In order to elucidate the role of the renin angiotensin system and ACTH in the biosynthetic pathway of aldosterone (Ald) and cortisol (F), 18 healthy subjects (13 males and 5 females aged 18 - 37) were studied during the changes in circadian rhythm of various plasma corticoids before and after dexamethasone (dexa) administration during daily life. Furthermore, changes in various plasma corticoids after exogenous angiotensin II (A II) and ACTH injection during pre and post dexa administration were investigated.
The results were as follows :
1. Diurnal variation of various corticoids.
a. Under usual active life, progesterone (P), deoxycorticosterone (DOC), corticosterone (B), 17-OHprogesterone (17-OHP), deoxycortisol (S) and F demonstrated distinct circadian rhythm. However, a few subjects failed to show the same results. Ald demonstrated no circadian rhythm and plasma renin activity (PRA) remained unchanged.
b. After suppression of endogenous ACTH by continuous 2 mg/day dexa administration during usual active life, S, F and B were significantly suppressed in the 2nd, 4th and 7th day causing complete disappearance of the circadian rhythm, but slight suppression of P, DOC, Ald and 17-OHP were observed.
2. Changes in various corticoids during continuous A II (5 - 20 ng/kg/min) infusion.
a. A II induced significant increment of Aid. But precursors like P, DOC and B either remained unchanged or diminished gradually. 17-OHP, S and F remained either unchanged or declined. PRA was suppressed slightly.
b. A II infusion after 2mg dexa administration raised Ald significantly. Furthermore, B also showed an increment, although within the range of low concentration. The remaining corticoids were not affected by A II infusion.
3. Changes observed in various corticoids after 250pg ACTH injection.
a. ACTH caused a significant increment of various corticoids within 30 mins after injecttion which maintained a high value until 180 mins. The increment of 17-OHP, S, F and B was associated with a strong effect, while P, DOC and Ald were found slightly weak. The maximum response of Ald and F was observed within 30-60 mins and within 90 mins respectively. PRA remained unchanged.
b. The response to ACTH after 2mg dexa administration showed an abrupt elevation of 17-OHP, S, F, P, DOC, B and Ald within 30 mins. Later, most of the corticoids showed a gradual increase with plateau formation except Ald. The responses of these corticoids demonstrated no remarkable differences without dexa administration.
c. ACTH injection after a continous 2mg/day dexa administration for 7 days demonstrated a remarkable decrease of S, F and B compared to the responses during 2mg dexa administration. Almost no change to slight decrement was noticeable in 17-OHP, P and DOC. Ald responded to a greater or the same degree. PRA remained unchanged.
With the above results, the author concluded as follows.
Under the daily activity, corticoids failed to demonstrate circadian rhythm in a few subjects. After dexa administration, degrees of suppression varied depending on the type of corticoids. By A II infusion under dexa administration, B responded slightly, but A II failed to increase corticoids before DOC in biosynthetic pathway of Ald. Various corticoids responded in different degrees to ACTH. Furthermore, the suppressive response of the corticoids to ACTH after 7 days of dexa administration also varied with the type of corticoids.

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Five healthy men were given 1.0 gr of metyrapone p.o. (MTP test) with and without 0.5 gr of 1- dopa p.o. (DOPA-MTP test) or 10 mg of metoclopramide p.o. (MTC-MTP test) and plasma 11-deoxycortisol, cortisol, ACTH and pregnenolone (only in MTP test) were determined before and hourly after the drug administraing for 6 hrs. In the MTP test, plasma 11-deoxycortisol increased significantly at 1 hr with peak at 5 hrs, whereas the significant increases of pregnenolone and ACTH were not seen until 2 hrs, although plasma cortisol levels were reduced definitely at 1 hr. Thus, the increase of 11-deoxycortisol in the MTP test should be divided into two phases; the increase in the phase II (later than 3 hrs) is due to pituitary ACTH reserve, and that in the phase I (until 2 hrs) is due to unknown mechanism other than pituitary reserve. In the DOPA-MTP test, the 11-deoxycortisol rise was significantly larger than that in the MTP test. The increases in 11-deoxycortisol at 4 hrs and 5 hrs in the MTC-MTP test were significantly smaller than those in the MTP test. The cortisol/ (cortisol + 11-deoxycortisol) ratio reached its lowest point at 3 hrs in the MTP test and at 2 hrs (a 1 hr advance) in the DOPA-MTP and MTC-MTP tests. However, there were no significant differences in the responses of cortisol and ACTH to metyrapone in these tests. This provides evidence for a discrepancy between plasma 11-deoxycortisol and ACTH responses in a single dose metyrapone test, as disclosed by the simultaneous administration of dopamine agonist or antagonist with metyrapone.

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Graves' immunoglobulins have been shown to displace labeled TSH bound to thyroid plasma membranes. However, TSH displacing activity (TDA) could not be detected in all of the cases. Its relationship to thyroid function and to clinical symptoms has not yet been clearly demonstrated. The present study was carried out to clarify the specificity and pathogenetic importance of TDA of IgGs mainly found in patients with Graves' disease.
IgG was purified using DEAE cellulose column chromatography. A radioreceptor assay was carried out according to the method of Smith and Hall. Human thyroid adenylate cyclase stimulator (HTACS) activity of IgG was determined according to the method of Orgiazzi et al. with slight modifications. Mean %B/F, calculated as percentage of B/F to that of the control obtained in the absence of IgG, was 98.5 ± 8.3 (SD), 94.7 ± 7.8, 90.0 ± 9.3 and 75.5 ± 11.0% in the presence of 1, 5, 10 and 20 mg/ml of normal IgGs, respectively. Since this decrease in %B/F with normal IgGs was considered as a nonspecific inhibitory effect of IgG itself, %B/F of sample IgG was corrected by dividing the obtained data by the mean %B/F of normal IgGs at each concentration, which was designated as corrected %B/F (C-%B/F). The percentage of patients with untreated Graves' disease who showed positive TDA increased as did IgG concentration up to 10 mg/ml of IgG. When assayed at 10 mg/ml of IgG, 40 out of 48 cases (83.3%) were TDA positive in untreated Graves' disease, 3 out of 18 (16.7%) in Hashimoto's thyroiditis, 2 out of 12 (16.7%) in thyroid neoplasm and 5 out of 9 (5 5.6%) in subacute thyroiditis.
All the untreated patients with Graves' disease including the displacing activity negative ones, showed a dose-dependent decrease in C-%B/F up to 10 mg/ml of IgG. On the other hand, the values obtained in most patients with thyroid neoplasm and Hashimoto's thyroiditis showed random dose-independent variations within normal ranges. In 5 out of 6 patients with subacute thyroiditis, dose-dependent decreases in C-%B/F were observed.
In patients with untreated Graves' disease, TDA was significantly correlated to the size of struma and to the values of Hertel's exophthalmometry. With antithyroid drug treatment, the numbers of TDA positive patients gradually decreased. Among the patients treated for more than a year, only 26% were TDA positive. Six out of 9 patients who had been TDA positive at the time of cessation of therapy relapsed immediately thereafter, while 8 out of 11 TDA negative patients remained euthyroid for more than 9 months. HTACS activity was positive in all of the 8 patients with untreated Graves' disease examined. IgG with stronger TDA tended to show higher HTACS activity.
On the other hand, serum IgG from a mother with nongoitrous hypothyroidism exhibited strongly positive TDA; C-%B/F being 4.1%. IgGs obtained from umbilical cord blood of her hypothyroid neonates (nonidentical twins) demonstrated positive TDA as strong as that of herself, which became undetectable in the surviving baby 6 months after birth, when her thyroid function became normal. Maternal IgG was devoid of HTACS activity, and it definitely inhibited thyroidal adenylate cyclase stimulation induced by TSH. The presence of a blocking antibody to TSH receptor in maternal serum and its trans-placental transfer to her children were suggested.
In summary, correlations between TDA and clinical symptoms, courses, prognosis and HTACS strongly suggested the pathogenetic role of anti-TSH receptor antibody in Graves' disease. However, the presence of a blocking antibody against TSH receptor in non-goitrous hypothyroidism suggested the diversity of anti-TSH receptor antibody concerning biological activity.

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The pressor response to angiotensin II (A II) has been shown to be enhanced, and aldosterone production to be attenuated following administration of indomethacin (Ind), a cyclooxygenase inhibitor, pretreatment. Since PGI₂ has been proposed to be a potent vasodepressor prostaglandins, with steroidogenic action, the present study was undertaken to evaluate the regulatory role of PGI₂ in the blood pressure and aldosterone responses to A II.
Ind was administered to suppress the production of endogenous prostaglandins (30 mg/kg. im. Ind-group) and PGI₂ was given after Ind pretreatment (2ng/kg/min. iv. Ind + PGI₂ group) to conscious male rabbits (2.5 - 3.5 kg). Baseline blood pressure readings were similar in three groups (control, Ind-, and Ind + PGI₂ -group). The increment of blood pressure by A II infusion were significantly enhanced in the Ind-group (12.6 ± 1.3 mmHg) compared with the control-group (8.6 ± 1.5 mmHg). This enhancement was diminished by PGI₂ infusion (7.0 ± 2.3 mmHg). PGI₂ infusion alone does not influence blood pressure in the dose of 0.5-4.0 ng/kg/min. Aldosterone response to A II was significantly attenuated in the Ind-group (1.5 ± 1.2 ng/dl) compared with the control-group (7.5 ± 3.1 ng/dl), but was restored in the Ind + PGI₂-group (4.5 ± 1.3 ng/dl).
It is suggested that circulating PGI₂ may play a role in the regulation of blood pressure by modulating the pressor response and the aldosterone response to A II.

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The purification of angiotensin I-converting enzyme (ACE) solubilized with Triton X-100 was carried out by a column chromatography of DEAE-cellulose and Sephadex G-200 and subsequently by SDS/polyacrylamide gel electrophoresis. Further, the subcellular distribution of the enzyme was studied in the aortic tissue by means of the ultracentrifugation method.
Treatment with Triton X-100 increased the enzyme activity by about 8 times, indicating that ACE may be a membrane-bound protein in aortic tissue. On chromatography with DEAE-cellulose, the enzyme was eluted at 3.0 (peak I), 8.0 (peak II), 11.0 (peak III) and 22.0 (peak IV) mohm^-1 in conductivity of NaCl gradient. On Sephadex G-200 gel filtration, peak I showed two active fractions of ACE activity (ACE-A and ACE-B). Peak II showed an elution pattern which was similar to that in peak I (ACE-C and ACE-D). Both peak III and peak IV showed only one active fraction (ACE-E for peak III and ACE-F for peak IV).
The molecular weights of these ACEs were estimated to be 460000 for ACE-A, 266000 for ACE-B, 440000 for ACE-C, 220000 for ACE-D, 217000 for ACE-E, and 119000 for ACE-F on Sephadex G-200 gel filtration. On SDS/polyacrylamide gel electrophoresis, all the ACEs prepared by Sephadex G-200 showed only a single major protein band at the same distance of migration, and the molecular weight was found to be 112000.
The optimal pH and temperature for these ACEs were identical and were 8.3 for pH and 37°C for temperature. The activities of enzymes A to F were almost completely inhibited by various ACE inhibitors; bradykinin potentiator c (10^-3M), EDTA (10^-4M), o-phenanthroline (10^-3M) and arg-pro-pro (10^-3M).
Subcellular distribution of ACE activity revealed a high percentage in the microsomal fraction of the aorta.
From these results, it is suggested that the aortic angiotensin I-converting enzyme, which might be produced in the microsomal fraction, is present in a form of the tetramer, dimer and/or the monomer, of which molecular weights are 112000 to 119000. The converting enzyme may play a possible role in the local control of vascular tone by the conversion of inactive angiotensin I into active angiotensin II in the arterial wall.

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Recently many gut hormones have been found in the brain, and there is some evidence to suggest that pancreatic polypeptide-like immunoreactivity (PP-LI) is also present in the brain. Although in mammals, confirmative evidence is not yet shown. In the present paper we report the distribution and tissue localization of PP-LI in the canine brain by radio-immunoassay (RIA) and immunohistochemistry.
Normal, fasted mongrel dogs were used. Brain tissue was extracted by boiling water. High concentrations of PP-LI were found in the pituitary gland (3.67 ± 1.10 ng/g wet wt), substantia nigra (1.58 ± 0.36 ng/g wet wt), hypothalamus (0.74 ± 0.28 ng/g wet wt) and olfactory lobe (0.58 ± 0.21 ng/g wet wt). PP-LI was not detectable in the frontal lobe, parietal lobe, striatum, thalamus, hippocampus, pons, cerebellum and medulla oblongata. The amounts of PP-LI in the brain were more less than the amounts of PP present in the pancreas (duodenal lobe, 29.3 ± 1.1 μg/g wet wt).
The dilution curve of the brain tissue extracts showed parallelism with the standard curve of human and porcine PP on the RIA system. On Bio-Gel P-30 column chromatography, PP-LI from the pituitary gland and olfactory lobe eluted as a single peak coincided with highly purified bovine PP. In immunohistochemical study, PP-LI was found in the intermediate lobe and the stalk of the pituitary gland by means of anti-bovine PP antiserum.
These findings of the specific regional localization suggest that PP or PP-LI may have a physiological role in the central nervous system.

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It is well known that there are functional and morphological similarities between the salivary glands and the pancreas. Amylase, kallikrein and glucagon are present in both tissues. Morphological similarities of the two tissues have been observed by using a light and electron microscope.
In order to examine the pathophysiological relationship between the pancreas and the salivary glands, an immunohistochemical study using antisera against proline-rich peptide P-C, which was recently isolated from human whole saliva, was carried out on the human salivary glands and the pancreas. Peptide P-C like immunoreactivity was found not only in the salivary glands but also in the pancreatic islets. Furthermore, observation of serial thin sections immunostained with insulin, glucagon, somatostatin, PP antisera and antisera against peptide P-C revealed that peptide P-C like immunoreactivity-containing cells were identical to insulin containing B-cells. As the antisera against peptide P-C did not have any cross-reactivity to other kinds of peptides including insulin, glucagon, somatostatin, pp, VIP, human C-peptide and kallikrein, the present finding suggests that peptide P-C like immuno-reactivity is present in the B-cells independently of insulin and proinsulin. The finding seems to be a new addition to the lists of proof which support the presence of a pathophysiological relation between the salivary glands and the pancreas. Although it seems likely that peptide P-C like immunoreactivity in the pancreatic B-cells may play some role in the function of the B-cells, since this material was present only in the B-cells among four kinds of cells in the pancreatic islets, its exact pathophysiological role remains to be elucidated.

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Insulin receptors on erythrocytes were studied in normal and diabetic pregnant women to clarify the mechanism of insulin resistance in pregnancy. The assays of insulin receptors were performed according to the method of Kobayashi, which is a slight modification of the method of Gambhir. ¹²⁵I-insulin binding showed no significant differences between normal pregnant women during the first (n=18), the second (n=15) and the third (n=54) trimesters and nonpregnant controls (n=52). There were also no significant differences between the values before and after delivery (n=8). Reticulocyte counts significantly increased in pregnant women during the second trimester and during the later periods. There was a positive correlation between ¹²⁵I-insulin binding and reticulocyte counts in late pregnancy. These findings suggest that reticulocyte counts should always be considered in estimating erythrocyte insulin receptors in pregnancy.
Then, insulin binding in late pregnant women with reticulocyte counts below 10%0 (n=20) was studied. The value was slightly decreased as compared to that in the nonpregnant controls, but the difference was not significant. ¹²⁵ I-insulin binding in gestational diabetes (n=6) was decreased, but that in overt diabetes (n=4) was not. Patients with overt diabetes had been receiving insulin therapy. Insulin resistance in normal pregnancy cannot be explained by the changes of insulin receptors from this study. It may be due to some post-receptor abnormalities. But decreased insulin binding might be one of the factors that manifest or deteriorate gestational diabetes.

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It has been reported that the neonatal administration of monosodium glutamate (MSG) in rodents produces lesions of the arcuate nucleus and the ventromedial nucleus of the hypothalamus and results in obesity in adulthood. Though there are a large number of reports using MSG-treated animals, most of them deal with neuronal and neuroendocrinological pathophysiology within the central nervous system.
The present study, therefore, was designed to investigate the time-course of metabolic and hormonal changes in relation to the development of hypothalamic obesity induced by MSG in female Wister rats. In addition, responses of insulin and somatostatin to glucose loading were examined in rats with hypothalamic obesity.
MSG, 2mg per g of body weight, was subcutaneously injected for 5 consecutive days after birth. Body weight, plasma glucose and immunoreactive insulin (IRI) were measured at one- to four-week intervals in rats aged from 6 days to 11 months. The Lee index, plasma triglyceride and immunoreactive glucagon (IRG) were measured at one-month intervals from one to 11 months. Immunoreactive somatostatin (IRS) and gastrin (IRGa) were determined at the 7th month or later. All parameters in plasma described above were assayed on samples drawn from the jugular vein. In addition, IRI and IRS in the pancreas were measured until 6 months. Further, responses of glucose, IRI and IRS to intragastric glucose loading (3g/kg body weight) were examined in portal plasma at the 11th month. All parameters in the MSG rats were compared with those in age-matched female controls which received the vehicle alone.
Plasma IRI was elevated by 54% in the MSG rats even at the 6th day. Thereafter, the MSG rats showed a progressive increase in plasma IRI which was 96% and 237% higher at the 1st and 3rd month, respectively. Parallel with the change in plasma IRI, there was a constant 1.5-fold elevation of pancreatic IRI in the MSG rats aged between the 21st day and the 6th month, though pancreatic IRI decreased by 20% at 6 days. Pancreatic IRS also increased at the 21st day, but thereafter decreased by 48% at the 6th month. A progressive increase in plasma TG which was 37,139,203 and 279% higher at 2, 3, 4 and 5 months, respectively, was followed by a 2- to 3-fold elevation during following 6 months.
The MSG rats showed a constant 20% reduction in body weight for the first 2 months, but thereafter showed a progressive increase throughout the observation which was 17, 39 and 55% higher at 4, 7 and 11 months, respectively. The Lee index of obesity also progressively increased from 337 ± 4 at the 3rd month to 390 ± 6 at the 8th month, but thereafter maintained a constant 1.2-fold elevation. In contrast with the marked changes in plasma IRI, no significant differences in plasma glucose were found between the two groups for the entire period of observation. Plasma IRG also showed a constant 1.2- to 2.0-fold elevation irrespective of the duration of obesity.
Rats with obesity induced by MSG had markedly elevated levels of IRI and IRS not only in jugular plasma but also in portal plasma. In addition, responses of the two hormones to intragastric glucose loading were exaggerated in portal plasma, whereas response of plasma glucose was unchanged. Plasma IRGa was also observed to be increased by 38 - 81%.
These findings indicate that hyperinsulinemia is the primary factor in the development of hypothalamic obesity induced by MSG. Hypothalamic obesity induced by neonatal administration of MSG in the rat is characterized by hypersecretion of glucagon and somatostatin as well as insulin, and may provide a readily available and valuable animal model for studying the hypothalamico-pancreatic axis.

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