In order to elucidate the mechanism of the resumption of follicular activity and ovulation in rats, levels of FSH, LH and prolactin in plasma and pituitary gland and ovarian follicular development were quantified after removal of the litter on day 3 of lactation (day of parturition = day 0 of lactation). Such removal resulted in ovulation of 13 oocytes 4 days later, a number comparable with that found in normal cyclic rats. Plasma levels of prolactin were high during lactation but markedly decreased after removal of the litter. Although plasma concentrations of FSH and LH did not change during days 3–7 of lactation, there was an FSH surge between 24 and 30 h after removal of the litter. Plasma concentrations of LH also increased slightly but significantly by 24 h after removal of the litter and this value persisted during the following 2 days. Surges of FSH, LH and prolactin occurred at 17.00 h 3 days after pups were removed. Removal of the litter did not increase pituitary contents of FSH, LH and prolactin and a marked reduction in pituitary levels of FSH and LH, but not of prolactin, occurred at 17 00 h 3 days after removal of the litter.

A quantitative study of follicular development indicated that follicles larger than 401 μm in diameter were absent during days 3–7 of lactation. However, the number and size of antral follicles increased by 30 h after removal of the litter, probably due to the increases in plasma levels of FSH and LH, and follicles larger than 601 μm in diameter appeared 3 days after the young were removed. Although ovulation could not be induced by human chorionic gonadotrophin from days 3 to 5 of lactation, its administration 30 h after removal of the litter produced ovulation in all rats by the following morning.

These results indicated that a moderate increase in FSH, although below the amounts released at the preovulatory surge, together with basal levels of LH which were within the range observed on the day of dioestrus during the normal cycle were responsible for the initiation of follicular maturation after removal of the litter.

A selective surge of FSH with a small concomitant rise in LH occurred invariably in rats when ovulation was induced by injecting human chorionic gonadotrophin (HCG) at various reproductive stages such as day 15 of lactation and in 29-day-old immature rats as well as in dioestrous animals. No FSH surge occurred on day 3 of lactation or in 26-day-old immature rats in which ovulation could not be induced by HCG. The FSH surge occurred 6–18 h after HCG treatment regardless of the time of day of injection of HCG. Ovulation began by 12 h and was completed by 18 h after injection of HCG.

Pituitary responsiveness to luteinizing hormone releasing hormone (LH-RH) with respect to FSH release strikingly increased at 01.00 h on day 1 after HCG injection at 17.00 h of dioestrus (day 0) to levels similar to those of the group at 01.00 h of oestrus, when the greatest response was noted during the normal cycle. With regard to LH release pituitary responsiveness to LH-RH at 01·00 h on day 1 markedly increased but the response was only about half of the response at 01·00 h of oestrus and one third of the response at 17.00 h of pro-oestrus when the greatest response was noted during the normal oestrous cycle.

These results indicate that during ovulation the pituitary gland of the rat is highly responsive to LH-RH with respect to the release of FSH, for which secretory changes in the ovary after an ovulating dose of HCG may be responsible.

When 1·0 μg luteinizing hormone releasing hormone (LH-RH) was given i.v. three times at 1 h intervals from 17.00 to 19.00 h on the day of dioestrus (day 0) to regular 4 day cyclic rats, premature ovulation was induced the next morning (day 1) with the number of ova present comparable to normal spontaneous ovulation. The next spontaneous ovulation occurred on the morning of day 5, 4 days after premature ovulation induced by LH-RH.

Plasma concentrations of FSH and LH showed transient rises and falls within 1 h of administration of LH-RH; concentrations of FSH in the plasma decreased from 20.00 h on day 0 but markedly increased again from 23.00 h on day 0 to 02.00 h on day 1 and these high levels persisted until 14.00 h on day 1, with only a small increase of plasma LH during this period. The duration of increased FSH release during premature ovulation induced by LH-RH treatment was 6 h longer than the FSH surge occurring after administration of HCG on day 0. Surges of gonadotrophin were absent on the afternoon of day 1 (the expected day of pro-oestrus) and the surges characteristic of pro-oestrus occurred on the afternoon of day 4 and ovulation followed the next morning. The pituitary content of FSH did not decrease despite persisting high plasma levels of FSH during premature ovulation induced by either LH-RH or HCG on day 0.

The changes in uterine weight indicated that the pattern of oestrogen secretion from the day of premature ovulation induced by LH-RH to the day of the next spontaneous ovulation was similar to that of the normal 4 day oestrous cycle. When 10 i.u. HCG were given on day 0, an increase in oestrogen secretion occurred on day 2, 1 day earlier than in the group given LH-RH on day 0. This advancement of oestrogen secretion was assumed to be responsible for the gonadotrophin surges on day 3.

Similar numbers of fully developed follicles were found by 17.00 h on day 2 after premature ovulation induced by either LH-RH or HCG, suggesting that the shorter surge of FSH during premature ovulation induced by HCG had no serious consequences on the initiation of follicular maturation for the succeeding oestrous cycle in these rats.

Administration of LH-RH on day 0 had no direct effect on the FSH surge during premature ovulation. Secretory changes in the ovary during ovulation may be responsible for this prolonged selective release of FSH.

Laboratory of Veterinary Physiology, Tokyo University of Agriculture and Technology, Fuchu, Tokyo 183, Japan

(Received 2 May 1977)

When an amount of human chorionic gonadotrophin (HCG) sufficient to cause ovulation is given to 4-day cyclic rats on the day of dioestrus, premature ovulation is induced the next morning (Eto & Imamichi, 1955). The pattern of release of follicle-stimulating hormone (FSH) responsible for the initiation of follicular maturation of the next set of follicles (Schwartz, 1969; Welschen & Dullaart, 1976) after HCG-induced ovulation has not been previously evaluated. The present communication is concerned with this problem and indicates that a large amount of FSH is released within 12 h of administration of HCG, with only a small concomitant rise in the concentration of luteinizing hormone (LH).

Adult female Wistar rats were maintained under a 14 h light : 10 h darkness schedule (lights on 05.00 h), and those showing three or

To investigate the mechanism of the selective surge of FSH during the period of ovulation induced by human chorionic gonadotrophin (hCG) in dioestrous rats, inhibin activity in ovarian vein plasma was determined at varying time-intervals after treatment with hCG using the primary monolayer culture system of anterior pituitary cells.

Inhibin activity in ovarian vein plasma had already decreased 6 h after injection of hCG, when concentrations of FSH in the plasma were still low in three of four animals. Inhibin activity further decreased 12–18 h after hCG, when a selective surge of FSH occurred. Inhibin activity increased to the level before hCG treatment 24 h after the treatment, when ovulation was completed and the FSH surge terminated.

These results suggest that the selective surge of FSH occurs as a consequence of the decrease in inhibin secretion from the ovary, which is perhaps due to the ovulation dose of hCG altering the functional activity of the granulosa cells in the large Graafian follicles.

The effects of 3-methyl-4-nitrophenol (PNMC), a component of diesel exhaust, on reproductive function were investigated in adult male Japanese quail. The quail were treated with a single i.m. dose of PNMC (78, 103 or 135 mg/kg body weight), and trunk blood and testes were collected 1, 2 or 4 weeks later. Various levels of testicular atrophy were observed in all groups treated with PNMC. Sperm formation, cloacal gland area, and plasma LH and testosterone concentrations were also reduced in birds with testicular atrophy. To determine the acute effect of PNMC on gonadotrophin from the pituitary, adult male quail were administrated a single i.m. injection of PNMC (25 mg/kg), and plasma concentrations of LH were measured at 1, 3 and 6 h. This dose significantly lowered plasma levels of LH at all three time points. These results suggest that PNMC acts on the hypothalamus–pituitary axis, by reducing circulating LH within a few hours of administration and subsequently reducing testosterone secretion. In addition, in order to investigate the direct effects of PNMC on the secretion of testosterone from testicular cells in quail testes, cultured interstitial cells containing Leydig cells were exposed to PNMC (10⁻⁶, 10⁻⁵ or 10⁻⁴ M) for 4, 8 or 24 h. These quantities of PNMC significantly reduced the secretion of testosterone in a time- and dose-dependent manner. The present findings also suggest a direct effect of PNMC on the testis to reduce testosterone secretion. This study clearly indicates that PNMC induces reproductive toxicity at both the central and testicular levels, and disrupts testicular function in adult male quail.

To elucidate changing patterns of inhibin/activin subunit mRNAs in the ovary of the golden hamster (Mesocricetus auratus) during the oestrous cycle, inhibin/activin subunit cDNAs of this species were cloned and ribonuclease protection assay and in situ hybridization were carried out. Inhibin α-subunit mRNA was localized in granulosa cells of primary, secondary, tertiary and atretic follicles throughout the 4-day oestrous cycle. It was also expressed in luteal cells on days 1 (oestrus), 2 (metoestrus) and 3 (dioestrus). βA-subunit mRNA was localized in granulosa cells of large secondary (>200 μm) and tertiary follicles throughout the oestrous cycle. βB-subunit mRNA was confined to granulosa cells of large secondary and tertiary follicles. Both α- and βA-subunit mRNAs were also found in ovarian interstitial cells and theca interna cells of tertiary and atretic follicles in the evening of day 4 (pro-oestrus). A striking increase in βA-subunit mRNA levels was also observed during the preovulatory period. The expression pattern of βA-subunit mRNA during the preovulatory period is unique and not found in other species. An i.v. injection of anti-luteinizing hormone-releasing hormone (LHRH) serum before the LH surge abolished the expression of α- and βA-subunit mRNAs in ovarian interstitial cells and theca interna cells. The treatment also abolished the preovulatory increase in βA-subunit mRNA. Furthermore, administration of human chorionic gonadotrophin (hCG), which followed the injection of anti-LHRH serum, restored the expression patterns of α- and βA-subunit mRNAs. The present study revealed that the golden hamster showed a unique expression pattern of βA-subunit mRNA in response to the LH surge.