Octopaminergic modulation of the single Ca2+ channel currents in Kenyon cells isolated from the mushroom body of the cricket brain.

Octopamine plays an important role in mediating reward signals in olfactory learning and memory formation in insect. However, its target molecules and signaling pathways are still unknown. In this study, we investigated the effects of octopamine on the voltage-activated Ca2+ channels expressed in native Kenyon cells isolated from the mushroom body of the cricket (Gryllus bimaculatus) brain. The cell-attached patch clamp recordings with 100 mM Ba2+ outside showed the presence of dihydropyridine (DHP) sensitive L-type Ca2+ channels with a single channel conductance of approximately 21+/-2 pS (n=12). The open probability (NPo) of single Ca2+ channel currents decreased by about 29+/-7% (n=6) by bath application of 10 microM octopamine. Octopamine-induced decrease in Po was imitated by bath application of 8-Br-cAMP, a membrane-permeable cAMP analog. Pre-treatment of Kenyon cells with the octopamine receptor antagonist phentolamine blocked the inhibitory effect of octopamine on Ca2+ channels. Pre-treatment of Kenyon cells with H-89, a selective inhibitor of cAMP-dependent protein kinase (PKA) attenuated the inhibitory effect of bath applied octopamine on Ca2+ channels. These results indicate that DHP-sensitive L-type Ca2+ channel is a target protein for octopamine and its modulation is mediated via cAMP and PKA-dependent signaling pathways in freshly isolated Kenyon cell in the cricket G. bimaculatus.

Single K+ channels in endocrine cells dispersed from the cricket (Gryllus bimaculatus) corpora allata.

Single channel currents were recorded from cell-attached patches of endocrine cells of the adult male cricket corpora allata. Three distinct types of K+ channels were identified; a weak inward rectifier (Type 1), a strong inward rectifier (Type 2) and a weak outward rectifier (Type 3). The type 1 channel had a slope conductance of 191 +/- 9 pS (n = 4) at negative membrane potentials (Vm) and 101 +/- 6 pS (n = 6) at positive Vm. In addition, the channel showed fast open-closed kinetics at negative Vm and slow open-closed kinetics at positive Vm. The open probability (Po) of this channel was strongly voltage-dependent at positive Vm, but less voltage-dependent at negative Vm. The reversal potential was not modified significantly by the substitution of gluconate for external Cl- but was modified after N-methyl-D-glucamine (NMDG+) was substituted for external K+, according to the Nernst equation for a K+-selective channel. The type 2 channel had a slope conductance of 44 +/- 2 pS (n = 5) at negative Vm, but no detectable outward current was observed at positive Vm. This channel showed very slow open-closed kinetics at negative Vm and its Po was not voltage-dependent. The type 3 channel had a limit conductance of 55 +/- 12 pS (n = 3) at negative Vm and 88 +/- 10 pS (n = 3) at positive Vm. This channel showed slow open-closed kinetics at negative Vm and fast open-closed kinetics at positive Vm. The Po for the channel was voltage-dependent at positive Vm but was voltage-independent at negative Vm. These three types of K+ channels may be important for the control of the resting membrane potential, and may thus participate in the regulation of Ca2+ influx and juvenile hormone secretion in corpora allata cells.

A Na+-dependent electrogenic glutamate transporter current in voltage-clamped cells of corpora allata in the cricket Gryllus bimaculatus.

Application of L-glutamate (1 mM) to corpora allata cells of the adult male cricket Gryllus bimaculatus caused a membrane depolarization of 5.9+/-0.3 mV (mean +/- SE) from a resting potential of -62.2+/-1.3 mV (n=57). The underlying mechanism for this depolarization was studied by applying the two-electrode voltage-clamp technique. Application of L-glutamate (1 mM) elicited an inward current that peaked at 8.1+/-0.7 nA (n = 73) at a holding potential of-50 mV. Both L- and D-aspartate also induced an inward current of almost the same amplitude as L-glutamate, whereas D-glutamate failed to induce an inward current. Glutamate receptor agonists, such as kainate, quisqualate, alpha-amino-3-hydroxy-5-methyl isoxazole-4-propionic acid, and N-methyl-D-aspartate, were ineffective in eliciting inward currents. The glutamate-induced inward current did not reverse even when the holding potential was set to +40 mV. The replacement of extracellular Na+ with choline+ eliminated the inward current. These results strongly suggest that the current induced by glutamate is mediated by a glutamate transporter rather than a glutamate receptor. We further examined the effects of 12 amino acid analogs which are known to be selective inhibitors of the mammalian excitatory amino acid transporters (EAATs) on the corpora allata transporter. From the effects of these inhibitors, we conclude that the glutamate transporter expressed in corpora allata cells of the cricket is similar to the high affinity glutamate transporters cloned from human brain, especially EAAT1 and EAAT3. Unlike mammalian transporters, however, serine-O-sulfate has the most potent action, suggesting the unique feature of the glutamate transporter expressed in the corpora allata.

[Surgical results of emergency coronary artery bypass grafting: effect of left main trunk lesion].

Emergency coronary artery bypass grafting has higher risk than elective surgery. Furthermore, if a lesion is located at left main coronary artery, the frequency of cardiogenic shock or high risk patient would be greater, and operative results would be worse. Between January, 1989 and December, 1995, 45 patients who underwent emergency CABG were included. Age ranged 44 to 80 years (mean 67 +/- 7.6 years; 31 men, 14 women). Of 45 cases, 12 cases were patients with LMT lesion. Results were analysed by univariate analysis and multivariate logistic analysis. Of 45 emergency cases, 5 were operative death and 3 were hospital death. Mortality rate was 17.8%, which was significantly higher than the mortality of elective CABG (2.8%) during the same period (p < 0.001). A factor that influenced the mortality was acute myocardial infarction (AMI), which was confirmed by both univariate and multivariate logistic analyses. Odds ratio (relative risk) was 12.4 for AMI. Only one patient died in 12 cases with LMT (8.3%). This case was due to complication after catheter intervention possibly caused by MOF. Thus, the relative risk of LMT was not so high (p = n.s). Other factors showed no significant correlation. Although the emergency case of LMT lesion was generally severe, we could have the same result as other emergency surgeries when patients were revascularized as soon as possible.

Therapeutic effect of egualen sodium (KT1-32), a new antiulcer agent, on chronic gastritis induced by sodium taurocholate in rats.

We investigated the therapeutic effects of egualen sodium (KT1-32), a new antiulcer agent, on chronic erosive and atrophic gastritis induced by 5 months' administration of sodium taurocholate (TCA; 5 mM) in rats. The chronic gastritis was manifested by mucosal surface injuries (erosions), reduced mucosal thickness, reduction of the number of parietal cells, infiltration of inflammatory cells, and proliferation of collagenous fiber. Egualen sodium, (10-100 mg/kg, t.i.d.) administered orally to the rats for 2 weeks after the withdrawal of TCA, dose-dependently and significantly decreased the total length of erosions. The indicators of atrophic gastritis, i.e., reduced mucosal thickness and reduction in the number of parietal cells, were improved dose-dependently by the administration of this agent. Egualen sodium also reduced the inflammatory cell infiltration and the proliferation of collagenous fiber in the gastric mucosa in a dose-dependent manner. The reduced staining of neutral gastric mucus was improved by a high dose (100 mg/kg) of egualen sodium. The therapeutic effects of egualen sodium on experimental gastritis were superior to those of sofalcone and sodium guaiazulene 3-sulfonate. These results suggest that egualen sodium may be a promising agent for the treatment of erosive and atrophic gastritis.

Azulene derivatives as TXA2/PGH2 receptor antagonists--II. Synthesis and biological activity of 6-mono- and 6-dihydroxylated-isopropylazulenes.

In order to examine the correlation between activity and hydrophilicity of the side chain of sodium 3-[4-(4-chlorobenzenesulfonylamino)butyl]-6-isopropylazulene -1-sulfonate (KT2-962), a non-prostanoid TXA2/PGH2 receptor antagonist, one or two hydroxyl groups were introduced into the isopropyl moiety. A series of 6-hydroxylated-isopropylazulenes were synthesized by regioselective oxidation of 6-isopropylazulenes and their in vitro and in vivo antagonistic activities were studied. Both the primary and tertiary alcohols, monohydroxylated derivatives, exhibited potent biological activities comparable to unmodified 6-isopropylazulenes both in vitro and in vivo. In contrast, the activities of 1,2- and 1,3-diols of 6-substituted derivatives, markedly decreased, but recovered by O-isopropylidenation of the dihydroxyl moiety. These findings indicate that the moderate hydrophobicity of substituent at the 6-position of the azulene ring might be required for the activity and the size of the substituent at this position, not so rigid for keeping potent biological activity.

Pharmacological properties of KT3-671, a novel nonpeptide angiotensin II receptor antagonist.

We examined pharmacological profiles of KT3-671, 2-propyl-8-oxo-1-[(2'-(1H-tetrazole-5-yl) biphenyl-4-yl)methyl]- 4,5,6,7-tetrahydro-cycloheptimidazole, a newly synthesized nonpeptide angiotensin II (AII) receptor antagonist in various in vitro and in vivo studies. KT3-671 displaced specific binding of [125I]Sar1 Ile8-AII to AT1 receptor with a Ki value of 0.71 +/- 0.14 x 10(-9) M in rat liver membranes, but had no affinity for AT2 receptor in bovine cerebellar membranes (Ki > 10(-5) M). In isolated rabbit aorta, KT3-671 produced a parallel rightward shift in the concentration-response curve for AII with a pA2 value of 10.04 +/- 0.12, but had no effect on KCl-, norepinephrine (NE)-, and serotonin (5-HT)-induced contractions. In conscious normotensive rats, KT3-671 (0.3-10 mg/kg, p.o.) inhibited the AII-induced pressor response dose dependently. In renal artery-ligated hypertensive rats, KT3-671 (0.1-3 mg/kg, p.o.) decreased arterial blood pressure (BP) dose dependently. The hypotensive action of 3 mg/kg KT3-671 was maintained for at least 24 h. These results suggest that KT3-671 is a potent AT1 subtype-selective and competitive nonpeptide AII receptor antagonist and has an orally active antihypertensive effect without agonistic activity.

Nonpeptide angiotensin II receptor antagonists. I. Synthesis and biological activity of pyridine derivatives.

Substituted pyridines were synthesized as potential angiotensin II (AII) receptor antagonists. Substitution at the position 2 in the pyridine resulted in potent activity, and the optimal alkyl length was four carbons. The potency further increased with the introduction of a hydroxymethyl group at the position 4. One of the compounds, 2-butyl-6-chloro-4-hydroxymethyl-5-methyl-3-[[2'-(1H-tetrazol-5-yl )biphenyl-4-yl]methyl]pyridine 9 h (KT3-579) is a competitive AII antagonist with a pA2 value of 9.31, and is about 10 times more potent than Du Pont 753. It was found to be an AT1 specific antagonist with an IC50 of 3.09 nM.

[The effects of prostaglandin E1 infusion during cardiopulmonary bypass on cardiac and renal function in the early postoperative period].

Postoperative complications after cardiac surgery are mainly caused by the detrimental effects of cardiopulmonary bypass (CPB) on hemodynamics and humoral factors. We studied the effects of prostaglandin E1 (PGE1) infusion on CPB induced organ failure. Twenty-four patients who underwent coronary bypass surgery with normal preoperative cardiac and renal function were divided into two groups. PG group (n = 11) was given PGE1 (0.03-0.05 microgram/kg/min) during CPB while the control group (n = 13) was not. Hemodynamic studies with the Swan-Ganz catheter and renal function tests including serum creatinine (Cr), serum beta 2-microglobulin (s-BMG) and urine beta 2-microglobulin (u-BMG) measurements were performed 24 hours after surgery. After CPB, alpha-adrenergic agent requirement was higher in the PG group than in the control group. In the PG group, the mean cardiac index was slightly higher and the mean left ventricular stroke work index was lower than in the control group. Although the mean systemic vascular resistance index was lower in the PG group, the mean pulmonary vascular resistance index was higher because of the vasoconstrictive action of alpha-adrenergic agents. These differences were seen only immediately after surgery; hemodynamic parameters in both groups were at the same level 3 hour postoperatively. Immediately after surgery, creatinine in the PG group was significantly lower than in the control group. While there was no significant differences in s-BMG between the two groups, u-BMG at 6 and 12 hours postoperatively in the PG group were significantly lower than in the control group.(ABSTRACT TRUNCATED AT 250 WORDS)

Vasoinhibitory action of KT2-734, an antihypertensive agent, in isolated rat aorta.

In rat aorta, KT2-734 inhibited contractile responses to 5-hydroxytryptamine (5-HT) and KCl. KT2-734 inhibited the relaxing effect of verapamil, but not nifedipine. Similarly, verapamil, but not nifedipine, inhibited the vasorelaxing effect of KT2-734. KT2-734 relaxation was inhibited by endothelium removal but not by atropine and propranolol. Methylene blue, a guanylyl cyclase inhibitor, and NG-monomethyl arginine also inhibited the relaxation both in the presence and absence of endothelium. In the absence of endothelium, KT2-734 potentiated the relaxation induced by L-arginine, nitroglycerin and isoproterenol. In addition, M & B 22,948, a cGMP phosphodiesterase inhibitor, and theophylline inhibited and potentiated, respectively, KT2-734-induced relaxation. However, methylene blue inhibited the potentiation of isoproterenol relaxation by KT2-734 and that of KT2-734-relaxation by theophylline. KT2-734 caused increases in the level of cGMP without significantly affecting the cAMP level. These results suggest that KT2-734 may cause endothelium-independent relaxation mainly due to inhibition of cGMP-phosphodiesterase.

Design, synthesis, and pharmacology of 3-substituted sodium azulene-1-sulfonates and related compounds: non-prostanoid thromboxane A2 receptor antagonists.

A series of novel azulene-1 carboxylic acid derivatives 28-30, azulene-1 sulfonic acid sodium salts 41a-c, and related compounds were synthesized. These compounds were tested for TXA2 receptor antagonistic activity. The inhibitory concentrations (IC50) of these compounds for vascular contraction (TXA2 tau receptor) and platelet aggregation (TXA2 alpha receptor) induced by (15S)-15-hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5(Z),13(E)- dienoic acid (U-46619) were obtained. Azulene-1-sulfonic acid sodium salts 41a-c were over 3 times more potent than azulene-1-carboxylic acids 28-30. The most potent compound, 41b was 4 orders of magnitude more potent than a TXA2 antagonist, BM13,177, in inhibiting vascular contraction (tau receptor) and had an IC50 of 9.0 x 10(-10) M. Compound 41b was also found to be a tau receptor selective antagonist (IC50 of contraction/IC50 of aggregation = 378) and to have no TXA2 synthetase inhibitory activity at concentrations up to 10(-4) M and no partial agonistic activity at concentrations up to 10(-5) M in rabbit aorta (tau receptor) and up to 10(-4) M in rabbit platelet-rich plasma (alpha receptor). In a radioligand binding assay using rabbit gel-filtered platelets, compound 41b had a high-affinity binding for the TXA2 receptor. In an in vivo study, compound 41b inhibited U-46619-induced sudden death in mice at a dose of 0.3 mg/kg and its duration of action was over 8 h when administered orally at 3 mg/kg.

Pharmacologic properties of KT2-962 (6-isopropyl-3-[4-(p- chlorobenzenesulfonylamino)-butyl]-azulene-1-sulfonic acid sodium salt); a new TXA2/prostaglandin endoperoxide receptor antagonist.

Pharmacologic properties of KT2-962 (6-isopropyl-3-[4-(p-chlorobenzenesulfonylamino)butyl]-azulene+ ++-1-sulfonic acid sodium salt, KT) were studied in isolated rat aorta, rat tail artery, rabbit aorta, rabbit renal artery, and pig coronary artery. KT competitively inhibited the contractions induced by thromboxane A2 (TXA2) mimetic, U46619 (pA2 values 9.95, 8.85, 7.87, 8.49, and 9.12, respectively). KT also inhibited the contraction of rabbit aorta induced by prostaglandin2 alpha (PGF2 alpha, pA2 value 7.85) and the contraction of guinea pig ileum induced by LTD4 (pA2 value 5.48) but did not alter the contractions induced by norepinephrine (NE), Ca2+, serotonin, and histamine. KT did not alter the contractions of guinea pig ileum, which did not contract with U46619, induced by PGE2 and PGF2 alpha. KT inhibited the aggregations of rabbit platelets induced by U46619, arachidonic acid, and collagen (IC50 values 7.9, 140, and 16 microM, respectively) but not those induced by ADP. It also inhibited the specific binding of TXA2/PGH2 receptor antagonist, [3H]SQ29,548, to rabbit gel-filtered platelets with an IC50 value of 1.5 x 10(-8) M. In in vivo experiments with mice, oral administration of KT protected the U46619-induced sudden death with the minimum effective dose of 0.3 mg/kg and provided such protection for > 8 h at 1.0 mg/kg. These results indicate that KT is a new nonprostanoid type TXA2/PGH2 receptor antagonist that is orally effective and long acting.

Synthesis and anti-peptic activity of compounds related to the metabolites of sodium 3-ethyl-7-isopropyl-1-azulenesulfonate (KT1-32).

The metabolites of sodium 3-ethyl-7-isopropyl-1-azulenesulfonate (KT1-32, 1), a candidate as an anti-ulcer drug, and related compounds were synthesized. The effects of the compounds on anti-peptic activity were determined as compared to that of 1.

[Healing promoting effect of azuletil sodium (KT1-32) on experimental chronic gastric ulcers and acceleration of healing in SPF environment].

We examined the healing promoting effects of azuletil sodium on acetic acid and clamping cortisone-induced gastric ulcer in rats. For the experiments on clamping-cortisone gastric ulcer, we used not only conventional rats in conventional conditions but also specific pathogen free (SPF) rats on SPF environment in order to prevent infection. The following results were obtained. 1) In acetic acid ulcer, azuletil sodium (AZE) (greater than or equal to 90 mg/kg/day, p.o.) significantly decreased ulcer index. As estimated on the basis of stage analysis (Ulcer, Healing, Scar), AZE (greater than or equal to 30 mg/kg/day, p.o.) significantly promoted the healing of ulcers. 2) In clamping cortisone ulcer (conventional), AZE (100 mg/kg/day, p.o.) significantly promoted the regeneration of blood vessels. 3) In clamping cortisone ulcer (SPF), AZE at greater than or equal to 30 mg/kg/day and 100 mg/kg/day significantly increased the healing index and mucosal regeneration index, respectively. 4) In clamping cortisone ulcer (SPF), the infection that was observed in the conventional test was not seen at all and the acceleration of healing was observed. Furthermore, the extent of adhesion was also reduced, and the standard errors of various healing indices were smaller. From these results, it is concluded that AZE accelerated the healing of experimentally-induced gastric ulcers in rats.

Characterization of the vasoinhibitory actions of KT2-230, a new benzothiazepine vasodilator derivative on isolated rabbit vascular smooth muscles: an alpha-adrenergic- and serotonergic-receptor antagonist.

1. Pharmacological properties of KT2-230 (benzothiazepine derivative), a newly synthesized vasorelaxing agent, were studied. 2. In the anesthetized dogs, KT2-230 increased the femoral and vertebral blood flow without effect on systemic blood pressure. 3. In rabbit aorta, KT2-230, methysergide and phentolamine inhibited contractile responses to 5-hydroxytryptamine. 4. The response to norepinephrine was also inhibited by KT2-230 and phentolamine. Responses to histamine were not affected by KT2-230. 5. Responses to KCl and Ca2+ in K+ depolarized aorta in Ca2(+)-free medium were inhibited by a high concentration of KT2-230. 6. In rabbit iliac artery, KT2-230 inhibited the response to caffeine in Ca2(+)-free medium. 7. KT2-230 decreased total La3(+)-resistant Ca2(+)-binding at high affinity sites. 8. These results indicate that KT2-230 inhibits alpha 1-adrenoceptors and 5-HT-receptors and at high concentrations it inhibits slow Ca2(+)-channels. KT2-230 may inhibit the Ca2(+) release from caffeine- and agonist-sensitive Ca2+ stores.

Studies on anti-ulcer agents. II. Synthesis and anti-ulcer activities of 6-isopropylazulene-1-sodium sulfonate derivatives.

A series of alkylazulene-1-sodium sulfonate derivatives which has an isopropyl group at 6-position were synthesized, and their anti-ulcer activities were examined in Shay pylorus-ligated rats. The values of lipophilicity (log P) as a parameter of these new azulene derivatives were also examined in reference to the structure-activity relationship. The optimum value of log P, which showed maximal anti-ulcer activity, was about -0.46. Among the derivatives of azulene examined, 3-ethyl-6-isopropylazulene-1-sodium sulfonate (compound IXb:XT1-785) exhibited the most potent inhibitory action against Shay ulcer, and its anti-peptic activity was similar to that of 3-ethyl-7-isopropylazulene-1-sodium sulfonate (KT1-32). It also had more activity than guaiazulene sodium sulfonate (GAS). Furthermore, KT1-785 was extremely stable under heating as compared to GAS.

[Analysis of scintigrams using fuzzy theory].

Textural features for phantom images were extracted. Texture parameters which represent RI distribution--skew, energy, entropy and angular second moment were used. But, it was difficult to analyse the images using discriminant analysis for textural features, because textural features had statistical noise. Therefore fuzzy reasoning was adapted to analyse the images. Textural features for six kinds of images were showed using membership function. The possibility to the image was evaluated using the value of membership function on each images. Fuzzy reasoning could be done easily using max-min composition formula. The reasoning was found more suitable to analyse the images than discriminant analysis and will be considered useful for analysis of clinical scintigrams.

[General pharmacological properties of an anti-ulcer drug, azuletil sodium (KT1-32)].

Azuletil sodium (AZE, 100 mg/kg, p.o.) did not affect the general behaviors, spontaneous motor activity, pentobarbital-induced hypnosis and body temperature. Furthermore, it did not elicit anticonvulsant and muscle relaxant actions. However, AZE (300 mg/kg, p.o.) elicited a stiff gate and slightly inhibited the spontaneous motor activity and electroshock-induced convulsions. It had no influence on spontaneous EEG activities, even at 30 mg/kg, i.v. AZE inhibited acetic acid-induced writhing moderately at doses above 100 mg/kg. AZE at concentrations up to 10(-5) g/ml did not affect agonist-induced contractions of the isolated ileum, trachea, vas deference and uterus, but inhibited serotonin and oxytocin-induced contraction at concentrations above 3 x 10(-4) and 10(-5) g/ml, respectively; and it also depressed spontaneous movements of the ileum and uterus at concentrations above 3 x 10(-4) g/ml. AZE caused no changes in blood pressure (BP), heart rate (HR), left ventricular pressure, ECG, tracheal pressure (TP), femoral blood flow (FBF) and coronary blood flow (CBF) at doses up to 10 mg/kg, i.v. in anesthetized dogs, but it caused an increase or a decrease in BP, an increase in TP and an increase in CBF at 30 mg/kg, i.v. However, even at 300 mg/kg, p.o., it caused no changes in BP and HR in conscious rats. AZE moderately promoted the charcoal transport. AZE at doses up to 300 mg/kg, p.o. did not affect urine volume, urinary electrolyte excretion, blood glucose and prothrombin time. These results suggest that AZE at anti-ulcer doses of 10-100 mg/kg, p.o. does not have noticeable effects on general pharmacological properties, and there is no marked differences as compared with those of GAS.

Intracellular accumulation of isoproterenol enhances the lactate production in the perfused rat heart.

Effects of intracellular accumulation of isoproterenol (ISO) on lactate production were examined in perfused rat heart. The lactate production during ISO perfusion in rat heart was increased and subsequent addition of an inhibitor of catechol-O-methyl transferase (COMT) further enhanced the production, and the enhanced production was significantly reduced by uptake2 inhibitor. The perfusion with ISO free-medium in the heart with high intracellular accumulation of ISO produced lactate more than that in the low intracellular accumulation. The present experiments demonstrated that the enhanced lactate production is accompanied by intracellular accumulation of ISO in the perfused rat heart, and suggested that the accumulated ISO may activate intracellular beta-adrenoceptors in the rat heart.