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Objective To assess if epididymal sperm cryopreserved 24hours after exposure to oocytes in conventional IVF can be successfully used for intracytoplasmic sperm injection (ICSI) in a subsequent cycle. Design Case report. Setting University of California, Irvine, Center for Reproductive Health. Patients Two men with obstructive azoospermia requiring microsurgical epididymal sperm aspiration, IVF, and ICSI. Interventions Freezing of epididymal sperm 24hours after egg exposure in conventional IVF and subsequent use for assisted fertilization in a new cycle. Main Outcome Measure Frozen-thawed epididymal sperm survivability and maintenance of fertilization and pregnancy capacity. Results At the time of sperm aspiration procedure (cycle I) a total of 30 oocytes were available for insemination. Of these, 15 were used for conventional IVF resulting in 2 embryos (13%) and 15 were used for ICSI, resulting in 3 embryos (20%). Sperm was cryopreserved 24hours after conventional IVF and thawed 6months later in a new cycle. Upon thawing, sperm were still found to be motile and at this time (cycle II) only assisted fertilization was used. Of 27 oocytes injected, 12 (44%) produced normal, cleaving embryos. One singleton pregnancy with the birth of a healthy infant girl was achieved after the tubal transfer of 5 embryos. Conclusion The birth of a normal, healthy infant girl with epididymal sperm frozen 24hours after exposure to oocytes in conventional IVF emphasizes the value of freezing any aliquot of epididymal sperm, even if the motility is very low, to avoid additional surgery in the male. From a basic science standpoint, this observation may renew interest in the study of sperm cryopreservation after occurrence of acrosome reaction and hyperactivation.

Hysteroscopy is an excellent additional instrument for evaluating the uterine characteristics in infertile women. This article reviews the two main indications for hysteroscopy in infertile patients who are candidates for assisted reproductive techniques: (1) to evaluate the cervix and uterine cavity and (2) rule out any pathology or lesions that could have been missed by hysterosalpingography. Hysteroscopy has been proposed as an alternative method to traditional transabdominal transfer of gamete or embryo to the tube.

To use the protein-free medium blastocyst antihatching model to characterize physiological events that mediate hatching.In a series of four prospective experiments, our aims were to [1] test the efficacy of the antihatching model and assisted hatching; [2] exam the influence of initial in vivo developmental stage and late serum supplementation on hatching inhibition; [3] discount the role of zona hardness and physical expansion directly affecting hatching; and [4] provide evidence that the trophectoderm is directly responsible for secreting a zona lysin.University-based research laboratory.Culturing two- to eight-cell mouse embryos in serum-free human tubal fluid (HTF) medium significantly reduced hatching levels toor = 2%, however, hatching increased to 10.7% when initially culturing morula-stage embryos. Hatching was effectively rescued to control levels when embryos were placed in HTF with serum at the early blastocyst stage. There was no difference in blastocyst total cell numbers or zona pellucida digestion intervals between culture treatments. Finally, we showed that trophectodermal vesicles, devoid of inner cell mass, are capable of hatching under control conditions.The primary mechanism of blastocyst hatching is not physical expansion and abnormal zona hardness is not responsible for hatching inhibition. Certain extracellular precursors found in serum (e.g., amino acids) are required in culture medium upon cellular determination of trophectoderm (i.e., morula to blastocyst stage) to facilitate the intrinsic secretion of an undefined hatching factor.

The objective of this study was to determine the conception rate in infertile couples in which the female partner wasor = 40 years old and who had received ovarian stimulation treatment and intra-uterine insemination (IUI). It was a retrospective study of 77 patients who underwent a total of 210 treatment cycles. Protocols for ovulation induction included clomiphene citrate, human menopausal gonadotrophin (HMG) and clomiphene citrate plus HMG. Patients were monitored using transvaginal ultrasound, and two IUI were performed 24 and 48 h after the determination of urinary luteinizing hormone (LH) surge or human chorionic gonadotrophin (HCG) injection. A total of 11 pregnancies were reported, giving a pregnancy rate of 14% per patient and 5% per cycle. Eight spontaneous abortions occurred, giving a pregnancy wastage of 73%. In a previous comparative analysis of 543 patients39 years old receiving IUI and identical protocols of ovarian stimulation, 141 pregnancies were achieved, giving a pregnancy rate of 21% per patient and 10% per cycle. The miscarriage rate in that group was 18%. This report compares IUI results for womenor = 40 years with those obtained previously for younger women, and shows the very poor success rate in women40 years of age. This information will be important in the proper counselling of this group of patients, as well as indicating that a prompt recommendation for assisted reproductive treatment should be made soon after the failure of a few attempted cycles of ovarian stimulation treatment and IUI.

Although a number of different mechanisms have been suggested to account for the decline of fertility with age, the majority of studies agree that poor oocyte quality and reduced endometrial receptivity are the most important. In fact, the increased incidence of early pregnancy loss and chromosomal abnormalities of oocytes in older women, as well as the ability to reverse decreasing pregnancy rates by using oocyte donation, strongly support the evidence that oocyte ageing is the main factor responsible for decreasing fertility. Conversely, the lack of knowledge of the physiological variables that determine a successful nidation of a human embryo makes the analysis of uterine receptivity much more difficult. In order to evaluate the impact of the age of donors and recipients on pregnancy, implantation and abortion rates, we have retrospectively analysed 258 cycles from our programme of oocyte donation. Results were reviewed according to the following subclasses of age groups:or = 30, 31-35 and 36-39 years for donors, andor = 30, 31-35, 36-40, 41-45 and 46-53 years for recipients.(ABSTRACT TRUNCATED AT 250 WORDS)

To assess whether frozen ET to the fallopian tube is a possible alternative for cryopreserved embryos.Fifty-four patients (mean age 35 years) participated, in which their embryos were cryopreserved in 1.5 M propanediol at the pronuclear or two-cell stage. Each patient then underwent a steroid replacement cycle consisting of oral micronized 17 beta-E2 2 mg on days 2 to 4, 4 mg on days 5 to 7, 6 mg on days 8 to 10, and 8 mg from day 11 on. Serial ultrasounds were performed to evaluate the endometrium until an optimal thickness ofor = 10 mm triple layer was achieved. At this time, 100 mg IM progesterone was initiated and the zygote intrafallopian transfer (ZIFT) procedure was performed on the third day of P administration. The average number of embryos transferred was 4.4.Twenty-two clinical pregnancies resulted, giving a pregnancy rate of 41%. Eight miscarriages occurred and one ectopic pregnancy resulted, giving a live birth rate of 24%. Implantation rate was 10.8%. The highest chance of pregnancy was seen in patients who never had a previous IVF, GIFT, or ZIFT (61%).This is the first report of a series of frozen ETs to the fallopian tubes. These results indicate that tubal transfer may offer a protective benefit of the tubal environment and avoidance of endometrial trauma, and should be added to our armamentarium of replacing cryopreserved embryos.

The relative importance that endometrial development and embryo quality have on implantation rates achieved with assisted reproductive technology (ART) is the subject of controversy. Ovarian stimulation has been repeatedly mentioned as having a detrimental effect on endometrial receptivity (Paulson et al., 1990, Fertil. Steril., 53, 870-874). We compared pregnancy and implantation rates achieved with ART during stimulated cycles and hormonal replacement cycles, in patients matched for the following criteria: age35 years for the patient donating oocytes; transfer of at least two good quality embryos/oocytes and good quality transfer. All transfer cases performed during hormonal replacement cycles were done with donated oocytes. Comparison of results between techniques was not attempted due to potential differences in populations. The pregnancy and implantation rates achieved with each technique during stimulated and hormonal replacement cycles were not statistically different. In contradiction to previous results, our data suggest that differences in uterine receptivity between stimulated and hormonal replacement cycles in the age group studied are not of critical importance in embryo implantation. Good embryo quality appears to be the dominant factor in determining the success of ART.

To present data on IVF in severe male factor infertility using modified IVF laboratory methods and to compare them with the reported data of gamete micromanipulation.Retrospective, not randomized.University of California, Irvine, Center for Reproductive Health.Seventy-one patients with severe oligoasthenozoospermia defined as total motile count in the pretreatment sample of5 x 10(6). Two groups were identified, group I with total motile count between 1.5 and 5 x 10(6) and group II with total motile count1.5 x 10(6).Treatment of the semen samples with mini-Percoll and the modification of standard IVF techniques. These modifications include insemination with larger numbers of sperm, the use of culture tubes for insemination of oocytes, and pooling oocytes together in one or more culture tubes.Fertilization and pregnancy rates (PRs). Additionally, the number of patients achieving cryopreservation and patients with previous failure to fertilize are specified.Eighty percent of the patients achieved fertilization with an overall rate of 38% per oocyte. Fifty-three ETs were performed and 23 clinical pregnancies (43% per transfer and 35% per cycle) were achieved. Fifty percent of the patients had excess embryos for cryopreservation. In group I the fertilization rate was 54% with a PR of 56% per transfer and 48% per cycle. In group II the fertilization rate was 25% with a PR per transfer of 32% and 24% per cycle. Of 28 patients who had previous failure of fertilization, 25 fertilized and 8 pregnancies were established.These data demonstrate that by the modification of standard laboratory methods for IVF, monospermic fertilization, cleavage, and a high clinical PR can be achieved in cases of severe male factor infertility without having to resort to micromanipulation.

Previous experiences in subjects with other forms of third space fluid accumulation have shown that albumin is efficacious in preventing and correcting haemodynamic instability. Using a similar approach in an effort to increase the serum oncotic pressure and to reverse the leakage of fluids from the intravascular space, high risk subjects for severe ovarian hyperstimulation syndrome (SOHS) were treated with albumin. In a recent large study two high risk factors were identified, i.e. the number of oocytes and levels of serum oestradiol. Thirty-six women undergoing assisted reproductive techniques who presented both these factors, received intravenous albumin at a dose of 5% in Ringers lactate in doses of 500 ml during oocyte retrieval and 500 ml immediately thereafter in the recovery room. Daily measurements of urine output, serum and urine electrolytes, weight, abdominal girth, and haematocrit prior to and after oocyte retrieval revealed normal serum and urine electrolyte levels, and no signs of haemoconcentration. No patient in this study developed SOHS, and of course none had to be hospitalized. Vaginal ultrasound performed in the majority of the subjects revealedor = 100 ml of peritoneal fluid 48-72 h after oocyte retrieval. The only complication from the use of intravenous albumin was the appearance of a 'flu-like condition' (low grade temperature, nausea and muscle pains) developed by 12 women between days 3 and 5 after oocyte collection. Intravenous albumin had thus prevented the development of severe ovarian hyperstimulation syndrome in an assisted reproduction programme.(ABSTRACT TRUNCATED AT 250 WORDS)

The relative importance that endometrial development and the time of entrance of embryos to the uterine cavity have on their chances of implantation, is controversial. We review pregnancy rate (PR) and implantation rate (IR) on 186 patients selected based on the following criteria: 1) ageor = 35 for patients undergoing ovarian stimulation. 2) transfer of at least two good quality embryos and 3) quality of the transfer: optimum. 131 patients received embryos during a Stimulated Cycle (SC). 67 of them had uterine (UT) and 64 tubal transfers (TT), with PR of 38.8% and 53.1% (N.S.) and IR of 13.9% and 20.9% (p0.005) respectively. 55 patients received embryos in a Hormonal Replacement Cycle (HRC). 28 of them had UT and 27 TT. PR were 53.6% and 62.9% (N.S.) and IR 17.0% and 21.6% (N.S.) respectively. Our data suggests that: 1) the delayed entrance of embryos into the uterus and/or their atraumatic transfer into the tubes does not seem to improve their chances of implanting. 2) Endometrial differences between stimulated and hormonal replacement cycles seem not to be of critical importance after 48 to 72 hours after oocyte aspiration. 3) A detrimental effect of the endometrial milieu of stimulated cycles, that express only the first 48 to 72 hours after oocyte aspiration, is suggested.

Recent reports in the literature have focused on the increased risk of heterotopic pregnancy after the transfer of multiple concepti or oocytes. In an international collaborative patient registry between 1985 and 1989, 601 clinical pregnancies resulted from 2092 gamete intra-Fallopian transfer (GIFT) retrieval cycles. Five of the pregnancies were heterotopic (0.83%). After surgical intervention, all five cases of combined gestation resulted in live birth from intrauterine pregnancies. Routine vaginal ultrasonographic examination of the adnexa in patients who conceive after GIFT may help early diagnosis of heterotopic pregnancy. If the diagnosis is made early, conservative treatment may preserve the future fecundity of the patient and more intrauterine pregnancies may be salvaged.

Many couples fail to achieve a viable pregnancy with multiple attempts of Assisted Reproductive Technology (ART). Immunologic rejection of the embryo may be a factor in these couples. We evaluated shared human leukocyte antigen (HLA) antigenicity in 10 couples who failed multiple ART cycles and compared them with 10 couples who achieved a viable pregnancy with their first ART attempt. Study couples shared a statistically significant greater number of HLA antigens than control couples. Leukocyte immunotherapy is discussed. We conclude that couples who share multiple HLA antigens may have an immunologic basis for failing to achieve a viable pregnancy with multiple ART cycles.

Kastin and Schally proposed the possibility of the synthesis of inhibitory analogs of LHRH (Kastin et al., 1972). This hypothesis was shown to be correct and polypeptides capable of competing with LHRH for binding in the pituitary and decreasing the secretion of LH and FSH have been synthesized (Pedroza et al., 1977; Coy et al., 1979). Thus, dosages of [d-Phe2,d-Trp3,d-Phe6]LHRH in the range of 1.5 mg per animal completely suppressed spontaneous ovulation in proestrous rats (de la Cruz et al., 1976). These analogs were also found to be effective in inhibiting ovulation in hamsters and rabbits (de la Cruz et al., 1975; Phelps et al., 1977) and decreasing the response to LHRH in some non-human primates including baboons and chimpanzees (Hagino et al., 1977; Gosselin et al., 1979). Clinical studies performed in men with the antagonist [d-Phe2, d-Trp3,d-Phe6 ]LHRH showed that a 90 mgdose could suppress the LH and FSH release in response to 25 μg of LHRH for up to 24 h (Gonzales-Barcena et al., 1977). An antiovulatory effect was shown in women with the same dose of this antagonist (Canales et al., 1980).

Sperm penetration through bovine cervical mucus was determined and correlated with the sperm characteristics in three fractions of the split ejaculate and in the whole ejaculate from 40 men of infertile couples in which a female cause of infertility had been ruled out. Sperm concentration and motility were found to be significantly higher in the first fraction (n = 40) as compared with the rest of the fractions and the whole ejaculate, whereas normal sperm morphology was not significantly different among the various fractions and the whole ejaculate. The bovine mucus penetration (BCMP) test was found to be normal in a significantly high percentage (58%) of samples of the first fraction as compared with the other two fractions (32% and 17%, respectively) and the whole ejaculate (35%) (n = 17). Moreover, in the first fraction of the split ejaculate, the results of the BCMP test were correlated significantly with sperm concentration (P less than 0.01), motility (P less than 0.0001), and morphology (P less than 0.02). The data demonstrate the superiority of the first fraction of the split ejaculate in terms of cervical mucus penetrability and sperm characteristics.

A series of studies was performed in order to assess the antigonadotropic potency of an inhibitory analogue of LH-RH (IA-LH-RH) ( [N-Ac-D-Trp1, 3, D-p-Cl-Phe2, D-Phe6, D-Ala10]-LH-RH) in oophorectomized rhesus monkeys. I) Single administration of 1 mg IA-LH-RH im resulted in a prompt and significant decrease of serum levels of FSH and LH which lasted for at least 24 h. A specific rebound of serum LH to higher concentrations than baseline occurred in all animals at +48 or +72 h. II) Exogenous LH-RH at a dose of 150 micrograms was administered iv 2 or 18 h after the injection of 1 mg IA-LH-RH. Blood was drawn at 0, 10, 20, 30, 60, 120 and 180 min post LH-RH. The rise in LH and FSH levels observed in the control animals was not altered when LH-RH was administered 2 h after the antagonist, but it was significantly inhibited and delayed when LH-RH was given 18 h after the LH-RH inhibitory analogue. These results show that antagonistic analogues of LH-RH are potent inhibitors of gonadotropin concentrations in non-human primates. Their potential use in conditions in which inhibition of gonadotropin secretion and/or release is desired is discussed.

The effect of intramuscular injections of human chorionic gonadotropin (hCG) on subsequent in vitro prostaglandin (PG) production by the corpus luteum was studied in the rhesus monkey. Four monkeys received increasing doses of hCG from days 6 to 10 after ovulation. On day 11, laparotomy and luteectomy were performed. Five untreated animals served as controls. The production of prostaglandins F (PGF) and E (PGE) by the corpus luteum of the animals treated wit hCG was significantly lower than that of the controls (P less than .01). After hCG treatment, the decrease in PGF production was greater than that of PGE, resulting in a lower ratio of PGF:PGE production than in the controls (P less than .01). Histologic evaluation of the corpora lutea revealed regressive changes in the control group, whereas signs of active secretion were observed in the hCG-treated group. Progesterone concentrations in peripheral blood of the hCG-treated group were approximately fourfold higher than in the controls, and the hCG-treated group did not have a postmidluteal phase decrease. These results suggest that the ratio of production of PGF to PGE by the corpus luteum of the rhesus monkey may play a role in the control of its lifespan and steroidogenic capacity.

Regularly cycling rhesus monkeys received, from days 1 to 6 of the menstrual cycle (1) the potent luteinizing hormone-releasing hormone (LH-RH) agonist D-Trp6-LH-RH, 20 micrograms/day; (2) the potent LH-RH antagonist [N-Ac-D-Trp1,3,D-p-Cl-Phe2,D-Phe6-D-Ala10]-LH-RH, 1 mg/day; or (3) vehicle. Whereas control animals showed normal menstrual cycles, as determined by dates of ovulation, length of luteal phases, and hormonal profiles, animals treated with either analog of LH-RH exhibited disruption of the cycles. In animals from both groups, delayed ovulation was observed (LH-RH agonist, 22, 23, 17, 19, and 20 days of the cycle; LH-RH antagonist, 22, 22, 24, and 10 days of the cycle, and one animal remained anovulatory for 65 days). Monkeys treated with either LH-RH analog showed normal luteal phase lengths (15, 15, 16, 14, and 15 days, and 15, 16, 16, and 13 days, respectively) and serum progesterone concentrations. The results of this study suggest that, in the rhesus monkey, the administration of LH-RH analogs during the early follicular phase induces a temporary cessation of folliculogenesis demonstrated by a delay of ovulation, with subsequent normal luteal function.

Gamete intrafallopian transfer (GIFT) was performed in eight patients with premature ovarian failure (POF), using donated oocytes. The steroid replacement protocol consisted of the administration of increasing dosages of 17 beta-estradiol (E2) and progesterone (P). Hormonal replacement was maintained until day 100 of gestation. All patients underwent an evaluation cycle in which serum levels of E2 and P were monitored and an endometrial biopsy was performed on day 21 or 22. All cases of GIFT were performed between days 12 and 15. Six clinical pregnancies were achieved in eight cycles (75% success rate). Three patients delivered and three are in their second or third trimester. No ectopics or miscarriages occurred. These results offer a promising approach for the establishment of fertility in agonadal patients.

The concentrations in follicular fluid (FF) of beta hCG, progesterone (P), estradiol (E2), androstenedione (A) and testosterone (T) by RIA were determined, in order to study the follicular micro-environment in stimulated cycles. Patients received either clomiphene with hMG plus hCG or hMG plus hCG. With increasing volume of FF (obtained at laparoscopy), P significantly increased (p less than 0.04) and hCG (p less than 0.0004) and T (p less than 0.05) significantly decreased with a borderline decrease in A (p less than 0.07). Progesterone also increased with advancing oocyte maturity. Beta-hCG and P were positively correlated with E2 and negatively correlated with A levels. It is suggested that FF hCG and progesterone levels may be markers of follicular development in stimulated cycles.

This article describes the first series of patients to undergo gamete intrafallopian transfer (GIFT) as a treatment for infertility. Ten patients with the diagnosis of either unexplained infertility or male factor were treated with human menopausal gonadotropin and human chorionic gonadotropin before surgery by laparoscopy or minilaparotomy. Semen was collected 2.5 hours before oocyte pickup at surgery and treated by the technique of wash and swim-up. After gamete evaluation, one or two oocytes and 100,000 actively motile sperm were loaded into a catheter and introduced through the fimbria. The contents of the catheter were gently emptied at a site approximately 1.5 cm inside each fallopian tube. Patients received progesterone in oil, 12.5 mg/day, from day 4 after GIFT until up to 8 weeks of gestation. Four patients became pregnant: two pregnancies aborted; the other two pregnancies proceeded to the delivery of viable infants. GIFT may be considered as an alternative to in vitro fertilization in infertility cases in which at least one fallopian tube is patent.

Pituitary gonadotropin response to luteinizing hormone-releasing hormone (LH-RH; 150 micrograms, administered intravenously) was studied during pregnancy and the early postpartum period in rhesus monkeys. Challenge tests were performed approximately every three weeks during pregnancy and every two weeks after delivery, in breast-feeding and non breast-feeding animals. It was found that the response to the releasing factor was completely abolished as early as day 20 after conception. This absence of pituitary responsiveness to LH-RH continued throughout pregnancy. The return of the response during the puerperium occurred as early as day 15 post-delivery in non-breast-feeding monkeys, but remained abolished up of the response to LH-RH in FSH and LH concentration was observed, in that LH levels increased 70%-90%, whereas FSH levels increased only 20%-30%. Serum prolactin (PRL) levels in breast-feeding animals presented a slight increase the first week post-delivery and thereafter, and decreased gradually until they reached normal levels four weeks post-delivery. In contrast, concentrations of serum PRL in non-breast-feeding animals decreased abruptly to normal levels within the first week post-delivery. These results demonstrate that, due to its similarity to the human, the rhesus monkey is an excellent animal model in which to study pituitary function during pregnancy and the puerperium.

The performance of the gamete intrafallopian transfer (GIFT) technique, utilizing an individualized regimen of follicular induction and minilaparotomy in 45 patients with infertility of varying etiologies is reported. The induction regimen consisted of the administration of clomiphene citrate, 100 mg, from day 3 to day 7, and 150 IU FSH/LH (human menopausal gonadotropin [hMG]) from day 6 on. Human chorionic gonadotropin (hCG; 10,000 IU) was administered when at least two follicles measured 16 mm or more in diameter and when serum estradiol (E2) measurement revealed levels of 350 pg/ml per each main follicle. Up to two oocytes and 100,000 motile sperm were transferred to the fallopian tubes via the fimbria, using a catheter. Of the 45 cases, 13 became pregnant by clinical criteria (29 per cent). Of these 13 pregnancies, nine continued to term (69 per cent), three miscarried spontaneously (12 per cent) and one was an ectopic (7 per cent). Of the nine pregnancies that continued to term, five (55 per cent) were twins. Details of the GIFT procedure as well the preliminary non-human primate research studies that led to the development of the GIFT technique are discussed. It is concluded that GIFT is an excellent alternative to in vitro fertilization and embryo transfer (IVF; ET) in all cases of infertility that failed to conceive using conventional forms of therapy and in which the female partner presents at least one normal fallopian tube.