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D. C. JOHNSON

The induction of a persistent vaginal cornification—anovulatory syndrome (androgenization) following steroid treatment of the neonatal female rat has been well documented (reviewed by Johnson, 1972). The exact timing of a `critical period' for the induction of this syndrome has not been determined but, in males, the gonads must be removed within 48 hr of birth in order to prevent their effects upon the hypothalamic—hypophysial axis. The upper limit in time for androgen to have any permanent effects is generally considered to be Day 10. While early (Days 1 to 3) postnatal treatment with testosterone can seriously interfere with normal vaginal development, this is clearly the most efficient time for induction of androgenization (Alklint & Norgren, 1971). Nearly all comparative studies involving a variety of steroids have used Day 5 for treatment,

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D. C. Johnson

Summary. Rats were androgenized by injection of 50 μg testosterone propionate on the 5th day after birth and when adult were treated with 5 i.u. PMSG; some of the animals were mated. Serum was obtained daily and the concentrations of progesterone, 20α-dihydroprogesterone and prolactin, estimated by radioimmunoassays, were compared to values found for mated, but not ovulating, androgenized females and those for normal pregnant females.

Ovulation and luteinization of follicles occurred. The concentration of progesterone increased after the injection of PMSG and remained elevated for at least 10 days; mating did not alter the progesterone levels. The concentration of 20α-dihydroprogesterone was also elevated but the ratio of the level of progesterone to this steroid was generally greater than unity. Prolactin levels were elevated in the rats which ovulated. It is concluded that the corpora lutea induced in androgenized females by PMSG are functional and maintained.

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D. C. Johnson and S. Chatterjee

The present study was undertaken to determine whether the mechanism of embryo transfer is a factor in the action of epidermal growth factor (EGF) in initiation of implantation. Unilateral intrauterine infusion of 3 μl buffered saline, or saline containing 1.5 μg EGF, plus i.v. injection of 100 μg EGF 2 h later resulted in implantation sites in all animals within 48 h. In several animals implantation was also initiated in the non-injected uterine horn. Administration of indomethacin 1 h before the intrauterine injection completely blocked the effect of EGF but not that of 25 ng oestradiol. The results confirm that EGF can replace oestrogen for initiation of implantation provided that the uterine trauma associated with embryo transfer, that is puncture, is provided. The mechanisms involved remain to be resolved.

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P. C. TURNER and A. D. JOHNSON

Summary.

The efferent ducts of male rats were ligated and severed unilaterally to prevent passage of materials into the epididymis. The contralateral testis-epididymis complex served as control. Total lipid, free, total and esterified cholesterol, phospholipid and triglyceride were analysed on both contralateral control and isolated tissues at 1, 7, 14, 21, 28 and 35 days after surgery. Histological examination of the surgically isolated epididymides revealed the absence of spermatozoa in the caput at 1 day, corpus at 7 days and cauda at 21 days after surgery. With removal of spermatozoa from an area, lipid fractions tended generally to increase in concentration. Free cholesterol, as the percentage of cholesterol, was far lower than has been reported in the testis.

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S. K. Dey and D. C. Johnson

Summary. The effect of adrenal steroids upon implantation was evaluated by examining the efficacy of oestradiol-17β on the initiation of implantation in ovariectomized, ovariectomized plus adrenalectomized or hypophysectomized pregnant rats treated with progesterone. More oestrogen (× 5) was required in ovariectomized animals to obtain results equivalent to those obtained with the other animal models.

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S. K. Dey and D. C. Johnson

Summary. Mouse embryos recovered on the 4th day of pregnancy produced histamine, as evidenced by the14CO2 produced from carboxy labelled l-histidine, at the rate of 1·5 ± 0·3 (s.e.m.) pmol/embryo per hour. Most (83·2 ± 4·6%) of the embryos flushed from the oviducts on Day 3 of pregnancy (4–8-cell stage) developed into blastocysts within 48 h after being placed in culture. Inclusion of l-histidine hydrochloride (4·7 × 10−4 m) in the culture medium did not alter this development but dl-α-methylhistidine (3·8 × 10−4 m), an inhibitor of histidine decarboxylase, reduced the number of embryos developing into blastocysts to only 10·8 ± 6·8%. A combination of l-histidine and dl-α-methylhistidine in the medium prevented the growth-retarding effect of the latter compound. The results indicate that mouse embryos can produce histamine and suggest that this is necessary for normal development.

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M. A. KRAMEN and D. C. JOHNSON

Departments of Obstetrics and Gynecology and Physiology, University of Kansas Medical School, Kansas City, Kansas, U.S.A.

(Received 26th June 1974)

There has been little success in inducing a decidual reaction in the uteri of adult female rats given androgen neonatally. Burin, Thevenot-Duluc & Mayer (1963) reported a positive reaction after scratching the antimesometrial lining and giving progesterone (5 mg) daily for 8 days, but the incidence of success, the age of the animals, and the criteria for decidualization were not given. Barraclough (1967) reported that 25% of adult females given HCG to induce ovulation, and reserpine to sustain pseudopregnancy, produced a positive decidual reaction. All other reports are negative. The reasons for failure are not clear. Lack of oestrogen-binding capacity has been documented (McGuire & Lisk, 1969) and may play an important rôle; the availability of progesterone receptors in the uteri of such females is unknown.

In the present study,

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R. C. Hoversland, S. K. Dey and D. C. Johnson

Summary. Rabbit blastocysts were homogenized by sonication, and centrifuged at 105 000 g for 60 min. The pellet was resuspended and incubated in phosphate buffer containing [1β-3H]testosterone and a NADPH generating system. The amount of 3H2O produced was determined by liquid scintillation counting. Enzyme activity was calculated, after subtracting blank values obtained with boiled embryos, and expressed as pg testosterone aromatized per embryo per hour. Aromatase activity was undetectable to low on Day 5 and increased on Day 6 of pregnancy. There was a 10-fold increase in activity in Day-6 embryos cultured for 24 h, with a further 6-fold increase in activity in Day-6 embryos cultured for 48 h. The enzyme had an apparent K m of 0·77 μm and was completely inhibited by an aromatase inhibitor. The results clearly indicate that the rabbit blastocyst has an increasing capacity for aromatization of testosterone at about the time of implantation.

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S. Long, S. K. Dey and D. C. Johnson

Summary. An intravenous injection of 2-fluoro-oestradiol simultaneously with an implantation-inducing dose of oestradiol reduced the number of implantation sites in delayed implanting hypophysectomized rats maintained with progesterone. Administration of 2-fluoro-oestradiol 1 h before or after oestradiol had no effect. Furthermore, injection of as much as 500 ng 2-fluoro-oestradiol 48 h before administration of oestradiol failed to have any effect upon implantation, i.e. failure to block implantation was correlated with failure to induce the uterine refractory state. These results suggest that conversion of primary oestrogens to catechol oestrogens could be important for implantation as well as for the induction of the oestrogen refractory state in the uterus.

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J. T. WU, Z. DICKMANN and D C. JOHNSON

Summary.

The beneficial effect of progesterone on blastocyst survival in the uterus was shown in rats hypophysectomized on Day 1 of pregnancy. Treatment with progesterone (2 mg/day) on Day 1, Day 1 and Day 6, or Days 1 to 5 could maintain most blastocysts for only 5 days following the last injection; during the next 5 days, many disintegrated in the uterus. Daily injections of 2 mg progesterone, however, could maintain most blastocysts for as long as Day 53.

In hypophysectomized rats, the developmental potential of blastocysts deteriorated rapidly even though the rats received 2 mg progesterone daily. By Day 20, only 23% of blastocysts were capable of developing into full-term fetuses compared with 52% in rats ovariectomized on Day 4 and injected daily with 2 mg progesterone. By Day 52, none of the blastocysts from either group of animals developed to term although many still retained the ability to implant.