We have measured vitamin D₃ hydroxylation in control, streptozotocin-induced diabetic, insulintreated diabetic, and glibenclamide-treated rats fed a vitamin D-deficient diet, using in vivo and in vitro isolated kidney cells. The diabetic animals were used in vivo and in vitro experiments 59 and 81 days after streptozotocin injection, respectively. In both studies, the conversion of [³H] 25 OHD₃ to [³H] 1, 25-(OH) ₂D₃ was found to be reduced significantly in diabetic rats, whereas it returned nearly to control levels after insulin treatment. On the other hand, the coversion of [³H] 25 OHD₃ to [³H] 1, 25-(OH) ₂D₃ was enhanced significantly in glibenclamide-treated diabetic rats in vitro. Furthermore, in diabetic rats in vitro the conversion of [³H] 25 OHD₃ to [³H] 24, 25-(OH) ₂D₃ was accelerated but this was suppressed with insulin treatment. It is speculated, therefore, that chronic insulin deficiency produced an impaired renal 1 α-hydroxylase activity and an enhanced renal 24-hydroxylase activity. Insulin treatment improved these abnormal activities of 1 α-hydroxlase and 24-hydroxylase. The oral administration of glibenclamide enhanced renal la-hydroxylase activity.
These alterations in vitamin D₃ metabolism may be related to deranged mineral homeostasis and skeletal morphology stemming from chronic insulin deficiency.

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The factors affecting lecithin cholesterol acyltransferase (LCAT) activity, the changes in LCAT activity and the mechanism of this phenomenon in diabetics, were investigated and lipoprotein-X (LP-X), regarded as the marker of an upset in the balance of the LCAT-lipids system was measured in diabetics to determine whether the LCAT-lipids system balance was maintained or not.
In untreated diabetics both the levels of LCAT activity and hyperlipidemia were significantly higher than those of the normal control group. Passive correlations were found between LCAT activity and lipids: total cholesterol, triglyceride and phospholipid (r=0.637, r=0.582, r=0.561 respectively). Conversely, negative correlations were found between LCAT activity acceleration rate and these lipids. Though the administration of glucose had no influence on the LCAT activity and LCAT acceleration rate, the infusion of 10% soybean oil emulsion raised LCAT activity. No parallel changes were found between LCAT activity and exogenous lipids that were to become the substrate of LCAT.
No LP-X was detected in diabetics, and it was only after long-term administration of 10% soybean oil emulsion in large quantities that the results turned positive Neither did the administration of glucose (50g) nor a single infusion of 10% soybean oil emulsion (1ml/kg) show any LP-X
It may be concluded from the above that
(1) Lipids significantly affect LCAT activity.
(2) In untreated diabetics LCAT activity increases and the mechanism of this phenomenon is that an increase in lipids rather than in glucose causes LCAT activity to rise.
(3) The balance of the LCAT-lipids system in diabetics is maintained.

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The present investigation was conducted to determine the effect of exogenously given insulin on feeding behavior. Female Wistar rats were individually housed on a 12-12 light-dark cycle and maintained with Purina chow diet and tap water available ad lib. Rats were injected s.c. with NPH insulin (8 U/day) just before the light cycle (L-NPH) or dark cycle (D-NPH). Their behavior was carefully analyzed, by means of the Gunma University type automatic apparatus for continuous and direct measurement of eating, drinking and ambulation. The following results were obtained:
1) Blood sugar level: There was no difference in the degree of hypoglycemia during L-NPH and D-NPH after insulin administration.
2) Food Intake: The increase in food intake in the L-NPH-treated group was times greater during the light cycle than in the D-NPH-treated group and saline controls. In contrast, the increase in food intake in the D-NPH-treated group was 1.5 times greater during the dark cycle than in the L-NPH-treated group and controls.
3) Drinking: The drinking volume increased immediately in each phase after insulin administration. The periods which affected the increased volume of drinking appeared to be longer during L-NPH than D-NPH.
4) Ambulation: Both insulin-treated groups showed depressed ambulatory activity, except for L-NPH during the light cycle.
5) Body weight: The degree of weight gain was greater in the L-NPH-treated than in the D-NPH-treated group.
In conclusion, there is a significant difference in the animal behavior of feeding and ambulation according to the time of insulin administration. This may be related to the difference in weight gain following insulin treatment.

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Using human insulins (HI), we studied the effect of mixing short-and intermediate-acting insulins on the absorption kinetics of short-acting insulins.
In one set of experiments, the serum immunoreactive insulin (IRI) and the blood glucose responses to the subcutaneous injection of a mixture of short-and intermediate-acting HI were compared with those of separrte injections in rabbits. The prompt injection of Actrapid HI mixed with Monotard HI (1: 1, 1: 3) in one syringe caused a prolongation of time to reach a maximum level in serum IRI and a delay of time to produce a nadir level in blood glucose. The prompt injection of regular HI mixed with NPH-HI (1: 1) in one syringe caused a decrease of maximum level in serum IRI and a delay of time to produce a nadir level in blood glucose. The prompt injection of regular HI mixed with NPH-HI (1: 3) in one syringe caused no change of serum IRI kinetics.
In another set of experiments, in order to assess the effect of zinc ions on the absorption kinetics of Actrapid HI, the serum IRI responses to the subcutaneous injecton of a mixture of Monotard HI supernatant (1000×g, 20 min) and Actrapid HI were investigated. Changing the Monotard HI supernatant: Actrapid HI ratio from 1: 1 to 3: 1 resulted in a decrease of the maximum level andα prolongation of time to reach a maximum level in serum IRI.
These in vivo data indicate that the absorption of Actrapid HI may be retarded when mixed with Monotard HI as the proportion of the latter increases and that the absorption of regular HI may be reduced when mixed with NPH-HI (1: 1)

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The dynamics of insulin release were examined in perfused monolayer-cultured B cells from neonatal rat pancreases that been maintained for 7 days in glucose-depleted TCM 199 containing 10% fetal bovine serum (FBS), 5.5 mM galactose and 0.1 M 2-deoxy-glucose. The perfusion chamber was made by placing a culture plastic sheet containing culture cells on each side of a siliconrubber sheet with a round hole in the center and fixing the sheets in place with square stainless steel plates. The basal perfusion medium used was glucose-depleted TCM 199 containing 0.5% FBS. The results are as follows:
1) On day 7, a biphasic, adult-like response to a single dose of 16.7 mM glucose was observed in cultured B cells in the glucose-depleted TCM 199. Likewise, significant increases in the second phase secretion were caused by either 10 mM leucine or 10 mM 2-ketoisocaproate Moreover, 10 mM β-aminobicyclo (2, 2, 1) heptane-2-carboxylic acid, which is a non-metabolic leucine analogue, also produced a biphasic pattern. However, 10 mM valine produced only a limited response in the first phase.
2) When exposed to linear gradient stimulation by glucose, leucine or 2-ketoisocaproate, these B cells secreted insulin in a dose-dependent fashion.
3) In the presence of 10 μM forskolin or 1 mM 3-isobutyl-1-methylxanthine, stimulation with 16.7 mM glucose produced a significant increase in either insulin release in the second phase or cAMP release, and the recovery of cellular cAMP also was markedly increased.
4) The rates of glutamine oxidation were significantly elevated by addition of either leucine 2-ketoisocaproate or β-aminobicyclo (2, 2, 1) heptane-2-carboxylic acid at 10 mM, whereas 10 mM valine had no effect.
From these results, it is suggested that even under culture conditions free of glucose, neonatal rat B cells in monolayers survive and mature during culture.

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A study was made on the effects of an H1 receptor agonist (2-pyridylethylamine dihydrochloride) and an H₂ receptor agonist (impromidine trihydrochloride) on the release of insulin from perfused rat pancreas and the isolated pancreatic islets of the rat. To elucidate the relationship of histamines to the adrenergic nervous system an α-blocker (phentolamine) or a β-blocker (propranolol) was administered. It was revealed that the administration of pyridylethylamine (0.1β M) significantly inhibited insulin secretion induced by 8.4mM glucose while that of phentolamine (1β M) mitigated this inhibitory action.
From the perfused rat pancreas, insulin secretion from the perfused rat pancreas was not affected by impromidine (0.1β M), but was slightly increased by phentolamine (1β M) added to the perfusate prior to the administration of impromidine, and was slightly decreased by propranolol (2β M) addition. The administration of pyridylethylamine (0.1β M) and impromidine (0.1β M) had no effect on the release of insulin from the isolated islets in response to 8.4-mM glucose.
Thus it was suggested that histamines influence insulin secretion in the pancreas, and that their effects are exterted at least in part through the adrenergic nervous system.

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An autopsy case of juvenile-onset diabetes in a patient who died of acute necrotizing pancreatitis following massive hyperlipidemia is reported. A 23-year-old woman was first admitted to the hospital in 1978 because of eruptive xanthoma which appeared at the age of 19. She had no family history of diabetes or hyperlipidemia but her parents were cousins. She was diagnosed as having type III hyperlipidemia and non-insulin-dependent diabetes mellitus. Both lipoprotein lipase (LPL) and hepatic triglyceride lipase (HTGL) activities were decreased. Restriction of energy intake resulted in marked improvemnt of the hyperglycemia and hyperlipidemia together with increases in LPL and HTGL activities. On the second and third admission, she was treated with glibenclamide and insulin. Through-out the course, levels of fasting blood glucose, triglyceride, cholesterol and broad beta band changed in parallel, while LPL and HTGL activities varied inversely with these parameters. On Sept 14, 1982, (age 27), she was admitted to the hospital because of severe abdominal pain and vomiting since the previous evening. On admission, she was stuporous, and blood pressure was 76/mmHg. The abdomen was distended and tender. Xanthoma was seen on the lower legs. The plasma was milky and the triglyceride level was 9, 000 mg/dl, cholesterol 1, 000 mg/dl, and chylomicron 8, 500 mg/dl. The blood glucose level was 556 mg/dl, urine ketone was positive, and the pH was 7.322. The urine amylase level was 39, 369 units. Treatments with continuous insulin infusion, fluids and heparin, and replacement transfusion were ineffective. She died on the next day. The autopsy revealed acute hemorrhgic pancreatitis. The pancreas was enlarged and there was hemorrhage and coagulation necrosis in the body and tail, and fat necrosis in the interlobular portions of the pancreatic head and adjacent adipose tissue. Veins and capillaries at the site of coagulation necrosis were filled with thrombi including cholesterol crystals. Lymphatics in the serosa of the stomach, and the small and large intestines were filled with chyle. The acute hemorrhgic pancreatitis was probably due to massive hyperchylomicronemia to which she was predisposed by the combination of subnormal LPL activity and diabetes mellitus, but the direct trigger of the terminal episode is unknown.

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Little is known about changes of ambulatory activity in diabetic animals. The present study was undertaken to determine such activity in streptozotocin (STZ)-induced diabetic rats. In addition, the turnover of dopamine in the rat striatum, which has been thought to be closely related to locomotor activities, was investigated.
The results of the present study were as follows:
Experiment 1. Ambulatory activity was gradually decreased following increase in the blood glucose level. The degree of the decrease was signficantly related to the elevation of blood glucose level (r=-0.76, p< 0.05)
Experiment 2. We measured the turnover rate of dopamine as DOPAC (3, 4-dihydroxyphenylacetic acid)(ng/g)/DA (Dopamine)(ng/g). In diabetic rats, dopamine turnover rate was significantly decreased (STZ-Gronp 0.102± 0.003 vs. Control Group 0.112± 0.003 (p< 0.05)). The ratio of the decrease was also significantly correlated with the elevation of blood glucoseluevel (r==0.693, p< 0.01).
It is suggested that the change of dopamine turnover in the striatum may play an important role, at least in part, in ambulatory activity.

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Changes in the subsets of spleen lymphocytes in non-treated and Cyclosporin (Cs)-treated NOD mice were anaylsed using immunofluorescent techniques by FACS. The number of spleen cells in non-treated NOD mice aged 120 days was significantly more decreased than that in ICR mice of the same age. All of the percentages of Thy 1.2, Lyt 1 and Lyt 2 positive cells in non-treated NOD mice were significantly increased compared with those in ICR mice. Moreover, both percentages of Thy 1.2 and Lyt 1 positive spleen cells in Cs-treated NOD mice were significantly decreased compared with those in non-treated NOD mice, while no significant difference was observed in the percentage of Lyt 2 positive spleen cells between non-treated and Cs-treated NOD mice. The ratio of Lyt 1+ cells/Lyt 2+ cells in Cs-treated NOD mice was significantly more decreased than that in non-treated NOD mice. These results suggest that Cs may exert its suppressive effect on development of insulitis in NOD mice by altering the compartmentalization of lymphocyte subsets.

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