Summary. Luteolysis in Shiba goats was spontaneous (N = 5) or induced by prostaglandin F-2α (N = 5). Blood sampling and the test for oestrous behaviour were carried out at 2-h intervals, and the time of follicular rupture (ovulation) was determined by culdoscopic observations performed every 2 h around the periovulatory period. In both groups plasma LH concentrations showed a temporary but significant increase during the abrupt fall in plasma progesterone concentrations at luteolysis. This LH rise may be responsible for the preovulatory development of antral follicles and the increase in oestradiol secretion from them. The total number of antral follicles (spontaneous luteolysis 3·6 ± 0·6, induced luteolysis 4·2 ± 0·7; mean ± s.e.m.) and the number of ovulations (1·8 ± 0·4, 2·4 ± 0·2) did not differ significantly between the two groups. There was also no significant difference in the timing of oestrus, LH surge and ovulation following the two modes of luteolysis. The interval from luteolysis to the peak of the LH surge averaged 65 h (range 56–72 h). The period of oestrous behaviour coincided with the acrophase of the LH surge and lasted for 22 h (12–28 h). Ovulations occurred 21 (16–24) h after the LH peak and 7 (2–12) h after the end of oestrus.
Y. Akazome and T. Mori
In chicken embryos, there is a difference between the sexes in the onset of lutropin receptor mRNA expression in the gonads. The effects of oestrogen on lutropin receptor expression were studied to investigate the mechanism controlling this difference. Lutropin receptor mRNA expression was detected in the ovaries of sesame oil-treated control female embryos on day 12 of incubation, while no expression was found in the testes of the male controls. Oestradiol administration to genetically male embryos before sexual differentiation resulted in gonadal sex reversal which was characterized histologically by the proliferation of cortical cords and the presence of lacunae. Lutropin receptor expression was detected in the feminizing testis on day 12 of incubation. Administration of aromatase inhibitor (CGS 16949 A) to genetically female embryos before sexual differentiation inhibited the formation of cortical cords, although a relatively weak expression of lutropin receptor was detected. These results indicate that early expression of the lutropin receptor is regulated by oestrogen.
Y Akazome, T Abe, and T Mori
The gonad is an endocrine organ secreting sex hormones and also a target of pituitary gonadotrophins. The expression of mRNAs encoding LH receptor (LHR), FSH receptor (FSHR), P450c17 and P450aromatase in the developing gonads of embryos between day 4 and day 6 of incubation was determined using a RT-PCR to elucidate the chicken gonad as a target organ of gonadotrophins. Although expression of mRNAs encoding LHR, FSHR and P450c17 was detected at day 4 of incubation in both sexes, mRNA encoding P450aromatase appeared at day 6 in female embryos only, indicating that mRNAs encoding gonadotrophin receptors can be identified before sexual differentiation. Quantitative PCR analysis revealed that expression of mRNA encoding LHR and FSHR remained low in male gonads from day 4 to day 6 of incubation, whereas they increased on day 6 in female gonads. The sexual dimorphism in the expression of mRNAs encoding LHR and FSHR was confirmed in the sexually differentiated gonads of embryos at day 12 of incubation (LHR in ovary ratio LHR in testis = 7 ratio 1; FSHR in ovary ratio FSHR in testis = 9 ratio 1).
M Nishihara, Y Takeuchi, T Tanaka, and Y Mori
The hypothalamic gonadotrophin-releasing hormone (GnRH) pulse generator governs intermittent discharges of GnRH into the pituitary portal circulation and, consequently, modulates the pulsatile pattern of gonadotrophin secretion. Electrophysiological correlates of pulsatile gonadotrophin secretion have been demonstrated in the mediobasal hypothalamus of monkeys, rats and goats by recording multiple unit activity. A temporal coincidence between characteristic increases in multiple unit activity and gonadotrophin pulses in the circulation is seen under a variety of physiological and experimental conditions in all three species examined, providing evidence that hypothalamic multiple unit activity originates in the GnRH pulse generator. During a preovulatory gonadotrophin surge induced by oestrogen in ovariectomized animals or occurring spontaneously in intact animals, GnRH pulse generator activity is decelerated, suggesting that it is not involved in generating the gonadotrophin surge. The gonadotrophin surge may be generated by an oestrogen-responsive neuronal complex intrinsically different from the GnRH pulse generator, the electrical operation of which remains unknown.
S. Natsuyama, Y. Noda, K. Narimoto, Y. Umaoka, and T. Mori
Summary. The development of mouse pronuclear-stage embryos in media containing various concentrations of thioredoxin was monitored and the influence of antithioredoxin immunoglobulin G (IgG) and heat-treated thioredoxin on the thioredoxin-induced effects was evaluated. A significant increase in the number of four-cell embryos (76·3%) and blastocysts (37·3%) was observed when embryos were cultured in the medium containing 50 μg thioredoxin ml−1 compared with the rates (55·8 and 3·8%, respectively) in the basic medium. The number of blastocysts increased significantly to a maximum of 70·2% at 500 μg ml−1. The biological activity of thioredoxin was evident after dialysis, but was markedly impaired by the addition of anti-thioredoxin IgG to the culture medium. Treatment at 60°C for 5 min did not affect the enzymatic and biological activity of thioredoxin. More severe heat treatment (121°C for 30 min) attenuated the enzymatic activity to 40% of its initial value and reduced the biological activity (number of blastocysts, from 77·8 to 51·6%).
These results indicate that the effect of thioredoxin on the two-cell block is due to the thioredoxin molecule itself, and suggest that disulfide formation within or between proteins resulting from oxidative stress is one of the major causes of the two-cell block.
Keywords: thioredoxin; embryo; mouse
Y. Mori, M. Tanaka, K. Maeda, K. Hoshino, and Y. Kano
Summary. Ovariectomized Shiba goats carrying an oestradiol implant (4–10 pg/ml) were kept under a short-day light regimen (10L:14D; Group 1, N = 4) or a long-day regimen (16L:8D; Group 2, N = 4). Plasma LH concentrations were lower (P < 0·05) in Group 2 than in Group 1 between Days 40 and 200, suggesting an enhanced negative feedback effect of oestradiol on LH secretion under a long-day regimen.
On Days 30, 60, 100, 149 and 279, an LH surge was induced by i.v. infusion of oestradiol for 48 h; the infusion rate was gradually increased from 0·5 (0 h) to 4·1 (48 h) μg/h, thereby mimicking the preovulatory increase of oestradiol secretion. The duration and magnitude of the induced LH surge were indistinguishable between the groups. The latency from the onset of oestradiol infusion to the LH surge was relatively constant in Group 1, 41·1 ± 0·9 h (mean ± s.e.m., n = 17) but was shorter in Group 2 (19·7 ± 3·7h, P < 0·05) on Day 149; less oestradiol was therefore required for induction of the LH surge (27·4 vs 89·7 μg, P < 0·01), suggesting an increased sensitivity to the oestradiol positive feedback under a long-day regimen.
These results might be interpreted to indicate that the hypothalamic–pituitary axis of the goat becomes hypersensitive to the positive as well as the negative feedback effect of oestradiol under long-day conditions.
Y. K. Oh, T. Mōri, and T. A. Uchida
Summary. The vaginal plug of the Japanese greater horseshoe bat is composed of an outer, thick, hard and homogeneous layer originating from the stratum disjunctum of the vaginal mucosa and a thin, soft and opaque central core containing accidentally trapped dead spermatozoa. The vaginal plug appeared to be expelled just before or after ovulation in spring. Immediately after copulation in autumn, uterine spermatozoa were dead but few leucocytes were present. As time passes after copulation, however, the infiltration of leucocytes into the uterus increased gradually, and extensive leucocytic phagocytosis of dead spermatozoa occurred during hibernation.
I. Takai, T. Mori, Y. Noda, and T. Nishimura
Summary. Rabbit antiserum raised against isolated human ova produced an intense immunofluorescence and a precipitate on the outer surface of human and porcine zonae pellucidae. The antiserum reacted with none of 22 human tissues nor with any of 4 body fluid components by immunofluorescence or immunodiffusion analysis, but the antiserum agglutinated AB erythrocytes after absorption with O erythrocytes. The anti-zona activity was achieved by higher titres for human than for porcine zonae. Immunofluorescence on porcine zonae was completely abolished by absorption with porcine ova, whereas a weak but definite fluorescence remained on human zonae. These findings indicate that the human zona pellucida consists of at least three distinct components; (1) a specific antigen(s) shared by human and porcine zonae, (2) an antigen(s) specific to human zonae, and (3) a non-specific antigen(s) associated with the blood group substances.
T. S. Choi, M. Mori, K. Kohmoto, and Y. Shoda
Summary. Mouse oocytes matured in vitro in chemically defined medium were not penetrated by spermatozoa. The time required for dissolution of the zona pellucida of such oocytes by α-chymotrypsin was much longer than that for ovulated oocytes. Addition of fetal calf serum to the medium for maturation of oocytes improved the incidence of sperm penetration and shortened the time of enzymic dissolution of the zona pellucida. These results suggest that the low rate of fertilization of oocytes matured in vitro is mainly due to qualitative changes of the zona pellucida, which could be overcome by a factor or factors in fetal calf serum.
Y. K. Oh, T. Mōri, and T. A. Uchida
Summary. After the mating season of the Japanese greater horseshoe bat in mid- or late October, only the right ovary maintained a single Graafian follicle throughout hibernation until early April. During this time the ovum was in prophase of meiosis I (resting stage) with many large lipid droplets as a nutrient source. In synchrony with stigma formation, there was resumption of meiotic activity, separation of the cumulus oophorus from the granulosa layer and dispersion of the follicle cells just before ovulation in spring. The block to polyspermy seemed to reside in the zona pellucida, because no spermatozoa could be detected in the perivitelline space of the 6 fertilized ova examined, although a second spermatozoon was recognized in the zona pellucida of 3 ova.