Segments Of Epididymis Research Articles

Aldose reductase and macrophage migration inhibitory factor are associated with epididymosomes and spermatozoa in the bovine epididymis.

During the epididymal transit, mammalian spermatozoa acquire new surface proteins necessary for male gamete function. We have previously shown that membranous vesicles, called epididymosomes, interact with spermatozoa allowing the transfer of some proteins to sperm surface within the epididymal lumen. The protein composition of those vesicles has been investigated to document the mechanisms of protein transfer from epididymosomes to spermatozoa. Electrophoretic analysis revealed that protein composition is different from the epididymal soluble compartment as well as from similar vesicles present in the semen. Protein association with epididymosome is very strong as revealed by resistance to extraction with detergent. Matrix-assisted laser desorption ionization time-of-flight as well as immunodetection techniques have been used to identify some proteins associated to epididymosomes and spermatozoa. An aldose reductase known for its 20alpha-hydroxysteroid dehydrogenase activity and the cytokine (macrophage migration inhibitory factor) have been identified. These two proteins have been immunolocalized in principal cells of the epididymal epithelium, a more intense signal being detected in the distal epididymal segment as well as in the vas deferens. Database search revealed that these two proteins are characterized by the lack of a signal peptide. These results are discussed with regard to a possible apocrine mode of secretion of these proteins acquired by spermatozoa during the epididymal transit.

Gene expression is differentially regulated in the epididymis after orchidectomy.

The epididymis is the site for the transport, maturation, and storage of spermatozoa. Regulation of epididymal structure and function is highly dependent on the ipsilateral testis. At the molecular level, however, few studies have been undertaken to determine which genes are expressed in the epididymis under testicular regulation. The goal of this study was to identify genes for which expression is regulated after orchidectomy, both throughout the epididymis and in a segment-specific manner. Microarrays spotted with 474 rat cDNAs were used to examine gene expression changes over the first 7 d post orchidectomy in the initial segment, caput, corpus, and cauda epididymidis of the adult Brown Norway rat. Using k-means cluster analysis, we show that four patterns of gene expression are activated in each epididymal segment over the first week following orchidectomy. Transient up-regulation of gene expression in the epididymis after orchidectomy is described for the first time. Potential androgen-repressed genes, including Gpx-1, show increased expression in the epididymis after orchidectomy. Several glutathione-S-transferases and calcium-binding proteins decline throughout the epididymis after orchidectomy, indicating that these may be novel androgen-regulated epididymal genes. Other genes coding for metabolism-associated proteins, transporters, and alpha-1 acid glycoprotein show segment-specific regulation in the epididymis after orchidectomy. Finally, we describe the expression of the previously uncharacterized heat shock proteins, and apoptosis-associated genes in the epididymis after orchidectomy. Thus, gene expression in the epididymis is differentially affected over time after orchidectomy. These results provide novel insight into androgen-dependent and segment-specific epididymal function.

Distribution of new human β-defensin genes clustered on chromosome 20 in functionally different segments of epididymis

Human β-defensins are a family of cationic peptides that share a pattern of six conserved cysteine residues. We describe the cloning and characterization of the cDNAs of five novel β-defensin genes ( DEFB25–DEFB29) clustered on chromosome 20p13, which were identified using a bioinformatics approach. Expression analysis revealed the occurrence of the transcripts in only a few organs, with the highest abundance in the male genital tract. Examination of β-defensin expression in human epididymis by real-time quantitative RT-PCR revealed a distribution along the functionally different segments of the epididymal duct. In situ hybridization for one of the cDNAs shows mRNA restriction to the epithelial cell layer of the epididymis, known to secrete factors responsible for sperm maturation. We suggest that the novel peptides carry out physiological functions in the male genital tract that may not be directly related to bacterial growth inhibition in host defense.

Stimulation of hypoxia-inducible factor-1 alpha (HIF-1alpha) protein in the adult rat testis following ischemic injury occurs without an increase in HIF-1alpha messenger RNA expression.

Hypoxia-inducible factor 1 (HIF-1) is a transcription factor composed of alpha and beta subunits. Stabilized from proteasome degradation and activated by hypoxia, HIF-1 stimulates expression of hypoxia-sensitive genes that mediate oxygen homeostasis in many tissues. Our hypothesis is that HIF-1 is involved in the cellular response to hypoxia in the ischemic testis. Goals of this study were to determine if HIF-1alpha mRNA is expressed in the testis, epididymis, and accessory sex glands of adult Sprague-Dawley rats and to determine if HIF-1alpha mRNA and protein expression in the testis is affected by experimentally induced ischemia. Total RNA from reproductive organs of adult rats was analyzed by relative reverse transcription-polymerase chain reaction (RT-PCR) analysis. HIF-1alpha mRNA showed equal expression in testis, all segments of epididymis, ductus deferens, accessory sex glands, and penis. To examine the effects of ischemia on HIF-1alpha mRNA and protein expression in the testis, rats were subjected to unilateral testicular ischemia by placing a ligature around spermatic artery or ischemia-inducing experimental torsion and reperfusion. RT-PCR revealed that HIF-1alpha mRNA expression at all times of ischemic treatment and reperfusion was unchanged compared with normoxic controls. HIF-1alpha protein was detected by immunoblot analysis of nuclear protein extracts from normoxic testes. Steady-state levels of HIF-1alpha protein were stimulated by 15 min of ischemia and showed a 2-fold increase at 30 min and 1, 3, and 6 h. HIF-1alpha protein was also elevated by experimental torsion and reperfusion compared with normoxic controls. These results support the hypothesis that HIF-1 may play a role in the cellular response to hypoxia in the ischemic testis.

Dynamic changes in gene expression along the rat epididymis.

In the epididymis a series of complex, sequential events transform immature, spermatozoa into mature, motile sperm with fertilizing ability. These events are not intrinsic to germ cells but rather are a direct result of exposure to, and interaction with, the environment created by the epididymal epithelium. Regional differences along the epididymis are essential in the establishment of the environment required for sperm maturation. Although parts of this process have been identified, the molecular basis for the segment-specific differences and how they contribute to the process of sperm maturation, are not yet resolved. The identification of genes expressed in a region-specific manner will provide valuable insight into the functional differences between the regions. To characterize gene expression in the different regions of the epididymis, microarrays containing 1176 rat cDNAs were used to examine gene expression in the initial segment, caput, corpus, and cauda epididymidis of the adult Brown Norway rat. Overall, the cauda epididymidis expressed the most genes and the corpus epididymidis the fewest. A small percentage of genes (3%) were expressed highly (greater than fivefold the average expression on the array) along the tissue. Segment-specific gene expression for genes expressed at high levels was observed in all epididymal segments except the corpus epididymidis. Of the genes on the array, 36% were expressed in all four epididymal segments; expression changes that were a minimum of twofold in either direction between adjacent segments are discussed. The expression of cathepsins and oxidative stress-related genes was investigated. Six of the eight cathepsins on the array (B, C, E, H, L, and K) were expressed above twofold background and showed different levels of expression along the duct with cathepsin K showing the most dramatic change (i.e., a decrease of 87% between the initial segment and the corpus epididymidis). There was also differential expression along the epididymis of many genes associated with oxidative stress defenses. Using the power of expression array technology, we have identified novel transcripts expressed in a segment-specific manner and been able to assess how the expression of several selected gene families is modulated along the epididymis.

Sobre a morfologia dos dúctulos eferentes epididimários no cão (Canis familiaris, L.)

The structure of the ductuli efferentes in the epididymis of the dog were studied by light microscopy. The efferent ductules connect the rete testis and the initial segment of the epididymis. Their epididymal segment is lined by a low simple cuboidal or columnar epithelium, which show basically nonciliated and ciliated cells. The morphological characteristics of ductuli efferentes are discussed in this report.

Gene Expression is Selectively Affected along the Epididymis after Orchidectomy.

INTRODUCTION. The epididymis is the site where immature spermatozoa from the testis acquire motility and the potential for fertilization. Although androgens decline more than 90% within 12 hours after orchidectomy, there is a wave-like pattern of epithelial principal cell apoptosis along the epididymis that progresses over 7 days (1); this coincides with the clearance of luminal fluid from the epididymal duct. The apoptotic response, detected by the TUNEL method, shows a peak at 2-3 days in the caput epididymidis and at 6 days in the cauda epididymidis. We hypothesize that orchidectomy has a dual effect on gene expression in epididymal cells; the first response is to the decreased serum androgen concentration while the second is to alter the content of the epididymal lumen. METHODS. Male Brown Norway rats were bilaterally orchidectomized and either sacrificed immediately or 2 days later. Epididymides were removed, sectioned and flash-frozen at –80. RNA was extracted from the caput and cauda epididymal segments, reverse transcribed, and probed on cDNA arrays (Clontech TM Rat Stress Toxicology II arrays, n=3 arrays/treatment group/time point). Each array contained 475 rat cDNAs related to the stress-toxicology gene families in addition to control cDNAs. Arrays were analyzed using GeneSpring TM software. Only genes having twice background levels were considered to be expressed; changes in gene expression of at least twofold will be discussed. RESULTS. On arrays using RNA from the control caput epididymidis, 195 genes were detected. Following orchidectomy, 73 genes are selectively up-regulated while only 6 genes were selectively down-regulated. Of those that increase, 6 are apoptosis-related including caspase-3, 5 are related to xenobiotic metabolism. Five genes are heat shock proteins, 12 are related to general cellular metabolism and 12 are related to transcription. Of the 6 genes that decrease, no single family of genes had more than 2 responders. The response to orchidectomy in the cauda epididymidis differs remarkably from that of the caput epididymidis. Of the 256 genes expressed in control, only 12 genes are up-regulated after orchidectomy; only one of these, clusterin, is apoptosis-related. The major trend after orchidectomy is toward a decrease in gene expression, with 103 genes being selectively downregulated. Of these genes, 3 are cell-cell adhesion receptors, 6 are related to xenobiotic metabolism, 29 are related to cellular metabolism and 9 are related to transcription. None of the genes down-regulated are related to the apoptosis.

Maturation-dependent glycoproteins containing both N- and O-linked oligosaccharides in epididymal sperm plasma membrane of rhesus monkeys (Macaca mulatta).

Glycosylation is one of the important post-translational modifications of sperm plasma membrane proteins during the maturation of epididymal spermatozoa that results in the development of motility and fertilizing capability. The aim of the present study was to identify and characterize the maturation-dependent asparagine-linked (N-linked) and serine- and threonine-linked (O-linked) glycoproteins of the epididymal spermatozoa of rhesus monkeys. The presence of N- and O-linked glycoproteins was confirmed by treatment of sperm membranes with N-glycosidase F and O-glycosidase. The major maturation-dependent sperm membrane glycoproteins identified on blots of SDS-PAGE-fractionated proteins of purified sperm plasma membranes from five segments of epididymis, probed with biotinylated lectins and Vectastain-ABC reagent included O-linked 170, 150, 86 and 60/58 kDa glycoproteins; N-linked 68, 56, 48 and 38 kDa glycoproteins and N- and O-linked 116 kDa glycoprotein, all of which exhibited marked differences in the degree of glycosylation between immature and mature sperm surfaces. These glycoproteins can be used as markers of sperm maturation in the epididymis of rhesus monkeys, during the screening of antifertility agents acting at the epididymis, or may be developed as potential sperm antigens. The 100% inhibition of fertility in female rats and rabbits immunized with major maturation-dependent 116 kDa glycoprotein showed the significance of glycosylation changes in the maturation status of epididymal spermatozoa. This 116 kDa protein can be used as a marker parameter of sperm maturation in the rhesus monkey, which is often the preferred animal model for preclinical studies. These results will contribute to the identification of an appropriate animal model for the development of male contraceptives in humans.

Andrographis paniculata (Nees) selectively blocks voltage-operated calcium channels in rat vas deferens

The possible blockade of voltage-operated calcium channels (VOCs) by Andrographis paniculata dried extract in vas deferens smooth muscle was investigated in rats. The tissues were incubated in Ca 2+-free Kreb’s solution and stimulated with KCl (40 mM) to produce depolarisation of the membrane. The isometric contractile response to cumulative concentrations of CaCl 2 was effectively blockaded by 0.2 and 0.4 mg/ml A. paniculata. In other experiments, the maximum contractile response induced by norepinephrine was not antagonised by 0.2, 0.4 or 0.8 mg/ml A. paniculata. The possible blockade of Ca 2+ entry by A. paniculata was evaluated with 45Ca 2+ uptake in vas deferens treated with reserpine (5 and 2.5 mg/kg) 48 and 24 h before the experiments. Epididymal segments were incubated with Ca 2+-free Kreb’s solution with KCl, 25 and 50 mM. The influx was completely blockaded with 0.4 mg/ml A. paniculata. These results suggest that A. paniculata selectively blockades VOCs, hence inhibiting the 45Ca 2+ influx.

Sex differences in the pharmacological activity of venom from the white-tailed spider ( Lampona cylindrata)

This study compared the pharmacological activity of venom from male and female white-tailed spiders ( L. cylindrata). In guinea-pig ileum, male L. cylindrata venom (1–10 μg/ml) caused dose-dependent contractions. The response to venom (5 μg/ml) was significantly inhibited by mepyramine (0.5 μM). Venom (5–50 μg/ml) from female L. cylindrata had no contractile activity in this tissue. However, female L. cylindrata venom (50 μg/ml) inhibited electrically-evoked twitches of guinea-pig ileum. This inhibitory effect was attenuated by 8-phenyltheophylline (10 μM) or by prior exposure of venom to adenosine deaminase. In the rat vas deferens, male (5 μg/ml) and female (50 μg/ml) L. cylindrata venom inhibited electrically-evoked twitches. 8-Phenyltheophylline (20 μM) significantly attenuated the response to female L. cylindrata venom, while the histamine H 2- and H 3-receptor antagonists ranitidine (10 μM) and thioperamide (0.2 μM) significantly attenuated the response to male L. cylindrata venom. Male L. cylindrata venom (5–20 μg/ml) caused dose-dependent contractions in the epididymal segment of the rat vas deferens. The response to male L. cylindrata venom (10 μg/ml) was significantly inhibited by prazosin (0.3 μM) but was unaffected by depleting monoamine stores with reserpine. Male L. cylindrata venom (5–15 μg/ml) caused dose-dependent increases in rate and force of rat atria which were significantly inhibited by propranolol (5 μM) but not by reserpine. Female L. cylindrata venom (50 μg/ml) had no effect in atria. In the anaesthetised (pentobarbitone, 100 mg/kg, i.p.) rat, male L. cylindrata venom (10–300 μg/kg, i.v.) caused dose-dependent depressor responses while venom (up to 1 mg/kg, i.v.) from female L. cylindrata had no effect on arterial pressure. A histamine content of 5 and 0.01% (dry weight) was detected in venom from male and female L. cylindrata, respectively. Venom from male L. cylindrata was found to contain 56 pg noradrenaline/μg whereas venom from the female contained negligible noradrenaline. The results of this study show the presence of histamine and noradrenaline in venom from male L. cylindrata. Although devoid of significant quantities of these amines, female L. cylindrata venom has activity at adenosine receptors.

Effects of short-term methylmercury exposure on metallothionein mRNA levels in the testis and epididymis of the rat.

Methylmercury (MeHg) is a widespread environmental contaminant that causes reproductive dysfunction in men. Metallothioneins (MTs) are low-molecular-weight proteins that can bind heavy metals and protect the cell from metal toxicity. MT levels are increased by exposure to metals and physiological stressors. Although MTs have been identified in the testis and epididymis, little is known about their distribution and regulation in the epididymis or the effects of MeHg on MT levels in male reproductive tissues. The objective of this study was to determine whether MT I, II, and III mRNA are present in the epididymis, if their relative levels differ between epididymal segments, and if MeHg alters cellular mRNA levels for MT I, II, and III in the testis and epididymal segments of the rat. Northern blot analysis was done on total cellular RNA isolated from each of the four epididymal segments (initial segment [IS], caput [CT], corpus [CS], and cauda [CA] epididymidis) using a cDNA probe for MT I and MT II. MT I transcripts were present in all epididymal segments. The lowest mRNA levels were observed in the IS; these levels were 4-fold less than in the CT and CS and 5.5-fold less than in the CA. MT II mRNA levels were similar in the IS and CT but were eightfold higher in the CS and CA. A cDNA probe for MT III was generated by reverse transcription-polymerase chain reaction using testicular RNA. MT III mRNA was detected only in the IS and CT and not in the CS and CA. To assess whether exposure to MeHg alters MT mRNA levels, rats were exposed for 14 days to one of five MeHg doses (0, 25, 50, 100, and 200 [microg/kg/day] via a subdermal osmotic pump. No changes were observed in either body weight or in the weights of the testis, epididymis, seminal vesicles, or ventral prostate between MeHg-treated and control rats. Serum testosterone levels were significantly decreased only at the highest MeHg dose. In the testis, MeHg treatment resulted in 2.5- to 7-fold increases in MT I mRNA levels. There were no changes in either MT II or MT III mRNA levels. In the initial segment of the epididymis, MT I mRNA levels were significantly increased only at the 50 microg/kg/ day dose, whereas there were no significant differences in MT II mRNA levels. In the caput epididymis, MT I mRNA levels were significantly lower at the 50 and 100 microg/kg/day dose. MT II mRNA levels were also lower, with the exception of the 50 microg/kg/day dose. Although MT III mRNA levels were lower at the two lower doses, levels were not different from controls in the two highest doses tested. In the corpus epididymidis, MeHg did not alter MT I mRNA levels, and MT II was higher only in the 50 microg/kg/day group. In the cauda epididymidis, MT I mRNA levels were decreased in a dose-dependent manner by up to 63%. MT II levels were unaltered. Together these data indicate that exposure of adult rats to MeHg can modulate MT mRNA levels in both the testis and epididymal segments. Furthermore, changes in MT mRNA levels following exposure to MeHg differ between epididymal segments, suggesting either differences in MeHg accumulation or differences in MT modulation.

Distribution of immune cells in the epididymis of the aging Brown Norway rat is segment-specific and related to the luminal content.

Remarkable changes occur during aging in the testis and epididymis of the Brown Norway rat. A dramatic increase in the number of halo cells, which are present in the epididymal epithelium and originate from the immune system, is found in animals of increasing age. Halo cells have been postulated to be either lymphocytes or monocytes. We hypothesized that halo cells are a mixture of different immune cells and that their relative composition changes with age. To verify this hypothesis, markers for helper T lymphocytes, cytotoxic T lymphocytes, B lymphocytes, and monocytes-macrophages were used to identify the major categories of immune cells in the epididymides of Brown Norway rats ranging in age from 3 to 24 mo. The numbers of immunocompetent cells in the epididymis were determined in relation to age, epididymal segment, and luminal content. We found that monocytes, helper T lymphocytes, and cytotoxic T lymphocytes belong to the population of halo cells. In addition, a segment-specific increase with age in the number of these immune cells was noted. Finally, we report a segment-specific recruitment of cytotoxic T lymphocytes and monocytes-macrophages in the epididymal epithelium of aged rats whose epididymal lumen contained few spermatozoa. We postulate that accumulation of damaged epithelial cells and antigens of germ cell origin, leaking through a dysfunctional blood-epididymis barrier, may contribute to the active recruitment of immune cells with age.

Recent biochemical approaches to post-testicular, epididymal contraception.

Results from recent animal models with implications for putative human male contraceptives acting on the epididymis are reviewed. Inducing sterility by enhancing sperm transport through the epididymis has not been achieved. The induction of infertility in males of several species is easier to achieve by direct actions of drugs on sperm function (e.g. inhibition of sperm-specific isoenzymes of the glycolytic pathway by chloro-compounds) than by indirectly reducing amounts of epididymal secretions normally present in high concentration (e.g. alpha-glucosidase, L-carnitine). The former show promise for the clinic since human spermatozoa are susceptible to inhibition. On the other hand, the infertile male mice of the c-ros knock-out model demonstrate the influence of even a small region of the epididymis on fertility, so that targeting the as yet unknown epididymal factors presumably secreted in limiting amounts by this epididymal segment, is a new lead for a contraceptive. Targeting a specific sperm protein acquired in the testis, but depleted in the epididymis by toxicants that induce rapid infertility, may also lead to the discovery of new contraceptives, but these will require developing new means of organ-specific delivery of contraceptive drugs.

Effects of ethanol ingestion on epididymal glycosidases and fertility in the rat.

Epididymal glycosidases play a role in sperm maturation by modifying sperm surface glycoproteins. To study the effects of ethanol on epididymal sperm maturation, ethanol (3 g/kg body weight as 25%, v/v) was administered to a group of rats by gastric-intubation twice daily for 30 days. In another group, rats were also treated with alcohol for 30 days but were then withdrawn from treatment for 30 days to assess the reversibility of ethanol-induced effects. Ethanol-induced changes in epididymal tissue and sperm glycosidases, cauda epididymal sperm motility and the fertility of rats were assessed. Ethanol treatment caused a marked decrease in the specific activities of glycosidases in both tissues and spermatozoa from epididymal segments. Cauda epididymal sperm motility and the fertility of ethanol-treated rats were significantly impaired compared to control rats fed an isocaloric diet. These changes are likely to be the consequence of direct and indirect effects of ethanol mediated through subnormal testosterone and dihydrotestosterone. Most of these changes were found to be reversible. The present study suggests that impaired activity of sperm glycosidases may be one of the factors responsible for defective sperm motility and fertilizing potential in ethanol-treated rats.

The characteristics of endothelin receptor subtypes on muscle contraction and neuro-transmission in rat vas deferens.

We observed endothelin (ET)-induced contractile responses on prostatic and epididymal segments, as well as the facilitation of an electrically stimulated tone on prostatic segments of isolated rat vas deferens. In both segments, the selective ET(B)-receptor agonists, IRL 1620 and sarafotoxin S6c, produced only a small contraction or no contraction at a concentration of 1 microM. The rank order of contraction potencies (pD2 value) was ET-1 = ET-2 > ET-3 >> sarafotoxin S6c = IRL 1620. The maximum responses of ET-induced contractions in the prostatic segments were larger than those in the epididymal segments. The contractile response to ET-3 was antagonized by pretreatment for 30 min with BQ-123 (10 nM), a selective ET(A) receptor antagonist, and BQ-788 (1 microM), a selective ET(B) receptor antagonist. The contractile responses to ET-1 were antagonized by pretreatment with BQ-123 (10 microM), but not with BQ-788 (1 microM). The ET-3-induced facilitation on the twitch response to electrical stimulation in the prostatic segment of the vas deferens was antagonized by BQ-123 (0.1 microM) and BQ-788 (1 microM). The ET-1-induced facilitation was antagonized by pretreatment with BQ-123 (3 microM), but not with BQ-788 (10 microM). These results suggest that in rat vas deferens the ET(A) receptors are divided into BQ-123-sensitive ET(A1) and BQ-123-insensitive ET(A2) subtypes, and the production of a contractile response of smooth muscle as well as the facilitation of neurotransmission are accomplished through mediation by ET(A1)- and ET(A2)-subtypes.

Stability and transcriptional regulation of gamma-glutamyl transpeptidase mRNA expression in the initial segment of the rat epididymis.

Gamma-glutamyl transpeptidase (GGT) is an enzyme believed to play a role in the protection of maturing spermatozoa in the epididymis. Our previous studies have shown that four GGT mRNAs (I-IV) transcribed from the single-copy rat GGT gene are differentially expressed and regulated in the rat epididymis. In particular, the normal expression of GGT mRNA(IV) in the epididymal initial segment is dependent upon the presence of testicular factors. The objective of this study was to test the hypothesis that the decreased expression of GGT mRNA(IV) in the initial segment following the in vivo removal of testicular factors by efferent duct ligation (EDL) is due to a decrease in stability and/or transcription rate. The stability of the GGT mRNAs was evaluated by measuring the rate of mRNA decay. These stability studies showed that GGT mRNA(IV) exhibited a rapid initial decay that slowed at later times to a decay rate similar to that of GGT mRNAs(II,III). The decay rates were not different following sham-operation or EDL, and thus the stability of GGT mRNAs were not influenced by the in vivo loss of testicular factors. Results of transcription analysis revealed that the transcription rate of GGT mRNA(IV) in the initial segment fell by approximately 68% following a 12-hour EDL. Additionally, secondary-structure models indicate two families of folding patterns for GGT mRNA(IV), which could be the reason for the two decay regimes detected in the stability study. Thus, the decreased expression level of GGT mRNA(IV) in the initial segment following the in vivo loss of testicular factors is a function of a decreased transcription rate and intricate decay kinetics.

Alpha-Glucosidase activity in the rat epididymis under different physiological conditions.

The estimation of alpha-glucosidase activity in semen is widely used as a marker of epididymal function. In the present studies, glucosidase activity was evaluated in the different segments of the rat epididymis under various physiological conditions. In addition, the effect of two known male antifertility agents, gossypol and alpha-chlorohydrin, on enzyme activity was evaluated. Enzyme activity was absent from the epididymis of rats aged 10 and 20 days but became detectable at 30 days of age when the adult pattern of distribution (highest activity in the caput epididymis) was established. Enzyme activity was reduced significantly in all segments of the epididymis at 7 days after castration and a significant decrease in activity was also observed following the administration of either gossypol or alpha-chlorohydrin. These findings are consistent with a role for alpha-glucosidase in sperm maturation in the epididymis.

Sperm maturation in rhesus monkey: changes in ultrastructure, chromatin condensation, and organization of lipid bilayer.

The changes in ultrastructure, lipid organization, chromatin decondensation, and denaturation of rhesus monkey spermatozoa during epididymal maturation were studied. This study would provide background information that would be useful to evaluate adverse effects, if any, caused by the use of contraceptive agents. Adult sexually mature rhesus monkeys were castrated under ketamine anesthesia. The epididymis was divided into initial segment, caput, corpus, and cauda epididymides. To study changes in lipid organization of the sperm plasma membrane during epididymal transit, spermatozoa from different epididymal segments and ejaculated spermatozoa were exposed to merocyanine 540 (MC 540). The changes in chromatin denaturation and decondensation were assessed by using the nucleic acid-specific fluorochromes, acridine orange, and ethidium bromide, respectively, prior to and after exposure to dithiothreitol (DTT). Testicular spermatozoa (approximately 40%) showed localization of MC 540 mainly in the midpiece, whereas remaining sperm did not localize MC 540. Spermatozoa from the initial segment of the epididymis showed uniform distribution of MC 540 localization in the head and midpiece. A pattern of localization of MC 540 similar to mature caudal and ejaculated sperm in which the staining was restricted to the acrosome and the midpiece first appeared in a small percentage of caput spermatozoa and was completed during transit through the corpus epididymidis. Mature spermatozoa from cauda epididymidis, vas deferens, and ejaculate did not undergo chromatin denaturation even after exposure to 10 mM DTT, unlike sperm from testis, initial segment, and caput epididymidis. Spermatozoa exposed to DTT showed chromatin decondensation; maximum decondensation was seen in testicular sperm and a decrease in the percentage of sperm, showing decondensation, occurred during epididymal transit. Ultrastructural studies showed that spermatozoa undergo structural changes during sperm maturation. The present study shows that rhesus monkey spermatozoa undergo reorganization of the plasma membrane lipids and stabilization of disulfide linkages during epididymal transit. The results would be of use in evaluating the action of potential male contraceptive drugs on epididymal spermatozoa.

Nucleotide sequence of the complementary DNA encoding equine clusterina

Abstract Two complementary DNA (cDNA) clones encoding equine clusterin were isolated from a stallion testis cDNA library. Nucleotide sequence analysis of the longer, near‐full length clone revealed a 1670 base pair (bp) fragment that contained 74 nucleotides of the 5’ untranslated region (UTR), an open reading frame of 1350 bp that encodes a 21 amino acid (aa) leader polypeptide segment as well as a 429 aa mature equine clusterin protein, and a 3’ UTR that is 225 bp long followed by a 21 bp polyadenylation signal. The equine clusterin nucleotide sequence is approximately 80% similar to other mammalian clusterins which have been cloned and sequenced. Northern blot analysis showed an approximate 1.8 kb equine testicular and epididymal mRNA transcript that hybridized to the equine cDNA probe. Differential expression of the equine clusterin transcript occurred between epididymal segments.

Autoradiographic distribution and physiological REGULATION OF 2-[ 125I]iodomelatonin binding in rat epididymis

Autoradiographic study was conducted to localize 2-[ 125I]iodomelatonin binding in the rat epididymis. In the peripubertal (6 weeks old), postpubertal (8 weeks old) and adult (3 months old) rats, intense specific 2-[ 125 I]iodomelatonin labelling of the corpus epididymidis was observed. The intensity of 2-[ 125I]iodomelatonin binding in the distal epididymal segment was significantly decreased in orchidectomized rats but the effect could be reversed with testosterone replacement. The intensity of 2-[ 125I]iodomelatonin binding in the distal rat epididymal segment did not show any diurnal rhythmicity when mid-light period and mid-dark period levels were compared, and was unaffected by constant lighting. Our data suggest androgen-dependent expression of 2-[ 125I]iodomelatonin binding sites, independent of lightinduced changes in circulating melatonin, in the rat corpus epididymidis. A novel role of melatonin and its receptor in the regulation of the functions of rat corpus epididymidis is strongly implicated.