Unilateral compensatory hypertrophy of the gonad after hemicastration has been extensively studied in males and females (reviews by Grant, 1956; Edgren, Parlow, Peterson & Jones, 1965; McClaren, 1966; Peppler, 1969). The mechanisms responsible for the compensatory growth are, however, as yet, undefined. One theory suggests that the loss of a gonad results in a decrease in circulating steroids which reduces their negative feedback effect on the hypothalamic-hypophysial axis and consequently triggers an increased outflow of gonadotrophins. An alternative view is that the remaining gonad is stimulated by more gonadotrophin because of a decrease in the rate of utilization. With either of these possibilities the amount of circulating gonadotrophin should increase over the normal intact level. However, presently available bioassay techniques have not shown any changes in plasma or pituitary gonadotrophins in the hemiovariectomized female rat (Edgren et al. 1965; McClaren, 1966). Previous experiments with parabionts (Johnson, 1966) indicated that unilateral
D. C. JOHNSON
The serum concentrations of FSH, LH and prolactin, as well as oestradiol and progesterone, were measured by immunoassays at various intervals up to 35 weeks after parabiotic union of an anovulatory androgenized female and a castrated male rat of the Fischer-344 strain. Modest increases in FSH (28%) and larger increases in LH (128%), compared to levels found in single adult animals, produced large increases in follicular growth, ovarian weight and oestrogen production in the female partners; thecal and interstitial elements of the ovaries were not heavily stimulated. Prolactin concentrations in the sera increased to exceedingly high levels and massive pituitary tumours were produced in all of the female partners. Development of cystic follicles with loss of granulosa-cell layers and the formation of early stages of granulosa-cell tumours progressed with time in union. Serum LH levels were as high in parabiotic as in single castrated male rats within 2 weeks, but concentrations of FSH remained much below those of the single animals until 27 weeks after castration. The results are consistent with the view that the ovaries of androgenized female rats of the Fischer strain produce some factor which prevents the post-castration rise in FSH of parabiotic male partners.
D. C. JOHNSON and TATSURO SASHIDA
Pregnant mare serum gonadotrophin given intravenously to immature rats caused a maximal (×70) increase in ornithine decarboxylase activity (ODC) at 3 h; enzyme activity declined to about ten times the control level by 9 h and a second rise began after about 20 h. Anti-PMSG given 30 min after PMSG reduced the peak response by 70%. Actinomycin D, or cycloheximide, completely prevented an increase in ODC when given with PMSG, but only cycloheximide lowered the enzyme activity when given 18 h later.
Ovine FSH plus LH also produced a peak in ODC at 3 h but the activity decreased quickly and by 9 h it was at the control level. Secretion of endogenous FSH and LH, induced by hourly injections of LH releasing hormone (LH-RH) increased ODC to the same extent as did the exogenous hormones; ODC was still higher than in the controls 4 h after the last dose of LH-RH.
Increased endogenous levels of FSH and LH did not consistently raise ovarian cyclic AMP content and the increases found were much less than those obtained after injection of PMSG or FSH + LH.
The results indicate that increased ODC is induced and maintained by the continual presence of gonadotrophin. The dependence of increased ODC upon increased cyclic AMP cannot be unequivocally determined because of important differences in the timing of the responses and the difficulty in determining biologically significant changes in cyclic AMP.
D. C. JOHNSON and E. WITSCHI
Immature male rats were (1) exposed to X-rays, or (2) made cryptorchid, or (3) subjected to a combination of the two and placed in parabiosis with castrated males. Pairs were autopsied at various intervals up to 46 weeks and effects upon gonads and accessory sex organs evaluated.
Androgen production in response to endogenous gonadotrophins from a castrated animal is greater in irradiated than cryptorchid testes. The stresses imposed upon irradiated, intra-abdominal testes greatly reduces their capacity to produce androgen in response to gonadotrophins.
D. C. Johnson and M. Sen
The cytochrome P450 responsible for androgen synthesis by the placenta during the second half of pregnancy in the rat was studied in intact and hypophysectomized animals. The two activities of P45017α, 17α-hydroxylase and C17,20-lyase, were limited to the junctional zone.
C17,20-Lyase activity was greater with progesterone than with 17-hydroxyprogesterone as substrate. Although the apparent Michaelis constants were similar, progesterone had a higher maximum velocity than 17-hydroxyprogesterone. Regardless of substrate, C17,20-lyase activity was greater with NADPH than with NADH as an electron donor, and there was no additive effect using both cofactors.
Administration of human chorionic gonadotrophin (hCG; 10IU at 09.00 and 21.00 h on days 13 and 14 and at 09.00 h on day 15) to intact females resulted in more than a 50% reduction of enzyme activity when measured on day 15. The same dose of hCG given to hypophysectomized animals with delayed implantation, i.e. pituitary removal on day 3 and implantation induced by oestrone 5 days later, had no effect on placental enzyme activity, but increased that in the ovary. Administration of ovine LH by osmotic minipump (days 11–15) to intact females resulted in abortion in all animals. The same treatment to animals hypophysectomized on day 11 produced abortion in three of four rats; enzyme activity was greatly reduced in the single animal with placentas. In contrast, infusion of LH into hypophysectomized animals with delayed implantation increased placental enzyme activities. However, administration of 20IU pregnant mare's serum gonadotrophin to the hypophysectomized animals on day 11 produced a large increase in ovarian follicular development and ovarian P45017α activities, and resulted in abortion in three of five animals within 96 h.
The results indicate that the P45017α of the junctional zone of the rat placenta has characteristics similar, if not identical, to those of the ovary with regard to preference of substrate and electron donor. However the enzyme activity in the placenta did not increase to the same extent as that in the ovary when exposed to gonadotrophins with LH activity, suggesting that the control mechanisms may be different.
Journal of Endocrinology (1990) 125, 217–224
R. H. NAQVI and D. C. JOHNSON
An increase in ovarian weight in immature rats after the injection of chorionic gonadotrophin (HCG) was used to measure variations in endogenous follicle-stimulating hormone (FSH) after steroid treatment. A single injection of several steroids (testosterone, androstenediol, androstenedione, oestradiol benzoate) given 12–96 hr. before treatment with HCG caused a 30–200% increase in ovarian weight. This was not a direct effect of the steroids since hypophysectomy abolished the response, and administration of the compounds concurrently with HCG was ineffective. Within certain limits an increase in the duration of pretreatment enhanced the ovarian response while an increase in the dose of steroid had little effect.
Pretreatment with testosterone propionate did not change pituitary FSH activity, indicating that the increase in circulating FSH was due to an increased production of hormone. On the other hand, pituitary FSH in animals treated with oestradiol benzoate was significantly lowered within 72 hr. suggesting an increased release of FSH.
R. S. MALLAMPATI and D. C. JOHNSON
Changes in serum concentrations and pituitary content of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and prolactin, determined by radioimmunoassay, were studied after oestradiol benzoate (OEB) administration to gonadectomized male and female rats and to gonadectomized females androgenized with 50 μg testosterone propionate on the 5th day post-partum. Oestradiol benzoate (0·05–50 μg) was given once a day for 7 days beginning 3 weeks after removal of the gonads. A rise in serum LH concentration was produced by 0·05 μg OEB in all animals, but a reduction was found with the larger doses. Serum FSH increased in females injected with 0·05 μg OEB, but males and androgenized females had decreased FSH levels with all doses. The prolactin concentrations increased in the sera of all animals treated with 0·05 μg or 0·5 μg oestrogen. Males showed no further increases with larger doses. A very large increase was found in females given 5 μg but 50 μg did not have this stimulatory effect. Dose-dependent increases in serum prolactin were noted in androgenized females. Pituitary prolactin content increased in females and androgenized females treated with oestrogen; a slight, but significant, increase was found only in the males given 50 μg. Pituitary LH was increased only in the animals receiving a large amount of oestrogen (5 or 50 μg). Glandular FSH increased in the males and androgenized females given 0·5 μg OEB, but in all animals a reduction in this gonadotrophin was found after giving 50 μg of the oestrogen. The results indicate that androgenization alters the normal female hypothalamo-hypophysial response to oestrogen but it does not induce the response pattern of the male.
D C Johnson and L H Crane
A study was designed to compare the effects of exogenous and endogenous oestrogens upon the expression of steroid 17α-hydroxylase/C17,20-lyase (CYP17) in the immature hypophysectomized rat ovary. C17,20-lyase activity was measured ex vivo using [21-14C]progesterone, while serum concentrations of androstenedione indicated in vivo activity. Immunocytochemistry was used to localize the enzyme within the ovary. Activity of CYP17 increased dramatically and remained high in thecal and interstitial cells after injection of equine chorionic gonadotrophin, even though serum oestradiol (OE2) levels exceeded 2 nmol/l. Production of comparable serum levels by the use of Silastic capsules containing OE2 greatly suppressed but did not stop, expression of the enzyme induced by repeated doses of human chorionic gonadotrophin (hCG). Subcutaneous implantation of Silastic capsules (1 cm) containing diethylstilboestrol (DES) stimulated follicular growth and increased ovarian weight by 67%, 5 days later. Injection of 50 IU hCG at various times after removal of the implant produced time-dependent changes in CYP17 activity and serum androstenedione levels, when measured 30 h later. Although the initial effect of removal of DES was an increase in CYP17 activity, delaying injection of hCG resulted in a reduced response. The results indicated that: (1) endogenous oestrogens do not inhibit CYP17 expression; (2) exogenous oestrogens only reduce the number of thecal/interstitial cells expressing CYP17 when they are exposed to hCG; (3) pretreatment with oestrogen removes the ovarian interstitial but not the thecal cell expression of CYP17 in response to hCG; and (4) oestrogens can be stimulatory for CYP17 expression in thecal cells.
Journal of Endocrinology (1995) 145, 59–67
D. C. JOHNSON and R. S. MALLAMPATI
Release of immunoreactive LH and FSH was induced in immature intact female rats by repeated injections of synthetic luteinizing hormone releasing hormone (LH-RH). Altering the dose of LH-RH (5, 10, 20, 50 ng) and the frequency of administration (every 10, 20, 30 or 60 min) over a period of 2 h produced a variety of serum LH and FSH concentrations and ratios. When the dose was a constant 20 ng but the frequency of injections was either 20 or 30 min, a steady state in serum gonadotrophin concentrations was reached within 1 h and the level remained the same during the second hour. When given every 10 min, 20 ng LH-RH produced a much higher concentration of both LH and FSH during the second hour of stimulation. Examination of the gonadotrophin levels after each injection of LH-RH showed that the pituitary response was variable in spite of a constant stimulus.
R. L. MORRISON and D. C. JOHNSON
Male rats were castrated on the day of birth and 5 days later half were given 2·5 mg. testosterone propionate (TP) subcutaneously (androgenization). When 30 days old, single animals were treated with graded doses of TP for 10 days. At the same time 57 males were united in parabiosis with normal intact males, and treated for 10 days with androgen.
Androgenization resulted in increased sensitivity of the accessory sex organs to subsequent treatment with TP. Also, the excessive secretion of gonadotrophin by the castrated animals, as measured by androgen production in intact parabiotic partners, was more effectively inhibited by TP in androgenized than in non-androgenized males. The results are consistent with the interpretation that early androgen treatment sensitizes both the male target organs and the hypothalamo-hypophysial system to androgen.