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Intravenously injected lipopolysaccharides (LPS) rapidly induce pulmonary platelet accumulation (PPA) and anaphylaxis-like shock (ALS) in mice. Macrophages reportedly release catecholamines rapidly upon stimulation with LPS. Here, we examined the involvement of macrophage-derived catecholamines in LPS-induced PPA and ALS. A catecholamine or Klebsiella O3 (KO3) LPS was intravenously injected into mice, with 5-hydroxytryptamine in the lung being measured as a platelet marker. The tested catecholamines induced PPA, leading to shock. Their minimum shock-inducing doses were at the nmol/kg level. The effects of epinephrine and norepinephrine were inhibited by prazosin (α1 antagonist) and by yohimbine (α2 antagonist), while dopamine's were inhibited only by prazosin. Use of synthetic adrenergic α1- and/or α2-agonists, platelet- or macrophage-depleted mice, a complement C5 inhibitor and C5-deficient mice revealed that (a) α2-receptor-mediated PPA and shock depend on both macrophages and complements, while α1-receptor-mediated PPA and shock depend on neither macrophages nor complements, (b) the PPA and ALS induced by KO3-LPS depend on α1- and α2-receptors, macrophages, and complements, and (c) KO3-LPS-induced PPA is preceded by catecholamines decreasing in serum. Together, these results suggest the following. (i) Catecholamines may stimulate macrophages and release complement C5 via α2-receptors. (ii) Macrophage-derived catecholamines may mediate LPS-induced PPA and ALS. (iii) Moderate PPA may serve as a defense mechanism to remove excess catecholamines from the circulation by promoting their rapid uptake, thus preventing excessive systemic effects. (iv) The present findings might provide an insight into possible future pharmacological strategies against such diseases as shock and acute respiratory distress syndrome.

The purpose o f this report is to show the importance of CCS as a climate change mitigation technology by comparing it with other climate change mitigation technologies currently being developed. In order to objectively evaluate those technologies including CCS currently being researched and developed, the study conducted and compared these technologies in term of their potential in reducing GHG emissions at two set points in the future. The authors believe that the result of the study can be an effective way to promote public acceptance (PA) of CCS technology. The study was conducted with support from the ‘R&D project of CO2 Geological Storage Technology’ which is subsidized by Japan’s METI. In the result of evaluation, it shows that in order for the CCS technology to become practical, risks must be more clearly identified and the economic viability must be improved. Therefore, preparing frameworks and building social systems that support CCS technology would be inferred to become critical elements. These analysis results can be re-assessed when situations change for each subject technology whenever appropriate and will help make it possible to deal with changing reality in a flexible manner.

Carbon Dioxide Capture and Storage (CCS) is a technology for reducing the amount of CO 2 to be emitted into the air, by storing the CO 2 . However, a CCS project itself emits some CO 2 . Therefore, to estimate CCS’s net CO 2 reduction effect, it is effective to apply life cycle assessment (LCA) to CCS projects. In this paper, we assumed a number of CCS project models to be feasible in Japan, and estimated the amount of CO 2 emissions to store one ton of CO 2 by applying LCA to the CCS projects. As a result, depending on the model case, 50% or more of the CO 2 amount to be stored was emitted. The study also revealed that approximately 80% of the CO 2 emissions at CCS-applied facilities derived from the energy consumed to re-heat the liquid absorbent that absorbed CO 2 . The study also indicated that a substantial amount of CO 2 emissions can be reduced if heat integration is conducted between the production equipment and the CCS equipment for energy utilization, such as bleeding CO 2 from a power station and other production equipment.

Carbon dioxide Capture and Storage (CCS) technology, which is one of the technologies against global warming, applied to the separation and capture of CO2 gas and finally storage underground is attracting attention. There are several technologies being developed for CO2 separation and capture, namely the chemical absorption method, physical absorption method, adsorbing separation method and membrane separation method. These are being developed to apply to postcombustion capture, oxyfuel postcombustion capture, etc. With regard to the CO2 storage technologies, the method of injection into an aquifer is being developed as the major one, and studies on issues for actual technical application are being conducted in the form of engineering study on the assumed model area. In this report, our expectations toward actual application of the CCS technology, the trends of the CCS technology and other related matters are introduced. Copyright © 2007 Institute of Electrical Engineers of Japan. Published by John Wiley & Sons, Inc.

The present study clearly demonstrated the effects of complete cerebral ischemia and selective ischemia on brain choline metabolism in ddY mice The complete cerebral ischemia mouse was produced by cervical dislocation, and the selective ischemia mouse was produced by bilateral occlusion of the common carotid arteries. The following results were obtained 1 Complete cerebral ischemia increased brain choline contents rapidly and remarkably for 10 minutes After 10 minutes, the rate of increase of brain choline contents suddenly fell. 2 The increased level of choline contents by complete cerebral ischemia differed among the brain regions (cerebral cortex, hippocampus, medulla oblongata, cerebellum) tested 3 Selective ischemia increased choline contents in the cerebral cortex and hippocampus but not in the medulla oblongata or cerebella. 4. As with complete ischemia, hypoxia induced by N_2 gas inhalation increased choline contents in all brain regions. 5. Hypoglycemia induced by insulin administration did not increase brain choline contents However, the ischemia under a hypoglycemia state produced a greater choline increase than ischemia alone 6 Complete cerebral ischemia decreased the contents of both phosphatidylcholme (PtdCh) and glycerophosphocholine (GlyCh) in the mouse brain. The present results suggest that a main cause for the ischemic increase of brain choline contents is hypoxia by cessation of blood flow, and a low energy state caused by hypoglycemia partly contributes to this increase Furthermore, it is suggested that the ischemic increase of brain choline depends on the choline accumulation that results from decomposition of both PtdCh and GlyCh

Introduction: To measure levels of phosphatidylcholine (PtdCh) in various mouse tissues, we developed a rapid and precise method using high-performance liquid chromatography (HPLC) with electrochemical detection (ECD) and an immobilized enzyme column. To generate an example data set, the effect of methoxamine (an α 1 -adrenergic agonist) on the PtdCh levels was examined by this method in the artery and the submandibular gland of the mouse in vivo. Methods: Under our modifications of the method of Zapata et al. [J. Neurosci. 18 (1998) 3597], the mixture of lipophilic choline metabolites (PtdCh, lyso-PtdCh, and sphingomyelin) extracted by chloroform from the tissue homogenate was dried without prior separation and hydrolyzed with free choline by a 1-N perchloric acid solution containing ethylhomocholine (an internal standard for choline assay) at 90 °C for 1 h. Subsequently, the hydrolyzed mixture was injected directly into the HPLC system for PtdCh assay. Results: The present method permitted PtdCh assay within 5 min in one chromatographic run. Recovery of an authentic PtdCh sample was 99% ( n =10). The within-run coefficients of variation for choline derived from PtdCh in the same tissue samples were 0.6% ( n =10) and 1.3% ( n =30). Under the present method, the lowest and highest PtdCh values in tissue samples were about 2 μmol/g (eye ball) and 29 μmol/g (spinal cord), respectively. Methoxamine significantly decreased PtdCh levels and increased free choline levels in mouse artery and submandibular gland. Discussion: Under the present sample processing procedure, the choline values originating from lyso-PtdCh and sphingomyelin were much less than those originating from PtdCh hydrolysis. Thus, it was possible to inject the hydrolyzed mixture directly into the HPLC system for PtdCh assay. Since the present method provides simple, rapid, and highly reliable PtdCh determination, it is suitable for routine assay of PtdCh in a large number of samples.

Antigen I/II protein is a cell surface protein antigen of Streptococcus mutans and is implicated in the adhesion mechanism to tooth surfaces. We have identified the antigen I/II homologous gene (paaA) from Streptococcus cricetus, a member of mutans streptococci. To clarify the mechanism of paaA expression, we sequenced the flanking regions of the paaA gene utilizing the gene-walking method in this study. In the upstream region of paaA, three homologous genes found in the corresponding region of Streptococcus sobrinus were identified. One of the corresponding genes in S. sobrinus was par, which has been characterized as a transcription represser of the antigen I/II (spaA) gene. Therefore, the homologous gene in S. cricetus could have a role in transcription of paaA. In the downstream region of paaA, the paaA homologous gene was found and designated as paaB. The paaB gene was interrupted by a novel insertion sequence element (ISScr1), a member of the IS982 family. Southern hybridization analysis of S. cricetus par indicated that no gene homologous to S. cricetus par was found in the oral streptococci used in this study. Furthermore, the downstream region of antigen I/II (spaA) in S. sobrinus was sequenced using the gene-walking method. No similar gene to antigen I/II gene was found in the region. It was suggested that the paaA might be regulated by the par in S. cricetus.

Whether free choline levels are changeable in vivo in response to different types of autonomic agonists was examined in several mouse organs. Upon one subcutaneous injection of isoproterenol, phenylephrine and pilocarpine, choline levels in whole organ decreased, increased and decreased, respectively, in various organs within 30 min and returned to initial levels in a day. In the three major salivary glands, a delayed choline elevation also appeared on day 2 after one isoproterenol injection and subsided by day 6. Only in the three salivary glands more choline was accumulated after 10 once-a-day injections of isoproterenol than after one isoproterenol injection. Neither phenylephrine nor pilocarpine induced comparable choline accumulation in any organs examined. Isoproterenol injection repeated at a 2-day interval augmented the subsequent, delayed choline elevation. Examination with dobutamine and the adenylyl cyclase activator 6-(3-dimethylaminopropionyl)forskolin suggested that isoproterenol-induced immediate choline lowering was downstream of cAMP synthesis and linked to cAMP more tightly than the choline accumulation, though both choline changes occurred via β1-adrenergic receptors. Choline levels in the salivary glands also changed depending on the form of diet given and particularly in the parotid gland in parallel with gland weights. These results provide the first evidence for the autonomic control of intracellular choline levels; intracellular choline levels might be an integral part of the autonomic signalling pathway.

柴胡加竜骨牡蛎湯 (SRT) および柴胡桂枝乾姜湯 (SKT) は臨床使用において, 虚実の区別はあるものの, お互いに類似した精神・神経症状が目標とされている。本研究においては, 両薬方の中枢神経に対する影響を明らかにするため, 脳内モノアミン類とその代謝に及ぼす影響について, マウスを用いて比較検討した。1) SRTおよびSKTの単回投与は線条体のドパミン作動性神経伝達物質の含量を増大し, 代謝を促進した。2) SRTの反復投与は視床下部および海馬のドパミン作動性神経系の伝達物質の代謝を促進し, アドレナリン作動性神経系の伝達物質の代謝を抑制した。一方, SKTの反復投与は海馬のドパミン代謝を促進し, セロトニン代謝を抑制した。従って, 両薬方のドパミン作動性神経系の充進作用とセロトニン作動性神経系の抑制作用が, 精神・神経症状の調節に関与しているかも知れないことが示唆される。

The present study re-investigated our previous findings that the memory of mice was enhanced by a non-convulsant dose of lidocaine, but was impaired by a convulsant dose of lidocaine in an experiment using a multiple maze. The effects of post-training administration of lidocaine on memory in mice was examined using a passive avoidance task. Twenty-four hours after the training, the test trial latency was measured. Male ddY mice treated with lidocaine 80 mg/kg immediately after the training exhibited a shortened test trial latency and a decreased number of mice showed the maximal latency of 300 sec in passive avoidance task using electric shock as a reinforcement. The mice treated with lidocaine 40 mg/kg immediately after training exhibited a lengthened test trial latency and an increased number of mice showed the maximal latency of 300 sec in passive avoidance task using a dislike for water as the reinforcement. The results supported our previous findings.

To investigate the alteration of the cholinergic neurotransmitter system in hepatic encephalopathy (HE), we measured the concentrations of choline (Ch) and acetylcholine (ACh) in the brain of mice with thioacetamide (TAA)-induced acute liver failure using a high-performance liquid chromatography with electrochemical detection (HPLC-ECD). The concentrations of Ch and ACh in TAA-treated mice decreased in most regional brain areas as compared to control mice. In particular, Ch significantly decreased in the cortex (P < 0.05), hippocampus (P < 0.01), hypothalamus (P < 0.05), midbrain (P < 0.01) and medulla oblongata (P < 0.01). Similarly, ACh significantly decreased in the striatum (P < 0.01), hippocampus (P < 0.01), midbrain (P < 0.01) and medulla oblongata (P < 0.01). Our findings suggest that the suppression of the cholinergic neurotransmitter system may play a role in the pathogenesis of HE in acute liver failure.

This report deals with three different maze methods using spontaneous learning behavior. Forty-eight mice housed in an apparatus with a multiple maze mastered the maze task on the 5th day after the start of housing. Then they were divided into four groups, and two of the groups were treated with AF64A (3.5 nmol, i.c.v.) or trimethyltin (TMT, 3 mg/kg, p.o.), and the other two groups were treated with the vehicle as the respective control. Seven days after the treatment, their memory retrieval was tested. Subsequently, the same mice were housed in the apparatus with a T-maze. After the finish of the experiment using the T-maze, they were housed in the apparatus with an eight arm radial maze. The control groups mastered the T-maze task with a 3-sec delay on the 4th day after the start of housing and the radial maze task on the 10th day after the start of housing. Both the treatments lowered the performance in all maze tasks. These results show that the mice housed in the apparatus with maze learned to negotiate the maze spontaneously, and the apparatuses are useful for estimating memory in mice with little effort.

As a results of years of studies on pyridylsulfonylureas we have found that novel compounds bearing substituted carbamoyl moiety on the 3-position of the pyridine ring were quite safe for corn (Zea mays). After studying the structure-activity relationships of substituents on the carbamoyl moiety and the heterocycles attached to the urea bridge, we confirmed that 2-(4,6-dimethoxypyrimidin-2-ylcarbamoylsulfamoyl)-N,N-dimethylnicotinamide, SL-950 (nicosulfuron) is the most effective against both grass weeds including perennial species and broadleaves at 40 to 80 g a.i./ha. SL-950 is now under development by Ishihara Sangyo Kaisha, Ltd. We established four novel routes to the syntheses of the key intermediates, 2-sulfamoyl-N-substituted nicotinamides.

It is well known that the characteristics of mastication are important for the maintenance of our physical well-being. In this study, to assess the importance of the effects of food hardness during mastication, we investigated whether a long-term powdered diet might cause changes in emotional behavior tests, including spontaneous locomotor activity and social interaction (SI) tests, and the dopaminergic system of the frontal cortex and hippocampus in mice. Mice fed a powdered diet for 17 weeks from weaning were compared with mice fed a standard diet (control). The dopamine turnover and expression of dopamine receptors mRNA in the frontal cortex were also evaluated. Spontaneous locomotor activity, SI time and dopamine turnover of the frontal cortex were increased in powdered diet-fed mice. On the other hand, the expression of dopamine-4 (D4) receptors mRNA in the frontal cortex was decreased in powdered diet-fed mice. Moreover, we examined the effect of PD168077, a selective D4 agonist, on the increased SI time in powdered diet-fed mice. Treatment with PD168077 decreased the SI time. These results suggest that the masticatory dysfunction induced by long-term powdered diet feeding may cause the increased SI time and the changes in the dopaminergic system, especially dopamine D4 receptor subtype in the frontal cortex.

柴胡加竜骨牡蛎湯 (研究用エキス原末; 以下柴竜と略す) の中枢モノアミン関連物質に及ぼす影響について, マウスを用いて検討した。柴竜は50mg/kgまたは400mg/kgを経口的に単回または反復 (1日2回, 7日間) 投与した。脳の摘出, 分割は常法に従って行い, モノアミン関連物質の測定はHPLC-ECDを用いる方法で行った。主な結果は, 次の通りである。大脳皮質では, 50mg/kgの単回および反復投与でDOPAC, HVA含量の増大; 視床下部では, 50mg/kgの単回投与でNE含量の減少, 反復投与でNE含量の減少とDOPAC含量の増大; 線条体では, 50mg/kgの単回投与でDA, DOPAC, HVA, 5-HIAA含量の増大, 反復投与ではそのほかにNE, MHPG含量の増大, 400mg/kgの単回投与でNE, 5-HIAA含量の増大と反復投与でNE含量の増大; 海馬では, 50mg/kgの反復投与でMHPG含量の増大, 400mg/kgの反復投与でHVAの減少が認められた。以上のことから, 柴竜は400mg/kgよりも50mg/kg投与のほうがマウス脳内モノアミン含量への影響が強く, その効果は脳部位によって異なることが示唆される。

Mice were housed in a cage containing a T maze. A watering place was located at the entrance of the maze. The right and left arms of the maze each had two exits, one of which led to the home cage where food was placed, while the other led to the watering place via a bypass. The exit leading to the home cage in either the right or left arm was alternately closed every 90 min. One-way swinging doors were inserted at the entrance arm and between each bypass and the watering place. The mice were given a cholinergic neurotoxin—ethylcholine mustard aziridinium ion (AF64A)—(8 nmol) or saline as a control into the left ventricle 2 weeks before they were housed in the apparatus. Those mice housed in this apparatus mastered the alternation task at a 5-sec delay on day 3 in the sham group and on day 4 in the AF64A group. When a longer delay (5–90 sec) was introduced for the mice that mastered the alternation task at 5-sec delay, the AF64A group made significantly more errors than did the sham group at 60- and 90-sec delays. These results show that the apparatus is useful in estimating working memory in mice with little effort.

Mice were housed in a cage with a maze. A water tap was placed at the entrance of the maze. The exit of the maze connected with another cage (home cage). Food was placed in the home cage. Three different multiple mazes (types 1–3) were placed. 1) Mice were housed for 10 h a day in the apparatus and then removed to a normal cage for fasting. One trial per day was carried out after fasting for 13 h. In each trial, a mouse was put at the entrance of the maze and then the number of errors and the time till it reached the home cage was counted. Mice reached a learning criterion at Trial 2. 2) Administering scopolamine (0.125–0.5 mg/kg) 30 min before Trial four disturbed the maze work dose dependently in a type 3 maze, the most complex maze among the three, but did not in type 1 and 2 mazes. 3) Administering scopolamine (0.25–1.0 mg/kg) 30 min before Trial 11 to the mouse of the type 3 maze did not disturb the maze work. These results show that a mouse housed in a cage with a maze learns the maze without explicit training and scopolamine can differentially effect performance based upon the degree of training.

Publisher Summary A high-resolution model with a Lagrangian method is used to simulate the distribution and concentration of CO 2 injected into the mid-depth ocean over timescales ranging from a week to a few years. A high-resolution model with a Lagrangian tracer enables effects of eddy activities on dispersion of particles to be well represented. Comparison of Lagrangian with Eulerian tracers shows that the Lagrangian tracer avoids artificial diffusion and enables CO 2 maximum concentration at specific sites to be predicted, which helps in assessing CO 2 impact on the marine ecosystem. Ensemble experiments using high-resolution models explicitly deal with dispersion by advection due to mesoscale eddies which cannot be achieved by coarse-resolution models with implicit eddy diffusion, and enable the distribution of CO 2 maximum concentrations to be predicted. A high resolution model is demonstrated with this method is a powerful tool for assessing the effects of CO 2 injection on the marine environment.

Publisher Summary The purpose of this chapter is to analyze the benefits of CO2 ocean sequestration for mitigation of global warming. “Benefit” here means reduced economic damage from global warming due to implementing sequestration. From the start of implementation of ocean sequestration, the rate of increase in the atmospheric concentration of CO2 is lowered compared to that if the technology is not adopted. This in turn begins to reduce the global mean temperature. This lowering of the rate of increase of atmospheric CO2 concentration and reducing of global mean temperature continue after completion of sequestration. This results in lowering of the peak values reached by both atmospheric CO2 concentration and the global mean temperature rise, mitigating economic damage induced by global warming. A novel method is proposed to calculate the benefit of CO2 ocean sequestration, which is expressed in dollars per ton of carbon sequestered. CO2 ocean sequestration is simulated by means of a simple atmosphere-ocean-land box model that accounts for the principal biogeochemical processes that control the global carbon cycle. Three scenarios are considered to reduce future atmospheric CO2 concentrations from business-as-usual forecasts to CO2 double stabilization. Atmospheric CO2 concentration and the increase in global mean temperature are simulated in cases with and without sequestration. The benefit is the difference between damage with and without sequestration. The result is that the benefit of CO2 ocean sequestration some hundreds of years in the future ranges from several hundreds to thousands of dollars per ton of carbon, well exceeding present-day estimated costs.

The flanking region of the antigen I/II gene, paaA, in Streptococcus cricetus was examined using the gene-walking technique. In the region downstream of the paaA gene, another antigen I/II gene designated as paaB was found. The paaB gene was disrupted at the alanine-rich region (A region) by a novel insertion sequence element, ISScr1. ISScr1 is a member of the IS982 family and is composed of a 962-bp sequence and duplicated target DNA (the sequence 5'-TAGCTAAAT-3') resulting from its insertion. To clarify the structural divergence of the two antigen I/II proteins (PAaA and PAaB), computational analysis of the paaB gene was performed and the two structures were compared. The amino acid sequence homology indicated that PAaB resembled PAaA, but the middle region showed little similarity to that of PAaA. Phylogenetic analysis showed that PAaB was better classified in a major group with S. mutans PAc and S. gordonii SspA and SspB than with PAaA. The transcriptional expression of paaA and paaB was demonstrated by reverse transcription (RT)-PCR. In the region upstream of the paaA gene, three genes homologous to the genes located in the region upstream of the S. sobrinus antigen I/II gene (pag) were found. Of the three genes, ORF3 showed homology to the par gene encoding a transcriptional repressor for the pag gene in S. sobrinus. Therefore, ORF3 was designated the par gene of S. cricetus. Southern hybridization revealed that the par gene of S. cricetus was not found in other oral streptococci examined in this study.

Publisher Summary The CO2 Ocean Sequestration Project in Japan was started as a national project in 1997. The implementation of this project has been entrusted to two organizations by the New Energy and Industrial Technology Development Organization (NEDO). The Research Institute of Innovative Technology for the Earth (RITE) was appointed to one of these key roles and has been working on RD the lethal CO2 concentration limit value will be identified for the mid-depth plankton of the site by the method developed in the first phase of the NEDO R&D program, and will then used in the food-web based assessment model of the site. The opportunity of the ocean experiment is open to the international partners, and the verified dilution factor attainable by moving-ship concept, coupled with the established assessment methodology, will be submitted to the international forum to obtain the legal acceptance of this technology as a measure of CO2 emission reduction.

Introduction: To measure the levels of phosphocholine (PCh) and glycerophosphocholine (GPCh) in the tissues and organs of mice, we developed a simple and rapid method using high-performance liquid chromatography (HPLC) with electrochemical detection (ECD) and an immobilized enzyme column. Methods: Under our modifications of the separation procedure of Klein et al. [Neurochem. Int. 2 (1993) 293], PCh and GPCh in the hydrophilic phase of the homogenate samples were selectively hydrolyzed into free choline by alkaline phosphatase and a 0.4-N perchloric acid solution, respectively, and the resulting hydrolyzed mixtures were directly injected into the HPLC system for analysis. Results: The present method permits PCh or GPCh assay within 5 min in one chromatographic run. Recoveries from tissue samples were 97% for PCh and 101% for GPCh. The percentages of the crossover reaction to the authentic PCh and GPCh were 0.4% and 3.8%, respectively. The within-run coefficients of variation for choline derived from PCh and GPCh in the tissue samples were 1.2% and 1.4%, respectively. Discussion: The method is effective and has been applied to the measurement of PCh and GPCh levels in several tissues of mice.

1. The purpose of this study was to demonstrate that the adrenergic nervous system regulates the in vivo choline levels in the mouse major salivary glands. 2. Methoxamine (alpha1-adrenoceptor agonist, 2.5-20 mg kg-1, s.c.) elevated choline levels dose-dependently and the effect of methoxamine (10 mg kg-1) was completely inhibited by the alpha-adrenoceptor antagonist phentolamine (5 mg kg-1, i.p.) but not by the beta-adrenoceptor antagonist propranolol (3 mg kg-1, i.p.). 3. In contrast, isoprenaline (beta-adrenoceptor agonist 0.25-20 mg kg-1, s.c.) lowered choline levels and the effect of isoprenaline (2 mg kg-1) was inhibited by propranolol, but not by phentolamine. 4 Noradrenaline (1-4 mg kg-1, s.c.) manifested both the alpha- and beta-adrenergic actions depending on its dose. Noradrenaline at 1-2 mg kg-1, lowered choline levels and the effect of noradrenaline (1 mg kg-1) was inhibited by propranolol, but not by phentolamine. On the other hand, noradrenaline (4 mg kg-1) elevated choline levels and the effect was blocked by phentolamine, but not by propranolol. 5. Tyramine (5-80 mg kg-1, s.c.) elicited the release of noradrenaline from sympathetic nerve terminals and induced essentially the same effects on the choline levels as noradrenaline. Tyramine (10 mg kg-1) lowered choline levels and the effect was inhibited by propranolol, but not by phentolamine. However, tyramine (80 mg kg-1) elevated choline levels and the effect was inhibited by phentolamine, but not by propranolol. 6. These results suggest that choline levels in the salivary glands may be under separate alpha- and beta-adrenergic control and suggest a possibility that the neurotransmitter noradrenaline released for sympathetic nerve terminals can manage the dual control of choline levels in some autonomic organs in a characteristic dose-dependent manner.

One group of mice was injected subcutaneously with 20 mg/kg body wt isoprenaline each day for 10 days; another group (control) was injected with saline. Half the animals of each group were kept untreated for a further 10 days for restoration. Chronic administration of isoprenaline caused enlargement of parotid (4-fold) and submandibular glands (1.7-fold) but had no effect on sublingual glands. Concomitantly, noradrenaline and acetylcholine contents were, in parallel, increased in parotid, decreased in sublingual, and unchanged in submandibular glands. Under these conditions, pilocarpine- or isoprenaline-induced salivation was not affected but phenylephrine-induced salivation was augmented; the protein component patterns of saliva characteristic of the three sialogogues were also changed. In addition, secretory proteins whose synthesis was induced by isoprenaline were found to be secreted by stimulation with different types of sialogogues. Most changes were reversible. These results indicate that continued beta-adrenoceptor stimulation not only causes broadly altered forms of saliva, probably by involving, in part, alpha-adrenoceptor hypersensitivity, but also changes the activities of both sympathetic and parasympathetic nerves to salivary glands in parallel, though the extent differs among the three glands.

Acetylcholine and noradrenaline concentrations in the submandibular, parotid and sublingual glands, and pilocarpine-, isoproterenol- and phenylephrine-induced salivation, were estimated in streptozotocin (STZ)-induced diabetic mice. Diabetic mice showed significant increases in acetylcholine and noradrenaline (expressed as nmol/gland) in sublingual and submandibular glands, respectively. The total volume of crude whole saliva in diabetic mice in response to pilocarpine and isoproterenol but not to phenylephrine was significantly reduced. These results suggest that alterations in the neurotransmitter levels and secretory function in the salivary glands occur rapidly after the induction of STZ diabetes, and that the secretory function appears to be more susceptible to effects of diabetes in the early stages than the autonomic nervous system. Since the alterations in neurotransmitter concentrations in diabetic salivary glands were slight and partial, it seems that they are unrelated to the markedly reduced salivation in response to pilocarpine and isoproterenol observed in these short-term diabetic mice.

Since administration of a powdered extract (TSS) of Danggui-Shaoyao-San (Toki-shakuyaku-san in Japanese) alone to naive mice had no influence on ACh levels in the brain, the present study examined the effect of TSS on the central cholinergic nervous system using mice treated with scopolamine (0.5 mg/kg) or mecamylamine (0.05 mg/kg), which affects the cholinergic nervous system. TSS was suspended in a 5% carboxymethylcellulose solution and mice were orally given single or repeated (twice a day, for 14 days) administration of TSS at 50 or 500 mg/kg. Results on spontaneous locomotor activity showed that (I) single administration of TSS at 50 or 500 mg/kg to naive mice significantly inhibited vertical and horizontal locomotor activities, while repeated administration of TSS at 50 mg/kg significantly stimulated both activities; (2) in mice treated with scopolamine, repeated administration of TSS at 500 mg/kg significantly inhibited the scopolamine-induced increase in locomotor activities, whereas in mice treated with mecamylamine, single or repeated administration of TSS at 50 and 500 mg/kg did not show any influence on the mecamylamine-induced decrease in locomotor activities. Regarding the step-down passive avoidance responses: single administration, but not repeated administration, of TSS at 50 and 500 mg/kg significantly inhibited scopolamine-induced shortening of step-down latency. In mice treated with mecamylamine, TSS did not exert any influence on the step-down latency. As for ACh contents, single or repeated administration of TSS at 50 or 500 mg/kg to naive mice had no influence on the levels of ACh in the cerebral cortex, corpus striatum or hippocampus. However, the levels of brain ACh in mice treated with scopolamine showed a decrease and a single administration of TSS at 500 mg/kg significantly inhibited this scopolamine-induced decrease in ACh levels. These results indicate that TSS ameliorates dysfunction of the central cholinergic nervous system and scopolamine-induced decrease in ACh levels in mouse brain, but has no influence on ACh levels in naive mice. Thus, it suggests that TSS may be a useful therapeutic agent in Alzheimer's disease and senile dementia.

The effects of the extract powder (CggT) from Chaihu-Guizhi-Gajiang-Tang (Saiko-keishi-kankyo-to, in Japanese) on the monoamines and their related substances and the acetylcholine in mouse brain were examined. 1) A single administration of CggT significantly increased the levels of HVA and 5-HIAA in the cerebral cortex, hypothalamus, corpus striatum and hippocampus at 75 mg/kg, and those in the hypothalamus, corpus striatum and hippocampus at 750 mg/kg. 2) The repeated administration of CggT significantly increased the level of 5-HT in the hippocampus at 75 mg/kg, and the levels of 5-HT in the corpus striatum and of NE and 5-HT in the hippocampus at 750 mg/kg. 3) The level of ACh was significantly increased in the hypothalamus alone after single administration of CggT. These findings suggest that CggT stimulates function of the dopaminergic and serotonergic nervous systems in mice, but not most of the NEnergic and cholinergic nervous systems.

The salivary glands of 5-week-old mice and rats were divided into submandibular, parotid, and sublingual and analysed to determine basal levels of the neurotransmitters noradrenaline (NE) and acetylcholine (ACh) and the possible neurotransmitters dopamine (DA) and 5-hydroxytryptamine (5-HT), using a combination of high-performance liquid chromatography coupled with coulometric and amperometric detection and a direct injection technique employing crude homogenate supernatants. In both species, levels of NE were higher in the submandibular than in the other salivary glands, whereas levels of ACh were higher in the mouse submandibular and the rat sublingual glands than in other glands. In all the salivary glands, levels of DA were markedly lower than those of other target substances. Levels of 5-HT were similar in all salivary glands. These results show that in mouse and rat salivary glands, species differences in neurotransmitter distribution are relatively small, whereas there are considerable differences in distribution between the salivary glands.

Male ddY mice were housed in a cage with an eight arm radial maze apparatus for 6 h a day. A water bottle was placed at the central platform. The end of each arm ran to the home cage through a guillotine door (G). Food was placed at the home cage. During the housing in the apparatus, one G was raised and the remaining seven G were lowered. The raised G was changed every 45 min in random order. Mice housed in this apparatus learned efficient strategy to return the home cage by trial and error. When they chose the arm in which the G was lowered on their way to the home cage from the platform, they returned to the platform, then chose a different arm until they were able to enter the home cage. The mice housed in this apparatus mastered the radial maze task on the 7th day. When scopolamine (SCO) was injected, SCO butylbromide had no effect on performance, but SCO hydrobromide (0.2 and 0.4 mg/kg) impaired working memory, dose dependently. These results show that the apparatus is useful for ease in estimating working memory in mice without the use of severe food or water deprivation.

For the rapid assay of norepinephrine (NE) and its major metabolite, 3-methoxy-4-hydroxyphenylglycol (MHPG) in the mouse brain, we developed a simple method using isocratic HPLC with coulometric detection. This method permits NE and MHPG assay with 5 min in one chromatographic run. Within-run coefficients of variation for NE and MHPG in the working standard solution were 0.8% and 0.6% ( n = 50), respectively. The detector responses were linear from 0.025 to 100 pmol for NE and from 0.05 to 100 pmol for MHPG in the working standard solution. Using this method, the NE and MHPG concentrations were measured in discrete brain areas of the mouse prior treatment with or without α -methyl- p -tyrosine or N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP4).

The effects of nefiracetam [DM-9384; N-(2,6-dimethyl-phenyl)-2-(2-oxo-pyrrolidinyl)acetamide] and of phosphatidylcholine on a step-up active avoidance response, locomotor activities and regional brain cholinergic and monoaminergic neurotransmitters in AF64A-treated mice were investigated. Intracerebroventricular (i.c.v.) injection of AF64A (ethylcholine mustard aziridinium ion; 8 nmol/ventricle) impaired acquisition and retention of the avoidance task, and increased vertical and horizontal locomotor activities. Regional levels of acetylcholine, noradrenaline, 3-methoxy-4-hydroxyphenylglycol (MHPG), 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) were significantly decreased and homovanillic acid (HVA) levels were increased in the hippocampus but not in the septum, cerebral cortex or striatum of AF64A-treated animals. Administration of nefiracetam (3 mg/kg, p.o.) twice daily for 9 days to AF64A-treated animals ameliorated the deficit in active avoidance response in addition to attenuating the increase in locomotor activities. In parallel with these behavioural effects, nefiracetam reversed AF64A-induced alterations in the hippocampal profiles of cholinergic and monoaminergic neurotransmitters and their metabolites. In contrast, administration of phosphatidylcholine (30 mg/kg, p.o.) twice daily for 9 days had no significant effect on the deficit in active avoidance response, despite significantly reversing the decrease in acetylcholine levels in the hippocampus. These results indicate that the effects of nefiracetam on AF64A-induced behavioural deficits are probably due to its ability to facilitate both cholinergic and monoaminergic neurotransmitter systems.

The effects of acute and chronic administration of MKC-231, a new choline uptake enhancer, and two other nootropic agents, linopiridine (Dup 996) and tetrahydroaminoacridine (THA) on working memory deficits and decreased hippocampal acetylcholine (ACh) content were studied in a delayed non-matching to sample task, using a T-maze, in ethylcholine aziridinium ion (AF64A)-treated mice. Treatment with AF64A (3.5 nmol, i.c.v.) produced memory deficits and decreased hippocampal ACh content. In acute behavioral experiments, MKC-231 and THA had no significant effect on AF64A-induced memory deficits at any doses tested (0.3, 1.0 and 3.0 mg/kg), whereas Dup 996, at a dose of 1.0 mg/kg, significantly improved memory deficits. In chronic experiments, MKC-231 improved memory deficit at all doses tested (0.3, 1.0, or 3.0 mg/kg p.o., once daily for 11 days) and Dup 996 did so only at a dose of 3.0 mg/kg, whereas THA did not improve memory deficit at any doses tested. In acute neurochemical experiments, MKC-231 and THA did not reverse the AF64A-induced hippocampal ACh depletion. Dup 996, however, further decreased hippocampal ACh content compared to that in the AF64A-treated group. In chronic experiments, MKC-231 significantly reversed hippocampal ACh depletion at doses of 0.3 and 1.0 mg/kg, whereas neither Dup 996 nor THA reversed hippocampal ACh depletion at any doses tested. These results indicate that MKC-231 improved the AF64A-induced working memory deficit and hippocampal ACh depletion, probably by recovering reduced high-affinity choline uptake and ACh release.

Male ddY mice were housed in a 4-, 6-, or 8-arm radial maze apparatus for 6 hr a day and then removed to a normal cage for fasting until the next day's trial. A water bottle was placed at the central platform. The end of each arm ran to the home cage through a guillotine door (G). Food was placed in the home cage. During the housing in the apparatus, one G was opened, and the remaining Gs were shut. The opened G was changed in random order during the housing period of 6 hr. At the beginning of the trial, all Gs were shut. The mouse was placed on the platform and was permitted to choose the arms until it chose all arms. When the mouse chose the last arm, the G was opened to allow stepping into the home cage. The mice housed in these apparatus learned to go to the home cage without entering already chosen arms within 4-6 days. Scopolamine (0.2 and 0.4 mg/kg, i.p.) impaired the maze performance. A 1- to 4-min delay impaired the performance slightly, but did not show any significant effect depending on the delay intervals. These results suggest that the apparatus is a useful and easy method for estimating working memory and the drug effects thereon in mice.

1. The effect of alpha-sialyl cholesterol (alpha-SC; alpha-D-N-acetylneuraminyl cholesterol) on disturbances of the central cholinergic system induced by ethylcholine mustard aziridinium ion (AF64A) and by scopolamine were studied by means of a step-down passive avoidance response and locomotor activities in mice. The levels of acetylcholine (ACh) in certain regions of the brain were measured to assess the neurochemical recovery promoted by alpha-SC. 2. Treatment with AF64A (2.5, 5 and 10 nmol, i.c.v.) impaired the 24 h retention latencies of animals in a dose-dependent manner, and scopolamine (0.5 mg kg-1, i.p.) also impaired the retention performance. Administration of alpha-SC (1 and 4 mg kg-1, p.o.) once daily for 13 days improved the retention performance in AF64A-treated animals in a dose-dependent manner, but not in the scopolamine-treated animals. 3. Treatment with AF64A (2.5, 5 and 10 nmol, i.c.v.) and scopolamine (0.5 mg kg-1, i.p.) increased vertical and horizontal locomotor activities. alpha-SC dose-dependently attenuated the increase in locomotor activities induced by 2.5 nmol of AF64A, but not the locomotor activities caused by 5 or 10 nmol of AF64A, or scopolamine (0.5 mg kg-1, i.p.). 4. The deficit retention performance of AF64A-treated animals was associated with depletion of ACh levels in the hippocampus, but not in the septum or cerebral cortex. Administration of alpha-SC to AF64A-treated animals dose-dependently reversed the depletion of ACh levels in the hippocampus. 5. The results indicate that alpha-SC had significant effects after oral administration of AF64A-treated animals. The behavioural recovery promoted by alpha-SC may be based on the reversal of ACh depletion in the hippocampus.

To clarify the pharmacological properties of ceruletide (CER) and cholecystokinin-octapeptide (CCK-8) with respect to vertical (VLA) and horizontal (HLA) locomotor activities of mice, effects of pretreatment with CER (0.5, 5, and 50 micrograms/kg, IP) and CCK-8 (5, 50, and 500 micrograms/kg, IP) on apomorphine (0.1 mg/kg, SC)- and clonidine (0.1 mg/kg, SC)-induced hypo-VLA and -HLA and on apomorphine (1 mg/kg, SC)-induced hyper-VLA and -HLA were examined. CER and CCK-8 had a dose-dependent inhibitory effect on VLA and HLA in intact mice. Pretreatment with CER had a biphasic effect (increase and decrease) on apomorphine- and clonidine-induced hypo-VLA, as well as an effect on apomorphine-induced hypo-HLA, a decreased effect on clonidine-induced hypo-HLA, and a decreased effect on apomorphine-induced hyper-VLA and -HLA. On the other hand, pretreatment with CCK-8 had no effect on apomorphine- and clonidine-induced hypo-VLA and -HLA and a decreased effect on apomorphine-induced hyper-HLA but not on hyper-VLA. These results suggest that for apomorphine- and clonidine-induced locomotion in mice CER has pharmacological properties different from those of CCK-8.

The effect of 3,3',5-triiodo-L-thyronine (T3) on working memory in ethylcholine aziridinium ion (AF64A)-treated mice was studied in a delayed non-matching to sample task using a T-maze. After behavioural testing was completed, mice were killed by microwave irradiation and regional brain levels of acetylcholine, aspartate, glutamate, glutamine, glycine, taurine, and gamma-aminobutyric acid (GABA) were measured by high-performance liquid chromatography with electrochemical detection. Treatment with AF64A (7 nmol, i.c.v.) produced a deficit in working memory performance in the non-matching to sample task at 30 s delay, and decreased acetylcholine, glutamate, and GABA levels in the hippocampus, but not in the septum and cerebral cortex. Administration of T3 (0.3 mg/kg, p.o., once daily for 6 days) to AF64A-treated animals improved the deficit in working memory performance and reversed the decrease in acetylcholine, glutamate, and GABA levels in the hippocampus. These results indicate that the deficit in performance induced by AF64A can be improved by T3 administration.

For simultaneous assay of the three monoamine neurotransmitters, norepinephrine, dopamine, and serotonin, and four respective metabolites in brain tissue, a rapid and simple method using high-performance liquid chromotography with coulometric detection is described. Because the present method permits the determination of these target substrates within 10 min or less in one chromotographic run, 150 samples can be analyzed using an autosampler and an integrator in a 24-h period. Within-run coefficients of variation for the target substrates in the standard solution and the whole brain sample were less than 3% and 2% (n = 40), respectively. The quantitative detection limits were 0.01-0.1 pmol. The present procedure was applied to measure the target substrates in several discrete brain areas in mice.

Dopaminergic supersensitivity in mice was induced by pretreatment with a single injection of haloperidol (4.8 mg/kg). After the pretreatment, further treatment with haloperidol (0.6 or 0.01 mg/kg) was made at varying intervals, and catalepsy, locomotor activity and homovanillic acid (HVA) were measured. The intensity of the supersensitivity was evaluated by enhanced apomorphine (1 mg/kg)-induced climbing behavior. Supersensitivity was displayed on the 2nd and the 4th day. The cataleptogenic effect of haloperidol (0.6 mg/kg) was significantly weakened on the 1st, 2nd and 4th days. The motor inhibitory effect of haloperidol (0.01 mg/kg) increased on the 1st, 2nd and 4th days. Homovanillic acid was measured in the striatum and the prefrontal cortex on the 2nd day. Haloperidol (0.6 mg/kg) increased the concentrations of HVA in both regions of the brain. The increase in the concentrations of HVA in the striatum was blunted after the pretreatment, but such tolerance did not develop in the prefrontal cortex. Haloperidol (0.01 mg/kg) did not influence the concentration of HVA in both regions. These results suggest that the behavioral effect of a small dose of haloperidol may be enhanced, rather than reduced, in the phase of supersensitivity.

The effects of 24-hr fasting on the vertical (VMA) and horizontal (HMA) locomotor activities, on cage climbing activity and on brain monoamine-related substances, were examined using male ddY mice. Both the VMA and HMA increased with fasting, but not the cage climbing activity. Methamphetamine (2 mg/kg, SC) increased the VMA and HMA in both the feeding and fasting mice, whereas with apomorphine (0.1 mg/kg, SC) both decreased. Furthermore, pretreatment with haloperidol (0.25 mg/kg, SC) showed no influence on the methamphetamine-induced VMA increase in both the feeding and fasting mice. However, pretreatment with haloperidol inhibited the methamphetamine-induced HMA increase in both the feeding and fasting mice and showed a higher level of HMA in fasting mice than in feeding mice. When measuring brain monoamine-related substances, the DA, NE, 5-HT, and 5-HIAA levels in the corpus striatum increased, whereas the 3-MT level decreased. The monoamine levels in the nucleus accumbens of fasting mice were the same as those in feeding mice, except for a decrease of the 3-MT level. These results suggest that the locomotor activity in fasting mice may be increased by a change in the sensitivity of dopaminergic neurons in the corpus striatum.