Trends in causes and treatment of infertility at Flinders Medical Centre, Adelaide, 1976-1989.

OBJECTIVE: To describe the causes of infertility among couples presenting to a fertility clinic over a period of 14 years. Changes in treatment methods over the same period are described. DESIGN: A retrospective analysis of records of patient age, date of first visit, duration and cause of infertility, treatment and outcome. SETTING: A fertility clinic set in a large public hospital. Patients are referred by both general gynaecologists and general practitioners. PATIENTS: From 1976 to 1989 (inclusive) 2895 couples who had engaged in unprotected intercourse for at least 12 months without conceiving attended the clinic. RESULTS: Three major diagnostic categories were considered; these embraced aetiological factors relating to semen, ovulation, and pelvic anatomy. Investigation of the causes of infertility after a couple's first visit showed these to be a semen defect in 22%, irregular cycles or anovulation in 29%, and a pelvic disorder in 57%. The incidence of semen defects as the cause remained constant between 1976 and 1989 while that of ovulation "problems" declined from 42% in 1976 to 13% in 1989. In contrast, the incidence of pelvic dysfunction increased from 40% in 1976 to 68% in 1989, predominantly due to an increased diagnosis of endometriosis. The principal treatments were ovulation induction/cycle regulation (51%), in-vitro fertilisation/gamete intrafallopian transfer (42%), and donor insemination (18%). CONCLUSIONS: Provided patient information is accurately and consistently recorded a Fertility Audit is a useful window on the activates of a Fertility Clinic and serves to highlight areas which warrant further investigation.

GIFT or IVF for patients with patent fallopian tubes?

The pregnancy rate in a group of 39 couples who underwent in vitro fertilization (IVF) with donor sperm was compared with that of couples undergoing gamete intra-Fallopian transfer (GIFT) over the same period (31 December 1986 to 30 June 1991). The diagnostic status of the female partners in each group was qualitatively similar. The women had patent Fallopian tubes, mild endometriosis or minor tubal adhesions. The male partners in the GIFT group had normal or marginally abnormal semen parameters which were considered to be comparable to the post-thaw quality of the semen used in donor insemination (DI). The mean age of the women in the DI group was 32 years (range 25-40) which was similar to that in the GIFT group [32 years (24-39)]. The women in the IVF group had undergone a mean of eight cycles of DI before proceeding to treatment with IVF. There was no difference between the pregnancy rate in the two groups (log-rank test). These observations advance the proposition that IVF should be reconsidered as an effective treatment for women with patent Fallopian tubes.

Perifusion of human granulosa-luteal cells: response to LH stimulation.

LH pulse frequency and amplitude vary significantly during the menstrual cycle; however, it is not clear what significance the secretory pattern has for the ovary. We have developed an in-vitro perifusion system in which luteinized human granulosa cells (GC) can be exposed to various patterns of gonadotrophin stimulation. GC were recovered following follicle aspiration for in-vitro fertilization, grown on Cytodex-3 for 6 days, and then perifused with medium containing LH (or hCG), delivered with differing pulse frequencies and amplitudes. When pulses of LH were applied to the cells, progesterone secretion rose initially and then fell to the baseline as the LH concentration declined. Pulsatile administration of LH, over a period of 10 h, stimulated progesterone secretion more efficiently than did continuous LH. Finally, delivery to the cells of pulses of hCG, a ligand known to bind to the LH receptor but with binding characteristics distinct from those of LH, failed to elicit pulses of progesterone.

Localization of relaxin binding sites in the rat uterus and cervix by autoradiography.

125I-labeled porcine relaxin was injected into 27-day-old rats treated with pregnant mare's serum gonadotropin (PMSG) and known target tissues for relaxin, the myometrium, endometrium and cervix, and putative control tissues, heart, thigh muscle and duodenum, examined for binding by autoradiography. Specific binding in the target tissues was demonstrated by simultaneous injection of excess unlabeled relaxin. Radioactivity was located and quantified by grain counts predominantly over the inner, circular muscle layer of the myometrium and the cervix and to a lesser extent over the outer longitudinal muscle layer of the myometrium and the endometrium. The route of injection, the circulation time, or counting grains in transverse or longitudinal sections of myometrium made little difference in these results. Ovariectomy decreased, but not significantly, the grain count in all of the target tissues studied and estrogen treatment of ovariectomized animals restored the numbers of grains to approximately that of intact PMSG-treated rats. The degree of binding of the cervix was approximately that of the circular myometrial muscle. This work confirms the presence of specific receptors for relaxin in the rat uterus and cervix of primed rats and it also suggests that the inhibitory action of relaxin upon the myometrium is primarily on the inner circular muscle layer.

Relaxin stimulates plasminogen activator secretion by rat granulosa cells in vitro.

Granulosa cells isolated from the ovaries of pregnant mare serum gonadotropin (PMSG)-treated immature rats were cultured with relaxin or FSH. Both hormones increased the secretion of plasminogen activator into the culture medium. Relaxin caused a dose-related rise in plasminogen activator but did not increase cAMP or progesterone levels in the medium of freshly harvested granulosa cells, although they were responsive to FSH. This ability of relaxin to stimulate plasminogen activator synthesis without progesterone or cAMP rises indicates that the pathways of post-receptor events leading to stimulation of plasminogen activator differ markedly from those of the gonadotropins. Relaxin is thus a fully characterized peptide hormone produced by the ovary with a well-defined action upon the granulosa cell and may have an intraovarian role in the events leading to ovulation.

Modulation of functional capacity of small ovarian follicles in the post-partum cow by prolactin.

This study was undertaken to examine the possibility that the prolonged anovulatory period frequently experienced by the post-partum cow is due to a disruption of function at the ovarian level promoted by the high, suckling-induced, blood prolactin concentrations. Fifteen cows, less than 35 days post partum, were allocated to three groups (1, 3 and 5) and given no hormonal treatment, prostaglandin plus pregnant mare serum gonadotrophin (PMSG) treatment or injected with 2-bromo-alpha-ergocryptine to reduce circulating prolactin levels. Ten synchronized cyclic cows were allocated to two groups (2 and 4) and given prostaglandin or prostaglandin plus PMGS treatment. All cows were ovariectomized 1 or 2 days after treatment of Graafian follicles less than 9 mm in diameter were selected after dissection from the ovaries. The follicles were cultured for 18 h with or without prolactin (1 microgram/ml) and steroid accumulation in the culture medium estimated. The follicles were then separated into theca and granulosa which were incubated for 40 min with LH (1 microgram/ml) or FSH (5 micrograms/ml). Cyclic AMP concentrations were estimated as an indication of tissue responsiveness to gonadotrophins. The secretion of oestradiol-17 beta, progesterone, testosterone or androstenedione during 18 h culture did not differ between follicles isolated from post-partum or cyclic cows. The presence of prolactin in the culture medium had no overall effect on steroid secretion although some specific effects within each group were noticed. Incubation with LH increased cyclic AMP levels in the theca but the granulosa did not respond. Likewise FSH increased cyclic AMP levels in granulosa preparations but not in theca. There were no differences in response between post-partum and cyclic cows, but exposure of the follicles to prolactin in vitro did significantly reduce the LH-induced increase in cyclic AMP levels in isolated theca. We have concluded that endogenous prolactin may modify but does not inhibit the resumption of ovarian function following parturition in the beef cow.

Hormonal and cellular interactions in follicular steroid biosynthesis by the sheep ovary.

Studies of isolated cell types from sheep follicles revealed several functional changes which occur during follicular maturation. Cyclic AMP production by granulosa cells from the smallest follicles studied (1-3 mm diameter) was stimulated by FSH but not by hCG, suggesting functional FSH receptors at this early stage of differentiation. Medium-sized follicles (4-6 mm) responded to both FSH and hCG. Granulosa cells were unable to synthesize androgens, but readily converted exogenous testosterone to oestradiol-17 beta. This conversion occurred to a limited extent in the cells from the smallest follicles, but was much greater in medium and large (greater than 6 mm) follicles. Oestradiol production by theca preparations from small follicles was barely detectable, but increased markedly with increasing follicle size. Androgen (androstenedione and testosterone) production by theca preparations was stimulated by hCG. This stimulation was short-lived, and levels declined to below control values after 6 h of culture. This decline could not be prevented by addition of cyclic AMP. The presence of granulosa cells with thecal preparations (i.e. follicle wall tissue) enhanced production of androgen by the theca, the effect being more marked for testosterone than for androstenedione. In-vivo studies in which granulosa cells and follicular fluid were removed during the preovulatory period suggested that granulosa cells and/or follicular fluid contributed to the oestradiol secreted into the ovarian vein during this period, but did not exclude a significant contribution by the theca as well.

Suppression of follicular cAMP and steroid response to gonadotrophins by pretreatment with gonadotrophin.

This study examines the effect of preincubation of sheep ovarian follicles with hCG or FSH on the ability of isolated theca and granulosa to increase cAMP levels in response to a second incubation with hCG or FSH. Secondly, the effect of preincubation of small follicles with hCG on androstenedione and testosterone production is examined in intact follicles. Preincubation with hCG resulted in a decrease in the ability of theca from large (4--6 mm in diameter) and small (1--3 mm) follicles and granulosa from large follicles to increase cAMP levels in response to a second exposure to hCG. Similarly, preincubation with FSH suppressed the response of isolated granulosa from small and large follicles to a second exposure to FSH. A partial, but significant inhibition of the response to the other hormone was observed after preincubation with both FSH and hCG. Intact small follicles pretreated with hCG were unable to increase testosterone secretion in response to a second exposure to hCG.

Effects of human chorionic gonadotrophin on the release of cyclic AMP and progesterone from the sheep ovary at different stages of the oestrous cycle.

The rate of release of cyclic AMP by sheep ovaries containing a corpus luteum was determined at different stages of the cycle before and up to 60 min after an intra-arterial (ia) injection of 500 IU human chorionic gonadotrophin (hCG). The median cyclic AMP concentration in arterial plasma and of ovarian venous plasma following hCG stimulation was 93.2 and 98.0 pmol/ml, respectively. The ovaries of ewes examined at Days 1 and 2 of the cycle showed no response to hCG, whereas in 2 sheep at Day 3, hCG caused a slight response, and in 13 sheep examined between Days 5-18, hCG caused a marked increase in cAMP release. In 5 of the sheep in which both ovarian veins were cannulated, only the ovary with a corpus luteum responded to hCG with an increased secretion rate of cyclic AMP and progesterone. The results indicate a lack of responsiveness in the newly formed corpus luteum to hCG.

Maturational changes in sheep ovarian follicles: gonadotrophic stimulation of cyclic AMP production by isolated theca and granulosa cells.

Theca and granulosa tissues isolated from sheep ovarian follicles of different sizes were incubated in the presence of human chorionic gonadotrophin (HCG; 5 IU/ml) or follicle stimulating hormone (FSH; 5 microgram NIH-FSH-S11/ml) for 40 min. Changes in the total amounts of cyclic 3',5'-adenosine monophosphate (cAMP) were used as an index of the responsiveness of these preparations to the hormones. Thecal tissue of both large (4-6 mm in diameter) and small (1-3 mm) follicles responded similarly to gonadotrophins. Granulosa cells from small follicles failed to respond to stimulation by HCG. FSH, however, consistently increased cAMP production in comparison with controls or cells treated with HCG. Granulosa cells of large follicles responded to both HCG and FSH.

Cyclic AMP in sheep ovarian follicles: site of production and response to gonadotrophins.

This study was undertaken (i) to establish a relationship between cyclic AMP (cAMP) production and the degree of LH and FSH stimulation; (ii) to determine the effects of various gonadotrophins on follicular formation of cAMP; and (iii) to identify the precise intrafollicular site of cAMP formation. The formation of cAMP increased rapidly in follicles exposed to LH. Maximum concentrations were reached after 90 min and were maintained for 180 min. Extracellular release of cAMP increased steadily throughout the 180-min experimental period. Tissue levels of cAMP increased proportionally and significantly when LH concentrations in the medium were increased from 0 to 200 mi.u. ml-1. Tissue levels of cAMP were significantly increased by HCG, prostaglandin E-2 and noradrenaline, but not by prolactin, prostaglandin F-2alpha, serotonin or melatonin. Cyclic AMP formation occurred predominantly in the thecal compartment; the membrana granulosa contributed less than 3% of the total amount of cAMP formed after gonadotrophic stimulation. A significant amount of cAMP from the thecal cells was released into the extracellular compartment and appeared to pass into the granulosa cells.