Inactive Testes Research Articles

Sexual maturation in male bowhead whales (Balaena mysticetus) of the Bering-Chukchi-Beaufort Seas stock

Since the mid-1970s, study has focused on reproductive biology of female bowhead whales, while little has been described for males. This study evaluates testicular morphology (mass and length) and histology in relation to body length to determine the onset of male sexual maturity. Mean testis mass and mean testis length were highly correlated. Body length and mean testis mass were significantly correlated and an inflection of increased testicular mass occurred at approximately 12.5-13.0m suggesting the onset of puberty, and also indicated by histologic findings. Biological variability and the fact that few male animals have been examined within this critical length cohort do not allow determination of the onset of maturity with higher precision. Too few mature males have been landed in spring to make statistical comparisons of testes mass with autumn-landed animals within specific size cohorts. Two large (15.7m and 17.7m) males landed in spring had relatively small inactive testes and were diagnosed as pseudohermaphrodites; body length and mean testis length and seminiferous tubule diameter (STD) were not correlated with the other ‘normal’ whales. The smallest male confirmed as mature based on the presence of spermatocytes was 12.7m. The largest testes measured (combined mass 203kg) were from a whale landed in autumn. Mean STD for individual whales ranged from 33.3-170.9m and increased with mean testis weight and whale length. The STD is similar within a testis regardless of region evaluated, with minor variability. Some variation was noted for transverse sections within a cross section for some whales but no pattern was evident.

Cell adhesion and immune response, two main functions altered in the transcriptome of seasonally regressed testes of two mammalian species.

In species with seasonal breeding, male specimens undergo substantial testicular regression during the nonbreeding period of the year. However, the molecular mechanisms that control this biological process are largely unknown. Here, we report a transcriptomic analysis on the Iberian mole, Talpa occidentalis, in which the desquamation of live, nonapoptotic germ cells is the major cellular event responsible for testis regression. By comparing testes at different reproductive states (active, regressing, and inactive), we demonstrate that the molecular pathways controlling the cell adhesion function in the seminiferous epithelium, such as the MAPK, ERK, and TGF-β signaling, are altered during the regression process. In addition, inactive testes display a global upregulation of genes associated with immune response, indicating a selective loss of the "immune privilege" that normally operates in sexually active testes. Interspecies comparative analyses using analogous data from the Mediterranean pine vole, a rodent species where testis regression is controlled by halting meiosis entry, revealed a common gene expression signature in the regressed testes of these two evolutionary distant species. Our study advances in the knowledge of the molecular mechanisms associated to gonadal seasonal breeding, highlighting the existence of a conserved transcriptional program of testis involution across mammalian clades.

Mediterranean Pine Vole, Microtus duodecimcostatus: A Paradigm of an Opportunistic Breeder.

Simple SummaryIn temperate zones of the Earth, some mammalian species reproduce seasonally whereas others do it continuously. Other species are summer breeders in the north and winter breeders in the south. Thus, the reproductive pattern seems not to be a species-specific but a population-specific trait. We investigated the reproduction pattern of the Mediterranean pine vole, Microtus duodecimcostatus, in the area around the city of Granada in Southern Spain, and found that individuals living in wastelands reproduce seasonally whereas those living in close poplar plantations (just 8 km apart) reproduce throughout the year, as did voles captured in wastelands and kept in captivity. These animals represent thus a paradigm of an opportunistic breeder as particular individuals stop breeding or not, depending on the environmental conditions they face at any moment. Sexually inactive male voles undergo complete testis inactivation and their sperm production is halted. The immune system in active testes is depressed, a phenomenon known as “immune privilege” that protect germ cells from autoimmune attack. We studied gene activity in active and inactive testes and our results indicate that such an immune privilege is lost in inactive testes, suggesting an important role for this process during testis regression.Most mammalian species of the temperate zones of the Earth reproduce seasonally, existing a non-breeding period in which the gonads of both sexes undergo functional regression. It is widely accepted that photoperiod is the principal environmental cue controlling these seasonal changes, although several exceptions have been described in other mammalian species in which breeding depends on cues such as food or water availability. We studied the circannual reproductive cycle in males of the Mediterranean pine vole, Microtus duodecimcostatus, in the Southeastern Iberian Peninsula. Morphological, hormonal, functional, molecular and transcriptomic analyses were performed. As reported for populations of other species from the same geographic area, male voles captured in wastelands underwent seasonal testis regression in summer whereas, surprisingly, those living either in close poplar plantations or in our animal house reproduced throughout the year, showing that it is the microenvironment of a particular vole subpopulation what determines its reproductive status and that these animals are pure opportunistic, photoperiod-independent breeders. In addition, we show that several molecular pathways, including MAPK, are deregulated and that the testicular “immune privilege” is lost in the inactive testes, providing novel mechanisms linking seasonal testosterone reduction and testis regression.

Chromosome abnormalities in dogs with disorders of sex development (DSD)

Disorders of sex development (DSD) caused by chromosome abnormalities are rarely diagnosed in dogs. In this report, there is a focus on five DSD cases in which the dogs had abnormal karyotypes. All animals were recognized by owners as females, however, these dogs had a large number of reproductive defects. Among these were abnormal external genitalia such as an enlarged clitoris, abnormal development of the labia, abnormal location of the vulva and urethral orifice, and other abnormalities were observed in four dogs. Gonadal histology assessments were conducted on three dogs and there were diagnoses of the presence of an ovary, inactive testes, and ovotestis with calcification in ovarian follicles. Results from cytogenetic analysis indicated there were the following karyotypes: (a) X trisomy in a mosaic form (79,XXX/78,XX); (b) Robertsonian translocation in a mosaic form (77,XX,rob/78,XX); (c) nonmosaic X/autosome translocation (78,X,t(X;A)); (d) X/autosome translocation in a mosaic form (78,X,t(X;A)/78,XX); and (e) leukocyte chimerism (78,XX/78,XY). The findings in the present study, emphasize that cytogenetic analysis is essential for elucidating the pathogenesis of DSD in dogs.

Blood-Testis Barrier and Sperm Delayed in the Cauda Epididymis of the Reproductively Regressed Syrian Hamsters.

The Syrian (golden) hamsters are seasonal breeders whose reproductive functions are active in summer and inactive in winter. In experimental facility mimicking winter climate, short photoperiod (SP) induces gonadal regression. The blood-testis barrier (BTB) of the sexually involuted animals have been reported to be permeable, allowing developing germ cells to be engulfed or sloughed off the epithelium of the seminiferous tubules. The expressions of genes related to the tight junction composing of BTB were investigated in the reproductive active and inactive testes. Claudin-11, occludin, and junctional adhesion molecule (JAM) were definitely expressed in the active testes but not discernably detected in the inactive testes. And spermatozoa (sperm) were observed in the whole lengths of epididymides in the active testes. They were witnessed in only cauda region of the epididymides but not in caput and corpus regions in animals with the inactive testes. The results imply that the disorganization of BTB is associated with the testicular regression. The developing germ cells are swallowed into the Sertoli cells or travel into the lumen, as supported by the presence of the sperm delayed in the last region of the epididymis. These outcomes suggest that both apoptosis and desquamation are the processes that eliminate the germ cells during the regressing stage in the Syrian hamsters.

Sexual development and maturity of the male brown banded bamboo shark,

The present study aimed to assess the reproductive biology of male brown banded bamboo shark (Chiloscyllium punctatum). Morphological and histological changes in the reproductive tract and changes in gonad steroid testosterone were assessed in 110 male C. punctatum collected from Bahrakan Creek between January 2018 and March 2019. The reproductive tract in male C. punctatum consisted of a pair of testes, efferent ducts, epididymidis, ductus deferens, seminal vesicles, Leydig gland and claspers. Four sexual development stages were determined: (1) immature, no calcified claspers (2) maturing, partially calcified claspers; (3) active and resting adults. Both adult resting and active stages had calcified claspers. In this study, immature male C. punctatum without calcified claspers were found from August to October, maturing C. punctatum with partially calcified claspers were found from November to January and adult C. punctatum caught from February to April were active with calcified claspers, whereas adult fish collected from May to July and during the rest of the year (until mid-January in the following year) were in the resting stage with inactive testes. The testis had a lobular structure and the spermatocysts were arranged diametrically (i.e. along the cross-sectional width of the testis) in each lobule. Plasma testosterone concentrations were associated with histomorphological changes in the reproductive tract. Testosterone concentrations peaked in March in active males and then decreased significantly in resting males. Overall, the variations in the reproductive organs during different sexual maturity stages were consistent with their roles in the production, movement and release of spermatozoa.

The Effects of Daily Melatonin Gavage on Reproductive Activity in the Male Syrian Hamsters.

The proper administration of melatonin has well been documented to induce testicular regression in seasonal breeding animals. The subcutaneous injections of melatonin in the afternoon, not in the morning, consistently occurred testicular involution in the male Syrian (golden) hamsters whose reproductive activity is regulated by the photoperiod. But the effects of daily melatonin via gavage have not been estimated. Golden hamsters housed in long photoperiod (LP) were divided into 5 groups: the control animals housed in LP or in short photoperiod (SP) and animals treated daily with low (15 μg), middle (150 μg), and high dosages (1,500 μg) of pure melatonin by using gavage in the evening for 8 weeks. As results, LP control animals had large testes and SP controls displayed small and entirely regressed testes. The animals treated with various dosages of melatonin showed collectively degenerating effects on the weights of testes, epididymides, and seminal vesicles in the middle and high dosage groups, with the individual differences as well. The high dosages induced testicular regression in more proportion than the middle dosages did. The low dosage had large testes like the LP control animals. The small and inactive testes shown in some animals of both middle and high groups presented the complete regression as those of the animals maintained in SP. These results strongly suggest that the administrations of melatonin lead to testicular involution in the male golden hamsters when it is administered through gavage.

Screening for structural variants of four candidate genes in dogs with disorders of sex development revealed the first case of a large deletion in NR5A1

Disorders of sex development (DSD) are important causes of infertility and sterility, and are risk factors for gonadal carcinogenesis. Many DSDs are caused by genetic factors, mainly sex chromosome abnormalities or mutations of genes involved in sexual development, as well as structural variants (SVs) - large deletions, duplications, and insertions, if these overlap genes involved in sex development. The aim of this study was to determine if there were SVs in four candidate genes - NR0B1 (DAX1), NR5A1, RSPO1, and SOX3 - using droplet digital PCR (ddPCR). There was study of two cohorts of dogs with DSD, including 55 animals with XX DSD and 15 with XY DSD. In addition, 40 control females and 10 control males were included in the study. Among cases, for which there were evaluations, a large deletion consisting of four exons of the NR5A1 gene was identified in a Yorkshire Terrier with a rudimentary penis, hypospadias, bilateral cryptorchidism, and spermatogenesis inactive testes. This is the first mutation in the NR5A1 gene leading to XY DSD phenotype to be reported in domestic animals. There were no SVs in the genes evaluated in the present study in the cohort of dogs with XX DSD. The results from this study provide evidence that the large structural variants of these genes are rarely associated with the DSD phenotype in dogs.

Germ cell desquamation-based testis regression in a seasonal breeder, the Egyptian long-eared hedgehog, Hemiechinus auritus.

Testes of seasonally breeding species experience a severe functional regression before the non-breeding period, which implies a substantial mass reduction due to massive germ-cell depletion. Two alternative mechanisms of seasonal germ-cell depletion have been described in mammals, apoptosis and desquamation (sloughing), but their prevalence has not been determined yet due to reduced number of species studied. We performed a morphological, hormonal, and molecular study of the mechanism of seasonal testicular regression in males of the Egyptian long eared-hedgehog (Hemiechinus auritus). Our results show that live, non-apoptotic, germ cells are massively depleted by desquamation during the testis regression process. This is concomitant with both decreased levels of serum testosterone and irregular distribution of the cell-adhesion molecules in the seminiferous epithelium. The inactive testes maintain some meiotic activity as meiosis onset is not halted and spermatocytes die by apoptosis at the pachytene stage. Our data support the notion that apoptosis is not the major testis regression effector in mammals. Instead, desquamation appears to be a common mechanism in this class.

The balance between apoptosis and autophagy regulates testis regression and recrudescence in the seasonal-breeding South American plains vizcacha, Lagostomus maximus.

Mammalian testis undergoes deep changes in their architecture and function during photoregression conditions in seasonal breeders. Particularly, the testicular mechanisms that regulate the transition between the active (functional) and inactive (regression) stage vary between species. The aim of the present study was to analyze the incidence of proliferation, apoptosis and autophagy in the testicular seminiferous ephitelium of a seasonal breeder, Lagostomus maximus, during the annual reproductive cycle. We observed that proliferating spermatogonia increased from the active testis until reaching the maximum peak in the activating testis. During the annual reproductive cycle, the quantity of apoptotic-TUNEL positive spermatogonia and meiotic germ cells was constant and this might be regulated by the members of the BCL2 family. Only in the activating testis, apoptosis of germ cells was almost undetectable. The analysis of the autophagic-related proteins BECN1 and LC3 showed their localization in Leydig cells and the germ cells in the active and activating testis. In the inactive testis, BECN1 and LC3 ceased to be immunolocalized within the seminiferous tubules and the mRNA expression of both regulators decreased. Moreover, the expression of BECN1 and LC3 and also the apoptotic index were up regulated in testicular cultures subjected to nutritional stress. These results suggest a possible interaction between apoptosis and autophagy in the active and activating testis (characterized by high metabolic requirement and nutrient), where autophagy could promote survival over cell death. In the inactive testis, the absence of autophagic-related proteins might explain the massive loss of germ cells, suggesting that autophagy plays new and key role in the alterations of the seminiferous epithelium during photoregression.

Seasonal and Ageing-Depending Changes of Aquaporins 1 and 9 Expression in the Genital Tract of Buffalo Bulls (Bubalus bubalis).

The presence of Aquaporins 1 (AQP1) and 9 (AQP9), integral membrane water channels that facilitate rapid passive movement of water and solutes, was immunohistochemically detected in the excurrent ducts collected from sexually mature buffalo bulls of proven fertility during the mating (late autumn-winter) and non-mating (late spring to the beginning of autumn) seasons. Furthermore, the research was performed also on the epididymal cauda of a senile buffalo bull with inactive testis. Aquaporins 1 and 9 were immunolocalized at distinct levels. In the efferent ducts, AQP1 immunoreactivity was strongly evidenced at the apical surface of the non-ciliated cells and weakly along the basal membrane of the epithelial cells. The latter reactivity disappeared during the non-mating season. No AQP1 immunoreactivity was detected in the epithelium of epididymis and vas deferens, whereas AQP1 was expressed in the smooth muscle layer of the vas deferens. Aquaporin 1 was present in the blood vessels and in small nerve bundles all along the genital tract. The supranuclear zone of the epididymal principal cells was AQP9 immunoreactive, limited to the corpus and cauda regions, and vas deferens. The samples collected in the two reproductive seasons showed a weaker AQP9 immunoreactivity during the non-mating season. A typical AQP9 immunoreactivity was noticed in the old buffalo examined. The tested AQP molecules showed a different expression pattern in comparison with laboratory mammals, primates, equine, dog and cat. In addition, seasonal differences were noticed which are possibly useful in regard to the comprehension of the morphophysiology of reproduction in the bubaline species, which are still a matter of debate.

Molecular basis of photoperiodic control of reproductive cycle in a subtropical songbird, the Indian weaver bird (Ploceus philippinus)

Less is known about genetic basis of photoperiodic regulation of reproductive cycle in subtropical birds. This study measured the expression levels of DIO2, DIO3, GnRH, and GnIH genes in Indian weaver birds subjected to short days (8h light:16h darkness, 8L:16D) and long days (16L:8D) for 48weeks. Whereas small, reproductively inactive testes were maintained under short days, weaver birds underwent testis recrudescence – regression cycle under long days. Relative expression levels of DIO2, DIO3, GnRH and GnIH genes were quantified by the real-time PCR (qPCR) in hypothalamus of birds (n=4) sampled at the beginning of the experiments, and after 10 and 48weeks of short and long day exposures. These sample times represented photosensitive unstimulated (day 0), and under long days the recrudescence (photostimulated, after 10weeks) and regression (photorefractory, after 48weeks) testicular phases. Birds under short days served as controls. The expression pattern of these genes corresponded with testicular phases. High and low GnRH and DIO2 levels were found in birds with large and small testes, respectively. By-and-large the converse was true for GnIH and DIO3 expression levels. Thus, after 10weeks of exposure, there was a significant difference in the mRNA levels between short and long day birds, with small and large testes, respectively. The results also suggest for a possible rapid switching between DIO2 and DIO3 and GnRH and GnIH expressions during testis maturation – regression cycle in Indian weaver birds.

The Expression of amh and amhr2 Is Associated with the Development of Gonadal Tissue and Sex Change in the Protandrous Black Porgy, Acanthopagrus schlegeli1

The protandrous black porgy, Acanthopagrus schlegeli, has a striking life cycle, with sex differentiation at the juvenile stage, mono-male development, a bisexual gonad during the first 2 yr of life, and a male-to-female sex change (with vitellogenic oocytes) at 3 yr of age. In the present study, we investigated the possible roles of amh and amhr2 in gonadal development in a nonmammalian model organism (protandrous black porgy), especially in relation to sex differentiation, testicular and ovarian growth, and sex change. Fish of various ages were treated with estradiol or an aromatase inhibitor to induce the fish to become female. Furthermore, a natural sex change (2(+)-yr-old [>2 yr and <3 yr] fish) and a nonchemical method to surgically remove one of the pair of gonads to examine the possible roles of amh in the natural sex change were conducted. We present integrative in situ hybridization, immunohistochemical, cellular, and molecular data describing these phenomena. During gonadal sex differentiation, an increase in amh and amhr2 expression was detected. Higher levels of amh and amhr2 transcripts were observed in the testicular tissue when compared to the ovarian tissue in the bisexual gonad of 1(+)-yr-old (>1 yr and <2 yr) fish. Transcripts of amh reached peak levels in November (prespermatogenesis period) and then declined to the lowest levels in January (spawning period). Chemical-induced ovarian tissue had very low amh transcript levels but high levels of amhr2. Active testes had significantly higher amh and amhr2 expression levels as compared to inactive testes. In contrast, no difference in the expression of amh and amhr2 between active and inactive ovarian tissues was found. Transcripts of amh were expressed in the somatic cells of the spermatogonia and vitellogenic oocytes, and amhr2 was expressed in the somatic cells of the spermatogonia. Transcripts of amh decreased in the testicular tissue 5 mo before occurrence of the sex change into a female. In contrast, testicular amh expression remained high if the fish remained male. Human chorionic gonadotropin regulated amh and amhr2 expression in the testicular tissue but not in the ovarian tissue. The present results suggest that amh plays important roles in early testicular and ovarian development, late ovarian growth (e.g., vitellogenic oocytes), and natural sex change in the protandrous black porgy.

Localization of the cytochrome p450 side-chain cleavage enzyme in the inactive testis of the naked mole-rat.

Spermatogenesis was histologically examined in non-breeding male of the naked mole rat (Heterocephalus glaber) using a light microscopy. Spermatogonia, spermatocytes and spermatids were confirmed in the seminiferous tubules. However, the spermatogenesis was disordered, and many spermatocytes and spermatids were sloughing. Sperms could not be seen in the lumen of the tubules. The characteristic accumulation of interstitial cells was the most noteworthy. In the immunohistochemistry for cytochrome p450 side-chain cleavage enzyme, immunoreactions were not entirely distributed in each interstitial cell, although positive reactions were scattered in the interstitial cell-mass. The findings indicate that few interstitial cells act as a testosterone-synthesizing apparatus in the characteristic structure with accumulated cell-mass. From the immunohistochemical data we suggest the possibility that spermatogonia and Sertoli cells may secrete 17 beta-estradiol. We also suggest that 17 beta-estradiol from spermatogonia and Sertoli cells may inhibit the interstitial cells from synthesizing and secreting testosterone and may suppress the later stages of the spermatogenesis to induce apoptosis of germ cells. The TUNEL methods demonstrated that cell death occurred in some spermatocytes in non-breeding males.

Influence of age and photoperiod on steroidogenic function of the testis in the golden hamster.

The golden (Syrian) hamster is a seasonal breeder, and exposure of adult animals to short days results in severe gonadal regression with morphological features that resemble the immature testis. The purpose of this study was to investigate testicular steroidogenic capacity in the golden hamster and to analyse the influence of age and photoperiod on this process. Hamsters aged 36 days were maintained on a long photoperiod (14L:10D), and adult animals were then exposed to a long or a short photoperiod (6L:18D) for 14 weeks (the period of time required to achieve maximal gonadal regression), to assess circulating levels and in vitro production of testosterone, dihydrotestosterone and androstane-3 alpha, 17 beta-diol. In peripubertal hamsters, androstane-3 alpha, 17 beta-diol was the main circulating androgen detected, whereas in active adult animals, testosterone showed the highest serum levels. In hamsters exposed to a short photoperiod, blood testosterone levels were significantly lower than levels in adult hamsters exposed to a long photoperiod. Exposure of adult hamsters to a short photoperiod produced a marked reduction in serum concentrations of dihydrotestosterone and androstane-3 alpha, 17 beta-diol, which was not accompanied by a decrease in testicular 5 alpha-reductase activity. In the in vitro experiments, active adult testes were less sensitive than inactive adult testes to stimulation of androgen production with hCG, but showed similar sensitivity to the gonads from hamsters aged 36 days. In accordance with circulating androgen concentrations, the principal androgens produced in the in vitro assays from peripubertal and normal adult testes were androstane-3 alpha, 17 beta-diol and testosterone, respectively. Unexpectedly, the main androgen produced from regressed testes under in vitro conditions was androstane-3 alpha, 17 beta-diol. Inactive gonads released more androstane-3 alpha, 17 beta-diol than did normal adult testes and total in vitro androgen production (testosterone + dihydrotestosterone + androstane-3 alpha, 17 beta-diol) from adult testes was not diminished by exposure to a short photoperiod. However, in spite of the significant increase detected in production of androstane-3 alpha, 17 beta-diol in vitro from regressed testes, inactive gonads produced less androstane-3 alpha, 17 beta-diol than did peripubertal testes. In summary, our studies suggest that testicular androgen biosynthetic capacity in adult hamsters exposed to short photoperiod is not reduced and these regressed testes represent an intermediate physiological state between peripubertal and active adult testes. The significant decrease detected in serum androgen concentrations during the involution phase could result from the absence of stimulating pituitary factors, together with a negative regulation of steroidogenesis by different non-steroidal signals originating within and/or outside of the testis.

Deferred seasonal increase in testes weight under poor nutritional conditions in a sub‐Antarctic population of rabbits (Oryctolagus cuniculus)

AbstractThe annual cycle of testes weight of adult male rabbits (Oryctolagus cuniculus) was studied in three populations on the sub‐Antarctic Kerguelen archipelago during 1984–85. This study was completed in April 1987 with an analysis of differences in diet quality between populations, assessed by the proportion of nitrogen and lignin in stomach contents. A marked annual cycle of testes weight was observed in all populations, with a rapid fall in testes weight from February to March. As in most rabbit populations studied elsewhere, testes growth resumed immediately in two populations (called Morne and Molloy). In contrast, testes growth was deferred by five to six months in the third population (Armor), subject to harsher environmental conditions. Moreover, a larger proportion of the Armor males had inactive testes during the reproductive season. The diet of the Armor population was characterized by low nitrogen and high lignin content, suggesting poor‐quality diet as an immediate cause of deferred testes growth. We propose that reduced testes size during adverse conditions might be adaptive by increasing survival probability through reduction of basal metabolism and limitation of agonistic and reproductive activities, behaviours which are costly in terms of high energetic expenditure. However, at the population level, the timing of the breeding season was not related to the cycle of male testes weight and thus seems to depend on females' condition.

Distribution of lectin binding in spermatogonia of Syrian hamsters in gonadally active and inactive states.

Lectin binding patterns in spermatogonia of Syrian hamsters in gonadally active and inactive states were examined by light and electron microscopy. After exposure to a short day cycle (SD), the testis weight and the diameter of seminiferous tubules decreased, reaching the minimum at 13 weeks. At that time, spermatogenesis was severely disrupted. In the animals kept exposed to an SD, spermatogenesis reinitiated spontaneously after 23 weeks. In the animals transferred to a long day cycle (LD) after exposure to an SD for 13 weeks, spermatogenesis reinitiated only 4 weeks later. As to the lectin binding, Dolichos biflorus agglutinin (DBA) bound specifically to spermatogonia. The number of DBA-positive spermatogonia per one seminiferous tubule increased until 13 weeks after exposure to an SD, and then gradually decreased. DBA bound to only type A spermatogonia in active testes, whereas it bound to type A, intermediate and type B spermatogonia in inactive testes. Moreover, SDS-PAGE and Western blot analysis of testes in active and inactive states indicated that DBA-binding bands (115 kDa, 76 kDa) in inactive testes were intense compared with those in active testes. The 82 kDa band was detected only in inactive testes. These results supported the finding obtained from lectin histochemistry. DBA-binding glycoprotein was detected in all types of spermatogonia in inactive testes, suggesting that this glycoprotein way concern the active/inactive state of spermatogenesis. The present study also indicated that DBA is a useful marker for spermatogonia in inactive testes of Syrian hamster.

Isolation and culture of testicular macrophages from a seasonally breeding species, Phodopus sungorus. Evidence for functional differences between macrophages from active and regressed testes

We have developed an isolation and cultivation protocol for testicular macrophages from the seasonally breeding Siberian hamster, Phodopus sungorus which allows serum-free culture. Macrophages were isolated from the active testes of adult Phodopus kept in long photoperiod (LD, 18hL:6hD), and from the inactive testes of animals with gonadal regression induced by exposure to a short photoperiod (SD, 6hL;18hD; for 10 weeks). The isolated cells were identified as macrophages by (a) electron microscopy, (b) non-specific phagocytosis, and (c) the presence of Fc-receptors. Treatment of the cultures with oFSH (at 0.5 and 1 micrograms ml-1 for 1 and 4 days) significantly stimulated lactate secretion by testicular macrophages from LD hamsters, as expected from studies in the rat. In marked contrast, oFSH did not affect lactate secretion by testicular macrophages from SD hamsters, or by peritoneal macrophages. To test possible macrophage-Leydig cell interactions, the influence of testicular macrophage-conditioned medium (TM-CM) from FSH-responsive LD macrophage cultures was assessed and compared with the effect of peritoneal macrophage-conditioned medium (PM-CM) on testosterone production by testicular parenchyma from LD hamsters. Unexpectedly, and in contrast to previous reports in rats, testosterone production by testes was inhibited significantly by TM-CM from cultures pretreated with either 0.1 or 0.5 micrograms FSH ml-1. FSH alone stimulated testosterone production (due either to LH contamination or to possible paracrine effects). PM-CM from cultures pretreated with FSH did have an inhibitory effect. It is concluded that: (i) There is indirect evidence for the presence of functional FSH receptors on macrophages from the active testes of Phodopus. (ii) Testicular macrophages from inactive testes of Phodopus are not responsive to FSH, implicating impaired macrophage function in the regressed testes of Phodopus exposed to SD. (iii) Testicular macrophages produce a factor which can inhibit testosterone production by incubated testicular tissue.

Effects of testosterone on a selected neuronal population within the preoptic sexually dimorphic nucleus of the Japanese quail

The effects of testosterone on the volume and cytoarchitecture of the sexually dimorphic nucleus of the preoptic area (POM) were investigated in male and female Japanese quail. It was confirmed that castration decreases the POM volume in males and that, in gonadectomized birds of both sexes, testosterone increases this volume to values similar to those observed in intact sexually mature males. This suggests that the sex difference in POM volume results from a differential activation by T so that this brain morphological characteristic is not truly differentiated in the organizational sense. This conclusion was extended here by demonstrating that males exposed to a photoperiod simulating long days and that are known to have high plasma levels of testosterone have a larger POM than short-day males that have inactive testes. Detailed morphometric studies of POM neurons revealed a structural heterogeneity within the nucleus. A population of large neurons (cross-sectional area larger than 70-80 microns2) was well represented in the dorsolateral but was almost absent in the medial part of POM. This lateral population of neurons was sensitive to variations of testosterone levels in males but not in females. The cross-sectional area, diameter, and perimeter of the dorsolateral neurons were significantly increased in males exposed to high testosterone levels (intact birds exposed to long days or castrated birds treated with the steroid). These changes were not observed in the medial part of the nucleus. Interestingly, the size of the dorsolateral neurons was not affected by testosterone treatments in females. These results suggest that the swelling of neurons in the lateral POM of males might be responsible for the increase in total volume of the nucleus, which is observed in physiological situations associated with a high testosteronemia. In addition, the sensitivity to testosterone of the dorsolateral neurons in the POM appears to be sexually differentiated. This differential response to testosterone might represent a truly dimorphic feature in the organizational sense and additional studies manipulating the early steroid environment should be performed to test this possibility.

Testicular steroid metabolism in juvenile bank voles ( Clethrionomys glareolus schreber) exposed to different photoperiods: An in vitro study

Juvenile male bank voles (18–22 days of age) were either sacrificed immediately (Group C) or subjected first to a long (18L:6D, lights on 0600–2400; Group L) or a short (6L:18D, lights on 0800–1400; Group S) photoperiod for 1 week. The animals were killed by decapitation, the gonads were excised, and minced, and the conversion of [4- 14C]pregnenolone (Δ 5P) and [4- 14C]dehydroepiandrosterone (DHA) to metabolites was studied in vitro. The radioactive steroids formed were separated and identified by thin-layer chromatography (TLC). The conversion of Δ 5P to C 19-steroids increased markedly from 35 (Group C) to 86% (Group L) during the first week in the long photoperiod whereas in the short photoperiod a decrease to 14.42% (Group S) was observed. The reduced production of C 19-steroids in the more inactive testes was accompanied by the accumulation of progesterone (Δ 4P) (52.98% Group S and 24.9% Group C) and small amounts of 17α-hydroxyprogesterone (17α-OH-Δ 4P) (2.5 and 4.5%, respectively), whereas in Group L only trace amounts of these metabolites were encountered. No marked differences in the metabolism of [4- 14C]DHA between the photoperiodic groups were observed. These results seem to indicate that at least in vitro marked changes in C 17-C 20-lyase and/or 17α-hydroxylase activities occur in this seasonally breeding species during testicular maturation and photoperiodically induced regression.