Fertilization by sperm injection in the rabbit

Whole rabbit spermatozoa and isolated sperm nuclei were microinjected directly into the ooplasm of hamster and rabbit ova. These injected sperm decondensed and formed male pronuclei during subsequent in-vitro culture. Injection of whole spermatozoa and sperm nuclei prepared by a protocol known to allow in-vitro capacitation of ejaculated spermatozoa yielded a significantly higher (P < 0.01) number of activated rabbit ova containing male pronuclei than did injection of uncapacitated epididymal sperm nuclei or ejaculated sperm nuclei. Rabbit ova fertilized by sperm injection were capable of undergoing normal-appearing cleavage division during 22 h of culture.     

Can spermatozoa with abnormal heads gain access to the ovum in artificially inseminated super- and single-ovulating cattle?

The collective efficiency of barriers in the female tract against spermatozoa with abnormal heads was studied. In Experiment 1, Day 6 ova/embryos were recovered nonsurgically from superovulated (n = 24) and single-ovulating (n = 44) cows following artificial insemination with semen of bulls selected for normal spermatozoal motility (> or = 50%) and high content (> 30%) of spermatozoa with misshapen heads, random nuclear vacuoles or the diadem defect. To assess characteristics of spermatozoa capable of traversing barriers in the female tract, accessory spermatozoa were classified morphologically (x 1250) and compared with those of the inseminate. Superovulated cows proved inadequate for assessment of accessory spermatozoa due to evidence of poor sperm retention in the zona pellucida; thus, only single-ovulating cows were used. Accessory spermatozoa (n = 479) from 31 ova/embryos recovered from 44 cows were more normal in head shape than those in the inseminate (76 vs 62%; P < 0.05). Spermatozoa with normal head shape, but with nuclear vacuoles appeared as accessory spermatozoa at the same frequency as they were found in the inseminate (20 vs 17%, respectively). Only sperm cells with subtly misshapen heads appeared as accessory spermatozoa. In Experiment 2, semen pooled from 4 bulls having large numbers of spermatozoa exhibiting a gradation from severely asymmetrically misshapen heads to subtly misshapen heads was evaluated. Again, the accessory sperm population (960 sperm cells recovered from 64 ova/embryos) was enriched with spermatozoa of normal head shape relative to the inseminate (53 vs 26%, respectively; P < 0.05). Sperm cells with only nuclear vacuoles and those with subtly misshapen heads were not different between the accessory and inseminate populations (11 vs 8%, and 20 vs 25%, respectively). We conclude that morphologically abnormal spermatozoa are excluded from the accessory sperm population based upon severity of head shape distortion.     

Effects on fertilization of rabbits of insemination with ejaculates treated with PG-dehydrogenase and antisera to PGE-2 and PGF-2 alpha

PGE-2 and PGF-2 alpha in rabbit semen were selectively inactivated by incubation with antisera, or most of the seminal prostaglandins were transformed into biologically inactive 15-keto-prostaglandins by prostaglandin-15-hydroxydehydrogenase(PG-15-HDH). These treated ejaculates were vaginally inseminated. Compared to the controls (738 eggs of which 94% were fertilized) a dose-dependent reduction of the fertilization rate was observed with the anti-PGF-2 alpha-treated ejaculates. A non-uniformly, but statistically significantly reduced fertility was found in the other 2 treatment groups. After incubation with higher doses of PG-15-HDH, some fertilization was accomplished with ejaculates showing an extremely weak forward progression or immotile spermatozoa. An improvement in sperm motility, however, was observed in ejaculates treated with antiserum to PGF-2 alpha. Seminal prostaglandins may not exclusively affect sperm motility. The observed influences on the fertilization rate after treatment of spermatozoa with antisera to PGE-2 and PGF-2 alpha or PG-15-HDH suggest that these are local effects in the female genital tract.     

Composition lipidique des spermatozoides humains et susceptibilité au stress oxydant avant et après migration dans le mucus cervical

Spermatozoa are particularly susceptible to damage induced by ROS, especially as their plasma membrane contains large amounts of polyunsaturated fatty acids. Mammalian sperm cells develop the capacity to fertilise ova during transport in the male and female reproductive tracts. The nature and quality of the micro-environment of the female reproductive tract are important factors for sperm selection, capacitation and subsequent acrosome reaction.In vitro experiments using capacitating media have shown remodeling of the lipid composition of the sperm membrane during these steps and the same approaches have also shown that a low level of ROS was necessary. The oxidative status of the female genital tract is therefore certainly of primary importance for the physiological maturation of the sperm cell. It has been previously reported that an inappropriate oxidative balance in the male genital tract (ie, an excessive ROS production overwhelming all antioxidant strategies) impairs the structure and several functions of sperm cells. This phenomenon may arise in the female genital tract, but has never been investigated. The present paper is a review of the literature on these subjects and also reports our results concerning the changes in semen lipid content during cervical mucus migration and the effect of cervical mucus polymorphonuclear (PMN) cells on sperm characteristics. We showed that the sperm levels of vitamin E, cholesterol, phospholipids, sphingomyelin and plasmalogen assessed by HPLC decreased after migration through cervical mucus. These modifications were observed in parallel with lipid enrichment of the cervical mucus, suggesting an efflux of cholesterol and lipids from sperm cells. The spermatozoa recovered postmigration in the cervical mucus were characterised by low levels of the various lipid classes. Spermatozoa that migrated in cervical mucus samples with a considerable quantity of polymorphonuclear leukocytes (PMN) also showed significantly increased levels of sphingomyelin, diacyl phospholipids and plasmalogens in comparison to spermatozoa that migrated in cervical mucus devoid of PMN. Finally, we also found that PMA-induced ROS production was significantly increased for spermatozoa treated with cervical mucus containing PMN.     

Efficacy of TEPA as a “sperm label” for competitive fertilization studies in the rabbit

In this paper, the conditions necessary to use TEPA [tris (1-aziridinyl)] effectively as a label for spermatozoa in competitive fertilization are established. The fertilizing ability of rabbit spermatozoa treated with 0 and 0.8 mg TEPA/ml was compared at insemination doses of 1, 5, 20, and 40 × 10⁶ spermatozoa. Fertility was assessed by collecting ova from 64 does 48 to 52 h after insemination. TEPA blocked all but 4% of the ova from developing when 1 × 10⁶ spermatozoa were inseminated, but fertility was reduced. When 5 × 10⁶ spermatozoa were inseminated following treatment with 0, 0.6 or 1.2 mg of TEPA/ml, the fertility was 83, 74 and 50% (P<0.05), and the percentage of ova containing more than four blastomeres was 83, 11 and 5% (P<0.05), respectively. The 0.6% TEPA level was selected for a competitive fertilization trial. Equal numbers of sperm from pure Dutch-color and albino sires were combined so that either both types were untreated, only the ‘albino’ semen was treated, only the ‘Dutch’ semen was treated, or both were treated. Does were inseminated with 5 × 10⁶ sperm and allowed to kindle. The litter sizes were 5.6, 3.1, 2.7, and 0 young, and the proportion of Dutch-color progeny was 63, 97, 0 and 0%, respectively, confirming the effectiveness of TEPA as a “label”. Only one of 60 young born resulted from fertilization by a TEPA-treated spermatozoon, demonstrating that few embryos fully escape the TEPA block. Thus, the TEPA concentration and sperm numbers were established to use TEPA effectively as a label for spermatozoa in competitive fertilization studies.     

Embryo quality and andrological study of two subfertile bulls versus five control bulls with normal fertility

Andrological studies and embryo morphology evaluation of superovulated cows were performed on 2 randomly selected subfertile dairy bulls whose semen was used for artificial insemination and on 5 control bulls with normal fertility. Neither sperm motility studies, nor sperm morphology or testicular measurements differed between the subfertile and the control bulls. Altogether 315 ova were recovered from 41 superovulated cows inseminated with semen collected from either the subfertile or the normal control bulls. The spermatozoa of one of the 2 subfertile bulls was shown to have a decreased ability to fertilize superovulated ova, while the other subfertile animal, the bull with the lowest noreturn rate, was found by chromosome analysis to have a reciprocal translocation (60, XY, rcp 20:24), causing embryonic death. We suggest that subfertile bulls should not be used in commercial embryo transfer programs nor in artificial insemination and that andrological studies on subfertile bulls with good sperm motility should include evaluation of 6- to 7-day-old ova from superovulated cows to determine if the fertilization rate is normal or impaired. A chromosome analysis should also be performed when a subjertile bull has a normal fertilization rate of ova.     

The antigenicity of the rabbit sperm acrosomes and the reversible sterility induced in the immunized rabbits

Rabbits were immunized against their alloantigens extracted from the sperm acrosome. The sequential method of extraction, first by MgCl₂ then by detergents, provided the plasma and outer acrosomal membranes and their contents in the first step and the inner acrosomal membrane and its components in the next step. Both groups of antigens appear to have produced monospecific antisera. The MgCl₂ extract antiserum lost its sperm immobilization and agglutination activities on absorption to its antigen. The detergent extract, under similar treatment, remained unaffected. Therefore, the agglutination and immobilization factors resided on the plasma membrane. The decrease in sperm concentration suggested that the immunogen affected the germinal epithelium. The reduction in the semen volume and seminal enzymes seems to indicate that the accessory sex glands were also affected by the treatment. The antiserum blocked the dissolution of the hamster and rabbit zona pellucida by the MgCl₂ extract. The normal serum had no such effect. Both the normal and immune sera inhibited acrosin equally well while the antisera inhibited other proteinases, arylsulfatase, and β-N-acetylhexosaminidase to a significantly greater extent. Immunized rabbits failed to impregnate females as long as the antibody titer remained high. As the titer declined these animals became fertile and the females delivered normal litters.     

Motility and fertility of alginate encapsulated boar spermatozoa

Ejaculated boar spermatozoa are vulnerable to cold shock. Prolonged storage of boar spermatozoa at low temperatures reduces survival rate, resulting in a bottleneck for the extension of artificial insemination in pig husbandry. This study evaluated whether alginate microencapsulization processing can improve the longevity of boar spermatozoa stored at 5 degrees C and the fertility of microencapsulated spermatozoa in vivo. Sperm-rich fraction semen from three purebred boars were concentrated and microencapsulated using alginate at 16-18 degrees C, and then were stored at 5 degrees C. Following storage for 1, 3 and 7 days, the microcapsule was taken out to assess sperm release under 37 degrees C incubation with or without 110 rpm stirring. The percentage of sperm released from microcapsules with 110 rpm stirring was higher than without stirring (81 versus 60%) after 24h of incubation. In another experiment, semen was also microencapsulated to evaluate the sperm motility. The motility of spermatozoa was assessed at 10 min, 8, 24, 32, 48, 56 and 72 h following incubation at 37 degrees C for nine consecutive days. The fertility of the free and microencapsulated semen was assessed by inseminating sows, and the reproductive traits (conception rate, farrowing rate, and litter size) were recorded. The motility of encapsulated spermatozoa was significantly higher than that of free semen after 8h incubation at 37 degrees C after storing for over three days (P<0.05). No significant difference existed in conception rate, farrowing rate, and litter size between the microencapsulated and non-encapsulated semen after four days of storage. In conclusion, microencapsulation can increase the longevity of boar spermatozoa and may sustain in vivo ova fertilization ability.     

Quantitative characteristics of Atlantic halibut, Hippoglossus hippoglossus L., semen throughout the reproductive season

The overall objective of the study was to investigate changes in quantitative parameters of Atlantic halibut (Hippoglossus hippoglossus L.) semen throughout the reproductive season in order to systematize the knowledge about biology of Atlantic halibut spermatozoa. Semen samples were collected from February to May from broodstock males kept under either a natural or 3-month advanced photoperiod regime. Spermatozoa concentration, semen pH and osmolality, as well as spermatozoa motility parameters were investigated. The use of catheterization of sperm was examined. Also, fertilization tests were performed. We found that spermatozoa concentration increases in a linear-like mode towards the end of the spawning season, which correlated with a decrease in a number of spermatozoa motility parameters, including actual percentage of motile spermatozoa (MOT), curvilinear velocity (VCL) and straight-line velocity (VSL) of spermatozoa. A breakpoint in MOT occurred when spermatozoa reached a concentration in the range of 17-20 x 10(9) spermatozoa/mL. The fertilization ability of sperm from males kept under natural photoperiod decreased in April. Survival of embryos at 80 degrees days produced by fertilizing eggs of single female with sperm from natural photoperiod males was 88, 76 and 41% on April 09 and 17, and May 01, respectively, whereas using sperm from 3-month delayed photoperiod males for fertilizing eggs from the same female on April 27 resulted in 80% of surviving embryos, not differing significantly from the data from April 09. Physical decomposition of spermatozoa was observed towards the end of the season and it was related to an increase in the whole semen osmolality. Catheterization of semen did not improve spermatozoa motility parameters, however, it reduced the variation in recorded values, especially in the case of pH, caused by contamination with feces or urine. Post-seasonal decrease in spermatozoa concentration was likely related to intensive ageing processes. Based on the present study and available data by other researchers, a model of changes of quantitative parameters in Atlantic halibut semen throughout the reproductive season is proposed.     

The expression of the aryl hydrocarbon receptor in reproductive and neuroendocrine tissues during the estrous cycle in the pig

The sprouted wheat (SW) contains the 6-methoxy-2-benzoxazolinone (6-MBOA), a phenol compound that stimulates reproduction in certain small wild herbivorous mammals. The objective of the present study was to evaluate the effect of short-term supplemental dietary SW on libido, semen and sperm characteristics of rabbit bucks. Five-month old New Zealand White pubertal rabbits (n=18) were randomly allocated to one of two treatments: supplementation or not (control) supplemented with SW. The experimental design was completely random with nine replications, experimental unit was one buck. Semen collection for each male was conducted once a week with two ejaculations during 20 weeks. The SW was given during four consecutive days prior to each semen collection. Analysis of variance was under a mixed model: treatment, ejaculate number and season were fixed and rabbit random effects. There was no effect of treatment (P>0.05) on reaction time, gel presence, volume, pH, sperm motility, sperm number per ml and sperm number per ejaculate. The percentage of normal alive spermatozoa was 13.5% greater in SW-supplemented bucks than in the control and the percentage of abnormal alive spermatozoa was 44.1% greater in the control than in the SW-supplemented bucks. The morphology of dead spermatozoa, integrity of acrosome, number of normal alive motile sperm and semen doses per ejaculate were not influenced (P>0.05) by SW supplementation. The proportion of presence of gel and semen volume in the first ejaculate was greater than the second ejaculate (+140% and +56.4%). However, the semen quality in the latter was greater (P=0.0001) than the former in terms of an increase in motility (+29.7%). Reproductive traits were more desirable (P<0.05) in winter than autumn. Dietary wilted SW as a source of biological 6-MBOA enhanced sperm characteristics in terms of a greater percentage of normal alive and lesser percentage of abnormal alive spermatozoa but did not affect the number of normal motile live sperm and suitable semen doses in rabbit bucks in autumn and winter.     

Entry of immotile spermatozoa into zona-free hamster ova

We studied six men whose spermatozoa were immotile and possessed a variety of sperm tail structural abnormalities by electron microscopy. The semen of all six subjects had a normal percentage of oval forms and sperm undergoing capacitation and acrosome reaction. Despite the absence of motility, when incubated sperm from these subjects was added to a microdrop of medium containing zona pellucida-free hamster ova, sperm penetration or entry into the cytoplasm of from 1–9% of the eggs was evident with phase contrast microscopy. This latter finding suggests that, at least in this system, oocytes actively facilitate sperm incorporation. Penetration was absent when sperm of fertile men were rendered immotile, though still viable, by heat treatment.     

平疣桑椹石磺生殖系统结构及精子储存场所

雌雄同体贝类精子的储存和利用规律一直是国内外贝类生物学研究的难点之一,本文利用活体解剖、显微观察、组织切片和扫描电镜技术,综合研究了平疣桑椹石磺(Platevindex mortoni)的生殖系统及精子储存场所。结果显示,其生殖系统包括生殖器本部、雌性生殖部分和雄性生殖部分。生殖器本部由两性腺、两性输送管、蛋白腺、黏液腺、支囊组成;雌性生殖部分包括输卵管、受精囊、阴道,位于身体中后方体腔内;雄性生殖部分包括输精管、刺激器、阴茎、阴茎鞘和阴茎牵引肌,位于身体前端右侧体腔内;其阴茎有阴茎鞘,阴茎表面布满倒刺。平疣桑椹石磺阴茎为直线状,无雄性附属腺。未交配的性成熟个体支囊内充满细长精子,受精囊内无精子;而交配后充当雌性个体的支囊内均为细长的自体精子,受精囊内有大量活力较强的粗短精子,其支囊为自体精子的存储场所,而受精囊为异体精子的存储场所。其精子储运情况为:两性腺内精子成熟后暂存于支囊,交配时通过输精管运输至阴茎,由阴茎输送精子至对方的阴道,异体精子进入受精囊内存储待用。     

In vitro Capacitation of Ejaculated Rabbit Spermatozoa

Prior to fertilization mammalian spermatozoa undergo physiological changes in the female reproductive tract. These changes are collectively known as capacitation. In essence capacitation is a further differentiation that the sperm cell must acquire beyond the maturational changes that it undergoes in the epididymis. Although capacitation in vitro has been easily achieved in rodents, its accomplishment in the rabbit is inconsistent and difficult. We report here successful in vitro capacitation of ejaculated rabbit sperm, assessed by in vitro and in vivo fertilization of rabbit ova. Sperm were used from pooled ejaculates collected from bucks of proven fertility since sperm collected from individual bucks resulted in significant differences in fertilization levels. Conditions favoring in vitro capacitation were: (1) extended incubation time of 12 h, (2) addition of 20% heated rabbit serum to incubation medium, and (3) an atmosphere of 5% CO2, 8% O2, and 87% N2 during incubation. In vitro capacitation of sperm under these conditions resulted in 67% fertilization compared to 89% for control sperm capacitated for 13 h in the uterus.     

Use of zona-free hamster ova to assess sperm fertilizing ability of bull and stallion

Zona-free hamster ova interacted with bull and stallion spermatozoa after treatment of ejaculated semen to capacitate the sperm cells. Sperm conditioning by prolonged incubation in BWW medium (18–26 hr) prior to insemination was effective for capacitation of bull and stallion sperm. Preincubation of bull sperm for 70 or 105 min in defined medium (DM) with NaCl content elevated to result in 350 m0sM/kg also led to penetration of hamster vitelli. More rapid sperm conditioning was possible and higher proportions of interacting vitelli followed insemination with bull or stallion sperm exposed to high ionic strength DM (380–390 m0sM/kg) for 10 min before incubation in isotonic DM prior to insemination, the treatment adopted in subsequent work. Initial efforts to assess relative fertilizing ability of freshly ejaculated semen from two fertile bulls (A and B) in A1 usage resulted in uniformly high ( > 90%) levels of sperm-vitelli interaction (for both) when the hamster ova employed resulted from superovulation with PMSG and HCG. Following use of ova from untreated hamsters sperm samples of bull A and bull B interacted with 53.8% and 84.9% (P < 0.05) of zona-free hamster ova, respectively. Conception data (60–90 day nonreturn rates) resulting from A1 with semen collected during the same interval but processed and stored in liquid nitrogen prior to use revealed an inverse relationship to proportions of vitelli interacting with fresh sperm; nonreturn rates were 69.3% and 66.3% for bull A and bull B, respectively. A similar treatment effected capacitation of frozen-stored bull semen to enable sperm-vitelli interaction. These findings encourage additional efforts to correlate testing of processed semen with fertility.     

In vitro fertility evaluation of cryopreserved ram semen and its correlation with relative in vivo fertility

The present study was designed to evaluate zona-free hamster ova assay conditions for cryopreserved ram semen and to investigate the correlation between ability to penetrate zona-free hamster ova and in vivo fertility. In vivo fertility was estimated for cryopreserved semen from 5 Merino rams using heterospermic insemination. Equal numbers of postthaw motile spermatozoa from a Merino ejaculate and pooled Suffolk ejaculates were mixed prior to insemination. Each Merino ejaculate was paired with the same pool of cryopreserved Suffolk semen. Relative in vivo fertility for each Merino ram was calculated as the proportion of offspring that were sired by the Merino (range 42 to 100%). These ejaculates also differed in their ability to penetrate zona-free-hamster ova (3.6 to 9.0 penetrated spermatozoa per ovum). Differences in penetration rate were correlated with in vivo fertility (P < 0.002, R2 = 0.69). Results of these studies suggest that the zona-free hamster ova bioassay may be a useful test in the assessment of cryopreserved ram sperm fertility.     

Fertilizing potential in vitro of semen from young beef bulls containing a high or low percentage of sperm with a proximal droplet

Fertilizing potential of semen containing a high percentage of sperm with a proximal droplet was evaluated using IVF. Design criteria: (a) specified semen with >100 x 10(6) sperm/mL with >40% progressively motile spermatozoa, after collection via electro-stimulation; (b) designated a droplet group, bulls whose semen contained >30% spermatozoa with a proximal droplet and <25% with other morphological abnormalities, and a control group, with <25% abnormalities of any type; and (c) stipulated evaluations at 11 to 13 mo of age and again -4 wk later. At the initial evaluation, when a bull was assigned to the droplet group, the next bull meeting control criteria was designated his pair; 15 pairs in four herds were studied. Semen was extended in egg-yolk citrate, cooled to 5 degrees C over approximately 2.5 h, and held at 5 degrees C. After 20 to 44 h, spermatozoa were processed by swimup, incubated with heparin, and co-cultured with oocytes (35 to 56 oocytes/sample; 18 h). Ova were observed for cleavage approximately 42 h after co-culture, and further development was evaluated on day 8. At first evaluation, cleavage rates were 18 and 46% for droplet and control groups (P < 0.01); semen had 34 to 70% and 0 to 12% droplet spermatozoa. For 10 of 15 droplet bulls, <10% of ova were cleaved whereas cleavage rate was >15% for all control bulls. At second evaluation, only three droplet bulls still had >30% of spermatozoa with a proximal droplet. Cleavage rates increased accordingly; only four droplet bulls had <10% cleaved ova and 10 had >34% cleaved ova. Three control bulls had <10% cleaved ova and nine had > or = 34% cleaved ova. Considering all 60 ejaculates, correlation between percentage of spermatozoa with a proximal droplet and percentage of cleaved ova was -0.49 (P < 0.0 1). Correlations between percentages of motile or normal spermatozoa in field evaluations and outcome in IVF were 0.28 and 0.52. Laboratory evaluations of spermatozoa concomitant with preparation for IVF revealed that only incidence of proximal droplets appeared related to outcome in IVF. We concluded: (a) semen from most yearling beef bulls with a high incidence of proximal droplet spermatozoa had severely compromised IVF fertility; (b) as these bulls matured, the incidence of proximal droplets decreased, and IVF fertilizing potential increased; and (c) semen containing >30% spermatozoa with a proximal droplet is strong evidence that fertilizing potential of the bull will be low until the incidence decreases.     

Semen from rainbow trout produced using cryopreserved spermatozoa is more suitable for cryopreservation

Oocytes from three female rainbow trout Oncorhynchus mykiss were inseminated separately with untreated or cryopreserved semen, which had been produced using either untreated (three males) or cryopreserved (three males) spermatozoa. In half of variants, the cryopreservation did not significantly affect fertilization efficiency. Regardless of whether the sperm donors were produced from cryopreserved or intact semen, insemination of oocytes with their intact sperm resulted in the same percentage of eyed embryos (94.4 and 94.3%, respectively). When eggs were inseminated with cryopreserved semen, the use of sperm from males produced with cryopreserved spermatozoa resulted in a significantly higher percentage of eyed eggs than in case of donors produced with intact sperm (89.6 and 81.7%, respectively). The production of rainbow trout using cryopreserved sperm does not appear to negatively affect reproductive abilities of male progeny and semen from donors, which were produced using cryopreserved sperm, is more suitable for cryopreservation than the semen from donors produced with intactspermatozoa.     

Evaluation of relative fertility of cryopreserved goat sperm

This study was designed to compare differences in the in vivo fertility of cryopreserved goat semen assessed by heterospermic insemination with differences in in vitro analyses. Five groups of does were inseminated with mixed frozen-thawed semen from different pairs of bucks. The percentage of offspring sired by each buck in the pair was compared with the relative ability of spermatozoa from that frozen-thawed ejaculate to penetrate zona-free hamster ova, relative post-thaw acrosomal integrity, ability to undergo an acrosome reaction during in vitro capacitation, and assessments of sperm motility. In 4 of the 5 different insemination pairs, the ratio of offspring born was other than 1:1. Acrosomal integrity, ability of spermatozoa to undergo an acrosome reaction, and parameters of sperm motility were not correlated with differences in relative fertility in this experiment using ejaculates from fertile bucks. The ability of spermatozoa to fuse with the oocyte plasma membrane was highly correlated with relative in vivo fertility (R(2) = 0.78, P = 0.04). This suggests that fusion with the oocyte plasma membrane is an event in the fertilization process in which significant variation exists among fertile bucks. Assessment of ability of spermatozoa to fuse with zona-free hamster ova may contribute to analysis of post-thaw fertility of frozen-thawed buck semen.     

Zinc does not inhibit the capacitation of human spermatozoa in vitro

The effects of zinc on human sperm motility, fertilizing capacity (as assessed by penetration of human spermatozoa into the zona pellucida-free hamster oocyte), and nuclear chromatin decondensation were investigated using spermatozoa from four fertile donors. Both sperm motility and the penetration of sperm into zona-free hamster ova were consistently impaired in media containing 1,000 μM zinc. Spermatozoa from one man were similarly affected at a concentration of 500 μM zinc, but no adverse effects were noted at this zinc concentration in experiments with other donors. Since decreased fertilizing capacity in response to zinc was always accompanied by a significant decline in both the percentage of motile cells and mean swimming speeds, it appears that all of these results reflect a general toxic effect on the cells. At lower concentrations (125–250 μM), zinc had no effect on human sperm motility nor their ability to undergo capacitation and penetrate zona-free hamster ova in vitro. For some donors, zinc (125–500 μM) stimulated both the attachment of spermatozoa to the hamster vitellus and the incorporation of spermatozoa into the hamster ooplasm. The decondensation of human sperm nuclear chromatin in sodium dodecyl sulfate was largely inhibited when zinc was added to the medium, but no significant changes in nuclear stability were apparent after capacitation in zinc-free medium. We conclude that zinc, when present in subtoxic concentrations, does not adversely affect the ability of human spermatozoa to undergo capacitation and penetrate zona-free hamster ova in vitro.     

Effects of heterologous sera on fertilized rabbit ova

Undiluted blood serum of various species was used to culture two-celled rabbit ova for 24 hours. It was found that there is an ovocidal factor present in the serum of man, sheep, cattle, goat, and fowl. The factor is lethal rather than inhibitory; exposure to it for 10 minutes will cause the death of the ova. This factor is unstable, thermolabile (destroyed at 55 degrees C. in 30 minutes), and of large molecular size. The strength or concentration of this factor varies according to the origin of the serum, increasing in the order man, sheep, cattle, goat, fowl. The blood serum of rabbit, horse, dog, guinea pig, rat, and pig contains no ovocidal factor against rabbit ova. The ovocidal factor differs from the spermicidal factor, which is present in all the sera of the different species studied with rabbit spermatozoa. Immunization of the guinea pig against rabbit ova is possible. Normal development of young rabbits was obtained by transplantation of ova cultured in the heated or normal serum of other species after 24 hours.