Antithrombotic effects of KW-3635, a newly synthesized thromboxane (TX) A-receptor antagonist, were studied in guinea pigs. In the extracorporeal circulation thrombosis model, the shunt was filled with thrombi, and reduction of platelet count and increase in plasma TXA₂ concentration were observed. KW-3635 (30 and 100 mg/kg, p.o.) inhibited the thrombus formation in the shunt and prevented the decrease in platelet count in the circulating blood without affecting the red blood cell count. BM13, 505 (30, 100 mg/kg, p.o.), another TXA₂-receptor antagonist, and ticlopidine (300 mg/kg, p.o.), an antiplatelet drug, also inhibited the thrombus formation, while aspirin (10, 300 mg/kg, p.o.) did not. Peripheral arterial occlusive disease was induced by injection of sodium laurate into the femoral artery in guinea pigs. Daily oral administration of KW-3635 (3 30 mg/kg) significantly prevented the progression of vascular lesions. BM13, 505 (3-30 mg/kg, p.o.) and ticlopidine (100 mg/kg, p.o.) also ameliorated the vascular lesions, whereas aspirin (10, 100 mg/kg, p.o.) did not. KW-3635 at concentrations up to 10^-4 M did not affect coagulation parameters in vitro. These results suggest that TXA₂ is involved in the pathogenesis of arterial thrombotic and ischemic disorders. KW-3635 may be useful for the treatment of thrombotic disease and peripheral arterial occlusive diseases.

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Structure-activity relationship of the chicken calcitonin gene-related peptide (cCGRP) was investigated and compared with human-α-CGRP (hCGRP) and rat CGRP (rCGRP) in the perfused mesenteric vascular beds of rats. In precontracted mesenteric vascular beds, cCGRP, hCGRP and rCGRP produced a concentration-dependent vasodilation. The vasodilator activities of cCGRP and rCGRP were equipotent and more potent than that of hCGRP. (Ala^{2, 7})cCGRP and (Des-1-Ala)-α-deamino cCGRP were 100 and 10-fold less potent than cCGRP, respectively. However, cCGRP[1-36] reduced vasodilator activity. The cCGRP [8-37] produced a vasoconstriction. Both cCGRP [8-37] and hCGRP [8-37] caused parallel shifts of the concentration-response curves of rCGRP to the right, but both peptides did not affect the maximum response and vasodilator responses to isoproterenol, these peptides being equipotent antagonists (pA₂ values: 7.6 and 7.4). In rat brain membrane preparations, cCGRP and its fragments and analogues, except for cCGRP[1-36], competed for specific binding sites with [¹²⁵I]-hCGRP. These results suggest that cCGRP has a potent vasodilator activity for which the disulfide bridge at position 2 and 7 of cCGRP is an important site and that position 37 is necessary for the binding of the peptide to the receptor. Also, cCGRP [8-37] is a competitive CGRP-receptor antagonist.

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The release and synthesis of neuronal histamine are regulated by histaminergic autoreceptors named as histamine H₃ receptors. The development of radiolabeled histamine H₃ antagonists is needed to characterize the binding of antagonists to these receptors. Here we describe the binding characteristics of a new histamine H₃-receptor antagonist, [³H]S-methylthioperamide (SMT), to rat tissues, and compare its binding with that of [³H](R)α-methylhistamine ((R)αMH), a selective histamine H₃-receptor agonist. The binding of [³H]SMT to the membranes of rat forebrain was found to be stereoselective, saturable, reversible and temperature-dependent. Saturation binding experiments indicated a single class of high affinity sites for [³H]SMT in forebrain membranes (K_D=2.1 nM, B_max=24.3 pmol/g of tissue at 4°C). The B_max was approximately 3 times that of [³H](R)αMH binding to rat forebrain membranes (K_D=2.5 nM, B_max=7.3 pmol/g of tissue at 25°C). Autoradiographic images of [³H]SMT binding in the brain were essentially the same as those of [³H](R)αMH. [³H]SMT also bound appreciably to peripheral tissues (the liver, adrenal, stomach, ileum, kidney, lung and bladder), whereas the [³H](R)αMH bindings to these peripheral tissues were negligible. These results indicate that [³H]SMT binds to H₃ receptors primarily in the central nervous system, and that it also has high affinity toward non-H₃ receptors, probably hemoproteins, in peripheral tissues.

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Adrenergic regulation of prostaglandin (PG) biosynthesis was investigated in 1-¹⁴C-arachidonic acid-prelabeled cultured rabbit gastric epithelial cells (RGECs). RGECs expressed adrenergic α₁-and β-receptors and muscarinic receptors. Norepinephrine facilitated the synthesis of PGs I₂ (determined as the stable metabolite 6-keto PGF_1α) and E₂ and hydroxyfatty acids, while epinephrine facilitated the synthesis of PGI₂ and hydroxyfatty acids, but not PGE₂. However, isoproterenol did not affect PG biosynthesis. The effects of norepinephrine and epinephrine on PG biosynthesis were markedly suppressed by the non-selective α-blocker phentolamine and/or the selective α₁-blocker prazosin. In combination with epinephrine, the selective α₂-blocker yohimbine or the β-blocker propranolol facilitated PGE₂ synthesis. Acetylcholine did not affect PG biosynthesis. These results indicate that norepinephrine and epinephrine act on PG biosynthesis as adrenergic agonists in these cultured RGECs and that an α₁-receptor couples PGI₂ and PGE₂ synthesis.

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We examined whether opening of the ATP-sensitive potassium (K_ATP) channels in the ischemic myocardium plays an important cardioprotective role during ischemia. Dogs were anesthetized with sodium pentobarbital (30 mg/kg, i.v.). Sixty minutes after treatment of the dog with glibenclamide (0.3 or 3 mg/kg, i.v.), the LAD was ligated. At 3 or 15 min after LAD ligation, left ventricular tissue was taken from the ischemic region to measure tissue metabolite levels. After ischemia, the tissue levels of ATP and creatine phosphate decreased to 49-74& and 26-34%, respectively, and lactate level increased to 380-660%. Ischemia (either 3 or 15 min) increased the levels of G6P and F6P and decreased the FDP level, indicating the inhibition of glycolysis. Glibenclamide at either dose decreased the level of blood glucose by 20-30% and increased the blood insulin level twice. The decrease in ATP and increase in lactate due to ischemia were significantly enhanced by glibenclamide at a dose of 3 mg/kg. The increase in G6P due to 15 min of ischemia were also enhanced significantly by 0.3 and 3 mg/kg of glibenclamide. Glibenclamide worsened the metabolic alterations produced by ischemia. These results suggest that KATP channels that can be inhibited by glibenclamide may perform some functions in the ischemic myocardium.

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KW-5092({1-[2-[[[5-(piperidinomethyl)-2-furanyl]methyl]amino]ethyl]-2-imidazolidinylidene} propanedinitrile fumarate) is a novel gastroprokinetic agent with acetylcholinesterase (AChE) inhibitory activity and acetylcholine (ACh) release facilitatory activity. The present study examined the effects of KW-5092 on gastrointestinal (GI) motor activity in dogs. In anesthetized dogs, KW-5092 at 0.03 to 1 mg/kg, i.v. dose-dependently enhanced the gastric antral and the colonic motor activity. Neostigmine, an AChE inhibitor, enhanced the motor activity at 0.03 and 0.1 mg/kg, i.v. Ranitidine, a histamine H₂-receptor antagonist with AChE inhibitory activity and ACh release facilitatory activity, enhanced the motor activity but decreased blood pressure at 1 to 10 mg/kg, i.v. In conscious dogs, KW-5092 at 0.03 to 1 mg/kg, i.v. or 1 to 10 mg/kg, p.o. dose-dependently enhanced the gastric antral, duodenal, ileal and the colonic motor activities. Neostigmine at 0.1 mg/kg, i.v. or 3 mg/kg, p.o. enhanced the duodenal, ileal and colonic motor activities, but induced excitement, slavering, vomiting and diarrhea. Ranitidine at 3 mg/kg, i.v. enhanced the gastric antral and colonic motor activities, but induced collapse or akinesia. The present results suggest that KW-5092 enhances the GI motor activity in a wide range from the gastric antrum to the colon and does not induce behavioral and cardiovascular side effects. KW-5092 may be a useful drug for the treatment of GI motility dysfunctions.

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Effects of acteoside (ACT) on crescentic-type anti-GBM nephritis in rats were investigated. When rats were treated with ACT from the 1st day after i.v. injection of anti-GBM serum, ACT inhibited the elevation of protein excretion into urine. In the ACT-treated rats, cholesterol and creatinine contents and antibody production against rabbit γ-globulin in the plasmas were lower than those of the nephritic control rats. Histological observation demonstrated that this agent suppressed hypercellularity and the incidence of crescent formation, adhesion of capillary wall to Bowman''s capsule and fibrinoid necrosis in the glomeruli. Furthermore, rat-IgG and C₃ deposits on the GBM were significantly less in the ACT-treated group than in the control nephritic group. When the treatment was started from the 20th day after i.v. injection of anti-GBM serum, by which the disease had been established, ACT resulted in a similar effect on the nephritic rats as stated above. These results suggest that ACT may be a useful medicine against rapidly progressive glomerulonephritis, which is characterized by severe glomerular lesions with diffuse crescents.

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A neurotrophic factor that supports the development of cholinergic retinal neurons was purified from media conditioned by a primary culture of embryonic hippocampal neurons. Retinal choline acetyltransferase (ChAT), which is located exclusively in amacrine cells, served as a marker for the development of retinal cholinergic neurons. In a serum-free control culture, retinal neurons from 17-day-old rat fetus displayed little increase in the enzyme activity and a low proportion of neurite-bearing cells (15-20%) within 7 days. The conditioned media, when added to the retinal neuron culture, dose-dependently increased ChAT activity and the number of neurite-bearing cells (40-60%), the maximum ChAT activity being ∼sixfold higher than that in the control. The fraction with these stimulatory activities was purified by Sephadex G-15 column chromatography and two times reverse-phase HPLC. The final fraction showed ∼3, 000-fold higher purification as compared with that in the Sephadex G-15 fraction. Gas-phase protein sequencing analysis of the final fraction yielded a peptide sequence: Tyr-Leu-Leu-Pro-Ala-Gln-Val-Asn-Ile-Asp. A synthetic peptide with this sequence dose-dependently stimulated ChAT activity in the retinal cell culture and dissociated cell culture of the septal nucleus. These findings suggest that the developing hippocampal neurons produce a neurotrophic peptide that stimulates the development of cholinergic neurons.

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We studied the effect of KW-3635, a selective thromboxane A₂ (TXA₂)-receptor antagonist, on experimental glomerulonephritis. The glomerulonephritis was induced in mice by the administration of rabbit anti-mouse glomerular basement membrane (GBM) antibody. It was characterized as proteinuria, changes of serum biochemical parameters and glomerular histopathological abnormalities. The administration of KW-3635 (30 mg/kg/day, p.o.) significantly ameliorated the proteinuria, elevation of serum urea nitrogen and the thickening of GBM. These data suggest that TXA₂ may play an important pathogenic role in the development and progression of glomerulonephritis.

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We investigated the possible renal protective effects of KW-3902 (8-(noradamantan-3-yl)-1, 3-dipropylxanthine), an adenosine A₁-receptor antagonist, against gentamicin (GM)-induced acute renal failure (ARF) in rats. ARF was induced by subcutaneous injection of GM at 80 mg/kg/day for 12 days. KW-3902 (0.001-0.1 mg/kg, p.o., twice daily) attenuated the increases of serum creatinine and urea nitrogen and the decrease of creatinine clearance in rats treated with GM. In contrast, furosemide and trichlormethiazide aggravated the GM-induced nephrotoxicity. These results suggest that KW-3902 can ameliorate the GM-induced ARF and that endogenous adenosine may be involved in GM-induced ARF via the adenosine A₁-receptor.

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We investigated the effect of cisapride, a gastroprokinetic agent, on the obstructed gastric mucosal hemodynamics induced by indomethacin using an organ reflectance spectrophotometry system in rats. Indomethacin (10 mg/kg, i.v.) reduced both the gastric mucosal blood volume and the gastric mucosal blood oxygenation. Pretreatment with cisapride (0.1 mg/kg, i.v.) prevented these deteriorations. The presently-clarified gastric mucosal protective effect of cisapride may contribute to its therapeutic efficacy.

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S-312, S-312-d, but not S-312-1, L-type calcium channel antagonists, showed anticonvulsant effects on the audiogenic tonic convulsions in DBA/2 mice; and their ED₅₀ values were 18.4 (12.8-27.1) mg/kg, p.o. and 15.0 (10.2-23.7) mg/kg, p.o., respectively, while that of flunarizine was 34.0 (26.0-44.8) mg/kg, p.o. Although moderate anticonvulsant effects of S-312-d in higher doses were observed against the clonic convulsions induced by pentylenetetrazole (85 mg/kg, s.c.) or bemegride (40 mg/kg, s.c.), no effects were observed in convulsions induced by N-methyl-D-aspartate, picrotoxin, or electroshock in Slc:ddY mice. S-312-d may be useful in the therapy of certain types of human epilepsy.

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