which established a normal spermatogonia/Sertoli cell ratio. FSH also elevated meiotic spermatocyte numbers 7-fold, notably at pachytene (28-fold), but induced only limited numbers of postmeiotic haploid cells (absent in hpg controls) that arrested during spermatid elongation. In contrast, T treatment alone had little effect on postnatal spermatogonial proliferation but greatly enhanced meiotic progression with total spermatocytes increased 12-fold (pachytene 53-fold) relative to hpg testes, and total spermatid numbers 11-fold higher than tg-FSH hpg testes. Combining tg-FSH and T treatment had no further effect on Sertoli or spermatogonia numbers relative to FSH alone but had marked additive and synergistic effects on meiotic cells, particularly pachytene (107-fold more than hpg), to establish normal meiotic germ cell/Sertoli cell ratios. Furthermore, tg-FSH had a striking synergistic effect with T treatment on total spermatid numbers (19-fold higher than FSH alone), although spermatid to Sertoli cell ratios were not fully restored to normal, indicating elevated Sertoli cell numbers alone are insufficient to establish a maximal postmeiotic germ cell capacity. This unique model has allowed a detailed dissection of FSH in vivo activity alone or with T and provided compelling evidence that FSH effects on spermatogenesis are primarily via Sertoli and spermatogonial proliferation and the stimulation of meiotic and postmeiotic germ cell development in synergy with and dependent on T actions.     

Dissociation of qualitative and quantitative effects of the suppression of testicular testosterone upon spermatogenesis

The present study examined the quantitative and qualitative effects of suppressed testicular testosterone concentration upon spermatogenesis. Adult male rats were given sc implants of 0.5- to 5-cm long testosterone filled silastic capsules (TC) for 10 weeks. Testicular testosterone concentrations were suppressed to below 5% of the control level in rats receiving 1.5- to 5-cm TC implants. Spermatogenesis was arrested at early spermatids in rats given 1.5-cm TC implants, but was maintained in those given 3- or 5-cm TC implants. The number of A1 spermatogonia was 10-15% higher in rats bearing 0.75- to 1.5-cm TC implants, while a 5-15% reduction in preleptotene spermatocytes was noted in rats receiving 1.5-cm or longer TC implants. Linear regression analysis revealed a significant negative correlation between the number of A1 spermatogonia and the percent yield of preleptotene spermatocytes. This result illustrates the presence of a negative feedback mechanism in the regulation of A1 spermatogonia. The lack of a correlation between the quantitative changes in germ cell number and the hormonal states suggests that factors or local mechanisms independent of the concentration of hormones are involved in the regulation of germ cell differentiation.     

The effect of testosterone, dihydrotestosterone and oestradiol on the re-initiation of spermatogenesis in the adult photoinhibited Djungarian hamster

The roles of testosterone (T) and its metabolites on hamster spermatogenesis are poorly defined. This study assessed the effects of T, dihydrotestosterone (DHT) and oestradiol (E) on the re-initiation of spermatogenesis in the adult Djungarian hamster. Hamsters raised under long photoperiods (LD, 16 h light:8 h darkness) were exposed to short photoperiods (SD, 8 h light:16 h darkness) for 11 weeks to suppress gonadotrophins. Groups of eight animals then received T, DHT and E for 5 weeks. Cell numbers were determined using the optical disector (sic). The number of Sertoli cells was suppressed in SD controls to 48% (P < 0.001) of LD control and restored either fully or partially by exogenous DHTand E (2.6- and 1.8-fold above SD levels) respectively, corresponding with a twofold elevation of serum FSH. The number of germ cells in SD animals was reduced (all P < 0.001) to levels reported. The number of type A spermatogonia increased in line with the rise in Sertoli cell number, by 2.6-fold (P < 0.01) and 1.8-fold (NS) above SD controls after DHT and E treatments respectively. DHT increased the number of type B spermatogonia/preleptotene spermatocytes, leptotene/zygotene and pachytene spermatocytes by 3.5-, 5.7- and 21-fold above SD (all P < 0.01) respectively, compared with a 2.2-fold (P < 0.01), 2.4-fold (not significant, NS) and 6-fold (NS) in E-treated animals respectively. Exogenous T had little effect on cell numbers or serum FSH compared with SD controls. Spermatids were rarely observed after steroid treatment. We believe this study suggests that steroids can regulate the re-initiation of early spermatogenic cells via a mechanism which includes FSH.     

Intratesticular injection of glycerol as a model for studying the quantitative relationship between spermatogenic damage and serum FSH

Intratesticular injection of glycerol (1,2,3-trihydroxypropane) was evaluated as an experimental approach for studying the relationship between spermatogenic activity, i.e. relative proportion of euspermatogenic and aspermatogenic tubules in the testis, and serum concentrations of FSH. Adult rats received a single intratesticular injection of either 400 microliters distilled water or five different doses of glycerol, ranging from 25 to 400 microliters, and were killed after 1 or 4 weeks respectively. Injection of glycerol caused focal destruction, so that the same testes contained intact tubules, tubules with spermatogonia and Sertoli cells only, and tubules devoid of cellular material. There was a close correlation (r = 0.896) between the frequency of intact tubules and the dose of glycerol, and a similarly strong correlation (r = 0.908) between acellular tubules and the dose of glycerol. A correlation existed between FSH and euspermatogenic tubules (r = -0.758, n = 122) and, conversely, between FSH and acellular tubules (r = 0.820, n = 122), while the correlation for tubules containing spermatogonia and Sertoli cells was only marginal (r = -0.055). The exact relationship between FSH and spermatogenic activity in the testis was used to estimate the minimal amount of spermatogenic tissue necessary for maintaining normal serum concentrations of FSH. Only the disruption of spermatogenesis in more than 30% of seminiferous tubules in each testis caused increased serum concentrations of FSH.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hormonal induction of all stages of spermatogenesis in vitro in the male Japanese eel (Anguilla japonica).

The importance of gonadotropins and androgens for spermatogenesis is generally accepted in vertebrates, but the role played by specific hormones has not been clarified. Under cultivation conditions, male Japanese eels (Anguilla japonica) have immature testes containing only premitotic spermatogonia, type A and early-type B spermatogonia. In the present study, a recently developed organ-culture system for eel testes was used to determine in vitro effects of various steroid hormones on spermatogenesis. After 9 days of culture in serum-free, chemically defined medium containing 11-ketotestosterone (10 ng/ml), a major androgen in male eels, type A and early-type B spermatogonia began mitosis, producing late-type B spermatogonia. After 18 days, zygotene spermatocytes with synaptonemal complexes appeared, indicating that meiosis had already started by this time. In testis fragments cultured for 21 days, round spermatids and spermatozoa were observed with spermatogenic cells at all stages of development. Addition of 11-ketotestosterone to the culture medium also caused a marked cytological activation of Sertoli cells. No other steroid hormones tested had such stimulatory effects. These results, together with our earlier observations, suggest the following sequence for the hormonal induction of spermatogenesis in eel testes; gonadotropin stimulates the Leydig cells to produce 11-ketotestosterone, which, in turn, activates the Sertoli cells leading to the completion of spermatogenesis. This is, thus, an example of an animal system in which all stages of spermatogenesis have been induced by hormonal manipulation in vitro.     

Restoration of spermatogenesis by high levels of testosterone in hypophysectomized rats after long-term regression

In order to correlate the levels of intratesticular testosterone and the status of spermatogenesis, the present study examined the spermatogenic responses of mature chronically hypophysectomized (HPX) rats to different regimens of testosterone (T) replacement, i.e. implantation of a 2 X 5 or 6 X 5 cm long testosterone capsule (TC), or injection of 25 mg or 100 mg of testosterone enanthate (TE) every 4 days for 90 days. These regimens restored the testicular testosterone of the HPX rats to 25-96% of that measured in normal control rats. The testis weight of untreated HPX rats was below 30% of the normal control values. It was restored to 60-80% of control values in the TC implanted rats and to 50% in those receiving TE injection. Complete spermatogenesis of HPX rats was restored in those receiving TC implants and 100 mg TE injections. It was incomplete in those given 25 mg TE injections. Quantitative evaluation of germ cells revealed that spermatogonial populations of HPX rats were restored to 70 and 50% of the normal levels in rats receiving TC implants and TE injection, respectively. Differentiation of these cells resulted in the population of preleptotene spermatocytes to the same extent as those observed in spermatogonia. The yield of spermatids in both TC- and TE-treated HPX rats was below 50% of normal controls. Ratios between two successive generations of germ cells in stage VII epithelium revealed both dosage and regimen effects upon the yield of meiotic cells and spermatids. These results suggest that both T concentration and the mode of T availability to the testis may be important for specific steps of germ cell development in different stages of the cycle of the seminiferous epithelium. The results of the present study demonstrate that as little as 25% of normal testicular T concentration is sufficient to support all stages of spermatogenesis. Failure to restore a normal germ cell number even in the presence of a normal testicular T concentration suggests the need of other factors for quantitative spermatogenesis. Furthermore, despite a higher testicular T concentration achieved by TE injections, the restoration of spermatogenesis was less pronounced by this regimen. This finding suggests that the consistency of testicular T may also be important for normal spermatogenesis.     

Induction of precocious maturation of spermatogenesis in infant rats by human menopausal gonadotropin and inhibition by simultaneous administration of gonadotropins and testosterone

This study was undertaken to determine if the initiation of spermatogenesis could be modified by the administration of gonadotropins and sex hormones in infant rats. Five-day-old rats were injected daily between the 5th and 11th days of life with test substances and killed on day 15. Administration of testosterone propionate (TP; 2.5 mg daily), human menopausal gonadotropin (hMG; 7.5 IU daily), or coadministration of both of these substances (TP + hMG) or administration of estradiol benzoate (15 micrograms daily) caused quantitative changes in premeiotic spermatogenesis, as measured by the mean cell counts per tubule cross-section. hMG caused an increased yield of type A1 spermatogonia (SgA1) from undifferentiated type A spermatogonia (UnA) and increased the yield of type B spermatogonia from SgA1. TP was not effective in stimulating first premeiotic spermatogenesis, and in contrast to hMG, it had a negative influence on the numbers of UnA and SgA1 and on the volume of Sertoli cell nucleus. Administration of TP + hMG or estradiol benzoate resulted in a significant increase in the numbers of UnA and SgA1, but inhibited cell differentiation. TP + hMG significantly reduced the rate of premeiotic spermatogenesis. The results demonstrate that precocious numerical stabilization of premeiotic spermatogenesis can be achieved by the application of hMG. TP applied alone was able to induce peripheral androgenic effects (seminal vesicle weight) 100% greater than those produced by administration of hMG, but was not able to stimulate seminal tubule function. TP applied together with hMG produced inhibition of spermatogenesis. This effect might be due to the inhibition of Sertoli cell function by the direct influence of testosterone. In contrast to testosterone, estradiol may play a stimulatory role in the multiplication of the reserve stem cells of the first spermatogenesis of the rat.     

Transferrin and sulfated glycoprotein-2 messenger ribonucleic acid levels in the testis and isolated Sertoli cells of hypophysectomized rats

Both FSH and testosterone act on Sertoli cells in the testis. It is possible that the action of these hormones on Sertoli cells results in an increased capacity for the cells to carry out their prescribed functions, among which are the synthesis and secretion of specific glycoproteins. Changes in the testicular levels of two specific mRNAs in hypophysectomized hormone-treated rats were determined by solution hybridization to cRNA probes. The mRNAs coding for transferrin and sulfated glycoprotein-2 (SGP-2), both of which are secretion products of Sertoli cells, decreased dramatically in the testis of hypophysectomized rats that were maintained for 20 days untreated with hormones. If hypophysectomy was done to rats at 20 days of age, daily injections for a subsequent 20 days with FSH or FSH in combination with testosterone partially maintained both transferrin and SGP-2 mRNA levels. Testosterone alone was ineffective in 20-day-old rats. In contrast, if hypophysectomy was performed on 40-day-old rats, daily injections of testosterone alone or in combination with FSH were most effective in maintaining higher levels of the specific mRNAs. When the Sertoli cells from rats hypophysectomized at 20 days of age were placed in cell culture, FSH again was most effective in the stimulation of transferrin mRNA above control levels. However, when the Sertoli cells from the rats hypophysectomized at 40 days of age were placed in culture, FSH was slightly stimulatory, but testosterone had no effect on the transferrin mRNA levels. Neither FSH nor testosterone affected the levels of SGP-2 mRNA in the cultured cells regardless of the age of the animal at the time of hypophysectomy. Additional in vivo studies were done in which the rats were hypophysectomized at 20 days of age, allowed to regress for 17 days, and then injected daily with hormones for 3 days. The levels of transferrin and SGP-2 mRNA in this experiment were stimulated by FSH alone or by a combination of FSH and testosterone to an extent similar to that in the cultured cells. These studies showed that FSH is most important in the younger rats and testosterone is most important in the older rats in the maintenance of specific mRNA levels. In addition, the level of stimulation observed with either hormone is different depending on whether the hormone is given in culture or in vivo.(ABSTRACT TRUNCATED AT 400 WORDS)     

Quantitative cytological studies of spermatogenesis in intact and hypophysectomized rats: identification of androgen-dependent stages

A stereological study of the numbers of germ cells in various stages of spermatogenesis was undertaken in testosterone-treated intact and hypophysectomized (HPX) rats. Adult Sprague-Dawley rats were given testosterone by Silastic implants, which either inhibited (3-cm length) or partially maintained (10 cm) spermatogenesis over a 13-week period. The numbers of nuclei of the various germ cell categories (spermatogonia, spermatocytes, and round spermatids) in the testes were estimated by profile counting and measurement of nuclear diameter. The numbers of elongated spermatids were determined separately in testicular homogenates. Testis weight, seminiferous tubule volume, and tubule diameter were significantly decreased in intact rats with 3- and 10-cm testosterone implants and in HPX rats, although they were partially maintained in groups with 10-cm implants compared to those in groups with 3-cm implants (P less than 0.05). The effect of 3-cm testosterone implants in the intact group was to suppress the number of spermatogonia to 57%, reduce the conversion of spermatogonia to spermatocytes to 85%, and reduce the conversion of round to elongated spermatids to 19% of the control value. This latter effect was largely overcome with 10-cm testosterone implants. In HPX rats, only 10-cm implants were effective in maintaining the conversion of round spermatids to elongated spermatids. However, testosterone alone was less effective in maintaining the conversion of spermatocytes to round spermatids, suggesting that a pituitary factor, probably FSH, was involved. It is concluded that testosterone has a major effect on the conversion of round to elongated spermatids. The conversion of spermatogonia to spermatocytes and the conversion of spermatocytes to round spermatids depend on the synergistic action of both FSH and testosterone. However, the effect of FSH is greatest on the conversion of spermatocytes to spermatids, i.e. meiosis.     

Dynamics of the follicle-stimulating hormone (FSH)-inhibin B feedback loop and its role in regulating spermatogenesis in the adult male rhesus monkey (Macaca mulatta) as revealed by unilateral orchidectomy

The purpose of this study was to document the morphological changes in the seminiferous epithelium that underlie the compensatory testicular hypertrophy observed in response to unilateral orchidectomy (UO) in the adult rhesus monkey and to describe the concomitant response in the endocrine feedback loops controlling testicular function in this species. Adult male monkeys were implanted with indwelling venous catheters; seven animals were then subjected to UO (data are presented from six) and three to sham UO. Profiles of circulating concentrations of FSH, LH, testosterone (T), inhibin B, and pro-alpha-C were monitored in 12-h series of sequential blood samples collected before, on the day of UO (day 0), and on days 1, 2, 4, 8, 16, 32, and 42 or 43 after UO. In the UO monkeys, the remaining testis was taken on day 44. Sertoli and germ cells in the removed and remaining testes were counted and expressed either as number per testis or, in the case of the differentiated spermatogonia (B1, B2, B3, and B4), as number per cross-section of the seminiferous tubule. UO was associated with a marked increase in the number of all germ cells more mature than undifferentiated spermatogonia (Ap) in the remaining testis. Sertoli cell number, on the other hand, did not change, and it is therefore reasonable to propose that the primary locus of the spermatogenic compensation was the differentiated spermatogonia. The additional finding that the relationship between the number of Sertoli cells and total germ cells in the remaining testis became robust (r = 0.92; P < 0.01 vs. r = 0.44; P > 0.05 for the removed testis) indicated that in the monkey, spermatogenesis does not normally operate at its ceiling. The increased drive to the seminiferous tubule of the remaining testis is hypothesized to be mediated by the sustained increase in FSH secretion that was observed after UO, although a role for increased testicular T production cannot be excluded. The stimulus for increased FSH secretion was presumably provided by the abrupt, 50% decline in circulating inhibin B levels. Interestingly, inhibin B secretion by the remaining testis was not dramatically affected by UO, and therefore, the deficit in circulating levels of this hormone and thus the error signal to FSH secretion were maintained for the duration of the experiment. In contrast, the changes in circulating LH and T concentrations were only transient, and within 48 h of UO, these hormonal parameters had returned to control values. The mechanisms by which the remaining testis rapidly acquires the capacity to double T production in the face of an unchanging LH drive remains to be determined. The foregoing body of evidence suggests that sperm output by the monkey testis is regulated by the circulating concentration of FSH and that in physiological situations, FSH secretion is insufficient to stimulate spermatogenesis to its ceiling. The results also indicate that FSH secretion is controlled by a feedback system in which the feedforward arm (FSH-inhibin B) is less robust than the feedback loop (inhibin B-FSH). Thus, a decrease in the inhibin B feedback signal results in a sustained increase in FSH secretion that drives the testes toward their spermatogenic ceiling, which is presumably set by Sertoli cell number.     

FSH receptor mRNA is expressed stage-dependently during rat spermatogenesis.

In situ hybridization was performed on testicular tissue from adult male Sprague-Dawley rats using cRNA antisense and sense probe of the monkey FSH receptor (FSHR) cDNA to determine the cellular site of synthesis, and possible stage-dependent expression of FSHR mRNA during the cycle of the seminiferous epithelium. Using antisense probe specific binding was first detected in Sertoli cells just prior to sperm release at stage VIII. The strongest specific hybridization signal was found during stages IX and X followed by a decrease of signal intensity in stages XI-XII. No specific binding was found in stages XIII-VII. The sequence of events with the maximum expression of FSHR mRNA in Sertoli cells in stages IX and X, before FSH-binding and FSH-stimulated cAMP production reach maximum values, coincides with a new wave of spermatogenesis and indicates an effect of FSH and spermatogenic cells on the regulation of FSHR mRNA expression.     

Inhibition of rat Sertoli cell aromatase by factor(s) secreted specifically at spermatogenic stages VII and VIII

The effect of spent medium recovered from cultured seminiferous tubules on the response of Sertoli cells to FSH was investigated. 4 pools of tubules at different stages of the spermatogenic cycle (VII–VIII, IX–XII, XIII–I, II–VI) were identified by transillumination and cultured for 20 h at 32°C. Then the culture media were added to Sertoli cell cultures and the FSH-dependent secretion of estradiol into the medium after 24-h incubation was measured by radio immunoassay.The spent media from tubules at stages IX–XII, XIII–I and II–VI did not significantly alter the FSH response of the Sertoli cells, while medium from stages VII–VIII inhibited the FSH dependent estradiol secretion in a dose-dependent manner. The inhibiting activity was not destroyed by heating at 60°C for 30 min, and it was not affected by repeated freezing and thawing or 24-h dialysis against fresh medium. The inhibition upon estradiol production was reversible after removal of the inhibitory agent(s) and subsequent culture of Sertoli cells under control conditions. The Sertoli cells had a good viability, as evidenced by trypan-blue exclusion and the undetectable levels of LDH in the medium after culture in the presence of the spent medium from stages VII–VIII.The inhibition of the FSH-induced aromatase activity in cultured immature Sertoli cells does not seem to be due to interference with FSH-receptor interactions or to an inhibition of adenylate cyclase because the dibutyryl-cAMP-induced increase in aromatase activity was also inhibited.These results are discussed in relation to the communication between Sertoli cells and germ cells in different stages of spermatogenesis and the possible physiological significance of the inhibitor(s) on the Sertoli cell activity even in the mature seminiferous tubule.     

Differential effects of FSH and testosterone on the maintenance of spermatogenesis in the adult hypophysectomized rat

In order to clarify further the role of FSH in the maintenance of spermatogenesis, adult rats were treated with purified human FSH (2 X 5 IU/day per rat), testosterone (1.5 cm silicone elastomer implant) or a combination of both hormones for 2 weeks following hypophysectomy. After hypophysectomy alone, no elongate spermatids were observed and the numbers of pachytene spermatocytes and round spermatids observed were reduced when compared with untreated controls. Testosterone supplementation alone qualitatively maintained the formation of elongate spermatids in most seminiferous tubules, whilst in FSH-treated rats increased numbers of round spermatids and pachytene spermatocytes were observed when compared with hypophysectomized animals. Formation of elongate spermatids, however, did not occur under FSH treatment alone. A combination of FSH and testosterone treatment maintained spermatogenesis in an almost quantitative fashion. Numbers of pachytene spermatocytes and round spermatids were maintained at about 80% of levels seen in intact control animals. Treatment with FSH or testosterone alone maintained testis weights at significantly higher levels than those seen in hypophysectomized controls (FSH, 0.79 +/- 0.05 g; testosterone, 0.81 +/- 0.07 g; hypophysectomized, 0.50 +/- 0.04 g). Animals treated with FSH and testosterone showed testis weights 20% below control values (1.22 +/- 0.05 vs 1.51 +/- 0.06 g; P less than 0.05). No increases in intratesticular or intratubular androgen concentrations or in testosterone: dihydrotestosterone ratios were observed in any of the hormone-treated groups when compared with hypophysectomized controls. In all hypophysectomized animals testicular androgen concentrations were reduced to less than 5% of control values. The results obtained in this study suggest that FSH is involved in the maintenance of spermatogenesis in the adult rat and that the effects of FSH are not mediated through changes in intratesticular androgens. Low levels of testosterone in combination with FSH can almost quantitatively maintain spermatogenesis in adult rats.     

Complete Sertoli cell proliferation induced by follicle-stimulating hormone (FSH) independently of luteinizing hormone activity: evidence from genetic models of isolated FSH action

Defining the gonadal effects of FSH distinct from those of LH remains difficult. We have characterized and compared the level of Sertoli and germ cell development in three mouse models recently created to isolate FSH activity from LH effects. Two models used LH-deficient hypogonadal (hpg) mice to selectively study either pituitary-independent transgenic (tg) FSH or ligand-independent activated tg FSH receptor (FSHR(+)) expression, and the third model used LH receptor (LHR)-deficient mice to isolate and examine endogenous mouse FSH effects. Stereological evaluation revealed tg-FSH or tg-FSHR(+) activity significantly increased total Sertoli cell numbers per testis in both hpg models relative to control hpg testes. Furthermore, tg-FSH dose-dependently restored hpg Sertoli cells to wild-type (wt) (non-hpg) levels, and LHR-/- testes also exhibited wt Sertoli numbers. Spermatogonial proliferation and meiotic development were enhanced by tg-FSHR(+) or tg-FSH. Despite producing normal Sertoli numbers, isolated tg-FSH activity only increased total spermatogonia and spermatocyte populations to 57 and 44% of wt, which was comparable to spermatogonia and spermatocyte numbers observed in LHR-null testes (45 and 34% of wt). Selective FSH activity initiated round spermatid formation in all three models. However, elongated spermatid formation was detected in tg-FSH and tg-FSHR(+) hpg testes but not in LHR-/- testes, which may reflect even lower intratesticular testosterone levels in LHR-null compared with hpg testes. FSH increased round and elongated spermatid numbers in hpg testes to 16 and 6% of wt without altering intratesticular testosterone levels, but failed to produce spermatozoa demonstrating the inability of FSH to complete spermatogenesis. These findings revealed that full Sertoli cell proliferation can be accomplished by FSH activity without LH requirement, and although postnatal mitotic and meiotic germ cell development can be promoted by FSH alone, LH-mediated effects remain a critical determinant for initiating the full complement of germ cells and final stages of postmeiotic development.     

Follicle-stimulating hormone-induced, adenosine 3',5'-monophosphate-mediated movement of immature rat sertoli cells in primary culture.

Sertoli cells dissociated from 10-day-old rat testes form colonies in primary culture in response to FSH. FSH and dibutyryl cAMP stimulated the attachment of Sertoli cells to an equal extent; however, FSH-treated cultures contained a small number of large colonies while dibutyryl cAMP-treated cultures contained a large number of small colonies. This relationship was not altered by the addition of a number of other peptide or steroid hormones. Extracellular cAMP levels and colony density were negatively correlated. Colony size at 24 h of culture was diminished in FSH-treated cultures by the addition of a cAMP antibody at 6 or 12 h of incubation. The addition of cAMP at 24 h to FSH-treated cultures caused a dose-dependent stimulation of colony size but not colony density at 48 h of culture. A point source of cAMP (4 x 10(-3) M in agar) inhibited migration of cells toward the agar spot. An agar spot on the dish substratum containing Sepharose-bound FSH exhibited a halo of cells next to the spot, with a zone lacking cells distal to it. Radioautographs of [125I]iodo-FSH-treated cultures exhibited a nonhomogeneous distribution of silver grains; colony size increased faster than the number of labeled cells. Taken together, the results suggest that 1) FSH is stimulating the attachment of Sertoli cells through an increase in intracellular cAMP, 2) FSH is promoting active aggregation of Sertoli cells in culture through a modulation of extracellular cAMP, and 3) cells with a large amount of bound FSH are acting as centers for aggregation.