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Administration of cisplatin and methotrexate significantly increased 5-hydroxytryptamine (5-HT) release from intestinal tissues isolated at 72 h after administration in rats. Daily administration with nafamostat mesilate, a potent serine protease inhibitor, significantly inhibited the release of 5-HT induced by methotrexate, but not by cisplatin, in a dose-dependent manner. When applied to isolated ileal tissues in vitro, nafamostat mesilate also significantly inhibited the release of 5-HT induced by methotrexate, but not by cisplatin, in a concentration-dependent manner. These results suggest that serine proteases are involved in the mechanism of the methotrexate-induced release of 5-HT from the rat small intestine.

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We achieved a unique light guide that functions as both a parallax barrier and a backlight. We fabricated a LCD using this unique light guide that can be switched between 2D and multi-view 3D modes. In addition to being based on a simple device that can be switched between 2D and 3D modes, our method provides extremely high-quality displays of 2D and 3D images.

The effects of anti-inflammatory drugs on ileal 5-hydroxytryptamine (5-HT) metabolic dynamics at 72 h after a single administration of cisplatin were investigated in rats. Cisplatin 5 mg/kg i.p. caused pathological changes, with an inflammatory response occurring 72 h after its administration. The inflammatory response was associated with the induction of cyclooxygenase-2, but not cyclooxygenase-1, in the ileal mucosa at 72 h after the cisplatin administration. Daily treatment with meloxicam 3 mg/kg s.c. ameliorated the cisplatin-induced mucosal damage, whereas dexamethasone 1 mg/kg s.c. did not. Cisplatin administration also caused a significant increase in cyclooxygenase-2 mRNA expression at 72 h after administration, which was blunted by dexamethasone, but not by meloxicam. Cisplatin increased the content of 5-HT and its metabolite, 5-hydroxyindoleacetic acid (5-HIAA), but had no effect on 5-HT turnover (5-HIAA/5-HT ratio). Meloxicam and dexamethasone did not significantly decrease 5-HT and 5-HIAA content. Cisplatin significantly decreased monoamine oxidase activity but increased tryptophan hydroxylase (TPH) activity and TPH(1) mRNA expression in ileal tissue. Meloxicam and dexamethasone significantly restored the decreased monoamine oxidase activity and inhibited the cisplatin-induced increase in tryptophan hydroxylase activity toward the control levels. These drugs also decreased the cisplatin-induced increase in TPH(1) mRNA expression. Neither cisplatin nor the anti-inflammatory drugs had significant effect on mRNA expression of the serotonin re-uptake transporter. These results suggest that the inflammatory response associated with cyclooxygenase-2 induction is involved in the opposite change in ileal tryptophan hydroxylase and monoamine oxidase activities in the delayed phase after single administration of cisplatin to rats.

We analyzed two disease model groups with rats infected by Japanese encephalitis virus (JEV), a 90-day group and a 180-day group after JEV infection. The time measured by the modified pole test showed that motor activities in these two groups were slower than those of age-matched control groups. Striatal dopamine (DA) levels were significantly decreased in all JEV-infected rats. Norepinephrine concentration in brain regions in the 180-day group was significantly decreased in the medulla oblongata and hypothalamus as compared with the control and 90-day group. Tyrosine hydroxylase-positive neurons were significantly decreased in both JEV-infected rat groups. These results suggest that DA decrease and pathological changes in JEV-infected model rats persist for a long time, at least up to 180 days, and this model will be useful for the evaluation of new anti-parkinsonian agents.

In neurons of dorsal motor nucleus of the vagus that is involved in the gastric motility and possibly emesis, application of 5-hydroxytryptamine produces membrane depolarization, and suppresses spike-repolarization and spike-afterhyperpolarization, suggesting divergent effects of 5-hydroxytryptamine through activating multiple subtypes of 5-hydroxytryptamine receptors. However, only the role of 5-hydroxytryptamine 2A receptors has been established to be responsible for the depolarization, and the mechanisms underlying the modulation of spikes remain unknown although a role of 5-hydroxytryptamine 4 receptors was implicated in modulations of spikes. There is now increasing evidence for the role of 5-hydroxytryptamine receptors in neurons involved in generating emesis following administration of anticancer drug. Since antagonists of 5-hydroxytryptamine 3/4 receptors are widely used as anti-emetic drugs, we have reevaluated the functional roles of 5-hydroxytryptamine 3/4 receptors of dorsal motor nucleus of the vagus neurons, especially in modulating transient outward currents that are presumed to be involved in spike-repolarization and spike-afterhyperpolarization. Whole-cell patch-clamp recordings were made from the dorsal motor nucleus of the vagus neurons, which were identified by a retrograde tracing method with dextran-tetramethylrhodamine-lysine injected into a bundle of abdominal vagus nerves. Under a voltage-clamp condition, dorsal motor nucleus of the vagus neurons expressed a prominent A-like current. The activation of 5-hydroxytryptamine 3 receptors reversibly increased the resting membrane conductance while the activation of 5-hydroxytryptamine 4 receptors led to an almost irreversible decrease in the A-like current. A long-lasting suppression of A-like current by transient activation of 5-hydroxytryptamine 4 receptors would result in a long-lasting increase in the excitability of dorsal motor nucleus of the vagus neurons, which might be involved in generation of the long-lasting facilitation of gastric motility or in generation of the long-lasting gastric relaxation through the activation of enteric non-adrenergic non-cholinergic neurons as implicated in the delayed emesis induced by anticancer drugs.

The effect of docosahexaenoic acid (DHA) on cyclooxygenase expression induced by interleukin (IL)-1β and phorbol 12-myristate 13-acetate (PMA) in rat vascular smooth muscle cells (VSMCs) was investigated in order to clarify the cellular mechanism of cardiovascular protective effects. DHA and eicosapentaenoic acid slightly enhanced IL-1β-induced cyclooxygenase (COX)-2, but not COX-1, expression, whereas arachidonic acid had no effect. DHA also slightly enhanced PMA-induced COX-2 expression. DHA stimulated both rapid and prolonged activation of p44/42, but not p38, mitogen-activated protein kinase (MAPK) induced by IL-1β and PMA. These results suggest that DHA enhances the COX-2 expression by selectively facilitating p44/42 MAPK activation in VSMCs. Keywords:: docosahexaenoic acid, cyclooxygenase-2, vascular smooth muscle cell

We investigated the effect of 5-hydroxytryptamine (5-HT) on nitric oxide (NO) production by vascular smooth muscle cells derived from 7–8 weeks old stroke-prone spontaneously hypertensive rats (SHRSP) and age-matched normotensive Wistar Kyoto rats (WKY). 5-HT significantly inhibited NO production and inducible NO synthase (iNOS) expression induced by interleukin-1β (IL-1β), which effect was greater in SHRSP cells than in WKY cells. The inhibitory effect of 5-HT was mimicked by α-methyl-5-HT, a 5-HT2 receptor agonist, but not by 5-HT1, 5-HT3 or 5-HT4 receptor agonists. 5-HT inhibition of NO production was dose-dependently reversed by sarpogrelate, a 5-HT2 receptor antagonist. Staurosporin, a protein kinase C (PKC) inhibitor, dose-dependently reversed the inhibitory effect of 5-HT, and phorbol 12-myristate 13-acetate, a PKC activator, dose-dependently inhibited the NO production in both WKY and SHRSP cells. Thus, 5-HT had an inhibitory effect on NO production and iNOS expression by vascular smooth muscle cells via 5-HT2 receptor subtype involving PKC activation, which effect was greater in SHRSP cells than in WKY cells. The enhanced inhibitory effect of 5-HT may have a pathophysiological relevance to vascular diseases in SHRSP.

Post-encephalitic parkinsonism has been well documented by now. Recently, an Indian group reported some post-encephalitic parkinsonism patients that resembled Japanese encephalitis cases with bilateral substantia nigra lesions that were detected by MRI. Post-encephalitis parkinsonism has been described following Coxsackie B virus, influenza A, poliovirus and measles virus infections. The possible involvement of virus infection has also been supported by experimental animal models. We have demonstrated pathological and to a certain extent clinical features consistent with Parkinson's disease following infection of rats with Japanese encephalitis virus (JEV). We evaluated new treatments with Parkinson's disease using this model. It was reported that tremor primarily involving the fingers, tongue and eyelids, muscle rigidity and masked face as the clinical features by JEV. Recently brainstem lesions produced by West Nile virus, which is a flavivirus like JEV, were documented by MRI. In the future, post-encephalitic parkinsonism may be found elsewhere in the world. In adult Fischer rats sacrificed 12 weeks after infection with JEV at the age of 13 days, neuronal loss with gliosis was confined mainly to the zona compacta of the bilateral substantia nigra, without lesions in the cerebral cortex and cerebellum. Furthermore, the most severe lesions were in the central part of zona compacta; the lateral cell groups were less affected. Thus, the distribution of the pathologic lesions in infected rats resembled those found in Parkinson's disease. JEV-infected rats showed marked bradykinesia. Significant behavioral improvement was observed upon administration of L-DOPA and monoamine oxidase (MAO) inhibitor. The findings suggest that JEV infection of rats under the conditions described may serve as a model of virus induced Parkinson's disease.

Patients with bulimia nervosa (BN) have disturbances of mood and behavior related to monoamine activities. There have been no reports concerning salivary concentrations of 5-hydroxytryptamine (5-HT) in patients with eating disorders such as anorexia nervosa (AN) and BN. In order to elucidate the involvement of 5-HT in eating disorders, 5-HT levels in the saliva of patients with BN were measured. Salivary 5-HT levels in patients with avoidant personality disorder (APD) were also measured as an active control. 5-HT levels in the saliva of patients with BN and APD were compared with those of 32 healthy volunteers. Simultaneously, salivary 3,4-dihydroxyphenylacetic acid (DOPAC) was measured using HPLC-ECD. 5-HT levels were 41.04 ± 14.41 in 11 patients with BN (10 women and 1 man, average age 19.6 years), 78.55 ± 45.55 in 5 patients with APD (3 women and 2 men, average age 19.2 years) and 8.91 ± 1.10 ng/ml in 32 healthy controls (25 women and 7 men, average age 21.7 years), respectively. This is the first report that salivary concentrations of 5-HT were significantly higher in BN (p < 0.05) and in APD than those in healthy controls. We found no significant differences among the salivary DOPAC levels in BN, APD and those of healthy controls. There were no significant differences in body mass index among the three groups. 5-HT concentrations in the saliva may be a useful marker of stress or eating disorders.

Cancer chemotherapy-induced nausea and vomiting have been demonstrated to involve humoral as well as neuronal mechanisms. A leading role of serotonin (5-hydroxytryptamine, 5-HT) in these mechanisms is supported by inhibition of the emesis by 5-HT3 receptor antagonists. We compared the effects of granisetron, a selective 5-HT3 receptor antagonist, and vagotomy on c-fos mRNA expression in the nucleus of the solitary tract (NTS) and the area postrema (AP) of the rat caudal brainstem by means of in situ hybridization. The expression of c-fos mRNA in the NTS and AP was significantly elevated 2 h after cisplatin administration. The induction of c-fos expression by cisplatin in the NTS was significantly inhibited by pretreatment with granisetron. In contrast, the c-fos expression in the AP did not differ between the cisplatin group and the granisetron-treated cisplatin group. The degree of the induction of c-fos mRNA expression in both the AP and NTS was similar between the vagotomy and sham operation groups. Our results suggest that the expression of c-fos mRNA in the NTS may be specifically controlled by 5-HT3 receptors and that nonspecific humoral factors, such as modulation of transcriptional activity, play an important role in c-fos expression in the AP after vagotomy.

We previously reported that exogenously administered isatin, an endogenous monoamine oxidase (MAO) inhibitor, significantly increased acetylcholine (ACh) and dopamine (DA) levels in the rat striatum. Selegiline [(-)-deprenil] was developed as a MAO-B inhibitor more than 30 years ago and widely used in the treatment of Parkinson's disease. Effects of isatin or selegiline were investigated in Japanese encephalitis virus (JEV)-induced post-encephalitic parkinsonism rats by a pole test for detecting motor activity and by the determination of biogenic amine levels. Motor activity of JEV-induced rats receiving isatin (100 mg/kg per day for 1 week, i.p.) or selegiline (0.2 mg/kg per day for 1 week, i.p.) was significantly improved compared with that of untreated JEV-infected rats. Both isatin and selegiline prevented the decrease in striatal DA levels in JEV-rats. The increased turnover of DA (DOPAC/DA) induced by JEV was significantly inhibited by isatin, but not by selegiline. These results suggested that exogenously administered isatin and selegiline can improve JEV-induced parkinsonism by increasing DA concentrations in the striatum.

Isatin, an endogenous monoamine oxidase (MAO) inhibitor, has an important role in the control of neurotransmitter concentration. We previously reported that exogenously administered isatin significantly increased acetylcholine (ACh) and dopamine (DA) levels in the rat striatum. In order to test the possibility of treating Parkinson's disease by isatin, we evaluated DA levels in the striatum and bradykinesia using a rat model of Parkinson's disease induced by the Japanese encephalitis virus (JEV). We have already reported that in adult Fischer rats infected with JEV at day 13, there was a marked decrease of tyrosine hydroxylase-positive neurons in the bilateral substantia nigra after 12 weeks. Effects of isatin were investigated in JEV-induced post-encephalitic parkinsonism rats by a pole test and high performance liquid chromatograph (HPLC) with an electrochemical detector (ECD). Isatin (100 mg/kg per day for 1 week, intraperitoneal injection) improved the bradykinesia observed in the JEV-induced parkinsonism rats. Dopamine (DA) concentrations in the JEV-infected rats were profoundly reduced in the striatum as compared with controls. Isatin also increased DA in the striatum of parkinsonism rats. These results suggest that isatin could be a possible treatment for Parkinson's disease as well as for post-encephalitic parkinsonism.

In this study, rat kaolin ingestion (pica) and serial urinary 5-hydroxyindoleacetic acid (5-HIAA) excretions were determined for three days after a single administration of low-dose cisplatin (5 mg/kg, intraperitoneal, i.p.). We found that cisplatin induced acute (first 24 hours after chemotherapy) and delayed phases (24-72 hours after chemotherapy) of kaolin ingestion in rats. An increase in urinary 5-HIAA excretion was also observed in the cisplatin-administered group. These results suggest that 5-HT is involved in delayed emesis, and that urinary 5-HIAA excretion can be used as a marker in the study of delayed emesis.

—Emesis caused by cytotoxic drugs is associated with an increase in the concentration of serotonin (5-hydroxytryptamine: 5-HT) in the intestine and the brainstem. 5-HT receptors in the central nervous system (CNS) and the gut participate in the induction of emesis. 5-HT1 receptors are negatively coupled to adenylate cyclase, but the 5-HT2C subtype is linked to phospholipase (PL) C activation. The 5-HT4 receptor, by contrast, is positively coupled to adenylate cyclase. 5-HT1A and 5-HT2A /5-HT2C agonists exhibit antiemetic properties. The 5-HT1D agonist reduces the emesis accompanying migraine headaches. Through use of 5-HT3 receptor antagonists, which selectively antagonize 5-HT3 receptors on the abdominal vagal afferent fibers, as much as 60% of the initial emetic response can be prevented. Inhibition of acute emesis also appears to be attained by blocking the initiation of the emetic reflex induced via 5-HT3 receptors by 5-HT released from enterochromaffin (EC) cells in the small intestine. Although 5-HT4 receptor antagonists have an antiemetic activity in some experimental models, the precise role of 5-HT4 receptors has not been fully elucidated. The present review aims to compare the involvement of 5-HT4 receptors with that of the 5-HT3 receptors in anticancer drug-induced emesis.

The abdominal vagus is the major nerve involved in the detection of emetic stimuli. Although afferent vagus nerves are considered to have polymodal properties, depolarization of the vagus nerve is mainly mediated by 5-HT in emesis. We studied direct drug effects on rat abdominal vagus nerve using the grease-gap technique. In this study, we also investigated reactions to various emetic stimuli such as dopamine (DA), prostaglandins (PGs) and digitalis. The abdominal vagus nerves rapidly depolarize with potassium chloride application, rapidly declining towards basal levels by the end of the application. Concentration-related depolarization evoked by DA (1 × 10-8 M to 1 × 10-4 M) showed a pattern, in contrast to potassium, where depolarization occurred approximately 30 s later. PGE1 and PGE2 showed a maximum response to concentration of 1 × 10-5 M and 1 × 10-4 M, respectively. The concentration-related curves of PGE1 are similar in PGE2, in that 1 nM elicited depolarization. Ouabain (1 × 10-9 M to 1 × 10-4 M) induced concentration-dependent depolarization responses on the isolated abdominal vagus nerve. The application of 100 μM ouabain induced prolonged depolarization. We confirmed that the peripheral role of the vagus nerve may be involved in the appearance of emesis caused by DA, PGs and digitalis. The grease gap technique of the isolated abdominal vagus nerve is a useful technique for screening the emetic agents.

The influence of tooth loss on the viability of cholinergic neurons was examined in rats. At 25th postnatal week, rats were divided into the three groups; a control group fed a solid diet, a soft diet group fed a powder diet and a molar crown-less group in which all molar crowns were removed and the powder diet was given. At 15 and 35 weeks post-treatment, the number of choline acetyltransferase (ChAT)-positive neurons in the nucleus of the diagonal band/medial septal nucleus (NDB/MS) was significantly smaller in the molar crown-less group than in the control group (P < 0.01). This was not the case in the pedunculopontine tegmental nucleus or (PPT) or in the trigeminal motor nucleus. Biochemical assay showed no statistically significant differences in choline concentrations in the hippocampus between the control and the molar crown-less group both at 15 and at 35 weeks post-treatment. Nevertheless, acetylcholine (ACh) concentration in the hippocampus of the molar crown-less group was significantly lower than that of the control group at 15 weeks post-treatment (P < 0.05). Taken together, a decrease of oral sensory information may have caused a reduction in the number of ChAT-positive neurons selectively in NDB/MS, which in turn caused a decline of ACh concentrations in the hippocampus.

The present study was undertaken to elucidate the effects of acute and subacute administration of docosahexaenoic acid (DHA) on blood pressure in SHRSP between the ages of 6 and 11 weeks. The continuous administration of DHA for 5 weeks in SHRSP significantly inhibited blood pressure rise as compared with levels in non-treated SHRSP. DHA-treated SHRSP showed significantly decreased urinary epinephrine contents as compared with those in non-treated SHRSP. DHA-treated SHRSP produced a significant decrease in adrenal epinephrine concentration as compared with those in non-treated SHRSP at the age of 11 weeks. DHA-treated SHRSP also demonstrated significant decreases in serum triglycerides, LDL and lipid peroxide levels as compared with levels in non-treated SHRSP. However, a single intraperitoneal administration of DHA did not exert any influence on blood pressure in SHRSP at the age of 11 weeks. These findings suggest that the anti-hypertensive effect induced by a 5-week, continuous administration of DHA appears to be associated with the amelioration of increased sympathoadrenal activity and serum lipid derangement during the developmental stage of hypertension in young SHRSP.The present study clarified that a 5-week, continuous administration of DHA had a protective effect against the development of hypertension in SHRSP.

The effect of docosahexaenoic acid (DHA) on nitric oxide (NO) production and inducible NO synthase (iNOS) expression induced by interleukin (IL)-1beta, and whether the effect of DHA is related to its effect on mitogen-activated protein kinase (MAPK) activation were investigated in cultured rat vascular smooth muscle cells (VSMCs). DHA and eicosapentaenoic acid (EPA), although less potent, increased the NO production induced by IL-1beta (3 ng ml(-1)) in a concentration-dependent manner (3 - 30 microM) Arachidonic acid had no significant effect. The stimulatory effect of DHA (30 microM) on the NO production was more obvious at lower concentrations of IL-1beta. IL-1beta induced iNOS protein and mRNA expressions, which were significantly potentiated by DHA. EPA (30 microM) had a tendency to increase the iNOS protein and mRNA expressions, but arachidonic acid had no effect. IL-1beta-induced iNOS protein expression was significantly inhibited by PD 98059 (10 microM), a selective inhibitor of p44/42 MAPK kinase, both in the absence and the presence of DHA. SB 203580 (10 microM), a selective inhibitor of p38 MAPK activity, had no significant effect, although had a tendency to inhibit slightly. IL-1beta increased the phosphorylation of p44/42 MAPK, while it did not apparently increase the phosphorylation of p38 MAPK. DHA significantly potentiated the IL-1beta-induced phosphorylation of p44/42 MAPK, while it had no significant effect on the phosphorylation of p38 MAPK. These results suggest that DHA increases NO production by potentiating iNOS expression induced by IL-1beta through mechanism involving p44/42 MAPK signalling cascade in rat VSMCs. The present study may contribute to the understanding of basic mechanisms underlying the beneficial effects of DHA on various cardiovascular disorders.

The present study evaluates juvenile stroke-prone spontaneously hypertensive rats (SHRSP) as an animal model of a developmental disorder, which is diagnosed according to hyperactivity-impulsivity and/or inattention. To characterize behavioural alterations, we studied motor activity, as well as emotional and cognitive behaviours in juvenile SHRSP, with and without methylphenidate, a psychostimulant. Ambulatory and rearing activities in the open-field environment were significantly higher in SHRSP than in Wistar-Kyoto rats (WKY). In the elevated plus maze task, the entries into open arms, as an index of impulsivity, were significantly increased in SHRSP. In the Y-maze task, spontaneous alternation behaviour, as an index of attention, was significantly lowered in the male SHRSP, but not in the female SHRSP, indicating that spontaneous alternation deficit is gender specific. Methylphenidate (0.01-1 mg/kg, i.p.) significantly attenuated locomotor hyperactivity at low doses and dose-dependently improved the spontaneous alternation deficit in SHRSP. Our findings reveal that juvenile SHRSP manifest problematic behaviours resembling a developmental disorder, attention-deficit/hyperactivity disorder (ADHD), namely hyperactivity-impulsivity and/or inattention. Methylphenidate alleviated the behavioural symptoms of hyperactivity and inattention. We propose that juvenile SHRSP are an appropriate animal model of a developmental disorder resembling ADHD, from behavioural and pharmacological perspectives.

The effect of CP-99, 994, a tachykinin NK(1) receptor antagonist, on abdominal vagal afferent nerve activity in the ferret was investigated. Substance P (1 microg/kg, i.v.) increased vagal afferent activity by 449.0+/-51.9% and this was reduced to 145.9+/-5.7% (p0.01) by pre-treatment with CP-99, 994 (1 mg/kg, i.v.), and to 149.5+/-1.5% (p0.001) by granisetron (1 mg/kg, i.v.), a 5-HT(3) receptor antagonist. In addition, the increase in vagal nerve activity induced by 5-HT (25 microg/kg, i.v., 552.0+/-57.0% increase from pre-injection level) was significantly reduced (401.3+/-10.6% increase from pre-injection level, p0.05) by CP-99, 994 (100 microg/kg, i.v.). These results provide evidence for an involvement of peripheral NK(1) and 5-HT(3) receptors in substance P-induced vagal afferent activation. While the functional consequences (if any) of such peripheral effects were not investigated, they could contribute either directly (e.g. by blockade of receptors on vagal afferents) or indirectly (e.g. modulation of 5-HT release or reduction of local inflammatory response) to the antiemetic effects of CP-99, 994 against cisplatin and other emetic agents acting primarily via the vagus.

The effects of Ni2+, a non-selective cation channel inhibitor, on 5-hydroxytryptamine (5-HT)- and angiotensin II (Ang II)-induced intracellular Ca2+ dynamics in rat aortic smooth muscle cells were investigated. Ni2+ (1 mM) significantly inhibited the transient increase in intracellular Ca2+ concentration ([Ca2+]i) induced by Ang II (100 nM) in aortic smooth muscle cells, as measured using fura-2. However, Ni2+ did not suppress the transient increase in Ca2+ influx induced by 5-HT (10 μM), while significantly suppressed the sustained increase. Ca2+ influx evoked by high KCl (80 mM), thapsigargin (TG) (1 μM) or depletion of intracellular Ca2+ store was almost completely suppressed by Ni2+. Ni2+ had no effect on 5-HT-induced inositol triphosphate production and Ca2+ release from the intracellular store(s). These results suggest that 5-HT, but not Ang II, induces transient Ca2+ influx through Ni2+-insensitive Ca2+ channels, which are distinguishable from the voltage-dependent or store-operated Ca2+ channels.

Although serotonin (5-HT) release from enterochromaffin (EC) cells is considered to be regulated by multiple receptor-mediated mechanisms, little is known about the signal transduction. Here, we introduce the methods to isolate the mouse ileal crypts, which consist of various types of cells including EC cells, and to analyze the intracellular calcium dynamics. Ileal tissue was inserted with a plastic straw and the smooth muscle layers were peeled off. The mucosa were digested with collagenase and then suspended by moderate pipetting. Ileal crypts were separated by stepwise filtrations through 2 different nylon-meshes. The isolated crypt contained 0-3 EC cells as identified by immunostaining using anti-5-HT antibody followed by confocal microscopy. Isolated crypts were attached to a coverglass by an adhesive material (Cell-Tak) and loaded with fura-2/AM. Intracellular calcium dynamics in EC cells were obtained by digital video-imaging analysis of fura-2 fluorescence followed by the identification of EC cells with immunostaining of 5-HT granules. By these methods, it was suggested that norepinephrine elicited a transient elevation of intracellular calcium concentration in EC cells via alpha 2-adrenoceptors. These methods could be also useful to analyze the signal transduction system in intestinal endocrine cells that contain various intestinal hormones such as gastrin, cholecystokinin or secretin.

Isatin (indole-2,3-dione), an endogenous inhibitor of monoamine oxidase (MAO), has several physiological properties for stress and anxiety. We previously identified isatin in the brain of stroke-prone spontaneously hypertensive rats (SHRSP) using gas-chromatography mass spectrometry. This study elucidated the effects of isatin on the ACh and DA levels of brain tissues in rats. Furthermore, we evaluated the effect of isatin on DA levels in a rat model of Parkinson's disease induced by Japanese encephalitis virus. Striatal ACh and DA levels significantly increased at 2 hours after isatin (50-200 mg/kg, i.p.) administration. Perfused through a microdialysis probe, isatin (10(-6)-10(-4) M) also produced a significant and concentration-dependent increase in the ACh and DA concentrations in the perfusate from the rat striatum. Furthermore, urinary isatin concentrations in patients with Parkinson's disease tend to increase according to the severity of disease. Isatin (100 mg/kg, i.p.) significantly increased striatal DA levels in a rat model of Parkinson's disease. These results suggest that urinary isatin may become a diagnostic marker for the clinical severity of Parkinson's disease and that endogenous isatin, a new biological modulator, may play a role in the regulation of the brain levels of ACh by increasing the level of DA under stress.

1. Granisetron and its combination with dexamethasone for the treatment of delayed emesis following cisplatin (CDDP) administration were investigated using ferrets. 2. CDDP-induced emesis was significantly inhibited in both the granisetron group and the combined granisetron and dexamethasone group during the acute and delayed phase in terms of total emesis, latency to first emesis and duration of emesis. 3. Food and water consumption in the combined group of ferrets was significantly increased as compared with the CDDP control group. 4. 5-Hydroxytryptamine (5-HT) level was increased in the ileum and the 5-hydroxyindole acetic acid (5-HIAA) level was increased in the area postrema of ferrets after 3 days of CDDP administration. It is suggested that the antiemetic activity of granisetron and/or dexamethasone is not related to 5-HT levels in delayed emesis. 5. Both granisetron and its combination with dexamethasone are effective in CDDP-induced emesis, but combination treatment is more effective than granisetron alone for the duration of emesis in the delayed phase.

We have recently shown that inferior performance in passive avoidance task is accompanied with decreased hippocampal choline (Ch) in stroke-prone spontaneously hypertensive rats (SHRSP) compared with normotensive control Wistar–Kyoto rats (WKY). We also reported that dietary docosahexaenoic acid (DHA) suppresses the development of hypertension and stroke-related behavioral changes, resulting in the prolongation of the life span of SHRSP. In this study, we examined the effect of dietary DHA on the cerebral acetylcholine (ACh) levels and learning performance in passive avoidance tasks in SHRSP. The arachidonic acid decreased and the DHA increased in plasma lipids dose dependently with dietary DHA treatments, which decreased the systolic blood pressure in SHRSP. Dietary DHA significantly restored the significantly inferior learning performance in passive avoidance response observed in control SHRSP (DHA 0%). Furthermore, the hippocampal ACh levels were correlated positively with the total response latency in passive avoidance tasks. These results suggest that cholinergic dysfunction in the brain of control SHRSP is responsible, at least in part, for the impaired learning ability and the dietary DHA ameliorates this performance failure.

Background and Purpose We investigated age-related changes in the central cholinergic systems in stroke-prone spontaneously hypertensive rats (SHRSP) to examine whether the regional and progressive cholinergic changes occur and are correlated with behavioral changes in the passive avoidance task. Methods Tissue levels of choline (Ch) and acetylcholine (ACh) were determined in the cerebral regions, including the hippocampus, of SHRSP (at two ages: 15 to 20 and 30 to 40 weeks) that had been tested in a passive avoidance task and were compared with those of age-matched controls, Wistar-Kyoto rats (WKY). With the use of in vivo microdialysis, high K + -stimulated release of hippocampal ACh, a functional parameter of the cholinergic system, was also determined in 15- to 20-week-old SHRSP. Results We found that 15- to 20-week-old SHRSP demonstrated a markedly lower level of hippocampal Ch than age-matched WKY. The decrease in the Ch level in 15- to 20-week-old SHRSP was observed in all regions examined; however, in the hippocampus a significant difference from WKY was subsequently observed at the age of 30 to 40 weeks. The hippocampal ACh release was markedly decreased by repetitive stimulation with high K + in 15- to 20-week-old SHRSP. Behavioral impairment in the passive avoidance task was observed in the two age groups of SHRSP, with significant and positive correlations between the hippocampal ACh levels and the response latency. Conclusions A decrease in hippocampal Ch level was observed in both 15- to 20-week-old and 30- to 40-week-old SHRSP, accompanied by performance failure in the passive avoidance task. The abnormal release of hippocampal ACh in response to the repetitive K + stimulation was also noted in 15- to 20-week-old SHRSP. Thus, cholinergic dysfunction in the hippocampal system may be responsible for behavioral abnormality in the passive avoidance task in SHRSP.

There are now abundant evidence to confirm the role of serotonin (5-HT) and in particular, 5-HT3 receptors in the control of cisplatin-induced emesis. Emesis caused by cisplatin is associated with an increase in the concentration of 5-HT in the intestinal mucosa and in the area postrema. The intestinal mucosa contains enterochromaffin (EC) cells that synthesize and secrete approximately 80% of all 5-HT produced in the body. A selective 5-HT3 agonist, 2-methyl-5-HT, induced a dose-dependent increase in 5-HT level from the ileum. Furthermore, a selective 5-HT4-receptor agonist, 5-methoxytryptamine also induced a concentration-dependent increase of 5-HT level. Both 5-HT3 and 5-HT4 receptors may be involved in intestinal 5-HT release. It is proposed that anticancer drugs cause 5-HT release from the EC cells and that the released 5-HT stimulates the 5-HT3 receptors on the afferent vagal fibers, resulting in their deporalization. Electrical stimulation of the abdominal vagal afferents is capable of inducing emesis, and abdominal vagotomy suppresses cisplatin-induced emesis. These results indicate that the vagus is the major afferent pathway involved in the detection of emetic stimuli.

This study was performed to assess the effect of BMY21190, an inhibitor of cAMP phosphodiesterase, on infarct size using a canine ischemic model that underwent a 90-min occlusion and a 6-hour reperfusion of the left coronary artery. The infarct zone/area at risk of the BMY21190 group was significantly smaller than that of the vehicle group (36.1 +/- 7.8%; 62.4 +/- 4.3%, respectively; p0.05). Myeloperoxidase activity, an indicator of neutrophil infiltration, was significantly correlated to infarct size (r = 0.6893, p0.02). Myeloperoxidase activity (0.14 +/- 0.07 U/100 mg tissues) measured in the area at risk from hearts of the BMY21190-treated group was significantly lower than that of the vehicle-treated tissue (0.40 +/- 0.08 U/100 mg tissue, p0.05). It is suggested that BMY21190 reduces infarct size through the inhibition of neutrophil infiltration in the canine model.

A number of compounds capable of inhibiting the aggregation of blood platelets have been reported in recent years. Synthesized compounds containing the imidazo [4,5-b] quinoline-Zone nucleus are a new series of structurally-novel inhibitors of platelet aggregation (38). These compounds are potent broad-spectrum inhibitors that inhibit platelet aggregation caused by thrombin, ADP, and collagen with roughly equal potency (8,15). BMY21190 (Fig. 1) inhibits ADPand collagen-induced aggregation of rabbit platelets with an efficacy comparable to prostacyclin (38). BMY21190 may also be capable of elevating the cAMP levels within platelets sufficiently to activate the CAMP-dependent protein kinase as does BMY20844 (60, Fig. 1). The active catalytic subunit is released by the binding of cAMP to the regulatory subunit of the protein kinase and then acts at many different steps in the biochemical pathways to inhibit platelet inhibition (13,58,59). Agents that elevate platelet cAMP levels, including inhibitors of platelet cAMP phosphodiesterase (PDE) (2,23,38,46,65), are known to inhibit platelet aggregation (21,38, 40). Platelet aggregation is generally regarded as an important event in the initiation of arterial thrombosis. The current interest in inhibitors of CAMP-PDE as therapeutic agents has largely arisen from the recognition that there are distinct PDE isozyme families and that tissues have different complements of isozymes. Nomenclature and subcellular localization of CAMP-PDE isozymes are to be found in articles from Beavo’s (4,28) and Weishaar’s laboratories (67,68). BMY20844 and BMY21190 have the characteristics of selective type I11 inhibitors of “low Km” one of which is high affinity for cAMP (38). The CAMP-PDE type 111 isozyme has been isolated from or found in platelets (17,46), cytosolic fractions of bovine cardiac tissue (19), and

Considering the importance of 5-hydroxytryptamine (5-HT) and cyclooxygenase (COX) products in vascular pathology, we investigated the effects of 5-HT on COX expression in rat vascular smooth muscle cells (VSMCs), and to provide mechanistic insights into these effects. VSMCs were enzymatically isolated from aortic media of Wistar rats. Incubation of VSMCs with 5-HT for 24 h stimulated prostaglandin I 2 production, but this stimulation was completely suppressed by NS-398, a selective COX-2 inhibitor. 5-HT induced transient COX-2, but not COX-1, protein and mRNA expression in concentration- and time-dependent manners. This effect of 5-HT was completely inhibited by sarpogrelate, a 5-HT 2A receptor antagonist. 5-HT-induced COX-2 expression was markedly blunted by Ca 2+ depletion; GF 109203X, a protein kinase C (PKC) inhibitor; PP2, an inhibitor of Src-family tyrosine kinase (Src); PD 98059, an inhibitor of extracellular signal-regulated kinase (ERK) activation; SB 203580, an inhibitor of p38 mitogen-activated protein kinase (MAPK); and SP 600125, an inhibitor of c-Jun N-terminal kinase (JNK). 5-HT activated ERK and p38 MAPK, followed by JNK activation. PP2 inhibited these activations, while GF 109203X inhibited only JNK activation. Furthermore, PD 98059 inhibited JNK activation. These results suggest that 5-HT induces COX-2 expression in rat VSMCs, and that PKC, Src, and MAPK activation are each essential for the full expression of COX-2 pathways.

1. An excitatory response to 5-hydroxytryptamine (5-HT) was measured from the afferent vagus nerve of anaesthetized rats. Measurements were determined by an extracellular recording from the whole nerve. 2. Intravenous bolus injection of 5-HT (1.56-100 micrograms kg-1) evoked a dose-dependent excitation of afferent vagus nerve activity. This response was blocked not only by a selective 5-HT3 receptor antagonist, GR38032F (10 and 100 micrograms kg-1), but also by a 5-HT2 receptor antagonist, ketanserin (10 and 100 micrograms kg-1). 3. Both a 5-HT3 receptor agonist, 2-methyl-5-HT (3.12-100 micrograms kg-1), and a 5-HT2 receptor agonist, alpha-methyl-5-HT (3.12-50 micrograms kg-1), produced a dose-dependent excitation of afferent vagus nerve activity. These excitatory effects were antagonized by GR38032F (10 micrograms kg-1) and ketanserin (10 micrograms kg-1), respectively. 4. A 5-HT1 like receptor agonist, 5-carboxamidotryptamine (50 micrograms kg-1), and a putative 5-HT4 receptor agonist, 5-methoxytryptamine (100 micrograms kg-1), failed to produce excitatory effects on the afferent vagus nerve. 5. These results suggest that the 5-HT-induced excitatory response of the afferent vagus nerve might be mediated not only via 5-HT3 receptors but also via 5-HT2 receptors in anaesthetized rats. It is unlikely, however, that either 5-HT1-like or putative 5-HT4 receptors are involved in the excitatory response of the afferent vagus nerve to 5-HT.

Cancer patients receiving cytotoxic drugs often experience severe nausea and vomiting. Effective antiemetic therapy has not been established. Cisplatin, a prototype of anti-cancer drugs, produces a dose-related emetic response in the ferret. Newly developed 5-HT3 receptor antagonists can prevent emesis induced by anti-cancer drugs. The enteric serotonin (5-HT) concentration of a cisplatin-administered group increased significantly compared with that of control frrrets. A pharmacokinetic study revealed that the efficacy of 5-HT3 antagonists depended on its concentration in the blood. Cytotoxic therapy stimulates the synthesis and release of neurochemical agents including 5-HT from the gut. The vomiting center may be activated directly by visceral afferent neurons or by input from the chemoreceptor trigger zone of the area postrema. It was demonstrated that 5-HT3 antagonists were useful for treatment of the emesis evoked by anti-cancer drugs. Further studied are required to evaluate the site of action of anti-emetic drugs and the precise mechanism of the vomiting induced by anti-cancer drugs.

The present study was undertaken to elucidate the anti-tachycardic effect of a direct-acting vasodilator, budralazine, using an electrophysiological technique. Normotensive male Wistar rats were used. Rats were anesthetized intraperitoneally with urethane and alpha-chloralose. Intravenous administration of budralazine (0.5-5.0 mg/kg) produced a dose-dependent reduction of mean arterial pressure in anesthetized rats. At doses of 0.5 and 1.0 mg/kg, budralazine induced bradycardia accompanied with a decrease in cardiac sympathetic nerve activity (ICNA). Preganglionic adrenal sympathetic nerve activity (ASNA) was also reduced by budralazine (1.0 mg/kg). A 0.5 mg/kg of budralazine neither influenced carotid sinus nerve activity nor augmented aortic depressor nerve activity (ADNA). On the contrary, at dose of 5.0 mg/kg, budralazine produced a tachycardia accompanied with increases in both ICNA and ASNA. The ADNA was decreased by budralazine (5.0 mg/kg) significantly. These findings suggest that the central sympathoinhibitory action of budralazine may be responsible for the anti-tachycardic effect of budralazine and baroreceptor-mediated tachycardia occurred after high dose of budralazine.

The effects of sphingosine 1-phosphate (S1P) on prostaglandin I(2) (PGI(2)) production and cyclooxygenase (COX) expression in cultured rat vascular smooth muscle cells (VSMCs) were investigated. S1P stimulated PGI(2) production in a concentration-dependent manner, which was completely suppressed by NS-398, a selective COX-2 inhibitor, as determined by radioimmunoassay. S1P stimulated COX-2 protein and mRNA expressions in a concentration- and time-dependent manner, while it had no effect on COX-1 expression. S1P(2) and S1P(3) receptors mRNA were abundantly expressed in rat VSMCs. Suramin, an antagonist of S1P(3) receptor, almost completely inhibited S1P-induced COX-2 expression. Pretreatment of VSMCs with pertussis toxin (PTX) partially, but significantly inhibited S1P-induced PGI(2) production and COX-2 expression. S1P also activated extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK). However, neither PD 98059, a selective inhibitor of ERK activation, nor SB 203580, a selective inhibitor of p38 MAPK, had a significant inhibitory effect on S1P-induced COX-2 expression, suggesting that the MAPK activation does not play main roles in S1P-induced COX-2 induction. S1P-induced COX-2 expression was inhibited by PP2, an inhibitor of Src-family tyrosine kinase, Ca(2+) depletion, and GF 109203X, an inhibitor of protein kinase C (PKC). These results suggest that S1P stimulates COX-2 induction in rat VSMCs through mechanisms involving Ca(2+)-dependent PKC and Src-family tyrosine kinase activation via S1P(3) receptor coupled to PTX-sensitive and -insensitive G proteins.

A single dose of isatin (indole-2,3-dione)(i.p.), an endogenous MAO inhibitor, significantly increased norepinephrine and 5-hydroxytryptamine concentrations in the rat brain and also significantly increased acetylcholine and dopamine (DA) levels in the rat striatum. Urinary isatin concentrations in patients with Parkinson’s disease tend to increase according to the severity of disease. We have developed a rat model of Parkinson’s disease induced by the Japanese encephalitis virus (JEV). The distribution of the pathological lesions of JEV-rats resemble those found in Parkinson’s disease. Significant behavioral improvement was observed in JEV-rats after isatin, L-DOPA and selegiline administration using a pole test. Both isatin and selegiline prevented the decrease in striatum DA levels of JEV-rats. The increased turnover of DA (DOPAC/DA) induced by JEV was significantly inhibited by isatin, but not selegiline. These findings suggest that JEV-infected rats may serve as a model of Parkinson’s disease and that exogenously administered isatin and selegiline can improve JEV-induced parkinsonism by increasing DA concentrations in the striatum.