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.     

Modification of the zona-free hamster ova bioassay of boar sperm fertility and correlation with in vivo fertility

These studies were designed to evaluate the ability of the zona-free hamster ova bioassay to detect differences in fertility of boar sperm. In the first study, sperm from two previously infertile boars were compared to sperm from seven previously fertile boars. The percentage of zona-free hamster ova penetrated by sperm from the previously infertile boars was significantly lower than the percentage of ova penetrated by sperm from previously fertile boars (18% of ova penetrated vs. 83%, P < .001). In the 14 ejaculates from the previously infertile boars that had ejaculate motilities of 50% or greater, the percentage of zona-free hamster ova penetrated continued to be lower than in ejaculates from the fertile boars. One of the two previously infertile boars consistently had a normal semen analysis. The only two observed manifestations of his reduced fertility were his zero conception rate and the limited ability of his sperm to penetrate zona-free hamster ova. In the second study, females were inseminated with equal numbers of sperm from two previously fertile males and the paternity of offspring determined at birth. The experiment was replicated with four combinations of six boars. A high correlation was observed between the percentage of offspring sired and the ability to penetrate zona-free hamster ova (R = .89). Neither morphology nor the ability of the sperm to undergo an acrosome reaction during in vitro incubation was correlated with fertility in the competitive mating situation. These results suggest the zona-free hamster ova bioassay can improve the in vitro fertility assessment of fresh boar semen.     

Evaluation of assay conditions for the zona-free hamster ova bioassay of boar sperm fertility

The ability to penetrate zona-free hamster ova may be a very useful test of fresh and frozen boar sperm fertility. These studies were designed to optimize assay conditions prior to evaluation of the accuracy of the bioassay in predicting boar sperm fertility. The ability to penetrate zona-free hamster ova was greater in sperm washed on a Percoll gradient than in sperm washed by dilution and centrifugation. Penetrating ability was greater in sperm from the sperm-rich fraction than from the whole ejaculate but did not differ among different aliquots of the sperm-rich fraction and did not decrease when the prewashing interval was increased from 15 to 85 min. Frequency of collection of ejaculates (1, 3, or 5 times per week) did not affect the penetrating ability of the sperm. Penetration rate was greater when sperm were coincubated with zona-free hamster ova at 39°C compared to 37°C. Sperm from an infertile boar had reduced penetrating ability compared to sperm from fertile boars (11% vs 93%, P < .001). These studies suggest that the zona-free hamster ova bioassay may be a useful assessment of fresh boar sperm fertility.     

Development of a zona-free hamster ova bioassay for goat sperm

In vitro conditions for a zona-free hamster ova bioassay of caprine sperm fertility were assessed. Washing the cryopreserved sperm by dilution and centrifugation resulted in greater ability to penetrate zona-free hamster ova than allowing the sperm to swim-up into medium. A 12-h preincubation of sperm prior to insemination of the zona-free hamster ova was optimal. Sperm had greater ability to penetrate the zona-free hamster ova when incubated in a Tris-buffered medium than when incubated in Ham's F-10 (95 vs 2%, P<0.001), although motility was not well maintained in the Tris-buffered medium. Ten million sperm/ml was sufficient for maximum penetration. The ability to penetrate zona-free hamster ova was positively but not significantly correlated with the sperm head ultrastructure, suggesting the two techniques may assess different aspects of sperm fertility.     

Do sire-dam interactions contribute significantly to fertility comparisons in heterospermic insemination trials

The percentage of offspring sired after heterospermic insemination of equal numbers of spermatozoa is believed to be a very sensitive measure of relative in vivo fertility of the inseminated samples. The objective of these trials was to evaluate whether there was a detectable male-female interaction in the fertilizing ability of spermatozoa. If there was such an interaction, we reasoned that the paternity of offspring from individual females in a heterospermic trial the second year would be similar to the paternity of offspring in the same individual females the first year if the same ejaculates were used. Five groups of ewes were inseminated with different combinations of semen (a single Merino ejaculate from one of five rams randomly paired with five different pools of Suffolk semen) in a heterospermic trial. Those ewes conceiving the first year were inseminated in a second breeding season with the same combination of semen used previously. The percentage of lambs sired by each ejaculate/pool of ejaculates was calculated for all lambs born from all ewes inseminated with each semen combination. These percentages would be the expected ratios of Merino-sired:Suffolk-sired lambs if there is no male-female interaction. Ewes in each group were divided into two subgroups: those conceiving only Merino-sired lambs the first year and those conceiving at least one Suffolk-sired lamb the first year. The ratio of Merino-sired lambs:Suffolk-sired lambs did not differ in either subgroup from those expected if there was no male-female interaction. These results are consistent with the absence of a male-female interaction in relative fertilizing ability of spermatozoa.     

The importance of porcine sperm parameters on fertility in vivo

It would be desirable to use semen parameters to predict the in vivo fertilizing capacity of a particular ejaculate. In animal production, an ejaculate is divided into multiple doses for artificial insemination (AI); therefore, it would be economically beneficial to know the functional quality (i.e., fertility) of the semen before it is inseminated. To identify a predictive assay of the fertilizing capacity of a porcine ejaculate, we performed 4 rapid assays of sperm quality (motility, viability, physiological status as assessed by chlortetracycline fluorescence, and ATP content) on samples from 9 ejaculates, before and after a thermal stress test (42.5 degrees C, 45 min). These parameters were subsequently correlated with in vivo fertility resulting from AI with 2 sperm doses, 3 x 10(9) or 0.3 x 10(9) motile cells in 70 mL (optimal or suboptimal sperm number per insemination, respectively) from these same ejaculates. No parameter was correlated to the fertility rates obtained after inseminating with the optimal semen doses, either before or after the thermal stress test (P > 0.05). However, with respect to the animals inseminated with the suboptimal semen dose, sperm motility (the percentage of motile spermatozoa as assessed visually by microscopy) prior to thermal stress was well-correlated to fertility rates (r = 0.783, P = 0.01). The percentage of spermatozoa displaying the chlortetracycline Pattern AR (acrosome reaction) was also statistically related to fertility (r = 0.05, P = 0.04), but the biological importance of this relationship is questionable given the small variation among ejaculates (range: 0 to 2%). No other sperm parameter was significantly related to fertility rates in this group (P > 0.05). These data, therefore, indicate that sperm motility is a useful indicator of sperm fertilizing capacity in vivo. Moreover, to identify a predictor of semen fertility it is critical that the number of spermatozoa used during insemination is sufficiently low to detect differences in sperm fertilizing efficiency.     

Prediction of fertility of bovine semen: Preliminary studies with the hamster egg penetration test

The ability of liposome-treated fresh and frozen spermatozoa from two bulls to interact with zona-free hamster oocytes was examined to show whether the in vitro test results would correspond with in vivo fertility as indicated by the 60 to 90 d nonreturn to service rates which, using frozen semen, were 77 and 59%, respectively. The motility of spermatozoa in washed suspensions was also rated. Hamster test results were obtained using three ejaculates from each bull both as fresh and frozen semen. The results with frozen semen corresponded with fertility. The averages of three hamster tests for oocyte penetration rates and mean number of spermatozoa per penetrated oocyte comparing spermatozoa from the bull with the higher fertility with spermatozoa from the bull with the lower fertility were 91% and 2.7 versus 56% and 1.4, respectively. Spermatozoa washed from frozen semen from the bull with the higher fertility interacted with hamster oocytes at the higher rate even when sperm motility was rated the same for both bulls. By contrast, fresh spermatozoa from the lower fertility bull interacted with hamster oocytes at a higher rate than spermatozoa from the higher fertility bull in six tests, comparing six ejaculates of fresh semen from both bulls. Comparing the higher fertility bull with the lower fertility bull, the average of six tests for oocyte penetration rates and mean number of spermatozoa per penetrated oocyte were 60% and 1.6 versus 89% and 3.0, respectively. This suggests that this hamster test cannot be used with fresh semen to predict relative levels of fertility of frozen semen. Also, the subjective rating of sperm motility did not correspond with the in vitro oocyte penetrating ability of the spermatozoa.     

Boar sperm plasma membrane protein profile: Correlation with the zona-free hamster ova bioassay

This study was designed to compare differences among porcine sperm plasma membrane proteins with the ability of spermatozoa to interact with zona-free hamster ova. Sperm plasma membrane vesicles were recovered from 24 ejaculates from 10 fertile boars, and from cauda epididymal spermatozoa from 3 fertile and 1 very subfertile boar. Solubilized sperm plasma membrane proteins were run on 1D SDS-PAGE gels, transferred to western blots, stained, and analyzed for quantity of protein per band by scanning laser densitometry. Variation in the quantities of individual sperm plasma membrane proteins in the 20 identified bands were statistically compared with the ability of spermatozoa from the same ejaculate to penetrate zona-free hamster ova. The percentages of plasma membrane protein present in 3 bands (90, 84 and 60 kD) were positively correlated with the ability of spermatozoa from the same ejaculate to fuse with zona-free hamster ova (P = 0.002, 0.01, 0.04; R = 0.53, 0.40, 0.38, respectively). The quantities of protein in 2 other bands (69 and 35 kD) were significantly but negatively correlated with the results of the zona-free hamster ova bioassay (P = 0.02, 0.01; R = -0.42, -0.37, respectively). The sperm plasma membrane profiles were quantitatively similar between the ejaculated samples and the fertile epididymal samples. Six epididymal sperm plasma membrane proteins were present in statistically different quantities in the subfertile boar sample and the 3 fertile controls. The 90 kD band positively correlated with the hamster ova bioassay in the ejaculated samples was not detected in the subfertile epididymal sperm plasma membrane sample. These results suggest that protein(s) in one or more of the 3 positively correlated ejaculated sperm plasma membrane protein bands may be involved in sperm-oocyte interaction.     

Some factors influencing the interaction of ram spermatozoa with zona-free hamster eggs

Variability in the interaction of ram spermatozoa with zona-free hamster eggs was recorded not only amongst individual males but also between the first and second ejaculates of the same male collected 30 min apart. Fusion ability also differed according to the conditions of gamete mixing. This ability decreased after in-vitro storage of undiluted ejaculates at room temperature but lasted for 48-192 h. The kinetics of sperm-egg fusion during the time of gamete incubation varied not only with the time of sperm storage in vitro but also with the ejaculate. When the semen was frozen, the ability of the spermatozoa to fuse was markedly reduced.     

In vivo fertility of rams in relation to sperm-zona pellucida binding and sperm-zona pellucida penetration of ovine oocytes

Sperm-zona pellucida (zona) binding and sperm-zona penetration have been suggested for use as in vitro bioassays of fertility since both are essential steps in the fertilization process. The correlations of sperm-zona binding and sperm-zona penetration with the in vivo fertility of sheep were investigated in this study. In vivo fertility was estimated from a heterospermic insemination trial using cryopreserved ram semen. Neither zona binding, zona penetration nor the ability to undergo an acrosome reaction was significantly correlated with the in vivo fertility of the rams (P = 0.78, P = 0.66, and P = 0.85, respectively). These results suggest that the zona binding and zona penetration bioassays may not be useful estimators for assessing cryopreserved ram sperm fertility.     

A chromosomal method to distinguish between X- and Y-bearing spermatozoa of the bull in zona-free hamster ova

The interspecific in-vitro fertilization system using zona-free hamster ova was adopted to distinguish chromosomally between X- and Y-bearing bull spermatozoa. Frozen semen samples were thawed and washed with modified BWW medium including 0.3% BSA (pH 7.2) to remove cryoprotective medium. Motile spermatozoa were recovered from the semen by the 'swim-up' method. These spermatozoa were treated with ionophore A23187 to facilitate capacitation. Adequate capacitation of spermatozoa was found by their movement patterns and the insemination was performed at the optimum time. The fertilized ova were cultured in Medium TC199 with 10% FBS including podophyllotoxin and vinblastine until they reached first cleavage metaphase. Chromosome slides were prepared by our gradual fixation-air drying method. The success rate of the method was 56% of the number corresponding to the zona-free ova used. Altogether 1116 spermatozoa from 4 different bulls were successfully analysed and the ratio of X- and Y-bearing spermatozoa was 542:574 (P greater than 0.3).     

Efforts to correlate laboratory with field observations on bull sperm fertility

This work represents efforts towards development of the zona-free hamster ovum sperm penetration assay for predicting relative levels of fertility for semen from individual bulls. Results reported here followed insemination of hamster vitelli with bull sperm, after frozen storage, with observations of sperm acrosomes and parallel inseminations of more than 1000 cows with semen from each bull. The average 75-day non-return rate for the four bulls was 74.0% (range 71.6 to 75.6). Laboratory studies indicated the following: the percentage of sperm with intact acrosomes varied from 55 to 73 between bulls, the percentage of motile sperm varied from 41 to 64, the percentage of sperm with progressive motility ranged from 24 to 40, the number of sperm interacting per (zona-free hamster) ovum ranged from 1.6 to 3.8, the number of sperm attached per ovum ranged from 1.4 to 2.9, the number of sperm within each penetrated ovum ranged from 1.5 to 1.8, the percentage of ova interacting with sperm ranged from 76 to 92, the percentage of ova penetrated ranged from 62 to 85, and the percentage of ova with male pronuclei ranged from 33 to 49. Although predictive ranking in the laboratory of these bulls with less than 4% variation in fertility levels was not possible, the zone-free hamster ovum test could be useful in identifying potentially subfertile bulls before they enter a young sire-sampling program.     

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.     

Effects of sperm preparation and bull fertility on in vitro penetration of zona-free bovine oocytes

The objectives of this study were to develop and validate a zona-free bovine oocyte penetration assay for detecting relative differences in bovine sperm fertility and to determine the effect of different sperm preparation methods on oocyte penetration. Oocytes were incubated with heparin-capacitated spermatozoa which either were or were not induced to acrosome-react with lysophosphatidylcholine. Heparin-capacitated spermatozoa treated with lysophosphatidyl-choline penetrated more oocytes and had more penetrations per oocyte than spermatozoa capacitated in heparin but not induced to acrosome-react with lysophosphatidylcholine. Spermatozoa stained with Hoechst 33342, fluorescein isothiocyanate or tetramethyl rhodamine isothiocyanate, alone or in combination, penetrated similar numbers and percentages of zona-free bovine oocytes as the similar to non-stained spermatozoa. When spermatozoa from the same ejaculate were stained with either fluorescein isothiocyanate or tetramethyl rhodamine isothiocyanate and competed in penetrating the same oocytes, the number of penetrations generated by the 2 differently stained spermatozoa was similar. Spermatozoa from bulls of differing in vivo fertilities were labeled with different fluorescent dyes, and their relative abilities to penetrate the same oocytes were assessed. Comparisons between spermatozoa from high and low fertility bulls demonstrated that high fertility spermatozoa had a significant oocyte penetrating advantage over low fertility spermatozoa in 13 of 16 paired competitions. We concluded that the results of the competitive penetration of zona-free bovine oocytes by fluorochrome-labeled spermatozoa from bulls of different fertilities were indicative of their relative in vivo fertility.     

Influence of sperm number and seminal plasma on fertility of progestagen-treated sheep in confinement

Progestagen-impregnated vaginal sponges + PMSG were used to synchronize oestrus in crossbred adult ewes which were inseminated 56 h after sponge removal with 0.5 ml diluted semen containing 400, 200, 100, 50 or 25 x 10(6) spermatozoa per insemination. The diluent was skim milk-citrate or pooled seminal plasma. There was no difference in reproductive performance due to the insemination medium. Fertility (no. of ewes lambing) after insemination of 400 or 200 x 10(6) spermatozoa was 68% and was similar to that observed after natural service at progestagen-induced oestrus. When less than or equal to 100 x 10(6) spermatozoa were inseminated, fertility fell markedly and the number of lambs per ewe inseminated decreased. A decrease in litter size also occurred. The data indicate that insemination of 200 x 10(6) spermatozoa, i.e. less than 10% of the number in a single ram ejaculate, allows normal conception rates in progestagen-treated ewes.     

Dilauroylphosphatidylcholine liposome effects on the acrosome reaction and in vitro penetration of zona-free hamster eggs by bull sperm: II. A fertility assay for frozen-thawed semen

Frozen-thawed sperm from five bulls with fertility rates ranging from 48% to 77% were treated with seven concentrations of dilauroylphosphatidylcholine (PC12) liposomes to induce an acrosome reaction (AR) that enabled sperm to penetrate eggs. Treated sperm were incubated with liposomes for 7 min prior to insemination of zona-free hamster eggs in vitro. Sperm and eggs were incubated 3 hr at 39°C prior to fixation, staining, and examination for sperm penetration and nuclear decondensation. The percentage of motile sperm immediately after thawing as well as after treatment with liposomes had a low correlation with sire fertility (r = .39 and ?.63, respectively). The percentage of sperm exhibiting an AR was more highly correlated with fertility (r ? ?.85). Similar correlations were found between fertility and the penetration rates of zona-free hamster eggs or the total number of penetrating sperm. When data for two high and for two lower fertility buils were each grouped to increase information per data point the correlation between the PC12 concentration giving the maximum proportion of eggs penetrated and fertility was r = .92 (P ≤ .05). The correlation between the PC12 concentration producing the most total sperm penetrating the eggs and fertility r = .97 (P ≤ .05). It was concluded that PC12 liposomes induced an AR in bull sperm frozen-thawed in egg yolk extender. Frozen-thawed sperm from low fertility bulls require less PC12 to induce the AR and to penetrate zona-free hamster eggs than do sperm from higher fertility bulls. These differences in lipid requirements may help to provide a quick, direct laboratory assay method to estimate the fertility of frozen bull semen.     

Correlations between heterospermic fertility and assays of porcine semen quality before and after cryopreservation

The relative fertilizing potential of frozen-thawed semen from four black and four white boars was determined following heterospermic insemination. A heterospermic index (HI) was computed for each of the 16 possible pairs of black and white boars. Correlation coefficients were computed between the HI and several in vitro tests of semen quality before and afttr cryopreservation of the semen. For the in vitro tests before cryopreservation, the HI were negatively correlated (-0.57) with spermatozoal motility before cooling the semen, but they were not correlated with spermatozoal motility after cooling to 5 degrees C. After freezing and thawing, the HI were correlated with the following in vitro tests: spermatozoal motility (0.50), spermatozoa with either normal or damaged apical ridges (0.31), spermatozoa with missing apical ridges (-0.51), spermatozoa filtered through sephadex columns (0.32), spermatozoa with acrosin-activity (0.38), percentage of maximal releasable glutamic oxalacetic transaminase (GOT) present extracellularly (0.54), spermatozoal intracellular GOT (-0.57), spermatozoa bound per zona-free hamster oocyte (0.64), and percentage of zona-free hamster oocytes penetrated (0.75). The HI were not correlated with the following in vitro tests after freezing and thawing: spermatozoa with normal apical ridges, damaged apical ridges and loose acrosomal caps, extracellular and maximal releasable GOT, and the number of penetrations per zona-free hamster oocyte. The multiple regression correlation coefficient between the HI and four selected variables from three in vitro tests was 0.94. This high correlation indicated that the fertilizing potential of the semen could be accurately predicted with four variables that appeared to measure different properties of the spermatozoa.     

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.     

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.     

Interaction of human spermatozoa with the human zona pellucida and zona-free hamster oocyte following capacitation by exposure to human cervical mucus

The functional capacity for sperm interaction with the human zona pellucida and zona-free hamster oocyte was tested after human spermatozoa were capacitated by passage through a column of human cervical mucus. The results were compared with those obtained when spermatozoa from an aliquot of the same semen sample were capacitated by the standard laboratory methods involving sequential washing by dilution and centrifugation of the semen. Washed-capacitated sperm suspensions were more successful than mucus-capacitated sperm in attaching to the zona-free hamster oocyte and in fusing with its oolemma. However, the ability of mucus-capacitated sperm to penetrate the human zona pellucida was equal to washed capacitated sperm. These experiments demonstrate an approach that may be useful in comparative studies of human sperm capacitation in vivo and in vitro.