In this communication, I have summarized our studies on the possible roles of cysteine sulfinic acid (CSA) in the central nervous system (CNS), from these observations, CSA was suggested to be a neurotransmitter. We reported the presence of CSA in the CNS and subsequently characterized Na⁺-dependent high affinity uptake and depolarization-induced release of CSA. Depolarizationinduced release of [¹⁴C] CSA from the preloaded hippocampal slices was specifically attenuated by benzodiazepines and GABA agonists. Synaptic membranes have a Na⁺-independent specific binding site for cysteic acid, an analogue of CSA, which may be a possible binding site for CSA. This binding site seemed to be distinct from that for glutamate. To assess CSA as a physiologically active candidate which is distinct from glutamate, two neurochemical experiments were performed: one experiment determined the enhancement by excitatory amino acids of depolarization-induced release of [³H]GABA from the preloaded slices, and the other one monitored the cyclic AMP formation by excitatory amino acids in hippocampal slices. In both studies, differences in the responses to the various antagonists indicate that CSA receptors are distinct from glutamate receptors. Furthermore, we proposed that excitatory amino acid receptors which are subsequently linked to adenylate cyclase are functionally related to the Cl^- channel.

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Latamoxef (daily 100 and 300 mg/kg, i.v.) was injected once a day for 8 days to 75% and 90% nephrectomized rats kept on a vitamin K-sufficient diet (500-600 ng/g) or a vitamin K-deficient diet (30-50 ng/g), and changes in ADPinduced platelet aggregation and prothrombin time were examined. The half-life of latamoxef was markedly prolonged and plasma latamoxef and N-methyl-tetrazolethiol (NMTT) concentrations increased, resulting in a delay of the total body clearance of the compounds. The ADP-induced platelet aggregation increased after nephrectomy, and latamoxef slightly but inconsistently decreased the aggregation. Prothrombin time did not change even in the 90% nephrectomized rats kept on an ordinary diet, but increased dose-dependently in the vitamin K-deficient nephrectomized rats, with the 90% nephrectomized animals showing larger increases of prothrombin time. These data suggest that NMTT or NMTT-containing antibiotics cause no hypoprothrombinemia even in partially nephrectomized rats when they are fed an ordinary diet containing vitamin K, but these compounds enhance the manifestation of hypoprothrombinemia in vitamin K deficiency. Further renal failure promotes the manifestation by increasing drug concentration in the blood. However, platelet aggregation in these animals is not significantly affected at the doses examined.

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Male neonates of Wistar strain rats were given s.c. 1-4 mg/kg/day of methamphetamine (MAP) for 7 successive days from days 6 to 12 after birth. The acquisition processes of the discriminative lever-press avoidance response were investigated from day 60 after birth. Effects of the postnatal pretreatments with MAP on saturation constants for specific bindings of ³H-spiperone (SPP) and ³H-WB4101, respectively, in 8 brain regions were also investigated at 100-120 days after birth. In addition, dopamine, noradrenaline and the levels of their metabolites were measured in the brain. No significant difference was detected in body weight, gross behaviors and avoidance learning between saline and MAP-pretreated groups. However, effects of MAP and apomorphine on the locomotor activity significantly increased in the MAP-pretreated group. Significant decreases in B_max and K_d values of ³H-SPP binding sites in the striatum were detected in the MAP-pretreated group, while significant decreases in Bmax values of ³H-WB4101 binding sites in the cortex and hippocampus as well as those in K_d values in the hippocampus were found in the treated group. Dopamine and noradrenaline levels significantly decreased in the MAP-pretreated group, but on the contrary, their metabolites levels significantly increased. These results suggest that postnatal pretreatments with MAP produce long-lasting behavioral changes associated with an impaired development of catecholaminergic neurons in the rat brain after maturity.

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The effects of tiapride on the convulsive seizures induced by pentylenetetrazole, strychnine, picrotoxin and bemegride, and on electric seizure are reported and compared with those of sulpiride, chlorpromazine, haloperidol and reserpine. The number of deaths and intensity of convulsion increased dosedependently and also with the increase in amplitude of electric shock. Tiapride and a similar compound, sulpiride, did not affect these seizures, whereas chlorpromazine potentiated strychnine-induced and electric seizure. Haloperidol and reserpine potentiated electric seizure, and chlorpromazine and reserpine tended to potentiate bemegride-induced seizure. Reserpine also tended to potentiate pentylenetetrazole-induced seizure. These results suggest that tiapride would be clinically safer than other drugs with anti-dopamine activity, except for sulpiride.

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Brain harmine was simply and sensitively determined by high performance liquid chromatography (HPLC), and the relationship between brain harmine concentrations and the jumping behavior induced by harmine and apomorphine was investigated. The concentration of harmine and the peak height in HPLC showed a good correlation, and the detection limit was 0.05 pmole harmine. In rats treated with 2, 5 and 10 mg/kg of harmine, the concentrations of harmine in the brain cortex were 9.0, 21.3 and 43.1 nmole/g tissue (wet wt.), respectively. The regional brain harmine concentrations and the jumping activity measuring by a scoring system increased with increasing doses of harmine administered, and there was a direct relation between the harmine concentrations and the jumping activity. This relation was observed in washed membranes in which about 40% of the harmine in whole homogenate was present. Tissue subfractionation showed that 30% of the harmine in whole brain homogenates was in the P₂ fraction, and of this, 70% was located in the synaptosomes. The brain homogenate did not metabolize harmine and no metabolite was detected in the brain 10 min after treatment with harmine and apomorphine. These results suggest that harmine itself, but not its metabolites, may be responsible for inducing the jumping behavior and that the jumping activity is correlated with brain harmine concentrations.

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We prepared an experimental system to study the effects of drugs on urinary bladder contraction and micturition simultaneously in rats anesthetized with urethane (1 g/kg, s.c.) and a-chloralose (25 mg/kg, s.c.). When Tyrode's solution was infused at a constant rate (0.8-1 ml/10 min) through a needle inserted into the bladder from the left ureter, the bladder pressure gradually and then steeply rose, and micturition took place. These changes in bladder pressure and micturition were constantly repeated. In this model, drugs which partially inhibited the bladder contractile force, e.g., atropine (0.01-1 mg/kg, i.v.) and hexamethonium (C6, 5 mg/kg, i.v.) increased the frequency of bladder contraction instead of decreasing the amount of solution excreted from the penis by bladder contraction. The rate of afferent discharges during bladder filling was increased after injection of atropine or C6, and this increase was considered to be responsible for the induction of the increase in the frequency of bladder contraction. Drugs which inhibited the bladder contraction and interrupted micturition, e.g., C6 (20 mg/kg, i.v.) raised the bladder pressure until the solution leaked from the penis. As phentolamine (5 mg/kg, i.v.) or propranolol (1 mg/kg, i.v.) did not facilitate bladder motility but rather inhibited it, the inhibitory action of sympathetic nerves on bladder motility was considered to be weak in rats. This model was useful for studying the effect of drugs on bladder motility and micturition reflex.

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Antiallergic asthma effect of trans-4-guanidinomethylcyclohexane-carboxylic acid p-tert-butylphenyl ester hydrochloride (NCO-650), a new antiallergic drug, was investigated in comparison with those of tranilast and disodium cromoglycate (DSCG) in anesthetized dogs. The asthmatic bronchoconstriction was induced by inhalation of Ascaris suum antigen (Asc-Ag) to naturally Ascarissensitive dogs. The airway resistance was determined using the modified KonzettRössler method. Both intravenous (1 and 5 mg/kg) and intraduodenal (10, 30 and 100 mg/kg) administrations of NCO-650 prior to the antigen challenge markedly inhibited the asthmatic bronchoconstriction induced by Asc-Ag inhalation. The antiasthmatic effect of NCO-650 was much stronger than that of DSCG (10 mg/kg, i.v.) and was about three-fold stronger than that of tranilast. On the other hand, when NCO-650 was administered after the antigen challenge, the agent had no inhibitory effect on the Asc-Ag induced bronchoconstriction. As for the effects on increased airway secretion at the time of asthmatic attack, NCO-650 inhibited the excessive secretions without any remarkable change in the viscosity of the secretions. NCO-650 had no effect on the bronchoconstriction induced by inhalation of acetylcholine, suggesting that NCO-650 appears to have no anticholinergic effect and thus no effect on the vagal reflex that occurred during the asthmatic responses. The above findings show that NCO-650 may be useful for the treatment of bronchial asthma as an orally active drug.

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Carbachol (CCh) at a concentration of 10^-7 M completely inhibited the twitch contraction of frog atrial muscle. However, when the preparation was treated with goniopora toxin (GPT), a selective inhibitor of Na channel inactivation, and the twitch contraction was augmented through the activation of Na-Ca exchange by this toxin, the reduction in contractile amplitude by CCh was only about 30% even at higher concentrations. The maximum rate of rise of the twitch contraction was not affected by CCh in GPT-treated preparations. The time course of configuration change of the twitch contraction after application of CCh in GPT-treated muscle indicated that the attenuation of amplitude preceded the shortening of duration. The residual contraction under the combined treatments with GPT and CCh was abolished by removal of external Ca⁺⁺ or addition of TTX. CCh only moderately shortened the action potential which was prolonged by GPT, and further addition of TTX abolished the action potential. From these results, we suggest that CCh does not influence the twitch contraction which is augmented via the activation of the Na-Ca exchange mechanism in frog atrial muscle.

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In mice, intraperitoneally injected chlord 1azepoxide and proglumide, both of which are regarded as cholecystokinin (CCK) receptor antagonists in the peripheral tissues, dose-dependently inhibited the satiety induced by 200 ng of intracisternally administered CCK octapeptide (CCK8). Intraperitoneally administered diazepam (1 mg/kg) and/or Ro 15-1788 (5 mg/kg), a benzodiazepine antagonist, both prevented the elevation in the pain threshold induced by 1 μg of CCK8. However, Ro 15-1788 did not antagonize the effect of diazepam that reversed the CCK-induced antinociception. Ro 15-1788 also inhibited the satiety induced by CCK8. From these results, it was considered that the antagonism, which was observed in the present work, of benzodiazepines and proglumide to CCK8 seemed to occur at the CCK receptor and not at the benzodiazepine receptor in the brain.

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Effects of tobramycin (TOB) alone and in combination with latamoxef (LMOX) on the stability of rat kidney lysosomal membranes were investigated. Rats were injected with doses of TOB (90 mg/kg/day, s.c.) alone, LMOX (2, 000 mg/kg/day, s.c.) alone or TOB (90 mg/kg/day, s.c.) and LMOX (2, 000 mg/kg/day, s.c.) for 5 consecutive days. The rat kidney lysosomes were isolated on the 1 st, 3rd and 5th days and incubated in a 0.25 M sucrose solution containing 1 mM EDTA (pH 7.0) at 37°C for 20 min. After incubation, the activity of N-acetyl-β-D-glucosaminidase (NAG) released from lysosomes was measured, and the percent NAG release was calculated as an index of the stability of lysosomal membranes. The percent releases of NAG from lysosomes of TOB alone-treated rats were 40 and 50% greater than those of normal rats on the 1st and 3rd days, respectively. On the other hand, treatment with TOB and LMOX suppressed the NAG release from lysosomes with TOB alone by about 80 to 100%. There were insignificant slight increases in the percent NAG release in LMOX alone-treated rats on the 3rd and 5th days. In addition, the in vitro study indicated that incubation of the lysosomal fraction from kidneys of normal rats with TOB (30 μg/ml) significantly increased the NAG release, compared with that of the non-treated lysosomal fraction. However, the preincubated mixture of TOB (30 μg/ml) and LMOX (50 μg/ml) in vitro significantly suppressed the release of NAG from Iysosomes by 85%. These results suggest that the suppression of the releases of NAG from lysosomes by the combination of TOB with LMOX may contribute to the protective effect of LMOX against TOB nephrotoxicity.

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TR K 100, a stable PGI₂ analogue structurally different from carbacyclines, was compared with other antiplatelet drugs for its effect on platelet functions using animal models. TRK-100 (10-300 nM) inhibited rat platelet aggregation induced by ADP (3 μM), collagen (12.5 μg/ml) and A23187 (10 μM), and its potency was about 1 /3-1 /7 that of PGI₂. TRK-100 (0.3-3 mg/kg, p.o.) dose-dependently inhibited rabbit platelet adhesion (ED50: 2.2 mg/kg), and its effect lasted over at least 5 hr. In contrast, aspirin and ticlopidine (both at 300 mg/kg, p.o.) showed only a slight inhibition (4-7%). In the thrombocytopenia induced by collagen injection in rats, TRK-100 (3-300 μg/kg, i.v.; 0.1-3 mg/kg, p.o.) dose-dependently inhibited a decrease in platelet number, and its ED50 was 0.48-0.62 mg/kg orally and 13.7-16.4 μg/kg intravenously, while the inhibition by aspirin and ticlopidine (both at 1000 mg/kg, p.o.) was 40 and 37%, respectively. In the experimental thread thrombosis in rats, TRK-100 (0.03-3 mg/kg, p.o.) dose-dependently inhibited thrombus formation, and its ED50 was 0.46 mg/kg, being 21 and 87 times as potent as aspirin and ticlopidine, respectively. These results reveal that TRK-100 has a potent antiplatelet activity and is orally and intravenously effective for a variety of thrombosis models, suggesting that it may have a therapeutic value as an antithrombotic drug.

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L-threo-3, 4-Dihydroxyphenylserine (L-DOPS), a norepinephrine (NE) precursor, 3 mg/kg, i.v., increased the cerebral blood flow (CBF) in both the striatum and hippocampus as well as the mean arterial blood pressure (MABP) in urethane-anesthetized rats, as NE infusion did. The L-DOPS induced increase in CBF was inhibited by benserazide (3 mg/kg/hour), a peripheral aromatic L-amino acid decarboxylase inhibitor, and propranolol (3 mg/kg, i.p.), a β-adrenoceptor blocker as well. These results suggest that the effects of L-DOPS may be attributed to the action of NE formed from L-DOPS, and the action may be mediated by stimulation of β-adrenoceptor.

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In the vascularly isolated and self -hemoperfused dog pancreas, secretin (0.025 clinical units) and adenosine (1.0 mg) administered intra-arterially (i.a.) increased tissue cyclic AMP level by about 46% and 37%, respectively. Although the increases in the nucleotide were not statistically different, only secretin stimulated exocrine secretion. The increase in cyclic AMP induced by adenosine was significantly reversed by pretreatment with theophylline (0.3 mg, i.a.). These results suggest that the effect of adenosine on cyclic AMP formation occurs mainly in the non-exocrine system of the dog pancreas through A₂/Ra-receptors.

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Following L-DOPA administration, DA metabolites were measured by both in vivo voltammetry and HPLC-ECD. In vivo, the amplitudes of the catechol oxidation currents in both the striatum and frontal cortex increased and reached a plateau from 60 min to 240 min after L-DOPA treatment. L-DOPA, DA, DOPAC and HVA levels measured by HPLC-ECD after L-DOPA treatment were increased in both regions, and the occurrence of the peak of each compound was dependent on its order in the metabolic pathway, i.e., 15 min for DA, 45 min for DOPAC and 60-180 min for HVA. However, NE remained unchanged. No indication of a rapid DA increase was apparent in the catechol oxidation current at 15 min; and thus, DA is unlikely to be a main contributor to the catechol oxidation current in the brain of rats treated with L-DOPA.

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The drug interaction between methamphetamine and quinidine/ cimetidine was investigated in terms of distribution of methamphetamine to the brain. The concentrations of methamphetamine and amphetamine in the brain after an s.c. Injection of methamphetamine were determined in rats pretreated with a single oral dose of quinidine or cimetidine. Both drugs markedly enhanced the increase in the concentrations of methamphetamine and its metabolite, amphetamine. These results suggest that the recent finding of the enhancements of the behavioral effect of methamphetamine by quinidine and cimetidine is due to the increase in levels of methamphetamine and amphetamine in the brain by these pretreatment drugs.

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Trypsin treatment of N-[p-(2-benzimidazolyl)phenyl]maleimide modified enzyme caused a marked reduction in Na⁺, K⁺-ATPase activity and in the amount of the α-chain, which contains the phosphorylation and ouabain binding sites. However, these preparations retained nearly 90% of the ouabain binding capacity and showed ouabain sensitive dynamic fluorescence changes accompanying the hydrolysis of ATP. The data showed that the three dimensional structure of Na⁺, K⁺-ATPase, which is important in the dynamic fluorescence change, is little affected in spite of extensive covalent bond splitting in the a-chain of Na⁺, K⁺ATPase.

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