Exposure of neonatal rats to anti-androgens induces penile mal-developments and infertility comparable to those induced by oestrogens.

We previously reported that oestrogen exposure in neonatal rats induced permanent infertility and malformed penis characterized by fat accumulation, which replaced most of the smooth muscle cells and cavernous spaces in the body of the penis, structures essential for erection. The objective of this study was to determine if reduced androgen production/action in the neonatal period, in the absence of exogenous oestrogen exposure, induces penile deformities similar to those caused by oestrogen. Male rats were treated from postnatal days 1-6 with GnRH antagonist antide (A, 10 mg/kg) or androgen receptor (AR) antagonist flutamide (F, 50 mg/kg) or F + A, with or without AR agonist dihydrotestosterone (DHT, 20 mg/kg). For comparison, pups received diethylstilbestrol (DES, 0.1 mg/kg), with or without DHT. Tissues were collected at ages 7 and 12 days and at adulthood. Flutamide alone decreased penile length and weight significantly (p < 0.05), but it caused neither fat accumulation, nor affected fertility (80% vs. 87% in controls). Antide alone reduced penile length and weight significantly, and induced fat accumulation in 4/11 rats and infertility in 13/14 rats. Conversely, all 11 F + A-treated rats, similar to all nine DES-treated rats, had fat accumulation and loss of smooth muscle cells and cavernous spaces in the body of the penis and were infertile. In addition, reductions in penile length and weight were higher than in rats treated with F or A alone. DHT co-administration mitigated penile deformities in the DES group, but did not in the F + A group. Testicular testosterone was reduced by 70-95% at 7 or 12 days of age in all treated groups, except in the F group, which had threefold higher testosterone than controls. Collectively, data unequivocally show that reduced androgen production/action in the neonatal period, in the absence of oestrogen exposure, induces permanent infertility and malformed penis similar to that caused by oestrogen.

Estrogen-induced developmental disorders of the rat penis involve both estrogen receptor (ESR)- and androgen receptor (AR)-mediated pathways.

This study tested the hypothesis that the estrogen receptor (ESR) pathway, androgen receptor (AR) pathway, or both mediate estrogen-induced developmental penile disorders. Rat pups received diethylstilbestrol (DES), with or without the ESR antagonist ICI 182,780 (ICI) or the AR agonist dihydrotestosterone (DHT) or testosterone (T), from Postnatal Days 1 to 6. Testicular T concentration, penile morphology and morphometry, and/or fertility was determined at age 7, 28, or 150 days. DES treatment alone caused 90% reduction in the neonatal intratesticular T surge; this reduction was prevented by ICI coadministration, but not by DHT or T coadministration. Unlike the T surge, coadministration of ICI and coadministration of DHT or T mitigated penile deformities and loss of fertility. Generally, ICI, DHT, or T treatment alone did not alter penile morphology; however, fertility was 20% that of controls in ICI-treated rats vs. 70%-90% in DHT- or T-treated rats. The lower fertility in the rats treated with ICI alone could be due to altered sexual behavior, as these males did not deposit vaginal plugs. In conclusion, observations that both an ESR antagonist and AR agonists prevent penile deformities and infertility suggest that both pathways are involved in estrogen-induced penile disorders. Observations that coadministration of ICI, but not DHT or T, prevents the DES-induced reduction in the neonatal T surge suggest that, although ICI exerts its mitigating effect both at the level of Leydig cells and penile stromal cells, DHT and T do so only at the level of stromal cells.

Role of estrogen in induction of penile dysmorphogenesis: a review.

In this review, we report permanent dysmorphogenesis of the penis and loss of fertility in adult rats treated neonatally with estrogen. Specifically, we report replacement of smooth muscle cells and cavernous spaces by fat cells in the corpus cavernosum penis, but not in the adjoining corpus spongiosum. Induction of these novel, region-specific phenotypes is dose-dependent, requires a critical window of exposure and associated with decreased testosterone and up-regulation of estrogen receptor alpha (ER alpha). The resistance of ER alpha knockout mice to develop these abnormalities implies an unequivocal role for ER alpha in mediating maldevelopment of the penis. Additionally, the prevention of estrogen-inducible penile abnormalities by ER antagonist ICI 182 780 implies that a functional ER-mediated pathway is essential for inducing penile abnormalities. Likewise, the ability of testosterone or dihydrotestosterone to negate these abnormalities suggests a role for an androgen receptor (AR)-mediated pathway. Taken together, these observations led us to hypothesize that neonatal estrogen exposure, via an ER-mediated pathway (direct action) or an AR-mediated pathway (indirect action through decreased testosterone) or both pathways, up-regulates ER alpha expression in stromal cells of the penis, which are then reprogrammed such that their differentiation into smooth muscle cells is inhibited and their differentiation into adipocytes is stimulated.

Estrogen receptor alpha mediates estrogen-inducible abnormalities in the developing penis.

Previously, we reported an association between estrogen receptor-alpha (ERalpha) upregulation and detrimental effects of neonatal diethylstilbestrol (DES) exposure in the rat penis. The objective of this study was to employ the ERalpha knockout (ERalphaKO) mouse model to test the hypothesis that ERalpha mediates DES effects in the developing penis. ERalphaKO and wild-type C57BL/6 mice received oil or DES at a dose of 0.2 microg/pup per day (0.1 mg/kg) on alternate days from postnatal days 2 to 12. Fertility was tested at 80-240 days of age and tissues were examined at 96-255 days of age. DES caused malformation of the os penis, significant reductions in penile length, diameter, and weight, accumulation of fat cells in the corpora cavernosa penis, and significant reductions in weight of the bulbospongiosus and levator ani muscles in wild-type mice. Conversely, ERalphaKO mice treated with DES developed none of the above abnormalities. While nine out of ten male mice sired pups in the wild-type/control group, none did in the wild-type/DES group. ERalphaKO mice, despite normal penile development, are inherently infertile. Both plasma and intratesticular testosterone levels were unaltered in the DES-treated wild-type or DES-treated ERalphaKO mice when compared with controls, although testosterone concentration was much higher in the ERalphaKO mice. Hence, the resistance of ERalphaKO mice to developing penile abnormalities provides unequivocal evidence of an obligatory role for ERalpha in mediating the harmful effects of neonatal DES exposure in the developing penis.

Estrogen-induced abnormal accumulation of fat cells in the rat penis and associated loss of fertility depends upon estrogen exposure during critical period of penile development.

We previously reported that diethylstilbestrol (DES) or estradiol valerate (EV) exposure at a dose of 0.10-0.12 mg/kg, or higher, per day, on alternate days, from postnatal days 2-12, resulted in abnormal penis development and infertility (H. O. Goyal et al., 2005, J. Androl. 26, 32-43). The objective of this study was to identify a critical developmental period(s) during which EV exposure results in the observed penile abnormalities. Male pups received EV at a dose of 0.10-0.12 mg/kg on postnatal day(s) 1, 1-3, 4-6, 1-6, 7-12, 13-18, 19-24, or 25-30. Fertility was tested at 102-115 days of age and tissues were examined at 117-137 days. Both penile morphology and fertility were unaltered in rats treated with EV after 12 days of age. Conversely, except in rats treated on postnatal day 1 only, none of the males treated prior to 12 days of age sired pups, and all had abnormal penises, including varying degrees of abnormal accumulation of fat cells and loss of cavernous spaces and smooth muscle cells in the corpora cavernosa penis, which were maximal in the 1-6-day group. Also, the preputial sheath was partially released or its release was delayed, and the weight of the bulbospongiosus muscle was significantly reduced. Plasma testosterone (T) in the 1-6- and 4-6-day groups and intratesticular T in the 4-6-day group were significantly lower. The testosterone surge, characteristic of controls in the first week of life, was suppressed in the 1-3-day group. Estrogen receptor alpha mRNA expression was enhanced in the body of the penis in the 1-3-day group, but not in the 13-18-day group. Hence, EV exposure prior to 12 days of age (as short as 1-3 days postnatal), but not after 12 days of age, results in long-term abnormal penile morphology, characterized by abnormal accumulation of fat cells in the corpora cavernosa penis and, consequently, loss of fertility.

Exposure of neonatal male rats to estrogen induces abnormal morphology of the penis and loss of fertility.

The research objectives are to determine whether estrogen-induced infertility is associated with abnormal morphology of the penis and if morphological alterations can be reversed by testosterone (T). Male pups received diethylstilbestrol (DES) on alternate days from postnatal days 2 to 12. They received T or empty implants at 180 days, were tested for fertility at 188 days, and terminated at 200 days. While 5/7 control males sired pups, only 1/6 did in the DES group, and 0/8 in the DES plus T group. In addition to reductions in penile length and weight, the novel structural change induced by DES, and not reversed by T, was a replacement of cavernous spaces by fat cells in the penis body. Hence, T substitution for 8 days at adulthood did not reverse infertility in rats treated neonatally with DES and provided evidence that infertility probably resulted from absence of cavernous spaces and/or accumulation of fat cells in the penis body.

Abnormal morphology of the penis in male rats exposed neonatally to diethylstilbestrol is associated with altered profile of estrogen receptor-alpha protein, but not of androgen receptor protein: a developmental and immunocytochemical study.

Objectives of the study were to determine developmental changes in morphology and expression of androgen receptor (AR) and estrogen receptor (ER)alpha in the body of the rat penis exposed neonatally to diethylstilbestrol (DES). Male pups received DES at a dose of 10 microg per rat on alternate days from Postnatal Day 2 to Postnatal Day 12. Controls received olive oil vehicle only. Tissue samples were collected on Days 18 (prepuberty), 41 (puberty), and 120 (adult) of age. DES-induced abnormalities were evident at 18 days of age and included smaller, lighter, and thinner penis, loss of cavernous spaces and associated smooth muscle cells, and increased deposition of fat cells in the corpora cavernosa penis. Fat cells virtually filled the entire area of the corpora cavernosa at puberty and adulthood. Plasma testosterone (T) was reduced to an undetectable level, while LH was unaltered in all treated groups. AR-positive cells were ubiquitous and their profile (incidence and staining intensity) did not differ between control and treated rats of the respective age groups. Conversely, ERalpha-positive cells were limited to the stroma of corpus spongiosus in all age groups of both control and treated rats, but the expression in treated rats at 18 days was up-regulated in stromal cells of corpora cavernosa, coincident with the presence of morphological abnormalities. Hence, this study reports for the first time DES-induced developmental, morphological abnormalities in the body of the penis and suggests that these abnormalities may have resulted from decreased T and/or overexpression of ERalpha.

Neonatal estrogen exposure of male rats alters reproductive functions at adulthood.

The effects of neonatal exposure to different doses of diethylstilbestrol (DES) on the reproductive functions of male rats at adulthood were evaluated. Sprague-Dawley rats (5-8/group) received sc injections of 25 microl olive oil containing DES (Sigma Chemical Co., St. Louis, MO) at a dose of 10 microg, 1 microg, 100 ng, 10 ng, or 1 ng per rat on alternate days from Postnatal Days 2-12. Control animals received olive oil only. All animals were allowed to develop until 83-91 days of age; however, when they were 70 to 80 days old, four male rats each from the 10 microg, 1 microg, 100 ng, and control groups were cohabited with untreated 60- to 70-day-old females (1:1) for 12 days. At the end of cohabitation, both mated and unmated male rats were weighed, and blood and tissue samples were collected and processed. Results revealed that although sperm motility patterns and sperm morphology were adversely affected in the 10- microg group, other reproductive parameters, including 1). daily sperm production (DSP)/testis; 2). absolute and relative weights of the testis, epididymis, and seminal vesicle; and 3). sperm numbers in both regions of the epididymis declined significantly in a dose-dependent manner in the 10- and 1- microg groups. Conversely, in the <1- microg groups, none of these parameters (except DSP/testis and weight of the epididymis in the 100-ng group, and sperm numbers in the epididymis of the 100- and 10-ng groups) was different from controls. Generally, plasma testosterone levels decreased in the 10- and 1- microg groups, FSH level increased in the 10-microg group, and prolactin and LH levels were unaltered. In the fertility study, although each male in the 1-microg, 100-ng, and control groups produced a copulatory plug and impregnated a female, none could do so in the 10-microg group. The mean number of pups per litter was reduced to eight in the 1-microg group, in contrast to 15 each in the 100-ng and control groups. In conclusion, exposure of neonatal male rats to DES altered sperm motility patterns, sperm fertility (as evident from the reduced number of pups in the 1-microg group), and sexual behavior (as evident from the absence of copulatory plugs in the 10-microg group) and reduced weights of reproductive organs, DSP/testis, and sperm numbers in the epididymis. Whether these alterations/reductions persist in older rats (6-8 mo of age) is under investigation.

Expression and molecular characterization of estrogen receptor alpha messenger RNA in male reproductive organs of adult goats.

The fact that male estrogen receptor alpha (ERalpha) knockout mice are infertile indicates a role for this receptor in male reproduction. Here, objectives were to evaluate ERalpha expression in male goat reproductive tissues at the transcriptional level using RNase protection assay (RPA) and in situ hybridization (ISH), and to clone a partial cDNA for caprine ERalpha using reverse transcription-polymerase chain reaction (RT-PCR). For RPA and ISH procedures, a radiolabeled antisense cRNA probe, generated in vitro from the ovine oER8 cDNA template, was employed. Evaluations were made on individual samples obtained from adult goats. Labeled cRNA sense probe was used as a negative control in ISH. A 530-base pair amplicon was generated by RT-PCR from efferent ductules (EDs), epididymis (EP), and testis, cloned from the ED and EP, and sequenced. The caprine ERalpha (cERalpha) cDNA displayed 81%-96% sequence identity with that of other species. A signal indicative of ERalpha mRNA was identified by both RT-PCR and RPA in all tissues, but was strongest in the ED. Compared with ED, ERalpha signal was sixfold lower in the EP, and 66-fold lower in the testis. Similarly, strong ERalpha expression was observed in ED epithelium, whereas little or no signal was detected in EP or testis by ISH. Thus, among different segments of the male reproductive tract and testis, the highest level of ERalpha mRNA expression was found in epithelium of the ED.

Diethylstilbestrol-treated adult rats with altered epididymal sperm numbers and sperm motility parameters, but without alterations in sperm production and sperm morphology.

In this study, we characterized estrogenic effects of diethylstilbestrol (DES) on reproductive parameters in male rats to identify a minimal dose level that alters epididymal and sperm functions but has little or no effect on sperm production and/or spermatogenesis. Adult rats (five animals/group) received s.c. injections of 0.2 ml of corn oil containing DES at a rate of 1.0 mg, 200 microg, 40 microg, 8 microg, 1.6 microg, or 320 ng x rat(-1) x day(-1) for 12 days. The control group received corn oil only. DES effects were similar in the 8-microg group and higher dose groups and included significant (P < or = 0.05) reductions in 1) absolute and relative weights of the head and body of the epididymis (EP), tail of the EP, and seminal vesicle, 2) numbers of sperm in both regions of the EP, and 3) motility characteristics in sperm collected from the tail of the EP. Conversely, no significant changes were observed in relative testis weight, daily sperm production, spermatogenesis, seminiferous epithelial height in stage VII, and sperm morphology. All of the above parameters in the 1.6-microg group (except seminal vesicle weight) and 320-ng group were comparable to those of controls. Plasma testosterone (T) level was reduced to an almost undetectable level in the > or = 8-microg groups and to a very low level in the 1.6-microg group (0.35 vs. 2.36 ng/ml in controls or 320-ng group), but LH level was unaltered. In a parallel fertility study, males received DES at a rate of 40, 8, or 1.6 microg x rat(-1) x day(-1) for 12 days prior to and 12 days during cohabitation (1:1) with untreated females. Of the 15 females cohabited with treated males (5 females/dose), none in the 40-microg and 8-microg groups and 1 in the 1.6-microg group formed a copulatory plug and delivered 8 pups, in contrast to 5/5 copulatory plugs and 13-15 pups/litter in the controls. DES at a rate of 8 microg x rat(-1) x day(-1) for 12 days reduced EP weights, sperm numbers in the EP, and sperm motility patterns but caused minimal to no alterations in daily sperm production, spermatogenesis, or sperm morphology. Factors other than T, or in addition to lower T, may be responsible for DES-induced reproductive disorders (despite lower T, sperm contents and sperm motility patterns in the EP were normal in the 1.6-microg group). Deficits in EP sperm functions and/or sexual behavior (as evident from absence of copulatory plugs) probably accounted for reduced fertility in treated males.

Postnatal differentiation of efferent ductule epithelium in goats: a light microscopic and ultrastructural study.

Caprine efferent ductule epithelium contains ciliated and nonciliated cells. The latter cells are divided into three types: type II cells contain PAS-positive granules, type III cells contain PAS-negative vacuoles, and type I cells lack both granules and vacuoles (Goyal and Williams, Anat. Rec. 220:58-67). The objectives of this study are i) to determine when the epithelium differentiates into ciliated and nonciliated cells, ii) to determine when nonciliated cells acquire characteristics typical for type II and type III cells, and iii) to relate developmental changes in the epithelium with those in the testis. Testes and efferent ductules were examined at the light and electron microscopic levels in goats from 1-25 weeks of age. Efferent ductule epithelium contained ciliated and nonciliated cells as early as week 1. While ciliated cells were differentiated at week 1, differentiation of nonciliated cells did not occur until week > or =15. Differential features in ciliated cells included the presence of cilia at the apical border and an aggregation of mitochondria in the apical cytoplasm. Those in nonciliated cells included the presence of i) an endocytotic apparatus at week > or =15, ii) PAS-positive granules at week > or =15, and iii) PAS-negative vacuoles at week > or =25. The seminiferous tubules developed lumens at 12-15 weeks. Hence, while differentiation of ciliated cells occurred much before lumen formation in the seminiferous tubules, that of nonciliated cells coincided with, or occurred soon after, lumen formation, suggesting a role for testicular fluid contents in their differentiation. The goat efferent ductules can be characterized morphologically mature by 25 weeks.

Postnatal differentiation of the ductus deferens, tail of the epididymis, and distal body of the epididymis in goats occurs independently of rete testis fluid.

Observations from extratesticular rete-ligated, mature goats indicated that epithelial morphology in the tail of the epididymis can be maintained without any input from testicular fluid (Goyal et al., Acta Anat., 1994;150: 127-135). Hence, the objective of this study was to determine whether the tail of the epididymis and/or other regions of the male excurrent ducts can differentiate prior to the appearance of lumen in the seminiferous tubules, which is an indicator for the onset of seminiferous tubular fluid secretion. Based on age and scrotal circumference (SC), 20 male goats were divided into four groups of five animals each: 1-4 weeks (SC, 6.5-7.5 cm), 7-10 weeks (SC, 8.5-11.0 cm), 12-15 weeks (SC, 11.0-14.0 cm), and 15-25 weeks (SC, 16.0-19.0 cm). Tissues were collected from the testis, six regions of the epididymis (proximal, middle and distal head; proximal and distal body; and tail), and the ductus deferens, and were processed for light and electron microscopic examination. Changes in epithelial height and cytological features associated with absorption (microvilli, pinocytotic and coated vesicles) and protein secretion (RER, Golgi body) were used as markers for differentiation. Differentiation of all of these features was comparable to that observed in the 15-25-week-old animals in the ductus deferens by > or = 1 week, in the tail of the epididymis by > or = 7 weeks, in the distal body of the epididymis by > or = 12 weeks, and in the proximal body of the epididymis and all three regions of the head of the epididymis by > or = 15 weeks. Seminiferous tubules developed lumens between 12 and 15 weeks. In conclusion, epithelial differentiation in the ductus deferens, tail of the epididymis, and distal body of the epididymis follows a time-dependent, spatial, ascending order and is achieved before lumen formation in the seminiferous tubules. Conversely, epithelial differentiation in all three regions of the head and the proximal body of the epididymis occurs simultaneously and after lumen formation in the seminiferous tubules.

Regulation of androgen and estrogen receptors in male excurrent ducts of the goat: an immunohistochemical study.

BACKGROUND: Since androgens and/or estrogens must bind with specific receptors in order to elicit a response at the target organ(s), it is important to understand factors that regulate expression of androgen receptors (AR) and estrogen receptors (ER). Hence, the objective of the study is to determine the relative significance between circulating androgen (CA) and luminal androgen (LA) in maintaining normal expression of AR and ER in male excurrent ducts. METHODS: Mature Nubian goats were subjected for 15 days each to the following treatments: (1) bilateral orchidectomy, (2) bilateral orchidectomy and testosterone treatment, (3) unilateral ligation of the extratesticular rete, and (4) unilateral orchidectomy. Tissues from different segments of the excurrent ducts were fixed in 4% paraformaldehyde and embedded in Paraplast-plus. Antigenic sites for AR and ER were immunolocalized using PG-21 rabbit antirat/human antibody and H-222 rat antihuman monoclonal antibody, respectively. The avidin-biotin horseradish peroxidase procedure was used to identify positive immunoreactivity. Negative controls included incubation of sections with irrelevant IgG in place of primary antibody. RESULTS: In intact animals, whereas AR were found in epithelial, connective tissue, and peritubular smooth muscle cells of the efferent ductules, regions I-V of the epididymis, and ductus deferens, ER were confined to nonciliated cells of the efferent ductules. Bilateral orchidectomy caused a severe loss of both AR and ER staining. Testosterone replacement to orchidectomized animals restored staining of both AR and ER to the intact level. Neither unilateral ligation of the extratesticular rete nor unilateral orchidectomy had any effect on AR or ER immunostaining. CONCLUSION: Circulating androgen alone, without any input from luminal androgen or other rete fluid contents, can regulate expression of both androgen receptor and estrogen receptor.

Immunolocalization of androgen receptor and estrogen receptor in the developing testis and excurrent ducts of goats.

BACKGROUND: Because of the significance of androgens and estrogens in prenatal and postanatal differentiation of the testis and excurrent ducts, it is important to understand the developmental pattern of androgen receptor (AR) and estrogen receptor (ER) in these organs. METHODS: Tissues from 1-23-week-old goats were fixed in 4% paraformaldehyde and embedded in Paraplast-plus. Antigenic sites for AR and ER were immunolocalized using the PG-21 rabbit anti-rat/human antibody and the H-222 rat anti-human monoclonal antibody, respectively. The avidin-biotin horseradish peroxidase procedure was used to identify positive immunoreactivity. Controls included incubation of sections with irrelevant IgG in place of primary antibody. RESULTS: Within the testis, immunostaining for AR in the nuclei of Sertoli cells increased gradually from mild at week 1 to strong at week > or = 19. In contrast, nuclei of peritubular myoid cells and Leydig cells exhibited moderate to strong reaction for AR in all animals. Germ cells were negative. Within the rete testis, efferent ductules, regions I-V of the epididymis, and ductus deferens, nuclei of all epithelial cells, peritubular myoid cells, and intertubular connective tissue cells expressed moderate to strong staining for AR at all ages. ER were confined to nonciliated cells of the efferent ductules, which displayed moderate staining in all animals, beginning from week 1. CONCLUSIONS: Nuclear AR staining, found in all testicular cells (except germ cells) and excurrent duct cells examined, was observed to change in an age-related manner only in Sertoli cells, where staining intensity increased between week 1 and week 19. Staining for ER, confined to nonciliated epithelial cells of the efferent ductules, was not affected by postnatal age.

Immunolocalization of receptors for androgen and estrogen in male caprine reproductive tissues: unique distribution of estrogen receptors in efferent ductule epithelium.

Androgens and estrogens affect physiological processes in the testis and male excurrent duct system. This study was designed to identify and characterize distribution of androgen receptors (AR) and estrogen receptors (ER) in the reproductive organs of the male goat. Tissues, including testis, efferent ductules, epididymis (regions I-V), and ductus deferens, were obtained from five mature Nubian goats, fixed in 4% paraformaldehyde, and embedded in paraplast. Antigenic sites for AR were unmasked by microwave treatment (four times, 5 min each) of tissue sections immersed in 10 mM citrate (pH 6) and were detected using the PG-21 rabbit anti-rat/human antibody. Antigenic sites for ER were identified using the H-222 rat anti-human monoclonal antibody after tissue sections were treated with pronase (0.5 mg/ml, 37 degrees C, 8 min). Avidin-biotin horseradish peroxidase procedures were used to identify positive immunoreactivity. Irrelevant IgG was substituted for primary antibody in negative controls. Positive nuclear immunostaining for AR was observed in all types of epithelial cells, peritubular smooth muscle cells, and intertubular fibroblasts of the intratesticular rete, efferent ductules, epididymis (regions I-V), and ductus deferens, as well as in Sertoli, Leydig, and peritubular myoid cells and intertubular fibroblasts of the testis. In contrast, nuclear immunostaining for ER was confined to nonciliated cells of the efferent ductules. Thus, AR-positive cells are ubiquitously distributed in caprine testicular and excurrent ductular tissues, and ER-positive cells are unique to the efferent ductules. The caprine model should be useful in studies designed to determine mechanisms through which androgens and estrogens regulate development and function of the testes and excurrent ducts.

Effects of androgen deprivation in the goat epididymis.

With the deprivation of both circulating androgen (CA) and luminal androgen (LA; orchiectomized goats), the epithelial height (EH) in regions I-IV of the epididymis was reduced to 28, 67, 58 and 56% of that of controls, respectively, but it was increased to 109% of that of controls in region V. Similarly, the volume density of epithelium (VDE) in regions I-V was reduced to 33, 49, 45, 41 and 70% of that of controls, respectively. Conversely, in the absence of LA only (extratesticular-rete-ligated goats), while both EH and VDE were reduced to almost 50% of those of controls in region I, they remained similar to those of controls in other regions. The morphological changes in the epithelium such as cytoplasmic regression, loss of stereocilia and disorderly arrangement of epithelial cells were maximal in region I, moderate in regions II-IV and minimal in region V. Testosterone treatment appreciably reduced the degenerative changes caused by orchiectomy in all regions except region I where the restorations were marginal at best. Hence, the results suggest a differential epididymal response to androgen deprivation. Whereas the LA and/or other rete fluid components seem essential for maintaining the epithelial structure of region I, the CA alone can maintain, at least partially, the epithelial structure of regions II-IV and almost completely that of region V.

Reexamination of the morphology of the extratesticular rete and ductuli efferentes in the goat.

The morphology of the extratesticular rete and ductuli efferentes was reexamined in serial cross sections collected from the entire mass of the efferent ductules and in longitudinal sections collected from the partially unraveled efferent ductules. The extratesticular rete forms a 3-4-mm-long sac-like dilatation, which, within the head of the epididymis, has a wide lumen (up to 4 mm) and gives off along its length numerous evaginations, which, in turn, make connections with the ductuli efferentes. The latter is a mass of 16-18 ductules lined by three types of nonciliated cells: type II cells are characterized by dense, periodic acid-Schiff (PAS)-positive granules; type III cells are characterized by empty-appearing, PAS-negative vacuoles; and type I cells lack both granules and vacuoles. The distribution of the three types of nonciliated cells varies along the length. Whereas only type I cells are present in the beginning portion of the efferent ductule, type II cells predominate in the middle portion and type III cells in the distal portion (near the epididymis). The transition from one cell type to the other type is gradual; thus there are short segments along the length that share characteristics first for type I and type II cells and then for type II and type III cells. These results demonstrate that different nonciliated cell types are not randomly distributed in the epithelium of the ductuli efferentes but, instead, gradually differentiate from type I to type II to type III cells along the length of each efferent ductule. Factors controlling this differentiation remain to be studied.

Regional differences in the morphology of the goat epididymis: a light microscopic and ultrastructural study.

The goat epididymis, based on morphological differences, was divided into five regions; regions I and II, and the proximal part of region III constituted the head; the distal part of region III and region IV, the body; and region V, the tail. The epithelium of all regions contained principal and basal epithelial cells and intraepithelial lymphocytes and macrophages. In addition, regions II to IV also contained a few apical cells. Clear cells were absent. The epithelium varied in height from the tallest in region I (88 +/- 33 microns) to the shortest in region V (38 +/- 5 microns). Conversely, the luminal diameter, thickness of smooth muscle wall, and luminal sperm concentration were highest in region V. The irregular epithelial height of regions I and IV accounted for a stellate lumen in contrast to the oval lumen of the other regions. Whereas the lumen of region I contained only a few sperm, those of regions II, III, and IV were filled with sperm. Principal cells were the only cell type that showed striking cytological differences between regions. While they contained absorptive features (canaliculi, pinocytotic and coated vesicles, and subapical vacuoles) in all regions, the principal cells of region II were filled with large, heterogeneous vacuoles (up to 5 microns in diameter), suggesting that they may be preferentially involved in transporting and digesting particulate material. Besides absorptive features, principal cells of all regions contained morphological correlates of protein synthesis such as highly developed Golgi complexes in the supranuclear area and numerous cisternae of RER near the Golgi body and in the infranuclear cytoplasm. The cisternae of RER were more developed in region IV, and in some instances, they were distended with flocculent material resembling newly synthesized protein. Unlike the protein synthesizing organelles, principal cells of all regions lacked morphological correlates of steroid hormone synthesis. These results are compared with previously published data on the regional differences in the epididymis of other species, especially with those of the rat and the bull, in an effort to understand the significance of the epididymis in sperm maturation.

Effects of age and season on the type and occurrence of sperm abnormalities in Nubian bucks.

Effects of age and season on type and occurrence of sperm abnormalities were examined in semen samples collected from 3 groups of Nubian bucks at ages of 4 to 9 months, 10 to 21 months, and 39 to 50 months. The average total percentage of sperm abnormalities at the onset of puberty (141 +/- 4 days) was 64.6 +/- 14.8% (head, 19.5 +/- 13.6%; middle piece, 17.2 +/- 9.3%; and proximal protoplasmic droplets, 14.6 +/- 10.5%), but this improved rapidly and was reduced to 12.5 +/- 7.5% by 8 months of age (head, 1.9 +/- 4.5%; middle piece, 4.6 +/- 2.8%). Further increase in age, at least up to 4 years, did not reveal a significant effect (P less than 0.05) on the type or percentage of total abnormalities. Similar to age, a comparison of data among seasons did not reveal a significant effect on the type or occurrence of sperm abnormalities in 10- to 21-month-old or 39- to 50-month-old bucks. Seemingly, Nubian bucks started producing good quality semen at 8 months of age, and season did not influence sperm abnormalities.