The purpose of the present study is to investigate the abnormality in gonadotrophin secretion profile in anovulatory conditions. The daily serum levels of FSH and LH were determined by the sensitive and specific double antibody radioimmunoassay in 7 women with normal menstrual cycles, a case of luteal insufficiency, 3 patients with anovulatory cycles and an anovulatory woman with oligomenorrhea. Determinations of FSH and LH were also made in 4 anovulatory women during their treatment cycles with clomiphene citrate.
The data of FSH and LH levels during normal menstrual cycles were obtained in relation to the day of LH surge and averaged. The mean level of LH in the first half of follicular phase (10.2±5.6 mIU/ml) increased toward the second half (20.0 ±5.6 mIU/ ml). At midcycle, the abrupt surge of LH level (113.1 ±20.0 mIU/ml) was demonstrated. Then the value decreased to a lower level (11.8±1.5 mIU/ml) throughout the luteal phase. The serum level of FSH stayed at a high level (5.9 ±0.3 mIU/ml) during the first two thirds of the follicular phase followed by the transitory drop (4.9±0.4 mIU/ ml) in the last one third of the follicular phase. The pattern of FSH showed the peak (14.1 +1.9 mIU/ml) which coincided with LH surge, then decreased to the lower level (3.7±0.4 mIU/ml) in the early luteal phase, followed by the gradual increase toward the end of this phase. The mean ratio of LH to FSH in the follicular, ovulatory and luteal phases were 2.9, 8.0 and 3.2 respectively.
The remarkable changes of profile in a case of luteal insufficiency were relatively low peak of LH (45.6 mIU/ml) at midcycle and decreased LH/FSH ratio (1.2) in the follicular phase. The duration of high basal body temperature (B.B.T.) was 10 days.
The peak levels of LH in 3 anovulatory cycles were found to be lower (28.3-37.0 mIU/ml) than the normal ovulatory peak. Two to five days' rise in B.B.T. was observed following the LH peak until the onset of the next menstruation. The mean ratios of LH to FSH before the LH peak in all cases were low (1.1, 1.2 and 1.5).
The serum levels of FSH and LH as well as the mean ratios of LH to FSH were constantly low during the first 30 days of the cycle in an anovulatory woman with oligomenorrhea.
By the administration of clomiphene citrate, ovulation was induced in two cases out of 4 anovulatory patients. The apparent findings in ovulatory cases were the two peaks of both LH and FSH. The first small peak was found during medication and the second peak comparable to ovulatory peak was demonstrated on 10 to 11 days after the initiation of clomiphene therapy. On the other hand, the above patterns of both LH and FSH were not observed in anovulatory cases, but only a moderate rise in LH level was noted.
In conclusion, the decreased ratio of LH to FSH in the follicular phase and low or no LH peak at midcycle compared to the normal menstrual cycle were constant findings in our mild anovulatory cases. It seems that the presence of LH as well as FSH in the follicular phase is necessary for the maturation of follicle and subsequent ovulation.

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Microanalytical modification of the serum total thyroxine measurement by the Sephadex-G25 column method was described. An accurate and precise measurement was possible, employing 10 μl serum specimens. Applying this method, T₄ concentration of murine serum obtained from orbital sinus was determined. On regular diet, it was 8.3μg/100 ml in average while that on low iodine diet and 0.5μg T₃ suppression was 0.9μg/100 ml, respectively. Simultaneous measurement of T₄ and blood radioactivity in prelabelled mice allowed the comparison of the effect of TSH on both indicators. The extent of the increment of T₄ by TSH was less when compared to that of blood radioactivity, indicating the heterologous nature of the radioactivity discharged from stimulated thyroid.

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It has been well-known that gonadotropin's function on the ovary is to induce such changes as ovulation, luteinization and its maintenance, and that gonadotropin also plays another important role in the metabolism of steroid hormones.
However, the mode of action of gonadotropin has not been clarified yet. As the first step to clarify the mode of action of gonadotropin, specific relations between HCG administered and rabbit ovary was studied in this experiment.
1) Intravenous administration of HCG in a rabbit
1500 I.U. of HCG was injected into the auricular vein of a rabbit and blood level of HCG was measured by radioimmunoassay during the lapse of time. Then 2.5μCi (S.A. 210μCi/μg) of ¹³¹I-HCG and 10 I.U. of HCG were combinedly administered to the rabbit and radioactivity in blood was recorded by a well-type gamma detector. As a result, both cases showed two-phase decrement curves with t¹/₂=37.9 min and t¹/₂=8.35 hr.
2) Suppression of ovulation by anti-HOG serum
Anti-HCG serum was administered following to the intravenous administration of 10 I.U. of HCG in a rabbit.
As a result, ovulation was suppressed when the administration of anti-HCG serum was made within 30 minutes.
By the administration after 30 minutes, however, ovulation was not suppressed. The above experiment showed that the matter of ovulation in a rabbit due to the HCG administration was brought into a inner part of ovary, within a short time, where antiHCG serum is unable to reach.
3) Distribution of ¹³¹I-HCG in organs
According to the paired labelled technique, 50μCi (S.A. 200-300 μCi/μg) of ¹³¹I-HCG, 250 μCi (S.A. 130-180 μCi/μg) of ¹³¹-human serum albumin, 10 I.U. of cold HCG and 10 mg of cold human serum albumin were mixed and administered to a rabbit. After a fixed time, the chest of the rabbit was opened and 5L of physiological saline was circulated from the artery to evacuate blood, and then organs were extirpated.
The value of ¹³¹I-HCG uptake per ¹³⁵I-albumin uptake was calculated. In ovary, the value started to rise from 60 minutes after the administration and the highest value of 8.20, compared to the other organs, was shown two hours later.
When anti-HCG serum was administered at the same time, it showed the low value of 0.18
In kidney and liver, a high value was observed in 15 to 30 minutes after the administration and then gradually decreased. A low value was maintained in skin, uterus and lung. From the above findings, it was considered that HCG showed specific distribution to ovary.
4) Effect of HCG on the activity of adenylcyclase in homogenate of rabbit ovary
At the 6th day after intravenous injection of 30-50 I.U. of HCG, the ovary of rabbits was extirpated to make homogenate. According to the method of Krishina (1968), the effect of HCG on the activity of adenylcyclase in the homogenate was evaluated. The effect was found to be more active at the ovary where the most luteinization takes place. When FSH was administered instead of HCG, the effect of adenylcyclase was not activated. Although it was activated by the administration of TSH, it may be attributable to contamination by LH, taking into consideration the purity of TSH employed in this experiment.

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Recently, many studies have been reported concerning the action mechanism of hormones, though the action mechanism of gonadotropin, especially the mechanism of ovulation induced by gonadotropin is not yet well known partially because of difficulty in purification. It was reported in our previous report that the synthesis of RNA, protein and DNA was increased in the ovary of immature rat after HCG administration.
In this paper, inhibition of ovulation was studied with cycloheximide or actinomycin to decide whether the enhancement of RNA and protein synthesis is an inevitable process in the ovary for ovulation. In order to clarify the role of catecholamines in the hypothalamo-pituitary system, studies on the ovulation after a single injection of PMS to immature rats were performed and then it was studied whether the RNA extracted from the ovary of rats treated with PMS exerts the biological action like PMS.
1) Female immature rats (Sprague Dawley strain, weighing 40-50 g) were used as experimental animals. Rats were administered with PMS (30iu) and then HCG (10iu) about 40 hours after PMS. 200 μg/100 gb.wt. of cycloheximide or 100 μg/100 g b.wt. of actinomycin was intraperitoneally administered at different intervals from the hormone injection. Ovulations were detected with the Rowlands technique.
Ovulations were almost always inhibited when cycloheximide was administered, from 30 min. before HCG injection to 5 hours after HCG injection but when cycloheximide was administered thereafter, ovulations could not be inhibited. When cycloheximide was administered within 5 hours after PMS injection, a tendency that ovulations were inhibited could be detected.
When actinomycin was administered from 30 min. before to 6 hours after PMS or HCG injection, ovulations were sometimes inhibited, but the ovulation rate was higher than in the case with cycloheximide administration. From these results, it was suggested that the ovulation was induced through the actions of RNA and protein which were synthesized after gonadotropin administration and the synthesis of RNA and protein inevitable for ovulation was finished within 5-6 hours after gonadotropin treatment.
When strophanthin G as an ATPase inhibitor was systemically administered after gonadotropin injection, in all cases ovulations occurred and both the number of ova and weight of ovaries were higher than in the strophanthin untreated cases. From these results, the active transport of Na+ or K+ with ATPase is suggested not to be related with the mechanism of ovulation.
2) PMS (30iu) was administered to female immature rats of Sprague Dawley strain. Ovulations were investigated about 68 hours after PMS injection. When reserpine or nialamide was administered following the schedule shown in the Table, ovulations with a single injection of PMS were inhibited. And when reserpine and dopa were injected, there was a case where the ovulation occurred. From these results, the process in the hypothalamo-pituitary system which involves LH release is suggested to need a moderate dynamics of catecholamines.
3) RNA of the ovary of PMS treated rats was extracted after the Scherrer and Darnell hot phenol method. The RNA was intraperitoneally administered to the immature rats according to the schedule shown in the Table. HCG (10iu) was injected after RNA administration. Ovulations were investigated about 20 hours after HCG injection. Neither ovulations nor weight gain of ovary could be detected in all cases.

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Ten patients who underwent various operations were examined in this study. Urinary total 17-OHCS and its fractions were determined with β-glucuronidase hydrolysis using thin layer chromatography. The method of analysis of fractions of urinary free 17-OHCS is outlined as follows : A 100 to 200 ml aliquot of 24 hour urine was extracted with dichloromethane without hydrolysis. The oillike low polar impurity was removed using column chromatography. The steroids were dissolved from column with ethanol and evaporated. The next process is exactly the same as the determination of total 17-OHCS. The recovery in this method was 72% on the average as in the fractions of total 17-OHCS. The following results were obtained.
The excretion volume of individual fractions of urinary free 17-OHCS was about 0.05 mg/day before operation. After operation, free comp. F increased significantly. In addition there was a limit to the amount of excretion of free comp. E, THF and THE. However, the excretion volume of THE in total evidently exceeded the other three fractions before operation. After operation, individual fractions increased in the following ascending order, THE, THF and comp. F, varying with the degree of stress. Therefore, the excretion pattern of fractions of urinary free 17-OHCS was quite different from that of fractions of total 17-OHCS that were hydrolyzed with β-glucuronidase.
The excretion volumes of free comp. F and comp. E were several tens percent of comp. F and comp. E calculated as fractions of total 17-0HCS. However, the excretion volumes of free THF and THE were only a few percent of THF and THE calculated as fractions of total 17-0HCS. It is clear that comp. F and comp. E are also conjugated with glucuronic acid though the degree of conjugation is much lower than their metabolites, THF and THE.
After operation, the above mentioned percentage increased in free comp. F and comp. E, while it decreased in free THF and did not change in free THE. It is clear that the rate of conjugation of free comp. F and comp. E with glucuronic acid diminish compared with control, and that the metabolism of cortisol shifts to THF pathway during surgical stress.

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HGH response to an insulin injection (0.1 u/kg) was evaluated in 62 male & 85 female subjects in an attempt to define its normal value, taking age, sex and the degree of obesity into consideration. Changes of serum FFA levels were also measured in order to compare the pattern of fat mobilization in obese and standard weight healthy subjects.
The following results were obtained :
1. The fasting level of HGH and HGH response to insulin-induced hypoglycemia are different with sex, age and the presence of obesity.
2. HGH response to insulin-induced hypoglycemia in standard weight healthy subjects aged in 20-29 was as follows : (M±SE, μg/ml)
fasting 15′ 30′ 45′ 60′ 120′ 180 male 4.4±1.6 7.4±2.0 28.8±5.2 66.0±6.4 70.6±6.3 36.0±6.4 9.2±1.7 female 9.6±1.5 6.2±1.1 15.4±2.3 38.3±6.4 32.0±5.6 14.1±4.1 3.2±0.4
3. It was observed that the fasting level of HGH was higher in the female than in the male, but in subjects over 45 years of age no difference was observed between the female and the male.
4. HGH response to insulin-induced hypoglycemia was significantly higher in the male aged in 20-29 than in the male aged over 30 and in the female of all age groups. In the group over 30 years of age, difference between the male and, the female was not clear.
5. There was not a significant correlation between HGH response and insulin sensitivity index, minimum plasma glucose level or FFA decrement.
There was, however, a significant correlation between FFA increment from 60 to 180 minutes after an insulin injection and HGH response. (γ=0.71, P<0.005)
6. It was observed that HGH response was lower in obese subjects than in standard weight healthy subjects, both the male and the female. It appears likely that obese subjects require less HGH for fat mobilization than standard weight healthy subjects.

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