12-O-Tetradecanoylphorbol-13-acetate (TPA) has been used as a potent tumor promoter in mouse skin. The mechanisms of TPA actions were studied by using several types of inhibitors. TPA-caused responses in mouse skin such as skin tumor promotion, epidermal ornithine decarboxylase (ODC) induction and skin inflammation were inhibited by various lipoxygenase inhibitors of the arachidonic acid cascade. Lipoxygenase inhibitors also inhibited TPA-caused ODC induction in isolated epidermal cells or cultured epidermal cells. Therefore, it is possible that these drugs inhibit TPA-caused ODC induction in mouse skin by directly acting on epidermal cells. TPA actions were also inhibited by either protein kinase C inhibitors, calmodulin antagonists or calcium blockers. These results suggest that arachidonic acid/lipoxygenase, protein kinase C and calcium-calmodulin systems play essential roles in the mechanism of tumor promotion by TPA.

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The present study was designed to examine the selectivity of ³H-CGP-12177 (4-(3-t-butylamino-2-hydroxypropoxy)-[5, 7-³H]benzimidazole-2-one hydrochloride) for β₁- and β₂-adrenergic receptors by the Scatchard and the displacement analysis. The plots of specific binding obtained from the Scatchard analysis using ³H-CGP12177 for the rat myocardium membrane were uniphasic when the non-specific binding was detemined by the use of 10 μM I-propranolol, and the K_d and B_max values were 408.53±67.20 pM and 12.27±0.83 fmoles/mg protein, respectively. On the other hand, two binding sites were observed in the displacement curve when I-metoprolol was used as a competitor. The existence of these two binding sites implied the selectivity of ³H-CGP12177 to β-adrenoceptors because ³H-CGP12177 was 1.8-fold more selective towards β₁-adrenoceptors than β₂-adrenoceptors. In addition, these two binding sites could be regarded as β₁-and β₂-adrenergic receptors from the evaluation of the binding characteristics of β-adrenoceptors by the displacement analysis using β-selective antagonists. Thus, ³H-CGP12177, a hydrophilic radioligand, was useful for the binding assay of β-adrenoceptors in rat myocardial membranes.

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(±)isofloxythepin (0.32-3.2 mg/kg, i.p.) significantly inhibited in a dose-dependent manner wet-dog shaking (WDS) induced by electrical stimulation of the rat hippocampus. In addition, both optical isomers of isofloxythepin inhibited WDS, with the (-)-isomer being almost 3 times more potent than the (+)-isomer. Other neuroleptics such as haloperidol, chlorpromazine, zotepine and sulpiride also reduced significantly the number of WDS. The inhibitory potency of haloperidol was comparable to that of (±)isofloxythepin, which was approximately 3 times more potent than that of chlorpromazine or zotepine. Sulpiride suppressed significantly WDS only at the high dose of 100 mg/kg. None of the drugs affected hippocampal afterdischarge. Inhibition of WDS produced by (±)isofloxythepin or haloperidol was antagonized by pretreatment with a dopamine receptor agonist, lisuride. The present results indicate that isofloxythepin shares with other neuroleptics an inhibitory effect on WDS; dopaminergic blocking action appears to be important in the inhibition of WDS induced by hippocampal stimulation.

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Effects of adrenalectomy (AD) on ethanol-induced gastric injury and prostaglandin (PG) protection on the damage were investigated in rats and compared with those of N-ethylmaleimide (NEM), a sulfhydryl (SH) blocker, and diethyl maleate (DEM), a SH depletor. Oral administration of 100% ethanol (1 ml) induced elongated bands of hemorrhagic lesions in the corpus mucosa of sham operated rats, and these lesions were significantly prevented by 16, 16-dimethyl PGE₂ (dmPGE₂, 10 μg/kg, s.c.). AD markedly enhanced the mucosal ulcerogenic responses caused by ethanol and abolished the protective effect of dmPGE₂; this agent rather worsened the lesions, which appeared throughout the corpus mucosa. AD by itself enhanced the microvascular permeability in the gastric mucosa without any effect on SH contents. These alterations caused by AD were significantly reverted by hydrocortisone treatment (10 mg/kg/day for 2 weeks, s.c.). On the other hand, a single injection of NEM (10 mg/kg, s.c.) similarly enhanced the vascular permeability, worsened the ethanol-induced lesion, and mitigated the protective effect of dmPGE₂ without altering mucosal SH contents, while DEM (1 ml/kg, s.c.) significantly reduced the mucosal SH levels and the lesions. These results suggest that AD worsened the mucosal lesions induced by ethanol, probably by enhancing the ricrovascular permeability, and this action may be due to a lack of steroid secretion but is not directly related to a mucosal SH deficiency.

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The effect of electrical stimulation of anterior parts of the hypothalamus on gastric acid secretion was examined in rats anesthetized with urethane. Electrical stimulation (0.5 mA, 2 msec, 10 Hz, for 10 min) of the lateral hypothalamic area (LHA) induced a marked increase in basal gastric acid output. Electrical stimulation of the preoptic area (PO), the paraventricular nucleus (PVN) and the anterior hypothalamus (AH) also induced increases in basal acid secretion. Electrical stimulation of the PO, PVN and AH inhibited the increase in acid secretion induced by electrical stimulation of the vagus nerve (1 mA, 0.5 msec, 3 Hz). These inhibitory effects were abolished by bilateral cutting of the splanchnic nerves. On the other hand, the stimulation of the LHA had no effects on vagally stimulated gastric acid secretion. Electrical stimulation of the anterior parts of the hypothalamus induces dual effects, vagally mediated increases in basal gastric acid output and splanchnic mediated inhibition of the vagally stimulated gastric acid output.

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To elucidate the toxic and protective mechanisms responsible for cephaloridine (CER)-induced nephrotoxicity, changes in renal formation of malondialdehyde and renal activities of xanthine oxidase, superoxide dismutase and catalase were mainly investigated for 15 days in rats that received single intravenous injections of CER in doses of 100 and 1, 000 mg/kg body weight. In the 100 mg/kg group, the above items determined remained within the control levels. In the 1, 000 mg/kg group, renal formation of malondialdehyde was observed to be accelerated with the following two stages: highly in the early stage (the 3rd hour to the 2nd day, especially at the 3rd hour) and more highly in the late stage (the 2nd to the 7th day). Concerning the other items determined, significantly different changes were hardly observed in the 1, 000 mg/kg group within the 12th hour of the early stage, while the rises in renal activities of xanthine oxidase and falls in renal activities of superoxide dismutase and catalase were observed in the late stage. These results suggested that the increment in malondialdehyde formation in the late stage might be explained enzymatically by both the rises in the activities of xanthine oxidase and the declines in the activities of superoxide dismutase and catalase and that those in the early stage did not relate directly to the above renal enzymatic systems.

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In order to establish an assay for the in vitro growth of myogenic cells, we investigated whether creatine kinase (CK) activity can be used as an index for the estimation of growth. The content of CK activity in a primary myogenic cell culture, almost all of which originated from myotubes, was shown to be linearly proportional to the content of proteins in the myotubes and increased concomitantly with myotube growth. CK activity can be easily and accurately assayed and used for routine assay of the cell growth. As an example of the application of CK activity for the bioassay of growth promoting substances, the relative potency (Pr) of turkey transferrin (Tf) to chick Tf in the promotion of chick myogenic cells was examined with a parallel line assay. The calculation limited that Pr=0.363, ranging from 0.356 to 0.370, at 95% safety. The results demonstrated the usefulness of CK activity for estimating cell growth, as well as the value of a statistical method for the assay of a growth factor in vitro.

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Effects of nifedipine (10^-8-10^-6 M), verapamil (10^-6-3×10^-5 M) and Bay k 8644 (10^-7 M) on the contractions induced by transmural nerve stimulation or agonists were investigated in the circular smooth muscle of guinea pig vas deferens using a ring preparation. Transmural stimulation (20 Hz for 20 sec) produced biphasic contractions consisting of initial phasic and secondary tonic components. Nifedipine preferentially inhibited the tonic component and norepinephrine (NE)-induced contractions over the phasic component and β, γ-methylene ATP (mATP)-induced contractions. Nifedipine suppressed the enhancing effect of NE on the mATP-induced contractions. The tonic component and NE-induced contractions were inhibited by verapamil in a similar dose range. Verapamil slightly inhibited the phasic component, but rather enhanced mATP-induced contractions. Bay k 8644 slightly increased the phasic component and mATP-induced contractions in the presence of prazosin. Bay k 8644 slowed the relaxation of the tonic component, thereby prolonging the tonic component. Bay k 8644 also prolonged the duration of NE-induced contractions. These results, except for the contradictory effects of verapamil between the phasic component and mATP-induced contractions, are consistent with the claim that ATP and NE are responsible for the generation of phasic and tonic component, respectively, in the circular muscle of guinea pig vas deferens.

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Effects of propranolol and atenolol on isoproterenol-induced responses of isolated atrial and papillary muscles from experimentally diabetic rats were examined. Male Sprague-Dawley rats were divided into the diabetic group (DM) which received streptozotocin 60 mg/kg, i.v. and the control group (C) which received vehicle i.v. At 6 weeks after, the right atrial or right ventricular papillary muscle was isolated, and the beating rate (R) in the atrium or isometric force development (F) and its first derivatives (±dF/dt) in the papillary muscle under pacing were recorded. Basal R was less frequent in DM than in C, but ED50 values for isoproterenol-induced chronotropy were not different between the two groups. Basal F and basal ±dF/dt were not different between the two groups, but isoproterenol-induced increases in F and ±dF/dt were less in DM than in C. ED50 values in F and +dF/dt were not different between the two groups. Propranolol and atenolol shifted the concentration-response curves in R, F and ±dF/dt for isoproterenol to the right in both groups. pA₂ values of propranolol and atenolol for each parameter were not different between the two groups. Results indicate that propranolol and atenolol exert the same beta-adrenoceptor blocking potency in both diabetic and non-diabetic hearts of rats.

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The effects of DN-9693, a synthesized phosphodiesterase inhibitor, on the secretion of pancreatic juice were investigated in preparations of the isolated and blood-perfused dog pancreas. DN-9693 injected intraarterially caused a dose-dependent increase in the secretion of pancreatic juice and decrease in the perfusion pressure. The threshold doses to increase the pancreatic secretion and to decrease the perfusion pressure were about 100 μg and 1 μg, respectively. Thus, the secretory response was less effective than the vascular response. The secretory activity of DN-9693 (0.3 mg) was approximately equal to that of 0.03 mg of 3-isobutyl-1-methylxanthine, 0.5 mg of papaverine, 5 mg of theophylline, 0.08 units of secretin and 0.2 units of cholecystokinin. The concentration of bicarbonate in the pancreatic juice induced by DN-9693 was increased, but protein concentration was not. DN-9693-induced pancreatic secretion was not modified by pretreatments with phentolamine, propranolol, atropine, sulpiride and cimetidine. Secretin-induced pancreatic secretion was significantly potentiated by infusion of DN-9693 (10 μg/min), but cholecystokinin-induced one was not. From these results, it is concluded that DN-9693 may produce an increase in pancreatic secretion by acting directly on the pancreatic exocrine gland of the dog, which might be mediated through an increase of intracellular cyclic AMP concentration by inhibiting phosphodiesterase activity.

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Flunarizine relaxed isolated canine arteries precontracted with prostaglandin (PG) F_2α, epithio-methano-thromboxane A₂ and K⁺; the relaxation was in the order of cerebral>renal>mesenteric=coronary arteries, when contracted with PGF_2α or the thromboxane A₂ analogue. Flunarizine-induced relaxation was unaffected by treatment with atropine, propranolol, cimetidine, chlorpheniramine, aminophylline and indomethacin, and by removal of endothelium. Under normoxia, flunarizine attenuated contractions elicited by Ca²⁺ in the K⁺-stimulated cerebral and mesenteric arteries that had been previously exposed to Ca²⁺-free media to a greater extent than that in PGF_2α-stimulated preparations. The Ca²⁺-induced contraction in cerebral arteries was more sensitive to flunarizine than that in mesenteric arteries. Contractions caused by PGF_2α in Ca²⁺-free media were not influenced by flunarizine. In cerebral and mesenteric arteries that had been previously exposed to Ca²⁺-free media and severe hypoxia and then stimulated by PGF_2α and Ca²⁺, reoxygenation produced a persistent contraction. Flunarizine suppressed the reoxygenation induced-contraction. It is concluded that flunarizine dilates cerebral arteries predominantly over the other arteries; the vasodilatation appears to derive from an interference with the transmembrane Ca²⁺ influx that occurs through a voltage-dependent process and, to a lesser extent, receptor-operated channels, but not with the Ca²⁺ release from stored sites. Contraction induced by reoxygenation is expected to be due mainly to the transmembrane influx of Ca²⁺, which is also suppressed by flunarizine.

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Progesterone enhanced the contractile effect of epinephrine on the ductus deferens of the guinea pig in vitro. In relation to this mechanical phenomenon, we examined the phosphatidylinositol metabolism. In the ³H-myoinositol labeled ductus deferens, radioactivity in phosphatidylinositol bisphosphate was about 2.6 times as high as that in phosphatidylinositol. Phosphatidylinositol and phosphatidylinositol phosphate were not changed by epinephrine (100 μM), but phosphatidylinositol bisphosphate was increased at 10 sec and 1 min after the administration of epinephrine (100 μM). Progesterone (100 μM) added 5 min before the administration of epinephrine increased the stimulatory effect of epinephrine on the phosphatidylinositol bisphosphate metabolism, but had no effect on the phosphatidylinositol and phosphatidylinositol phosphate metabolism. These studies suggest that progesterone expresses its activity not through the cytoplasmic progesterone receptor but through the epinephrine-mediated smooth-muscle contractile mechanism.

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Progressive passive Heymann nephritis (group II) was induced in rats by i.v. injections of rabbit antiserum against an antigen from the brush border of the proximal tubules of rat kidney following immunization with rabbit γ-globulin in Freund's complete adjuvant, and the process of the disease was compared with that of rats that received the antiserum alone (group I). The rats of group I showed proteinuria (30-50 mg/day) and plasma cholesterol content (80-90 mg/dl) slightly higher than the normal level from the 17th or the 22nd day after antiserum injection to the 90th day. The rats of group II revealed a heavy proteinuria (300-400 mg/day) and hypercholesterolemia (approx. 200 mg/dl) during the same period. In group II, there were the thickening of glomerular basement membrane (GBM) and spike formation. Moreover, granular deposits of rat IgG, rabbit IgG and rat C₃ were observed along the GBM. These changes were weaker in group I. When given orally, daily, azathioprine (20 mg/kg) and prednisolone (1 and 3 mg/kg) showed a beneficial effect on the nephritis in group II. The group II model closely resembles human glomerulonephropathy and may be useful for studying the effect of medication on glomerulonephropathy.

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In order to investigate the behavioral characteristics of the SART-stressed (repeated cold-stressed) animal, a model of dysautonomia, step-down passive avoidance performance was examined in SART-stressed mice. SART-stressed mice exhibited a shortened test trial latency and a decreased incidence of maximum latency of 300 sec, but no change in the training latency. These alterations were blocked by single administration of chlorpromazine or carpipramine prior to the training trial. Repeated, but not single treatments with neurotropin and hopantenate improved the impaired performance due to SART stress. On the other hand, alprazolam and diazepam were ineffective by either mode of administration. Thus, SART-stressed mice appear to have impairment in the process of acquisition of a passive avoidance task.

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Effects of S-adenosyl-L-methionine on Ca accumulation as well as the changes of electrolytes contents and glucose metabolism after transient ischemia were investigated. In the 4-vessel occlusion model of rats, brain Ca content increased to 200-250% of that in sham operated rats 1 day after 60 min transient ischemia. The change in the striatum was more severe than that in the cortex or hippocampus. Na content increased and K content decreased, and glucose, pyruvate and lactate contents increased significantly in the striatum 1 day after transient ischemia. SAM (100 mg/kg, i.p.) was injected at the end of occlusion and every hour for 5 hr thereafter. Treatment with SAM reduced Ca accumulation, decrease of K, and increase of lactate and pyruvate in the striatum significantly. The present results suggest that the inhibition of Ca accumulation might partly explain the beneficial effects of SAM on the cerebral dysfunction following ischemia.

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MK-733 was found to prevent an increase of serum cholesterol levels in cholesterol-fed rabbits, and lovastatin also markedly inhibited their increase. MK-733 and lovastatin inhibited the increase of very low density lipoprotein (VLDL) and low density lipoprotein (LDL) cholesterol, and it slightly affected the high density lipoprotein (HDL) cholesterol levels. MK-733 and lovastatin suppressed the increase of serum phospholipid levels and slightly affected the triglyceride levels. MK-733 suppressed the development of atherosclerosis in coronary arteries and aorta, and lovastatin also diminished their development.

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Dioxamide derivatives of 1, 3, 4-thiadiazole, synthesized earlier by Oke (1986), were screened for hypoglycemic activity. Alloxan-induced diabetic albino rats were used in the study. The structure-activity relationship of the dioxamide derivatives of 1, 3, 4-thiadiazole is discussed. The study shows that the dioxamide derivatives of 1, 3, 4-thiadiazole are more potent and of longer duration of action when compared with the oxamide derivatives of 1, 3, 4-thiadiazole, which were previously reported by Oke and Cherynk (1981).

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Time-dependent changes in the blocking effect of diltiazem on norepinephrine (NE)-induced vasoconstrictions were investigated in isolated canine pulmonary veins. Within 2-3 hr of the perfusion period, pretreatment with diltiazem did not affect the vasoconstrictor responses to NE. However, in the 8-11 hr perfused preparations, diltiazem suppressed the NE-induced vasoconstriction significantly. It may participate in the mechanisms for the appearance of the blocking effect in the following manner: 1) a proportion of alpha adrenoceptors which are sensitive to calcium entry are increased and/or 2) spare alpha adrenoceptors are decreased time-dependently.

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The brain histidine level in mice dose-dependently increased 1 and 2 hr after an i.p. injection of 0.5-1.5 g/kg of L-histidine. The treatment with 1.0 and 1.5 g/kg but not 0.5 g/kg of L-histidine significantly decreased the brain levels of tyrosine, phenylalanine and some other amino acids 1 hr later. A complete recovery or a rebound rise of amino acid levels was observed 2 hr after treatment. These results indicate that there is a change in the transport of amino acids into the brain after treatment with large doses of L-histidine.

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The effect of disulfiram on the urotoxicity induced by ifosfamide was studied in mice. Ifosfamide administered intraperitoneally to mice caused a dose-related increase in bladder weight within 48 hr of treatment. Disulfiram prevented ifosfamide-induced bladder damage when administered orally within 1 hr of ifosfamide treatment. The results indicate that disulfiram is an effective protective agent against bladder damage caused by ifosfamide treatment.

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The effect of diethylnitrosamine (DEN), a well-known experimental carcinogen, toward MAO-A and MAO-B activity of rat liver was investigated. The oxidations of both β-PEA (MAO-B) and 5-HT (MAO-A) were inhibited by DEN. The K_i values of DEN in the inhibition of rat liver MAO-A and MAO-B activity were determined. The kinetic data show that DEN is a competitive, MAO-B selective inhibitor and its inhibitory effect on MAO-B is about 4-fold more potent than that on MAO-A. DEN might change the proportions of the multiple forms of MAO activity in tumor cells.

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