Effect of photic changes and olfactory impairment on reproduction in female south Indian gerbil

Effect of exposure to constant light (CL), blinding and olfactory bulbectomy (OBX) on reproduction of adult and weanling female Tatera indica cuvieri was investigated. In adult females, CL induced changes in estrus cyclicity. Weanlings subjected to CL showed reduced ovarian weight. Blinding did not bring about changes in estrus cyclicity and reproductive organ weight (ovary and uterus) of either adults or weanlings. Estrus cyclicity of both adults and weanlings were affected consequent to OBX. In weanlings, OBX lowered the ovarian and uterine weight.     

Olfactory bulbectomy-induced changes in the reproductive behaviour of the south Indian gerbil, Tatera indica cuvieri

Male South Indian gerbils (T. indica cuvieri), both adults and weanlings, were olfactory bulbectomized (OBX) and changes in male reproductive behaviour were assessed. Consequent to OBX, majority of the adult males failed to ejaculate, and courtship behaviour has also been considerably reduced. All OBX weanlings were rendered incapable of ejaculation. However, maturational parameters, and organ weights (testes, epididymis and seminal vesicle) remained unchanged in OBX males.     

Comparison of male reproductive parameters in three rat strains: Dark Agouti, Sprague-Dawley and Wistar

The choice of experimental animal can have a large impact on experimental results, an example is the anecdotal evidence suggesting that Dark Agouti (DA) rats have a lower reproductive capacity than other rat strains. In this paper we report on an investigation into male reproductive characteristics in three rat strains--Wistar, Sprague-Dawley (outbred strains) and DA (an inbred strain). Reproductive organ weights, blood testosterone levels and sperm counts were measured in mature age-matched male rats. DA animals had significantly smaller testis weights than the Sprague-Dawley and Wistar animals, and this did not appear to be related to the overall smaller body mass of the DAs. There were no differences between the three strains in testicular histology or sperm counts (per gram testis). Although there was also no significant difference in epididymal sperm count, the DA animals had a much greater variability in sperm count than the other strains. There were no differences in relative (to body weight) epididymal, seminal vesicle or ventral prostate weights or in the blood testosterone levels. These results suggest that differences in reproductive capacity in DAs are neither the result of morphological differences in the reproductive organs nor in circulating testosterone levels. Sperm production appears to be normal but the lowered testicular weight and variability in epididymal sperm counts suggests that there are other factors in the testicular or epididymal environment which alter male reproductive function.     

Effect of different intensities of swimming exercise on testicular oxidative stress and reproductive dysfunction in mature male albino Wistar rats

Swimming exercise for 1, 2 and 3 hr for 5 days/week for consecutive 4 weeks, results in a significant reduction in testicular, epididymal, prostetic, seminal vesicle somatic indices; epididymal sperm count, sperm motility; preleptotine spermatocytes, mid pachytene spermatocytes and stage 7 spermatids; plasma levels of testosterone, luteinizing hormone; testicular delta5, 3beta-hydroxysteroid dehydrogenase, 17beta-hydroxysteroid dehydrogenase; testicular superoxide dismutase, catalase, glutathione peroxidase, glutathione-s-transferase and glutathione along with significant elevation in malondialdehyde in male albino rats. However, no significant change was noted in final body weight, spermatogonia-A and plasma level of follicle stimulating hormone. The results that oxidative stress develops with the increasing of exercise intensity, which may interfere in male reproductive activities.     

Sperm morphology of the neotropical harvestman Iporangaia pustulosa (Arachnida: Opiliones): comparative morphology and functional aspects

We describe herein the sperm morphology of the harvestman Iporangaia pustulosa. Adult males were dissected, the reproductive tract was schematized and the seminal vesicle was processed by light, transmission and scanning electron microscopy. The male reproductive tract is composed of a tubular testis, two deferent ducts, a seminal vesicle, a propulsive organ and a penis, similar to that observed in other Opiliones. The spermatozoa from the seminal vesicle are oval, aflagellate and immotile, presenting a nucleus surrounding an invagination of the cytoplasm, as well as a complex acrosome and projections on the cell surface. In the testis, spermatozoa are devoid of projections. In the seminal vesicle, they gradually acquire the projections with tufts adhering to it. Consequently, spermatozoa in various distinct stages of projection development can be found in the seminal vesicle. We believe that these projections (1) could help transport sperm along the male and perhaps female reproductive tracts; (2) are used to anchor the spermatozoa inside the female spermatheca in order to avoid mechanical displacement by the genitalia of other males and (3) may play a role in oocyte recognition. We propose that the evolution of aflagellarity in Opiliones is related to the unique morphology of the female reproductive tract. Since eggs are fertilized on the tip of the ovipositor just prior to being laid, there is no advantage favoring sperm mobility. Additionally, female sperm receptacles are small and males that produced small spermatozoa would have a higher chance of fertilizing more eggs.     

Evidence for a proteinase inhibitor binding component associated with murine spermatozoa

The supernatants of frozen-thawed murine epididymal sperm suspensions contain a heat-labile component capable of binding a low molecular weight, acid-stable proteinase inhibitor of seminal vesicle origin. The substance has a molecular weight of approximately 15,000 and can be isolated by affinity chromatography using the inhibitor as the ligand. Although the substance has no inherent enzymatic properties, it will decrease the activity of the seminal inhibitor in the standard N-benzoyl-DL-arginine-p-nitroanilide (BAPNA) assay. Enzyme-linked immunosorbent assays (ELISA) indicate that the substance, when bound to microtiter plates, is capable of binding the seminal vesicle inhibitor. Turkey egg white trypsin inhibitor will decrease the amount of the seminal inhibitor that will bind to the substance, while noninhibitor proteins, e.g., bovine serum albumin or insulin, have no effect. Turkey egg white and lima bean trypsin inhibitor will also decrease the amount of seminal vesicle inhibitor capable of binding to washed sperm. These data indicate the presence of an inhibitor acceptor site associated with murine epididymal spermatozoa.     

Ejaculated and epididymal mouse spermatozoa are different in their susceptibility to nuclease-dependent DNA damage and in their nuclease activity

Ejaculated mouse sperm retrieved from the uteri are more susceptible to DNA damage during freeze-drying and freezing without cryoprotection than epididymal sperm. This prompted us to speculate that a factor present in the uterus after mating, either male or female derived, was responsible for increased susceptibility of ejaculated sperm to DNA damage during preservation and that the differences between epididymal and ejaculated mouse sperm in response to stress originated from varying nuclease activity. We first exposed epididymal sperm to the uterine content from females mated to vasectomized males (UCSP), to the uterine content from unmated females in estrus (UC), and to the seminal vesicle fluid (SVF) and examined sperm chromosomes after intracytoplasmic sperm injection (ICSI). We found an increased incidence of chromosome breaks and extremely severe DNA breakage after exposure to UCSP and SVF, respectively, but the chromosomes were normal in sperm exposed to UC. Comet assay results verified that DNA damage after exposure to SVF was present in sperm before fertilization. Next, we examined nuclease activity in sperm and their associated components with a plasmid digestion assay. Nuclease activity was detected in isolated epididymal and ejaculated sperm, as well as in epididymal fluid and seminal plasma, and was much more pronounced in all samples originating from ejaculate. The combined results from the present study imply that there are intrinsic differences between the epididymal and ejaculated mouse sperm preparations in their susceptibility to nuclease-dependent DNA damage that originates from their nuclease activity. This nuclease activity was detected both in the sperm-free fraction of preparations and isolated sperm.     

Impact of feeding ethanolic extract of root bark of Cananga odorata (Lam) on reproductive functions in male rats

The 50% ethanolic extract of the root bark of C. odorata administered orally at the dose of 1g/kg body weight/day for 60 days resulted in decreased epididymal sperm motility and sperm count in male albino rats. Morphological abnormalities were also observed in the sperms. The testicular glycogen, the activities of 3beta hydroxy steroid dehydrogenase, glucose 6-phosphate dehydrogenase, malic enzyme, sorbitol dehydrogenase in seminal vesicle, fructose in seminal plasma and serum testosterone were significantly decreased in treated group. While testicular cholesterol level, the concentration of the fecal bile acids, urinary excretion of 17 ketosteroids, the activities of 17beta hydroxy steroid dehydrogenase, epididymal lactate dehydrogenase and that of testicular HMG CoA reductase were increased in treated group when compared to control. The results suggest that the ethanolic extract of C. odorata possesses the spermatotoxic effects in male albino rats.     

Characterization of acid and neutral alpha-mannosidases in bull semen and reproductive organs

Acid and neutral alpha-mannosidase activities were studied in the bull reproductive tissues, isolated spermatozoa, epididymal and seminal vesicle secretion and seminal plasma. The acid enzyme in the seminal plasma mainly derived from the epididymal secretion, while the neutral one was enriched in the sperm cells. The latter activity in the seminal plasma appears to be due to an enzyme released from the cytoplasmic droplets in the epididymis. The acid enzyme had a molecular weight of 220,000-320,000, pI 7.3-6.0 and an optimum at pH 4.0. It was sensitive to swainsonine but was stimulated by Zn2+. The neutral enzyme had a molecular weight of 360,000-460,000, pI 5.4-4.7 and showed double optima at pH 5.5 and 6.0-7.0. It was resistant to swainsonine but was markedly activated by Co2+ or Fe2+. The neutral enzyme was also more sensitive to thermal inactivation than the acid one.     

Slimmer or Fertile? Pharmacological Mechanisms Involved in Reduced Sperm Quality and Fertility in Rats Exposed to the Anorexigen Sibutramine

Sperm acquire motility and fertility capacity during epididymal transit, under the control of androgens and sympathetic innervations. It is already known that the acceleration of epididymal sperm transit time can lead to lower sperm quality. In a previous work we showed that rats exposed to the anorexigen sibutramine, a non-selective serotonin-norepinephrine reuptake inhibitor, presented faster sperm transit time, lower epididymal sperm reserves and potentiation of the tension of epididymal duct to norepinephrine exposed acutely in vitro to sibutramine. In the present work we aimed to further investigate pharmacological mechanisms involved in these alterations and the impact on rat sperm quality. For this, adult male Wistar rats were treated with sibutramine (10 mg/kg/day) or vehicle for 30 days. Sibutramine decreased final body, seminal vesicle, ventral prostate and epididymal weights, as well as sperm transit time in the epididymal cauda. On the contrary of the in vitro pharmacological assays, in which sibutramine was added directly to the bath containing strips of distal epididymal cauda, the ductal tension was not altered after in vivo sub-chronic exposure to sibutramine. However, there is pharmacological evidence that the endogenous epididymal norepinephrine reserves were reduced in these animals. It was also shown that the decrease in prostate weight can be related to increased tension developed of the gland, due to sibutramine sympathomimetic effects. In addition, our results showed reduced sperm quality after in utero artificial insemination, a more sensitive procedure to assess fertility in rodents. The epididymal norepinephrine depletion exerted by sibutramine, associated with decreases in sperm transit time, quantity and quality, leading to reduced fertility in this experimental model, reinforces the concerns about the possible impact on fertility of man taking sibutramine as well as other non-selective serotonin-norepinephrine reuptake inhibitors, especially considering the lower reproductive efficiency of humans compared to males of other species.     

Effects of hexavalent chromium on reproductive functions of male adult rats

Hexavalent chromium is an environmental contaminant which may be associated with reproductive abnormalities in male rats. In the present study, we examined the effect of hexavalent chromium on male reproductive function of rats. Male Wistar rats received a daily intraperitoneal injection of potassium dichromate (1 or 2 mg/kg body weight) for fifteen consecutive days. A decrease in testis weight and an increase in seminal vesicles and prostate weights were demonstrated after chromium treatment. Moreover, a dose-dependent increase in blood and testis chromium levels as well as an increase in FSH and a decrease in LH and testosterone serum levels were detected in treated rats. Histological analysis revealed pronounced morphological alterations with enlarged intracellular spaces, tissue loosening and dramatic loss of gametes in the lumen of the seminiferous tubules of treated rats. In addition, a decreased sperm motility and number of epididymal spermatozoa together with an increased sperm abnormality rate was found in chromium-treated rats in comparison to controls. In rats receiving the higher chromium dose, histological images presented considerably increased areas filled with seminal vesicle and prostate secretions. The mucosal crypts of seminal vesicles and the typical invaginations of prostate were altered. The results suggest that subacute treatment of potassium dichromate promotes reproductive system toxicity and affects testicular function of adult male rats.     

Heterogenous turnover of sperm and seminal vesicle proteins in the mouse revealed by dynamic metabolic labeling

Plasticity in ejaculate composition is predicted as an adaptive response to the evolutionary selective pressure of sperm competition. However, to respond rapidly to local competitive conditions requires dynamic modulation in the production of functionally relevant ejaculate proteins. Here we combine metabolic labeling of proteins with proteomics to explore the opportunity for such modulation within mammalian ejaculates. We assessed the rate at which proteins are synthesized and incorporated in the seminal vesicles of male house mice (Mus musculus domesticus), where major seminal fluid proteins with potential roles in sperm competition are produced. We compared rates of protein turnover in the seminal vesicle with those during spermatogenesis, the timing of which is well known in mice. The subjects were fed a diet containing deuterated valine ([(2)H(8)]valine) for up to 35 days, and the incorporation of dietary-labeled amino acid into seminal vesicle- or sperm-specific proteins was assessed by liquid chromatography-mass spectrometry of samples recovered from the seminal vesicle lumen and cauda epididymis, respectively. Analyses of epididymal contents were consistent with the known duration of spermatogenesis and sperm maturation in this species and in addition revealed evidence for a subset of epididymal proteins subject to rapid turnover. For seminal vesicle proteins, incorporation of the stable isotope was evident from day 2 of labeling, reaching a plateau of labeling by day 24. Hence, even in the absence of copulation, the seminal vesicle proteins and certain epididymal proteins demonstrate considerable turnover, a response that is consonant with the capacity to rapidly modulate protein production. These techniques can now be used to assess the extent of phenotypic plasticity in mammalian ejaculate production and allocation according to social and environmental cues of sperm competition.     

Semen-coagulating protein, SVS2, in mouse seminal plasma controls sperm fertility

Mammalian seminal plasma is known to contain a decapacitation factor(s) that prevents capacitation and thus, the fertility of sperm. This phenomenon has been observed in experiments conducted in vitro that assessed the inhibition of epididymal sperm fertility by seminal plasma or by the purified decapacitation factor. However, the phenomenon of decapacitation has not yet been characterized in vivo. In the present study, we demonstrate that seminal vesicle protein secretion 2 (SVS2), which is a 40-kDa basic protein and a major component of the copulatory plug, enters the uterus and interacts with ejaculated sperm heads after copulation. The SVS2-binding region of sperm changed from the postacrosomal region to the equatorial segment, while the sperm migrated through the uterus and finally disappeared in the oviduct. Furthermore, SVS2 reduced the fertility of epididymal sperm. The sperm treated with SVS2 decreased the percentage of fertilized oocytes from 60% to 10%. The capacitation state was assessed by protein tyrosine phosphorylation and the comprehensiveness of the acrosome reaction. SVS2 functioned to maintain sperm in the uncapacitated state and to reverse capacitated sperm to the uncapacitated state. We found that the fertility of ejaculated sperm is associated with SVS2 distribution in the female reproductive tract. These results indicate that SVS2 functions as a decapacitation factor for mouse sperm.     

Effects of age and repeated mating on male sperm supply and paternity in a parasitoid wasp

Post-copulatory paternity biases after female multiple mating are major constraints on both male and female reproductive systems. The outcome of paternity in certain situations is only controlled directly by male sperm stock. This was tested experimentally in the parasitoid wasp Anisopteromalus calandrae (Howard) (Hymenoptera: Pteromalidae), in which sperm stocks are small (several hundred) and the fertilizing efficiency of stored sperm is high (the ratio of sperm stored/fertilized eggs is about 0.75). Sperm in seminal vesicles and paternity of males of different status (virgin young, virgin old, or young previously mated) were measured after female single and double mating. The amount of sperm in the seminal vesicle differed according to male status (increasing from previously mated males to old males), but there was no difference in sperm stored by females after a single mating. In double mating experiments with two males of different status, paternity increased linearly with the relative amount of sperm in seminal vesicles. Paternity distribution conforms to 'a fair raffle' of sperm from both donors following complete mixing of sperm prior to fertilization. Thus, in a female multiple mating context, male fitness depends principally on their sperm stock, which in turn depends on life history parameters, such as age and previous mating.     

Effect of castration on epididymal sperm storage in male musk shrews (Suncus murinus) and mice (Mus musculus)

Reproductively mature male musk shrews and mice were bilaterally castrated. Epididymal sperm numbers and motility were assessed 0, 2, 4 and 6 weeks after surgery. Seminal vesicle weights and plasma concentrations of total androgens were also measured. In male musk shrews, 30% of the original epididymal sperm numbers were still present 2 weeks after castration and motile spermatozoa were present in 2 of 7 individuals. By 4 and 6 weeks after castration the numbers of spermatozoa remaining declined to about 10% and no sperm motility was noted. Seminal vesicle weights were maintained at about 30% of their original size even up to 6 weeks after castration. In male mice, epididymal sperm numbers, seminal vesicle weights, and androgen levels declined more dramatically after castration. Although androgen concentrations in gonadally intact male musk shrews were approximately 50% of the values in male mice, after castration the concentrations in musk shrews were approximately 2-fold higher than in mice at all times. The results suggest that post-castration retention of epididymal sperm and seminal vesicle weights in the male musk shrew as compared with male mice, is facilitated either by a relatively greater adrenal contribution to circulating androgen levels and/or greater target tissue sensitivity.     

Phosphorylation of sperm-binding proteins from seminal vesicle secretion by sperm surface protein kinase

Rat seminal vesicle secretion does not demonstrate protein kinase activity, either towards endogenous or exogenous proteins. When epididymal sperm were incubated in vitro with seminal vesicle secretion, three prominent secretory proteins were bound to the sperm. Two of these proteins were highly phosphorylated. Thus, selected sperm-binding proteins from accessory gland secretions are phosphorylated by sperm surface protein kinases.     

SERPINE2, a serine protease inhibitor extensively expressed in adult male mouse reproductive tissues, may serve as a murine sperm decapacitation factor

SERPINE2, one of the potent serine protease inhibitors that modulates the activity of plasminogen activator and thrombin, is implicated in many biological processes. In the present study, we purified SERPINE2 from mouse seminal vesicle secretion (SVS), using liquid chromatography and identified it by liquid chromatography/tandem mass spectrometry, and it showed potent inhibitory activity against the urokinase-type plasminogen activator. SERPINE2 was expressed predominantly in seminal vesicles among murine male reproductive tissues. It was immunolocalized to the SVS and mucosal epithelium of the seminal vesicle, epididymis, coagulating gland, and vas deferens. In the testes, SERPINE2 was immunostained in spermatogonia, spermatocytes, spermatids, Leydig cells, and spermatozoa. SERPINE2 was also detected on the acrosomal cap of testicular and epididymal sperm and was suggested to be an intrinsic sperm surface protein. The purified SERPINE2 protein could bind to epididymal sperm. A prominent amount of SERPINE2 was detected on ejaculated and oviductal spermatozoa. Nevertheless, SERPINE2 was detected predominantly on uncapacitated sperm, indicating that SERPINE2 is lost before initiation of the capacitation process. Moreover, SERPINE2 could inhibit in vitro bovine serum albumin-induced sperm capacitation and prevent sperm binding to the egg, thus blocking fertilization. It acts through preventing cholesterol efflux, one of the initiation events of capacitation, from the sperm. These findings suggest that the SERPINE2 protein may play a role as a sperm decapacitation factor.     

A time-course study of chronic paternal cyclophosphamide treatment in rats: effects on pregnancy outcome and the male reproductive and hematologic systems

We have found previously that daily treatment of male rats for 11 wk with low doses of the anticancer drug cyclophosphamide had no apparent effect on male reproductive organ weights, epididymal sperm counts, or serum hormones at the end of the treatment period; yet, upon breeding to untreated females, these males produced a high rate of post-implantation loss and fetal anomalies. The present study was designed to investigate the time course and dose response of the effects of chronic cyclophosphamide treatment on the male reproductive and hematologic systems. Male Sprague-Dawley rats were gavage-fed for 1, 3, 6 and 9 wk with saline (control), or 5.1 (low dose) or 6.8 (high dose) mg/kg/day of cyclophosphamide. After each of the treatment periods, males were mated to determine the effect on pregnancy outcome, then killed, and the effects on the male reproductive and hematologic systems were assessed. After 6 wk of treatment, a sharp increase in mortality was found between the 5.1 and 6.8 mg/kg/day doses of cyclophosphamide. The high dose of cyclophosphamide induced higher levels of pre- and post-implantation loss but fewer fetal anomalies than did the low dose. The low dose of cyclophosphamide did not affect reproductive organ weights; in contrast, the high dose caused decreases in epididymal, ventral prostate, and seminal vesicle weights after 3, 6, and 9 wk. Testicular and epididymal sperm counts were decreased in a dose-dependent manner after 3 wk; in addition, the high dose led to a decrease in epididymal sperm counts after 6 wk of treatment. Another rapidly proliferative tissue, the bone marrow, was dramatically affected by both doses of cyclophosphamide at all time points, with leukocyte counts decreasing to 40% of control by 1 wk. After 9 wk of treatment, effects on the male reproductive system were less marked, compared to earlier time points, whereas those on the hematologic system and pregnancy outcome persisted. Thus chronic low-dose treatment of male rats with cyclophosphamide not only had early and striking effects on the bone marrow and the pregnancy outcome but also affected the male reproductive system in a clear time- and dose-dependent manner.     

Effects of seminal vesicle removal on fertility and uterine sperm motility in the house mouse

The relative significance of the accessory glands of the male reproductive tract in fertility is unclear. To clarify the role of the seminal vesicles, fertility and uterine sperm motility were determined before and after removal of seminal vesicles in the house mouse. After removal of seminal vesicles, the pregnancy rate (number of females pregnant/number of females X 100) was reduced and the time to birth was increased, while the average litter size was not changed. Fertilization, determined by examining the oocytes 30 h after mating, was highly variable after matings with males whose seminal vesicles were removed; in some cases none of the oocytes were fertilized. The motility of sperm recovered from the uterus 1 h after matings with males before and after seminal vesicle removal and sham operations was analyzed using a videomicrographic system. The motility of uterine sperm was less progressive with more lateral displacement of the head about the trajectory and a less linear trajectory after removal of the seminal vesicles. Sham-operated animals showed no consistent changes in motility of uterine sperm. The changes in sperm motility could contribute to the reduction in fertilization since sperm motility is necessary for transport in the female reproductive tract and interaction with the oocytes.