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PubMed1989

In vitro fertilization in the sheep: effect of elevated calcium concentration at insemination.

Oocytes (n = 273), collected from superovulated ewes, were inseminated with in vitro capacitated spermatozoa from four rams [Crozet et al., 1987]. In each experiment, parallel insemination was performed using aliquants from a single ejaculate in either our standard fertilization medium DM-H-SS (a modification of Brackett's defined medium, buffered with HEPES and containing 20% v/v sheep serum) or in the same medium supplemented with calcium lactate (DM-H-SS + Ca). The measured total calcium concentrations were Ca++ T = 2.74 mM in DM-H-SS medium and Ca++ T = 8.74 mM in DM-H-SS - Ca; the ratio of free to total calcium in DM-H-SS was Ca++ F/Ca++ T = 0.85. Fertilization was assessed at 17 hours postinsemination. Variations in the ejaculates were observed for each of the four rams tested. When DM-H-SS--CA was used, the percentages of fertilized (75% vs. 50%) and monospermic (58% vs. 41% eggs were significantly enhanced compared with use of DM-H-SS. No improvement was observed in control medium DM-H-SS + lac containing neutralized lactic acid. Supplementing the fertilization medium with calcium had no apparent effect on the incidence of polyspermy. These experiments show that the fertilization rate achievable in vitro by individual ejaculates from various rams can be increased by raising the calcium concentration in the fertilization medium to a value much higher than that present in tubal fluids from estrous ewes. Extended incubation in such a high calcium concentration is unnecessary for in vitro capacitation of ram spermatozoa.

Animals↗
PubMed1987

Effect of oocyte quality and sperm characteristics on the number of spermatozoa bound to the zona pellucida of human oocytes inseminated in vitro.

Sperm characteristics and oocyte quality may play a role in in vitro fertilization. The objective of this paper is to analyze the effect of the quality of oocytes, the husband's semen characteristics, and category of the couple's infertility on the number of spermatozoa bound to the zona pellucida. One hundred eighty-one oocytes which failed to fertilize or failed to cleave were fixed in 2.5% glutaraldehyde 40 to 60 hr after insemination in vitro and examined under interference microscopy and the number of sperm bound to the zone pellucida was determined. The means +/- SD of sperm bound to mature, immature, and atretic oocytes were 51.0 +/- 50.7, 7.3 +/- 12.1 10.4 +/- 17.8, respectively. Fertilized mature oocytes (81.0 +/- 53.3) had a significantly higher number of sperm bound to zonae compared to unfertilized oocytes (41.8 +/- 47.3). It is concluded that the number of sperm bound to zonae is functionally important. The sperm motility and the number of motile sperm used to inseminate oocytes were significantly correlated with the number of sperm bound to zonae, whereas sperm morphology and sperm concentration did not correlate. This study supports the notion that sperm motility is the single most important factor influencing fertilization of human oocytes in vitro.

Female↗
PubMed1991

Insemination of bile duct carcinoma to the liver after insertion of percutaneous biliary endoprosthesis.

Percutaneous transhepatic biliary drainage is widely used to relieve bile duct obstruction which can be caused by bile duct or pancreas carcinomas. Although the incidence is low, insemination of carcinoma along the transhepatic catheter tract is considered to be a serious complication of percutaneous transhepatic biliary drainage. The authors present a case of intrahepatic insemination of bile duct carcinoma along the catheter that subsequently underwent a curative resection consisting of pancreaticoduodenectomy and right hepatic lobectomy. It is suggested that a percutaneous biliary endoprosthesis through the tumor should be avoided in patients in whom a possible curative resection can be considered. External biliary drainage should only be performed in order to minimize the manipulation of the tumor in such patients.

Adenocarcinoma↗
PubMed1983

Insemination of rabbit eggs is associated with slow depolarization and repetitive diphasic membrane potentials.

The plasma membrane of the rabbit egg allows only one sperm to enter the egg during fertilization, but the mechanism of this block to polyspermy is unknown. Electrophysiology and in vitro fertilization techniques were employed in this study to investigate the possibility that a voltage block to polyspermy exists in rabbit eggs. Ovulated zona-intact eggs had a mean membrane potential of -71 +/- 2.1 mV (interior negative). A stereotypic response occurred 12-135 min following in vitro insemination in 19 of 40 eggs. Association of this stereotypic response with the appearance of pronuclei suggested that the electrical response was related to some interaction of gametes. This response consisted of a slow transient 8 +/- 1.5 mV depolarization upon which were superimposed up to 36 repetitive diphasic insemination potentials. Each potential consisted of a brief 2.0 +/- 0.44 mV hyperpolarization followed by a slow 2.5 +/- 0.45 mV depolarization. The small amplitude of the stereotypic response when compared with the large variation of resting potentials suggested that the response was insufficient to block polyspermy by a mechanism dependent upon the magnitude of the rabbit egg membrane potential.

Animals↗
PubMed1992

Chronic stress increases estrogen and other steroids in inseminated rats.

Physical restraint, like many other stressors, can block early pregnancy, but the underlying physiological mechanisms have not been established. Exogenous estrogens in minute doses will also block early pregnancy. In the present study, female rats were exposed to 5 h of restraint daily for the first 5 days after insemination. A subset of animals was sacrificed after 1, 2, 3, 4, and 5 days of restraint, and blood was collected for radioimmunoassay. Blood was also collected from unstressed control animals on each of the first 5 days after insemination. Plasma concentrations of estradiol, as well as those of corticosterone and progesterone, were increased in the stressed animals as compared to the controls. The finding of significantly enhanced maternal estrogen suggests the possibility that estrogens mediate psychogenic pregnancy blocks.

Animals↗
PubMed1994

Estrogen antibodies reduce vulnerability to stress-induced failure of intrauterine implantation in inseminated mice.

In Experiment 1, inseminated mice were randomly assigned to either an undisturbed control condition or four conditions involving exposure to restraint stress on days 1 through 5 of pregnancy. Restrained animals received one of three doses of estrogen antibodies or just vehicle injections on each day of restraint. Restrained animals receiving vehicle only showed significantly fewer uterine implantation sites than did unrestrained controls, while restrained animals given estrogen antibodies showed more implantation sites than did vehicle-treated restrained animals. In Experiment 2, varied dosages of refined estrogen antibodies were administered to inseminated females concurrent to restraint-stress on days 1 through 5 of pregnancy. More females receiving the higher dosages of antibodies produced litters than did restrained females with just vehicle injections, and at the highest dose the number of litters was similar to that produced by undisturbed control females. These results converge with other evidence to suggest that stress-induced pregnancy blocks are mediated by estrogens.

Animals↗
PubMed1979

Estrus synchronisation and fertility in gilts using a synthetic progestagen (Allyl Trenbolone) and inseminated with fresh stored or frozen semen.

An orally active synthetic progestagen was administered at two dosage levels to synchronise estrus in gilts. Fertility following insemination with either fresh stored or frozen semen was determined by examining surgically recovered ova for cleavage, and numbers of spermatozoa attached to the zona pellucida, or enumeration of embryos in gilts slaughtered 30 days post insemination. There was no significant difference (P>0.05) between treated and control groups in the duration of estrus or in fertility as determined by cleavage of ova. A significantly (P<0.001) shorter interval to estrus and better synchronisation was obtained with both treatment groups than with the control group. The mean interval from the end of treatment to the onset of estrus for the untreated controls and the treated groups receiving 12.5 and 15 mg compound per day was 11.25 +/- 10.4 SD; 5.6 +/- 0.52 SD and 7.3 +/- 5.3 SD. Fresh semen yielded significantly (P<0.01) more cleaved ova than frozen semen.

Journal Article↗
PubMed1983

Clitoral stimulation and the effect of age, breed, technician, and postpartum interval on pregnancy rate to artificial insemination in beef cattle.

An experiment was conducted to assess the effect of clitoral stimulation on pregnancy rate to artificial insemination in 1,856 beef females. For unadjusted data, pregnancy rate to first service in nonstimulated females was 61% for cows and 59% for heifers, and that of females receiving clitoral stimulation was 69% for cows and 58% for heifers. After adjusting the data for the influences of age, breed, technician, and postpartum interval, clitoral stimulation increased first service pregnancy rate in cows (74+/-3% vs 59+/-3%, P<0.05), but not in heifers (53+/-5% vs 57+/-5%, P>0.10). Postpartum interval significantly influenced pregnancy rate, regardless of treatment, and clitoral stimulation was beneficial when applied to cows with relatively short postpartum intervals. Variation in the effect of clitoral stimulation on pregnancy rate was observed among technicians. Clitoral stimulation improved pregnancy rate more effectively in 3 to 4 year old cows than in cows 2 years old and cows 5 years of age or older. We concluded that clitoral stimulation at the time of artificial insemination was an effective means of increasing pregnancy rate in cows, but not in heifers.

Journal Article↗
PubMed1985

Fertility trial in Zebu cattle after a natural or controlled estrus with prostaglandin F2 Alpha, comparing natural mating with artificial insemination.

With the object of comparing reproductive efficiency obtained by natural mating and by artificial insemination (AI), not only following a natural estrus but also after an induced estrus with PGF2Alpha in Zebu cattle in the tropics, 244 adult cows were divided into 4 groups. Group I (N = 69) and Group III (n = 62) were injected with 25 mg of PGF2Alpha when a functional CL was found on rectal examination. Group I was inseminated and group III was served by natural mating, both groups within five days after injection. Groups II (n = 57) and IV (n = 56) were left untreated, group II being AI and group IV ran with a fertile bull for 22 days. Estrus detection was carried out only in the injected groups (I and III) for 15 minutes every three hours between 0600 and 1800. All information was analyzed by linear trigonometric models. The onset of estrus occurred on average 68.7 h after injection in group I and 59.5 h in group III. However only 46.3% and 54.8% of animals were detected in estrus in group I and III respectively, the difference being significant (P < 0.10). Conception rates were 18.6%, 29.8%, 19.3% and 33.9% for groups I, II, III, and IV respectively. A significant difference (P < 0.10) existed between the injected groups and the untreated ones.

Journal Article↗
PubMed1990

Failure of gonadotropin-releasing hormone or human chorionic gonadotropin to enhance the fertility of repeat-breeder cows when administered at the time of insemination.

Repeat-breeder dairy cows failing to conceive to two previous inseminations were injected with saline (2 cc i.m.), gonadotropin-releasing hormone (GnRH, 2 cc i.m.) or human chorionic gonadotropin (hCG, 5000 IU i.v.) at the time of the third and succeeding inseminations. This field study involved 240 cows at three dairies. Mean conception rate was high (49.6%) for repeat-breeders, and did not differ among farms (P>0.10) or treatments (P>0.10), varying from 44.0 to 51.9% for farms and from 44.4 to 54.2% for treatments. The percentage of repeat-breeder cows bred during diestrus was very low on each of the three farms (mean = 7.9%). For this and other reasons discussed in the study, hormonal therapy was not successful in improving the conception rate of repeat-breeder cows under field conditions.

Journal Article↗
PubMed1993

A field investigation of the effects of bovine viral diarrhea virus infection around the time of insemination on the reproductive performance of cattle.

During a study of methods of synchronizing estrus in Bos indicus cattle, blood was collected from 169 heifers and 38 cows 2 to 3 days prior to artificial insemination (AI), and then again at Day 51 and Day 210 after AI to determine the incidence of infection with bovine viral diarrhea (BVD) virus. Prior to insemination 53 and 68% of the cows and heifers, respectively, were seronegative to the BVD virus. At Day 51 after AI, 70 and 32% of the seronegative cows and heifers, respectively, had seroconverted; but between Day 51 and Day 210, only 17 and 3% of the seronegative cows and heifers, respectively, had seroconverted. The Day- 51 pregnancy rate of cows which were immune (seropositive) to BVD virus infection at the time of AI was similar to the rate of the cows which became infected around the time of AI. However, the pregnancy rate of the immune heifers (44%, n=54) was significantly (P=0.04) greater than the rate of the heifers which became infected around the time of AI (24%, n=37). It was concluded that infection of susceptible females with BVD virus around the time of AI may significantly lower the pregnancy rate.

Journal Article↗
PubMed1993

The relationship of semen quality to pregnancy rate and litter size following artificial insemination in the bitch.

Artificial insemination of 31 bitches with fresh, undiluted, semen resulted in 22 31 conceptions (71%) and the birth of 124 puppies. Inseminations using at least 220 x 10(6) spermatozoa of normal morphology resulted in a mean pregnancy rate of 81.5% (22 27 ). The pregnancy rates were significantly influenced by the total number of progressively motile or morphologically normal sperm cells per ejaculate (P < 0.05). The pregnancy rate was not influenced by the percentage of progressively motile or morphologically normal sperm cells per ejaculate (P > 0.1). Litter size was not influenced by sperm motility or morphology (P > 0.1).

Journal Article↗
PubMed1994

Kinetics of first polar body extrusion and the effect of time of stripping of the cumulus and time of insemination on developmental competence of bovine oocytes.

Kinetics of extrusion of the first polar body was examined as well as the effect of the time of stripping of the cumulus cells on this kinetics. In addition, the effects of time of stripping and time of insemination on developmental competence of the oocytes, as evaluated by the percentage of morulae and blastocysts, were studied. Polar body extrusion occurred in 80% of the oocytes between 12 and 18 h after the onset of maturation. The remainder of the oocytes did not extrude a polar body at all. Stripping of the cumulus at 12 h after the onset of maturation delayed polar body extrusion significantly by about 1 h. No significant differences were found in the percentage of oocytes that could be fertilized, and the percentage of oocytes that cleaved and developed to the morula and blastocyst stages, between oocytes that were stripped free of cumulus and inseminated at either 16 or 20 h after onset maturation. Oocytes that had extruded a polar body at either 16 or 20 h after onset maturation showed significantly higher percentages of cleavage and development than oocytes that had not extruded a polar body at those time points. However, the percentage of oocytes that could be fertilized was not affected.

Journal Article↗
PubMed1996

Laparoscopic insemination of frozen-thawed semen into one or both uterine horns without regard to ovulation site in synchronized Merino ewes.

Synchronized ewes (n = 217) were bred by laparoscopic insemination of frozen-thawed semen from 1 of 3 rams. The ewes were bred by either a double (110 ewes) or single horn (107 ewes) technique without regard to the site of ovulation. There was no difference in the percentage of ewes pregnant to either the single or double horn breeding technique. There was a significant effect of sire, with 1 ram producing a higher pregnancy rate in the ewes and 1 ram producing a significantly lower pregnancy rate when compared to the total pregnancy rate for all the ewes (P < 0.05). Thus, the single horn breeding technique is presented as an alternative technique for use in the commercial breeding of ewes by laparoscopic insemination of frozen-thawed semen.

Journal Article↗
PubMed2004

Fallopian tube sperm perfusion versus intrauterine insemination in unexplained infertility: a randomized, prospective, cross-over trial.

OBJECTIVE: To compare the results of fallopian tube sperm perfusion (FSP) versus standard intrauterine insemination (IUI) in patients with unexplained infertility undergoing controlled ovarian hyperstimulation (COH). DESIGN: Randomized, prospective, cross-over study. SETTING: Reproductive medicine unit of a university hospital. PATIENT(S): Fifty-six couples with unexplained infertility. INTERVENTION(S): COH was induced by recombinant FSH and monitored by serial transvaginal ultrasound. On the day of hCG administration during the first treatment cycle, patients were randomized to either IUI or FSP. Thereafter, in case no pregnancy was achieved, patients went on being treated with FSP and IUI in alternate cycles. A maximum of four treatment cycles per couple was performed. FSP was performed using a pediatric Foley's catheter inseminating 4 mL of sperm-enriched suspension; a Kremer-Delafontaine catheter delivering 0.5 mL of sperm suspension was used for IUI. MAIN OUTCOME MEASURE(S): Clinical pregnancy rate per cycle. RESULT(S): One hundred twenty-seven cycles (58 FSP, 69 IUI) were performed. The clinical pregnancy rate per cycle was 21.7% for IUI and 8.6% for FSP, respectively. No major adverse effects were recorded for either technique. CONCLUSION(S): After COH, FSP is less effective than IUI in couples with unexplained infertility.

Cross-Over Studies↗
PubMed2006

Influence of oral and subcutaneous bisphenol-A on intrauterine implantation of fertilized ova in inseminated female mice.

Intrauterine implantation of fertilized ova in inseminated females is sensitive to minute levels of natural estrogens. Bisphenol-A (BPA), a widely used chemical in the production of polycarbonate plastics and epoxy resins, can be estrogenic. Here we administered BPA during the period of implantation to determine levels of exposure required to terminate pregnancy in mice. Varied doses were given through either injection or ingestion. Subcutaneous injections during days 1-4 of gestation significantly reduced litter size at 3.375 mg/day and substantially reduced the proportion of females that were parturient at 10.125 mg/day. Uterine implantation sites were also significantly reduced in females sacrificed at day 6 after receiving 10.125 mg/day. Exposure to lower doses was without significant effect. When inseminated females' diets were supplemented on days 1-5 with peanut butter contaminated by 0.11-9.0% BPA, litter size and percent parturient were not affected. However, when the animals' diet was exclusively comprised of a mixture of BPA, peanut butter, and powdered chow during days 1-4, an average daily intake of 68.84 mg BPA terminated all pregnancies. No significant effects at lower doses of BPA were seen in number of births or other measures through either mode of administration.

Administration, Oral↗
PubMed1989

Incorporation and dispersal of sperm surface antigens in plasma membranes of inseminated sea urchin (Arbacia punctulata) eggs and oocytes.

Monoclonal antibodies have been generated to determine the fate of sperm-specific surface components of the sea urchin, Arbacia punctulata, subsequent to gamete fusion. Monoclonal antibody-7 (MAB-7) reacted with two polypeptide bands having apparent molecular masses of 33 and 35 kDa derived from the sperm plasma membrane; similar reactivity was not detected in egg preparations. Eggs and oocytes were prepared for electron microscopy at periodic intervals following insemination and reacted with MAB-7 followed by antimouse antibodies conjugated to colloidal gold. In samples prepared 30 to 60 sec postinsemination (PI) colloidal gold was confined to the surface of fused sperm with only a few gold particles associated with the egg plasma membrane. In later samples (2-10 min PI) label was present on increasingly greater areas of the egg/oocyte surface away from the site of gamete fusion so that by 20 min PI particles were located along one hemisphere of inseminated eggs and oocytes. These results demonstrate the incorporation of sperm plasma membrane polypeptides and their intermixing with egg plasmalemmal components subsequent to gamete membrane fusion. From measurements of labeled oocytes at different times PI, the diffusion coefficient for sperm surface polypeptides detected by MAB-7 was estimated to be 0.7 to 2.4 x 10(-9) cm2/sec.

Animals↗
PubMed1983

Oocyte recovery, in vitro insemination, and transfer into the oviduct after its microsurgical repair at a single laparotomy.

A method combining recovery of a preovulatory oocyte, its in vitro insemination, and transfer into the oviduct with tubal microsurgery at a single laparotomy is described. Of four women in whom tubal microsurgery was clinically indicated and the accompanying tubal transfer of an oocyte inseminated in vitro was performed, two became pregnant. It is not clear whether fertilization occurred in vitro or in vivo. One of the pregnancies miscarried in the fifth week after surgery; the other pregnancy continued without any complications. A normal male infant was subsequently delivered at term. The proposed method is discussed as a possible alternative to in vitro fertilization and embryo transfer in some cases of infertility and as a suitable supplemental procedure in some cases in which tubal microsurgery is indicated.

Adult↗