Changes in protein composition of the luminal fluids along the epididymis of the tammar, Macropus eugenii

Micropuncture samples of luminal fluid were collected from the rete testis and along the epididymis. Quantitative analyses showed that the ductuli efferentes reabsorb about half the protein leaving the testis. Considerable protein is secreted by the caput epididymidis (initial segment) and there is a net loss of protein from the corpus and cauda epididymidis. Denatured, polyacrylamide gel electrophoresis showed that there are 5 proteins in rete testis fluid which are not present in blood (Mr of 14,700, 22,800, 24,100, 43,000 and 44,800). One of these proteins (Mr 14,700) is lost from plasma in the ductuli efferentes and 2 (Mr 43,200 and 44,800) are lost in the corpus epididymidis. Twelve proteins appear in the epididymal plasma and are not present in rete testis fluid or blood: 6 appear in the caput epididymidis (Mr 30,000, 31,000, 32,300, 17,400, 18,700 and 21,400), 3 in the corpus epididymidis (Mr 12,800, 39,800 and 90,600) and 3 in the cauda epididymidis (Mr 10,900, 56,300 and 63,000). A protein with the same molecular weight as a blood protein (149,500) accumulates in the corpus and cauda epididymidis. None of the samples of luminal fluid contained particulate matter other than spermatozoa, indicating that the tammar is a useful animal for micropuncture studies.     

Immunolocalization and concentrations of inhibin alpha in the ovine testis and excurrent duct system

Inhibin was localized in the ovine testis, excurrent ducts, and accessory sex glands by using a rabbit antiserum against a synthetic polypeptide representing the first 30 amino acids of porcine inhibin alpha-subunit. Concentrations of inhibin in fluids entering and leaving the epididymis also were determined in a radioimmunoassay using the same antibody. In the testis, immunostaining of inhibin was conspicuous in the seminiferous epithelium. Leydig cells occasionally were stained and the tunica media of blood vessels always was stained. Intense staining was observed in the epithelia lining the rete testis and ductuli efferentes. Staining also was intense in the epithelium of the initial segment and proximal caput epididymidis, and became less intense along the length of the epididymis. These observations were consistent with concentrations of inhibin in rete testis fluid (8.2 pmol/ml) entering the ductuli efferentes and in cauda epididymal plasma (0.67 pmol/ml) leaving the epididymis. Epithelia of ampullary and vesicular glands and of some prostatic acini were positively stained, but bulbourethral glands were never stained. Adrenal cortex, some proximal convoluted tubules in the kidney, and transitional epithelium of the urethra also were stained. Based on radioimmunoassay data and fluid flow rates for the ram, it was concluded that almost all of the 328 pmol inhibin that enters the ductuli efferentes daily is endocytosed in the proximal parts of the excurrent duct system. The physiological role(s) for inhibin, or inhibin-like peptides, in the excurrent duct system remains speculative.     

Transport of IgG across the blood-luminal barrier of the male reproductive tract of the rat and the effect of estradiol administration on reabsorption of fluid and IgG by the epididymal ducts

In rats immunized systemically with tetanus toxoid the concentration of specific anti-tetanus-toxoid-specific IgG in fluid from the rete testis and cauda epididymidis were respectively 0.6% and 1.4% the concentration in blood serum. The extratesticular duct system reabsorbed 97% of the IgG and 99% of the fluid leaving the rete, but estradiol administration affected the site of reabsorption. In untreated rats, the ductuli efferentes reabsorbed 94% of the IgG and 96% of the fluid leaving the rete, whereas estradiol-treated rats reabsorbed 83% of the IgG and 86% of the fluid, and the ductus epididymidis fully compensated for these different effects of estradiol on the ductuli efferentes. The concentrations of IgG in secretions of the seminal vesicles and prostate gland were lower (0.1% and 0.3% respectively of the titers in blood serum) than in fluids from the extratesticular ducts, and were not affected by the administration of estradiol. RT-PCR showed that Fcgrt (neonatal Fc receptor, also known as FcRn) is expressed in the reproductive ducts, where IgG is probably transported across epithelium, being particularly strong in the ductuli efferentes (where most IgG was reabsorbed) and distal caput epididymidis. It is concluded that IgG enters the rete testis and is concentrated only 2.5-fold along the extratesticular duct system, unlike spermatozoa, which are concentrated 95-fold. Further, the ductus epididymidis can recognize and compensate for changes in function of the ductuli efferentes.     

Morphology of the epithelium of the extratesticular rete testis, ductuli efferentes and ductus epididymidis of the adult male rabbit

The fine structure of the epithelium lining the extratesticular rete testis, ductuli efferentes and ductus epididymidis of the rabbit has been investigated. In the ductuli efferentes the epithelium is composed of two cell types, principal cells and ciliated cells. The latter type is distinguished from principal cells by the presence of cilia projecting into the lumen and the position of the nucleus in the apical half of the cell. Principal cells in this segment are characterized by micropinocytotic vesicles on the surface plasma membrane and a variety of small dense bodies scattered throughout the cytoplasm. In the ductus epididymidis basal cells replace ciliated cells as the second cell type, but differences between various segments of the epididymis are related to the fine structure of the principal cells. In the proximal caput epididymidis (Nicander's region 1) the principal cells are tall with long microvilli. They typically contain a small Golgi apparatus and a cluster of dense bodies adjacent to the nucleus. In the distal caput epididymidis (Nicander's regions 2-5) the apical cytoplasm of principal cells is filled with numerous micropinocytotic vesicles and large multivesicular bodies; these features are interpreted as signs of absorptive activity. The multivesicular bodies are absent from the cytoplasm of principal cells in the corpus epididymidis (Nicander's region 6) and, instead, numerous elements of smooth endoplasmic reticulum, a large Golgi apparatus, lipid droplets and dense bodies characterize principal cells in this segment. Towards the proximal cauda epididymidis (Nicander's region 7), the number of dense bodies (lysosomes) in the cytoplasm increases considerably. In the globose cauda (Nicander's region 8), the principal cells are reduced in height, and in addition to the features described in region 7, are characterized by a concentric array of rough endoplasmic reticulum in the basal cytoplasm. These observations are discussed in relation to the role of the epididymis in promoting the maturation and survival of spermatozoa.     

Protein gene product 9.5 and ubiquitin immunoreactivities in rat epididymis epithelium

A quantitative immunohistochemical study was performed of the distribution of protein gene product 9.5 (PGP, a soluble protein localized in neurons and neuroendocrine cells as well as in some non-nervous cells) and ubiquitin along the rat epididymis. In the ductuli efferentes, PGP immunoreaction was observed in the whole cytoplasm of some columnar cells; a smaller number of columnar cells showed ubiquitin immunoreactivity with limited apical and basal cytoplasmic localization. In the proximal caput epididymidis, the whole cytoplasm of all columnar cells showed PGP immunoreactivity, ubiquitin immunostaining was negative in this region. In the middle and distal caput epididymidis and the distal cauda, the apical cytoplasm of some columnar cells and the whole cytoplasm of some basal cells showed immunoreactivity to PGP. In these regions, immunoreactivity to ubiquitin was positive in the supranuclear cytoplasm of some columnar cells but not in the basal cells. No immunoreactivity to PGP or ubiquitin was detected in the corpus epididymis and the proximal cauda. Double immunostaining revealed that all the epididymal ubiquitin immunoreactive cells were also PGP immunoreactive, whereas most PGP immunoreactive cells did not immunoreact to ubiquitin. In ubiquitin-PGP immunoreactive cells, the site of the PGP immunoreaction differed from that of the ubiquitin immunoreaction. PGP-ubiquitin immunoreactive cells also seemed to be immunoreactive to anti-AE1/AE3 keratin antibodies. The spermatozoal heads were immunoreactive to PGP antibodies in the epididymal regions from proximal caput to distal cauda but not in the ductuli efferentes. The findings suggest that non-ubiquitinated PGP immunoreactive proteins are secreted in the epididymis, mainly in the proximal caput, and attach to spermatozoa.     

Appendix epididymidis and aberrant ductules of the bull: light-microscopic and ultrastructural study

The appendix epididymidis and aberrant ductules possessed similar morphological characteristics. The epithelium was 31 +/- 3 microns in height and consisted primarily of ciliated and nonciliated cells, although a few lymphocytes were also present. The ultrastructure of major cell types showed most cell organelles in their cytoplasm. However, these organelles were poorly developed, suggesting that neither cell type performed either a secretory or an absorptive function. Although the vestigial organs and ductuli efferentes were similar in epithelial height and epithelial cell types, there were important morphological differences that were reliably used to differentiate between the two. First, the luminal diameter was significantly smaller in the vestigial organs (60 +/- 12 vs. 146 +/- 44 microns in the ductuli efferentes). Second, the nonciliated cells of the vestigial organs, unlike those of the ductuli efferentes, lacked both dense granules and vacuoles in the cytoplasm. Finally, the tubular cross-sections of the vestigial organs were closely packed and were located at the tip of the caput epididymidis in the case of the appendix epididymidis, and between the lobules of the ductuli efferentes in the case of the aberrant ductules.     

Regulation of the elemental composition of the epididymal fluids in the tammar, Macropus eugenii

Micropuncture, microanalytical and microelectrode techniques were used to study electrochemical aspects of 7 elements and fluid in the ductuli efferents and ductus epididymidis of the tammar. Rete testis fluid was isosmotic with blood and had a lower pH. It also contained lower concentrations of bicarbonate, sodium, calcium, magnesium, phosphorus and sulphur and higher concentrations of potassium and chloride than blood. The luminal fluid was acidified further during passage through the sperm ducts and all of the elements which were studied moved in or out of the lumen, usually against an electrochemical gradient. The ductuli efferents reabsorbed 87% of the fluid leaving the testis without changing the intraluminal concentrations of sodium, potassium and calcium, but the concentrations of magnesium, phosphorus and sulphur increased. The caput epididymidis reabsorbed about half the fluid entering it: sodium concentrations decreased and those of potassium and phosphorus increased. There was also some fluid reabsorption and an increase in the values of potassium and phosphorus in the corpus epididymidis. There was little net transport of fluid in the cauda epididymidis; sodium, chloride, magnesium and phosphorus concentrations decreased and potassium values increased. Studies involving filtration through a dialysis membrane of blood and fluid from the rete testis and cauda epididymidis showed that, whilst some of the calcium, magnesium, phosphorus and sulphur was associated with high molecular weight compounds in blood, the association was not significant in the reproductive fluids.     

Androgen receptor and 5 alpha-reductase activity in the ductuli efferentes and epididymis of adult rhesus macaques

We measured androgen receptors (AR) and 5 alpha-reductase activity (5 alpha RA) in the ductuli efferentes and epididymides from adult rhesus macaques. Tissue samples were either assayed biochemically for AR or stained immunocytochemically (ICC) with a monoclonal antibody against AR. To estimate 5 alpha RA, tissue microsomes were incubated with [1 alpha,2 alpha-3H]testosterone, and the [3H]dihydrotestosterone formed was quantified. We found significant regional differences in the levels of both 5 alpha RA and AR in the excurrent ducts. In general, both enzyme activity and AR levels were higher in the caput and corpus epididymis than in ductuli efferentes and cauda epididymis. With ICC, positive nuclear AR staining was detected in all epithelial cell types, whereas variable numbers of stromal cells were positively stained. Our data demonstrate that there are segmental differences in the concentrations of 5 alpha RA and AR in epididymis and suggest that there may be regional differences in the regulation of epididymal functions by androgen.     

Estrogen receptor in the ductuli efferentes, epididymis, and testis of rhesus and cynomolgus macaques

We obtained the testes, ductuli efferentes, and epididymides from adult rhesus and cynomolgus macaques and examined these tissues for estrogen receptors (ER) with immunocytochemistry (ICC) and a sucrose gradient assay. Both techniques employed monoclonal antibodies prepared against ER, and both showed that high concentrations of ER were present OFFy in the ductuli efferentes. Moreover, all specific staining was confined to the nuclei of the nonciliated, absorptive epithelial cells. The quantity of salt-extractable ER in the ductuli efferentes (834 +/- 161 [SEM] fmol/mg DNA [n = 8]) did not differ significantly from the amounts measured with the identical assay in oviducts and endometrium of estrogenized female macaques. Testes and epididymides of macaques had no specific staining by ICC and barely detectable amounts by biochemical analysis (7 +/- 4 [n = 3], 8 +/- 2 [n = 5], 33 +/- 16 [n = 3], and 6 +/- 3 [n = 8] fmol/mg DNA for testis and caput, corpus, and cauda epididymis, respectively). The functional significance of the high levels of ER in the ductuli efferentes of macaques remains to be determined.     

Changes in the morphology of spermatozoa during their passage through the genital tract in dairy bulls with normal and impaired spermat ogenesis

An investigation was conducted on changes in sperm morphology along the excurrent duct of bulls. The study comprised 20 bulls with proven fertility and normal semen picture, and 10 bulls with pathologic semen. The morphology of spermatozoa was evaluated on ejaculates and on postslaughter collected contents from the excurrent duct. The incidence of pathologic sperm heads decreased along the duct in both groups of bulls. The main decrease generally occurred in caput epididymidis. In bulls with pathologic semen, the decrease continued in lower regions of epididymis and was deferens. The rate and pattern of sperm removal seem to primarily depend on the quality of spermatozoa entering the excurrent duct. The removal was clearly selective and is assumed to be associated with phagocytosis of spermatozoa mainly in the efferent ductules and proximal part of caput epididymis. Proximal cytoplasmic droplets were present on almost all sperm in the efferent ductules. The incidence decreased during passage along the genital tract. Migration of cytoplasmic droplets from a proximal to a distal position took place between regions C and D of the caput epididymidis. The incidence of middle-piece abnormalities decreased during passage along the genital tract, while the incidence of sperm tail abnormalities increased in the corpus and cauda epididymidis in all bulls.     

Lectin staining of rat testis and epididymis after ligation of excurrent ducts at different levels

Seven rhodamine-conjugated lectins (PNA, RCA I, SBA, Con A, WGA, UEA I, and DBA; see Table 1) were utilized in studying the staining pattern of glycoproteins in rat testis and epididymis after ligation of ductuli efferentes (DE), corpus epididymidis (CE), and vas deferens (VD) for various time periods. Ductuli efferentes ligation caused a widening of seminferous tubules and detachment of spermatids with formation of multinuclear cells. These cells acquired a strong affinity for all lectins. Corpus epididymidis ligation also caused degeneration of spermatids with increased lectin staining in some tubules, but after 7 days another cell population close to the periphery of seminiferous tubules showed an increased nuclear affinity for some lectins followed by a clear degeneration and strong cytoplasmic staining with all lectins. Vas deferens ligation caused no degenerative changes in testicular spermatids. However, the peripheral cell population showed degenerative changes similar to those found after CE ligation. In both cases this was coincident with the formation of spermatic granulomas at the site of ligation. Ductuli efferentes ligation caused a gradual decrease of intratubular content in caput epididymidis, while the contrary was true after CE ligation. The latter was associated with intratubular accumulation of lectin-positive swollen cells and sperm aggregates as well as an increased lectin staining of narrow cells in initial segment and light cells in distal caput. After VD ligation an increased staining of light cells was initially found in distal cauda and distal caput, but, concomitant with distension of the tubules this reaction decreased.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immunohistochemical investigation of different cytokeratins and vimentin in the human epididymis from the fetal period up to adulthood

Summary The anatomical distribution of cytokeratins and vimentin was investigated by means of immunohistochemistry in the human epididymis. Epithelial cells of the ductuli efferentes and the corpus epididymidis were positive for cytokeratins and vimentin. The expression of epithelial vimentin decreased toward the cauda epididymidis, whereas cytokeratins remained unchanged. The epithelium of the ductus deferens was negative when antibodies against vimentin were used. With monoclonal antibodies to individual cytokeratins, the presence of cytokeratins 7, 8, 18, and 19 was demonstrated histochemically throughout the epithelium of the epididymis. Monoclonal antibodies specific for cytokeratin 17 allowed immunohistochemical differentiation between the ductuli efferentes and the ductus epididymidis.     

Distribution of carnitine and acylcarnitine in the hamster epididymis and in epididymal spermatozoa during maturation

The highest levels of carnitine and acylcarnitine were found in the cauda epididymidis, and spermatozoa from the cauda contained greater amounts of total carnitine (free carnitine plus acylcarnitine) than those removed from the corpus or caput epididymidis. Spermatozoa from the distal cauda contained significantly greater amounts of both free and total carnitine than those removed from the proximal cauda epididymidis. The acylcarnitine:carnitine ratio was 1.7 and 0.37 in caput and cauda spermatozoa, respectively and 1.7 and 1.3 in caput and cauda fluid, respectively. It is suggested that the accumulation of carnitine is involved in sperm maturation and that acylcarnitine serves as an energy substrate for epididymal spermatozoa.     

Structure and functions of the genital ducts of the male Port Jackson shark,Heterodontus portusjacksoni

Synopsis The genital ducts ofHeterodontus portusjacksoni consist of the sperm carrying ducts (the rete testis, ductuli efferentes, and initial and terminal segments of the ductus epididymidis) and the Leydig glands (anterior opisthonephros). The ducts are lined by a ciliated epithelium which maintains a barrier to the transport of solute between blood and the lumen of the duct. Spermatozoa, Sertoli cell bodies, Sertoli cell cytoplasts and cellular debris are released from spermatocysts into the longitudinal canal of the rete testis. However, only the Sertoli cell cytoplasts persist throughout the sperm ducts. The epithelia lining the initial segment of the ductus epididymidis and secretory tubules of the Leydig glands are specialized for protein secretion and (particularly the Leydig glands) must be the main source of luminal protein in the ductus epididymidis. The epithelium lining the terminal segment of the ductus epididymidis also secretes protein, reabsorbs fluid and sodium, and may carry out heterophagic digestion. Spermatozoa develop the capacity for motility in the extratesticular sperm ducts, but do not undergo structural changes. However, they form spherical bundles in the terminal segment of the ductus epididymidis. It is suggested that the reduction in ratio of sodium:potassium from 48:8 in the ductuli efferentes to 3:4 in the distal end of the terminal segment of the ductus epididymidis may favour sperm survival.     

Sperm maturation, fluid transport, and secretion and absorption of protein in the epididymis of the echidna, Tachyglossus aculeatus

Extratesticular sperm maturation in the echidna mainly occurs in the initial segment of the ductus epididymidis. The process involves the development of motility, migration and loss of the cytoplasmic droplet, a decrease in permeability to Congo red and the formation of sperm bundles. The spermatozoa are supported in the bundles by a matrix of electron-dense material; the bundles are very motile is undiluted samples of luminal fluids. Micropuncture studies of anaesthetized echidnas showed that the ductuli efferentes absorb 74% of the fluid and 46% of the soluble protein that enters them. The initial segment of the ductus epididymidis absorbs 83% of the fluid which enters it, and its secretions increase the concentration of protein in luminal fluid by 107%. Denatured, linear-gradient polyacrylamide gel electrophoresis of micropuncture samples showed that 1 protein (apparent M4 = 100 500) which is not present in blood plasma is present in rete testis fluid, and a glycoprotein which is present in rete testis fluid (apparent Mr = 78500) is absorbed by the ductuli efferentes. Six proteins which are not present in blood plasma are secreted into the initial segment of the ductus epididymidis; 5 are glycoproteins (apparent Mr = 48500, 39000, 32000, 20500 and 19000) and one (apparent Mr = 82500) is not. The most prominent electrophoresis bands corresponded to the glycoproteins with apparent molecular weights of 48500, 20500 and 19000.     

GPX5 is present in the mouse caput and cauda epididymidis lumen at three different locations

In mice, GPX5 is a secreted protein abundantly synthesized by the caput epididymidis. The protein is secreted as early as the initial segment of the caput and is found subsequently associated with the sperm plasma membrane in a sub-acrosomic localization. We show here that GPX5 is present in the caput and cauda epididymides lumens in three different locations: either free as a soluble protein in the caput epididymal fluid, weakly bound to caput sperm membranes, or, finally, associated to lipid-containing structures conferring to the protein a protective effect against proteolytic digestions. Within the cauda epididymidis, the amount of free GPX5 is low compared to the caput and the association with sperm membranes proved to be more solid. In both caput and cauda sperm samples, the association of GPX5 with the sperm membrane protects GPX5 from proteolytic cleavages. Protection against proteolytic digestions can be overcome by physical treatments of epididymal fluid and sperm samples such as ultrasounds or very acidic pH. These data suggest that complex phenomena and structures participate in the transfer and binding of the caput-secreted GPX5 protein to the sperm plasma membrane.     

Spontaneous electrical activity of the rat epididymis in vitro

Smooth muscle electrical activity was recorded with suction electrodes from the partly or completely uncoiled epididymal duct of the rat in vitro. The electrical activity of the cauda epididymidis consisted of one or few spikes followed by a plateau of 1-2 sec. The frequency of electrical activity declined from the thicker-walled initial segment of the thin-walled initial segment, was increased to the level seen in the initial segment in the thicker, major portion of the caput epididymidis, declined in the corpus and fell steeply in the cauda epididymidis towards the vas deferens. Electrical activity spread over long distances in the distal cauda and epididymal vas. Elsewhere in the epididymis activity remained synchronous only for a short period in short segments.     

Resorption versus secretion in the rat epididymis

Micropuncture samples were taken from the rete testis, caput epididymidis and cauda epididymidis of anaesthetized adult rats and assayed for total protein, sodium and potassium concentrations. Intraluminal sperm concentrations were determined and used to calculate the amount of fluid resorbed from the efferent duct and epididymal lumen. It was demonstrated that large amounts of protein (30.2 mg/ml cauda volume) and sodium (241.8 mequiv./l) and smaller amounts of potassium (19.4 mequiv./l) are resorbed from the rat epididymal lumen between the caput and corpus epididymidis. This occurs despite increases in intraluminal concentrations of protein (from 22 to 28 mg/ml) and potassium (from 16 to 50 mequiv./l). Resorption is an important aspect of epididymal control of the intraluminal environment.     

Microanatomy of the testes and testicular ducts of the butterfly lizard,Leiolepis ocellata Peters, 1971 (Reptilia: Squamata: Agamidae) during the active reproductive period

The microanatomy of the testes and testicular ducts (rete testis, ductuli efferentes, ductus epididymis and ductus deferens) of Leiolepis ocellata (Agamidae) was investigated using light microscopy including histochemistry. Each testis contains seminiferous tubules and interstitial tissues. The former house spermatogenic cells (spermatogonia A & B, preleptotene, primary and secondary spermatocytes, spermatids (steps 1–8) and spermatozoa) and Sertoli cells, while the latter comprise peritubular and intersitial tissues. The rete testis is an anastomosing duct, having intratesticular and extratesticular portions. The proximal region of ductuli efferentes has wider outer ductal and luminal diameters than those of the distal region. The convoluted ductus epididymis is subdivided into four regions (initial segment, caput, corpus and cauda), based on the ductal diameter, epithelium characteristics and cell components. The ductus deferens has the greatest diameter and is divided into the ductal and ampulla ductus deferens. The ductal portion is subdivided into the proximal and distal regions, based on the epithelium types and ductal diameters. The ampulla ductus deferens is a fibromuscular tube, having numerous mucosal folds projecting into the lumen. Spermiophagy is detectable in the ductus epididymis and ductus deferens. The present results contribute to improved fundamental knowledge on the microanatomy of the reptilian reproductive system.