RESUMEN
This paper was aimed to study the effects of Qizhiweitong particles (QZWT) on gastric motility in gastroparesis model rats, and to provide a theoretical and experimental basis for its clinical treatment. Rat gastroparesis model was established by bilateral injection of 6-hydroxydopamine into the substantia nigra in male Sprague-Dawley (SD) rats. The model rats received single gastric feeding of 1, 10, 30, 100, 200, 450, or 675 mg/kg QZWT or continuous administration of 675 mg/kg QZWT per day for 7 days. The gastric motility was measured by gastric emptying study and in vivo digital X-ray imaging system. The in vivo and ex vivo gastric longitudinal muscle contraction was recorded by PowerLab biological signal acquisition system. Gastric myoelectric signals were recorded by wireless implantable telemetry system. Protein expression levels of proinflammatory proteases in the myometrium were determined by Western blot. The results showed that the single administration of QZWT dose-dependently inhibited the contractile activity of isolated gastric strips from normal rats. The single administration of QZWT inhibited the in vivo contraction of gastric smooth muscle and gastric myoelectric signal in the control and model rats. The gastric emptying rate, in vivo and ex vivo gastric motility and gastric myoelectric signal in the model rats were significantly decreased compared with those in the control rats; While the continuous administration of QZWT markedly improved all the above indices of gastric motility function. The single administration of QZWT inhibited isolated gastric muscle strip contraction, and neither atropine nor nitric oxide synthase inhibitor pretreatments affected QZWT's inhibitory effects. The continuous administration of QZWT down-regulated the increased protein expression levels of nitric oxide synthase and cyclooxygenase 2 in the model group. These results suggest that, in clinical treatment, the single administration of QZWT may induce an analgesic effect by rapidly inhibiting gastric motility, while this effect is not related to acetylcholine or nitric oxide pathways. Long-term treatment with QZWT may ameliorate gastric motility through enhancing myoelectric activities, gastric smooth muscle contraction and gastric emptying, and this effect may partly be related to its anti-inflammatory effect.
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Medicamentos Herbarios Chinos , Gastroparesia , Femenino , Ratas , Masculino , Animales , Ratas Sprague-Dawley , Óxido Nítrico SintasaRESUMEN
Patients with Parkinson's disease (PD) often suffer from delayed gastric emptying, but the underlying mechanism remains unclear. We have shown previously that a PD rat model comprising bilateral substantia nigra destruction by 6-hydroxydopamine (6-OHDA rats) exhibits gastroparesis with alteration of neural nitric oxide synthase (nNOS) and acetylcholine in gastric corpus. However, changes in pyloric motility in the 6-OHDA rats have not been characterized. Solid gastric emptying test, immunofluorescence, Western blot, and in vitro pyloric motility recordings were used to assess pyloric motor function in the 6-OHDA rats. The 6-OHDA-treated rats displayed delayed solid gastric emptying and a lower basal pyloric motility index. In the 6-OHDA rats, high K+-induced transient contractions were weaker in pyloric sphincters. Electric field stimulation (EFS)-induced pyloric sphincter relaxation was lower in the 6-OHDA rats. NG-nitro-l-arginine methyl ester (l-NAME), a nonselective inhibitor of NOS, markedly inhibited the EFS-induced relaxation in both control and 6-OHDA rats. Pretreatment of tetrodotoxin abolished the effect of EFS on the pyloric motility. In addition, nNOS-positive neurons were extensively distributed in the pyloric myenteric plexus, whereas the number of nNOS-immunoreactive neurons and the protein expression of nNOS were significantly decreased in the pyloric muscularis of 6-OHDA rats. However, sodium nitroprusside-induced pyloric relaxations were similar between the control and 6-OHDA rats. These results indicate that the pyloric sphincters of 6-OHDA rats exhibit both weakened contraction and relaxation. The latter may be due to reduced nNOS in the pyloric myenteric plexus. The dysfunction of the pyloric sphincter might be involved in the delayed gastric emptying.NEW & NOTEWORTHY Reduced nitrergic neurons in pyloric myenteric plexus potently contributed to the attenuated relaxation in 6-hydroxydopamine (6-OHDA) rats, subsequently affecting gastric emptying. SNP could well improve the relaxation of pylori in 6-OHDA rats. The present study provides new insight into the diagnosis and treatment of delayed gastric emptying in patients with PD.
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Gastroparesia , Enfermedad de Parkinson , Animales , Gastroparesia/etiología , Humanos , Óxido Nítrico Sintasa de Tipo I/metabolismo , Oxidopamina , Píloro/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
In vivo administration of dopamine (DA) receptor (DR)-related drugs modulate gastric pepsinogen secretion. However, DRs on gastric pepsinogen-secreting chief cells and DA D2 receptor (D2R) on somatostatin-secreting D cells were subsequently acquired. In this study, we aimed to further investigate the local effect of DA on gastric pepsinogen secretion through DRs expressed on chief cells or potential D2Rs expressed on D cells. To elucidate the modulation of DRs in gastric pepsinogen secretion, immunofluorescence staining, ex vivo incubation of gastric mucosa isolated from normal and D2R-/- mice were conducted, accompanied by measurements of pepsinogen or somatostatin levels using biochemical assays or enzyme-linked immunosorbent assays. D1R, D2R, and D5R-immunoreactivity (IR) were observed on chief cells in mouse gastric mucosa. D2R-IR was widely distributed on D cells from the corpus to the antrum. Ex vivo incubation results showed that DA and the D1-like receptor agonist SKF38393 increased pepsinogen secretion, which was blocked by the D1-like receptor antagonist SCH23390. However, D2-like receptor agonist quinpirole also significantly increased pepsinogen secretion, and D2-like receptor antagonist sulpiride blocked the promotion of DA. Besides, D2-like receptors exerted an inhibitory effect on somatostatin secretion, in contrast to their effect on pepsinogen secretion. Furthermore, D2R-/- mice showed much lower basal pepsinogen secretion but significantly increased somatostatin release and an increased number of D cells in gastric mucosa. Only SKF38393, not quinpirole, increased pepsinogen secretion in D2R-/- mice. DA promotes gastric pepsinogen secretion directly through D1-like receptors on chief cells and indirectly through D2R-mediated suppression of somatostatin release.
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Células Principales Gástricas/efectos de los fármacos , Agonistas de Dopamina/farmacología , Pepsinógeno A/metabolismo , Quinpirol/farmacología , Receptores de Dopamina D2/agonistas , Células Secretoras de Somatostatina/efectos de los fármacos , Somatostatina/metabolismo , Animales , Células Principales Gástricas/metabolismo , Antagonistas de Dopamina/farmacología , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Dopamina D1/agonistas , Receptores de Dopamina D1/antagonistas & inhibidores , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/genética , Receptores de Dopamina D2/metabolismo , Vías Secretoras , Células Secretoras de Somatostatina/metabolismoRESUMEN
The cholinergic anti-inflammatory pathway (CAIP) has been proposed to regulate gastrointestinal inflammation via acetylcholine released from the vagus nerve activating α7 nicotinic receptor (α7nAChR) on macrophages. Parkinson's disease (PD) patients and PD rats with substantia nigra (SN) lesions exhibit gastroparesis and a decayed vagal pathway. To investigate whether activating α7nAChR could ameliorate inflammation and gastric dysmotility in PD rats, ELISA, western blot analysis, and real-time PCR were used to detect gastric inflammation. In vitro and in vivo gastric motility was investigated. Proinflammatory mediator levels and macrophage numbers were increased in the gastric muscularis of PD rats. α7nAChR was located on the gastric muscular macrophages of PD rats. The α7nAChR agonists PNU-282987 and GTS-21 decreased nuclear factor κB (NF-κB) activation and monocyte chemotactic protein-1 mRNA expression in the ex vivo gastric muscularis of PD rats, and these effects were abolished by an α7nAChR antagonist. After treatment with PNU-282987 in vivo, the PD rats showed decreased NF-κB activation, inflammatory mediator production, and contractile protein expression and improved gastric motility. The present study reveals that α7nAChR is involved in the development of gastroparesis in PD rats and provides novel insight for the treatment of gastric dysmotility in PD patients.
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Patients with Parkinson's disease (PD) have a higher incidence rate of duodenal ulcers. The mucus barrier provides the first line of defense for duodenal mucosal protection. However, it is unknown whether duodenal mucus secretion is affected in PD. In the present study, we used the rats microinjected 6-hydroxydopamine (6-OHDA) into the bilateral substantia nigra to investigate duodenal mucus secretion and potential therapeutic targets in duodenal ulcer in PD. Alcian blue-periodic acid-Schiff, transmission electron microscopy, immunofluorescence, duodenal mucosal incubation, and enzyme-linked immunosorbent assays were used. The 6-OHDA rats exhibited mucin accumulation and retention in duodenal goblet cells. Mucin granules were unable to fuse with the apical membranes of goblet cells, and the exocytosis ratio of goblet cells was significantly reduced. Moreover, decreased acetylcholine and increased muscarinic receptor 2 (M2R) levels were detected in the duodenal mucosa of 6-OHDA rats. Bilateral vagotomy rats were also characterized by defective duodenal mucus secretion and decreased acetylcholine with increased M2R levels in the duodenal mucosa. Application of the cholinomimetic drug carbachol or blocking M2R with methoctramine significantly promoted mucus secretion by goblet cells and increased MUC2 content in duodenal mucosa-incubated solutions from 6-OHDA and vagotomy rats. We conclude that the reduced acetylcholine and increased M2R contribute to the impaired duodenal mucus secretion of 6-OHDA rats. The study provides new insights into the mechanism of duodenal mucus secretion and potential therapeutic targets for the treatment of duodenal ulcers in PD patients.
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Acetilcolina/metabolismo , Mucosa Intestinal/metabolismo , Moco/metabolismo , Enfermedad de Parkinson Secundaria/metabolismo , Receptor Muscarínico M2/metabolismo , Animales , Duodeno/metabolismo , Masculino , Oxidopamina , Ratas , Ratas Sprague-DawleyRESUMEN
Constipation and defecatory dysfunctions are frequent symptoms in patients with Parkinson's disease (PD). The pathology of Lewy bodies in colonic and rectal cholinergic neurons suggests that cholinergic pathways are involved in colorectal dysmotility in PD. However, the underlying mechanism is unclear. The aim of the present study is to examine the effect of central dopaminergic denervation in rats, induced by injection 6-hydroxydopamine into the bilateral substania nigra (6-OHDA rats), on colorectal contractive activity, content of acetylcholine (ACh), vasoactive intestinal peptide (VIP) and expression of neural nitric oxide synthase (nNOS) and muscarinic receptor (MR). Strain gauge force transducers combined with electrical field stimulation (EFS), gut transit time, immunohistochemistry, ELISA, western blot and ultraperformance liquid chromatography tandem mass spectrometry were used in this study. The 6-OHDA rats exhibited outlet obstruction constipation characterized by prolonged transit time, enhanced contractive tension and fecal retention in colorectum. Pretreatment with tetrodotoxin significantly increased the colorectal motility. EFS-induced cholinergic contractions were diminished in the colorectum. Bethanechol chloride promoted colorectal motility in a dose-dependent manner, and much stronger reactivity of bethanechol chloride was observed in 6-OHDA rats. The ACh, VIP and protein expression of nNOS was decreased, but M2R and M3R were notably upregulated in colorectal muscularis externa. Moreover, the number of cholinergic neurons was reduced in sacral parasympathetic nucleus (SPN) of 6-OHDA rats. In conclusion, central nigrostriatal dopaminergic denervation is associated with decreased cholinergic neurons in SPN, decreased ACh, VIP content, and nNOS expression and upregulated M2R and M3R in colorectum, resulting in colorectal dysmotility, which contributes to outlet obstruction constipation. The study provides new insights into the mechanism of constipation and potential therapeutic targets for constipation in PD patients.
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Na+-K+-2Cl- cotransporter (NKCC) is expressed at exceptionally high levels in gastric parietal cells. Bumetanide, a potent loop diuretic that blocks NKCC, usually causes a decrease in gastric acid secretion. Endotoxaemia causes hypochlorhydria in vivo, in which lipopolysaccharide (LPS) plays an important role. This study aimed to investigate the effect of NKCC2 on gastric acid secretion and its alteration in LPS-treated mice. The scanning ion-selective electrode technique and real-time pH titration combined with RNA interference were used to determine the effects of bumetanide on gastric acid secretion. Immunochemistry and Western blotting were performed to investigate the changes in NKCC2 expression in LPS-treated mice. Immunoreactivity of NKCC1 and NKCC2 was mainly observed near the basolateral and apical membranes of parietal cells, respectively. Pretreatment with bumetanide reduced the histamine-stimulated H+ flux in the mouse gastric mucosa. The apical, but not the basolateral, addition of bumetanide inhibited forskolin- or histamine/3-isobutyl-1-methylxanthine(IBMX)-induced gastric acid secretion. In vivo treatment with NKCC2 siRNA inhibited forskolin-induced acid secretion. Upon histamine stimulation, the majority of NKCC2 was targeted to the apical membrane in the gastric mucosa and in primary cultured parietal cells. The expression of NKCC2 and vesicle-associated membrane protein-2 (VAMP2), but not that of H+/K+-ATPase, was decreased in the gastric mucosa of LPS-treated mice. Blocking apical NKCC2, but not basolateral NKCC1, by bumetanide inhibited secretagogue-induced gastric acid secretion, during which the membrane trafficking of NKCC2 may be necessary. The downregulation of NKCC2 and VAMP2 may be related to the reduced gastric acid secretion induced by LPS.
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Bumetanida/farmacología , Ácido Gástrico/metabolismo , Células Parietales Gástricas/efectos de los fármacos , Secretagogos/farmacología , Miembro 1 de la Familia de Transportadores de Soluto 12/metabolismo , Animales , Células Cultivadas , Regulación hacia Abajo/efectos de los fármacos , Femenino , Humanos , Lipopolisacáridos/farmacología , Masculino , Ratones Endogámicos C57BL , Células Parietales Gástricas/metabolismo , Ratas Sprague-Dawley , Proteína 2 de Membrana Asociada a Vesículas/metabolismoRESUMEN
Gastroparesis is a common symptom in Parkinson's disease (PD) and whether any change occurs in gastric smooth muscle cells (SMCs) of PD patients is unclear. We previously reported that rats with bilateral substantia nigra lesions induced by 6-hydroxydopamine (6-OHDA), referred to as 6-OHDA rats, manifest typical gastroparesis. In the present study, we further investigate the underlying mechanism. By means of an organ bath system and an implantable radiotelemetry system, both a weakened contractile force of gastric circular smooth muscle and gastric myoelectric activity were detected in the 6-OHDA rats and phasic and tonic contractions elicited by carbachol or high concentration of potassium were significantly reduced in gastric circular muscle strips. A thickened smooth muscle layer was observed under a light microscope and an ultrastructure of hypertrophic SMCs, with increased caveolae and decreased dense bodies, was observed under transmission electron microscope. Furthermore, the mRNA and protein expression levels of contractile markers (myosin light chain 20, myosin heavy chain 11 and α-smooth muscle actin) and the transcription factor serum response factor (SRF) were significantly decreased, while the TNFα and IL-1ß content was increased in the 6-OHDA rats. These results suggest that the decreased contractile force in 6-OHDA rats may be associated with the phenotypic abnormality observed in SMCs, which is due to downregulated contractile proteins induced by decreased SRF expression in the inflammatory muscular microenvironment.
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Gastroparesia/patología , Contracción Muscular , Miocitos del Músculo Liso/patología , Enfermedad de Parkinson/patología , Estómago/patología , Animales , Motilidad Gastrointestinal , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND AND PURPOSE: Dopamine protects the duodenal mucosa. Here we have investigated the source of dopamine in gastric juice and the mechanism underlying the effects of luminal dopamine on duodenal bicarbonate secretion (DBS) in rodents. EXPERIMENTAL APPROACH: Immunofluorescence, UPLC-MS/MS, gastric incubation and perfusion were used to detect gastric-derived dopamine. Immunofluorescence and RT-PCR were used to examine the expression of dopamine receptors in the duodenal mucosa. Real-time pH titration and pHi measurement were performed to investigate DBS. KEY RESULTS: H+ -K+ -ATPase was co-localized with tyrosine hydroxylase and dopamine transporters in gastric parietal cells. Dopamine was increased in in vivo gastric perfusate after intravenous infusion of histamine and in gastric mucosa incubated, in vitro, with bethanechol chloride or tyrosine. D2 receptors were the most abundant dopamine receptors in rat duodenum, mainly distributed on the apical membrane of epithelial cells. Luminal dopamine increased DBS in a concentration-dependent manner, an effect mimicked by a D2 receptor agonist quinpirole and inhibited by the D2 receptor antagonist L741,626, in vivo D2 receptor siRNA and in D2 receptor -/- mice. Dopamine and quinpirole raised the duodenal enterocyte pHi . Quinpirole-evoked DBS and PI3K/Akt activity were inhibited by calcium chelator BAPTA-AM or in D2 receptor-/- mice. CONCLUSION AND IMPLICATIONS: Dopamine in the gastric juice is derived from parietal cells and is secreted along with gastric acid. On arrival in the duodenal lumen, dopamine increased DBS via an apical D2 receptor- and calcium-dependent pathway. Our data provide novel insights into the protective effects of dopamine on the duodenal mucosa.
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Bicarbonatos , Dopamina , Animales , Cromatografía Liquida , Duodeno , Jugo Gástrico , Ratones , Fosfatidilinositol 3-Quinasas , Quinpirol/farmacología , Ratas , Receptores Dopaminérgicos , Receptores de Dopamina D1 , Espectrometría de Masas en TándemRESUMEN
Most Parkinson's Disease (PD) patients experience gastrointestinal (GI) dysfunction especially the gastroparesis, but its underlying mechanism is not clear. We have previously demonstrated that the neurons in the substantia nigra (SN) project to the lateral hypothalamic nucleus (LH) and the dorsal motor nucleus of vagus (DMV) receives the neural projection from LH by the means of anterograde and retrograde neural tracing technology. Orexin A (OXA) is predominately expressed in the LH. It has been reported that OXA can alter the gastric motility through the orexin receptor 1 (OX1R) in DMV. We speculated that this SN-LH-DMV pathway could modulate the motility of stomach because of the important role of LH and DMV in the regulation of gastric motility. However, the distribution and expression of dopamine receptors (DR) in the LH is unknown. In the present study, using a double-labeling immunofluorescence technique combined with confocal microscopy, we significantly extend our understanding of the SN-LH-DMV pathway by showing that (1) a considerable quantity of dopamine receptor 1 and 2 (D1 and D2) was expressed in the LH as well as the OX1R was expressed in the DMV; (2) Nearly all of the D1-immuoreactve (IR) neurons were also OXA-positive while only a few neurons express both D2 and OXA in the LH, and the DR-positive neurons were surrounded by the dopaminergic neural fibers; In the DMV, OX1R were colocalized with choline acetyltransferase (ChAT)-labeled motor neurons; (3) When the gastroparesis was induced by the destruction of dopaminergic neurons in the SN, the decreased expression of D1 and OXA was observed in the LH as well as the reduced OX1R and ChAT expression in the DMV. These findings suggest that SN might regulate the function of OXA-positive neurons via D1 receptor, which then affect the motor neurons in the DMV through OX1R. If the SN is damaged the vagal pathway would be affected, which may lead to gastric dysfunction. The present study raises the possibility that the SN-LH-DMV pathway can regulate the movement of stomach.
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Mosapride, a gastrointestinal prokinetic drug, is an agonist of 5-hydroxytryptamine (5-HT) receptor 4 that also reduces blood glucose. Whether 5-HT4 receptor is distributed in pancreatic islets and whether mosapride can directly stimulate insulin secretion is unclear. In the present study, the protein expression and cellular location of 5-HT4 receptor in pancreas was detected through western blotting and immunofluorescence. The acute effects of 5-HT4 receptor agonists, mosapride and prucalopride, on insulin secretion were investigated in vivo and in vitro in normal and alloxan-induced diabetes rats. The results indicated that 5-HT4 receptor immunoreactivity was co-existed in the islets insulin-immunoreactive cells of rat, mouse, pig and human. However the immunoreactive cells of insulin and 5-HT4 receptor and the protein expression of 5-HT4 receptor were significantly decreased in the pancreas of alloxan-induced diabetes rats. In normal rats, mosapride and prucalopride decreased blood glucose and increased insulin secretion during glucose tolerance test, in association with an increase in glucose-stimulated insulin secretion, which was abolished by the 5-HT4 receptor antagonist GR113808. In diabetes rats, mosapride and prucalopride failed to improve blood glucose and insulin levels in the group of 180mg/kg alloxan, but increased glucose-stimulated insulin secretion in the group of 120mg/kg alloxan in vitro. We conclude that 5-HT4 receptor is distributed in the islet ß cell. Activation of 5-HT4 receptor is able to stimulate insulin secretion directly, thereby reduce blood glucose. The study provides important experimental evidences for the 5-HT4 receptor regulating insulin secretion and acting as a potential drug target in diabetes treatment.
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Glucemia/metabolismo , Insulina/metabolismo , Islotes Pancreáticos/metabolismo , Receptores de Serotonina 5-HT4/metabolismo , Animales , Regulación de la Expresión Génica/efectos de los fármacos , Insulina/sangre , Secreción de Insulina , Islotes Pancreáticos/efectos de los fármacos , Masculino , Ratones , Terapia Molecular Dirigida , Transporte de Proteínas/efectos de los fármacos , Ratas , Agonistas del Receptor de Serotonina 5-HT4/farmacologíaRESUMEN
BACKGROUND: Parkinson's disease (PD) is characterized by dopaminergic neuron degeneration in the substantia nigra (SN) accompanied by pathology of the dorsal motor nucleus of the vagus (DMV). Gastroparesis is a common non-motor system symptom of PD in patients and in animal models. However, the underlying mechanism of this symptom is not clear. We previously reported on the expression of enhanced tyrosine hydroxylase (TH) and decreased choline acetyltransferase (ChAT) in the DMV of a PD animal model and colocalization of TH and ChAT with the dopamine receptors D1 and D2. We hypothesize that these receptors might contribute to the delayed gastric emptying observed in PD. OBJECTIVE: To investigate the distribution of D1 and D2 in gastric-projecting DMV neurons and alteration of their distribution in a PD rat model. METHODS: Retrograde tracing, double-labeling immunofluorescence techniques and western blotting were used. RESULTS: After injection of the retrograde tracer fluoro-gold (FG) into the gastric wall, FG-labeled gastric-projecting motoneurons were observed in the caudal and rostral parts of the DMV, and neurons with D1-, D2- and ChAT- immunoreactivity (IR) were widely colocalized in the DMV. Many TH-IR fibers were observed around the D1- and D2-IR neurons. Moreover, decreased D1 and enhanced D2 expression in the DMV was observed in 6-hydroxydopamine (6-OHDA) rats that were treated with a bilateral microinjection of 6-OHDA in the SN. CONCLUSIONS: The results indicate that dopamine receptors might affect the activity of gastric-projecting neurons in the DMV, their altered expression may contribute to the gastroparesis observed in PD.
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Neuronas/metabolismo , Enfermedad de Parkinson/metabolismo , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/metabolismo , Estómago/inervación , Nervio Vago/metabolismo , Animales , Colina O-Acetiltransferasa/metabolismo , Modelos Animales de Enfermedad , Dopamina/metabolismo , Masculino , Técnicas de Trazados de Vías Neuroanatómicas , Oxidopamina , Trastornos Parkinsonianos/inducido químicamente , Trastornos Parkinsonianos/metabolismo , Ratas , Ratas Sprague-Dawley , Tirosina 3-MonooxigenasaRESUMEN
AIM: To study the effects of entacapone, a catechol-O-methyltransferase inhibitor, on colon motility and electrolyte transport in Parkinson's disease (PD) rats. METHODS: Distribution and expression of catechol-O-methyltransferase (COMT) were measured by immunohistochemistry and Western blotting methods. The colonic smooth muscle motility was examined in vitro by means of a muscle motility recording device. The mucosal electrolyte transport of PD rats was examined by using a short-circuit current (ISC ) technique and scanning ion-selective electrode technique (SIET). Intracellular detection of cAMP and cGMP was accomplished by radioimmunoassay testing. RESULTS: COMT was expressed in the colons of both normal and PD rats, mainly on the apical membranes of villi and crypts in the colon. Compared to normal controls, PD rats expressed less COMT. The COMT inhibitor entacapone inhibited contraction of the PD rat longitudinal muscle in a dose-dependent manner. The ß2 adrenoceptor antagonist ICI-118,551 blocked this inhibitory effect by approximately 67% (P < 0.01). Entacapone increased mucosal ISC in the colon of rats with PD. This induction was significantly inhibited by apical application of Cl(-) channel blocker diphenylamine-2, 2'-dicarboxylic acid, basolateral application of Na(+)-K(+)-2Cl(-)co-transporter antagonist bumetanide, elimination of Cl(-) from the extracellular fluid, as well as pretreatment using adenylate cyclase inhibitor MDL12330A. As an inhibitor of prostaglandin synthetase, indomethacin can inhibit entacapone-induced ISC by 45% (P < 0.01). When SIET was applied to measure Cl(-) flux changes, this provided similar results. Entacapone significantly increased intracellular cAMP content in the colonic mucosa, which was greatly inhibited by indomethacin. CONCLUSION: COMT expression exists in rat colons. The ß2 adrenoceptor is involved in the entacapone-induced inhibition of colon motility. Entacapone induces cAMP-dependent Cl(-) secretion in the PD rat.
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Antiparkinsonianos/farmacología , Inhibidores de Catecol O-Metiltransferasa/farmacología , Catecoles/farmacología , Canales de Cloruro/efectos de los fármacos , Cloruros/metabolismo , Colon/efectos de los fármacos , Motilidad Gastrointestinal/efectos de los fármacos , Mucosa Intestinal/efectos de los fármacos , Nitrilos/farmacología , Trastornos Parkinsonianos/tratamiento farmacológico , Inhibidores de Adenilato Ciclasa/farmacología , Antagonistas de Receptores Adrenérgicos beta 2/farmacología , Animales , Catecol O-Metiltransferasa/metabolismo , Canales de Cloruro/metabolismo , Colon/metabolismo , Colon/fisiopatología , AMP Cíclico/metabolismo , Inhibidores de la Ciclooxigenasa/farmacología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Mucosa Intestinal/metabolismo , Transporte Iónico , Masculino , Oxidopamina , Trastornos Parkinsonianos/inducido químicamente , Trastornos Parkinsonianos/metabolismo , Trastornos Parkinsonianos/fisiopatología , Ratas Sprague-Dawley , Factores de TiempoRESUMEN
Parkinson's disease (PD) is a common neurodegenerative disorder that is often associated with weak tongue motility. However, the link between the degenerated dopaminergic neurons in the substantia nigra (SN) and lingual dysfunction remains unclear. In the present study, we investigated the localization of dopamine receptor 1 (D1) and dopamine receptor 2 (D2) and alternations in their expression in cholinergic motoneurons of the hypoglossal nucleus (HN) using double-label immunofluorescence, Western blotting and semi-quantitative reverse transcription and polymerase chain reaction (SqRT-PCR) in rats that received microinjections of 6-hydroxydopamine bilaterally into the SN (6-OHDA rats). The results revealed that a large population of choline acetyltransferase immunoreactive (ChAT-IR) neurons was distributed throughout HN and that almost all of the ChAT-IR motoneurons were also D1-IR and D2-IR. Several tyrosine hydroxylase (TH)-IR profiles were observed in a nonuniform pattern near the ChAT-IR, D1-IR or D2-IR somas, suggesting potent dopaminergic innervation. In the 6-OHDA rats, TH immunoreactivity in the SN was significantly decreased, but food residue was increased and treadmill occupancy time was shortened. In the HN, protein expression of TH and D2 was increased, whereas that of ChAT and D1 was decreased. A similar pattern was observed in mRNA levels. The present study suggests that dopamine may modulate the activity of cholinergic neurons via binding with D1 and D2 in the HN. Changes in the expression of ChAT, TH, D1 and D2 in the HN of 6-OHDA rats might be associated with the impaired tongue motility in PD. These findings should be further investigated.
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Neuronas Colinérgicas/metabolismo , Nervio Hipogloso/metabolismo , Neuronas Motoras/metabolismo , Enfermedad de Parkinson Secundaria/genética , Receptores de Dopamina D1/genética , Receptores de Dopamina D2/genética , Animales , Colina O-Acetiltransferasa/genética , Colina O-Acetiltransferasa/metabolismo , Neuronas Colinérgicas/patología , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Humanos , Nervio Hipogloso/patología , Masculino , Neuronas Motoras/patología , Oxidopamina , Enfermedad de Parkinson Secundaria/inducido químicamente , Enfermedad de Parkinson Secundaria/metabolismo , Enfermedad de Parkinson Secundaria/patología , Ratas , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/metabolismo , Transducción de Señal , Sustancia Negra/metabolismo , Sustancia Negra/patología , Lengua/inervación , Tirosina 3-Monooxigenasa/genética , Tirosina 3-Monooxigenasa/metabolismoRESUMEN
BACKGROUND: Parkinson's disease (PD) is a neurodegenerative disorder that is characterized by degeneration of dopaminergic neurons in the substantia nigra (SN). Destruction of the SN can lead to gastric dyskinesis accompanied by decreased expression of choline acetyltransferase (ChAT) and increased expression of tyrosine hydroxylase (TH) in the dorsal vagus complex (DVC), which includes the dorsal motor nucleus of the vagus (DMV) and nucleus tractus solitarius (NTS). However, it is unclear if the SN and DVC are directly connected. OBJECTIVE: To investigate the neural projection from the SN to the DVC in rats. METHODS: Retrograde and anterograde tracing techniques combined with double-labeling immunofluorescence technique were used. RESULTS: Destruction of the SN significantly decreases ChAT immunoreactivity (IR) and increases TH-IR in the DVC. After injection of the retrograde tracer fluoro-gold (FG) into the DVC, FG-labeled neurons were observed in the hypothalamic paraventricular nucleus (PVN), lateral hypothalamus (LH), inferior olive (IO), and locus coeruleus (LC). No FG-positive cells were observed in the SN or striatum. Furthermore, after injection of anterograde tracer biotinylated dextran amine (BDA) into the SN, BDA-positive fibers were observed in the caudate putamen (Cpu), globus pallidus (GP), LC, and LH but not in the DVC. CONCLUSION: This study is the first to demonstrate that neurons in the SN do not directly innervate the DVC in rats. The DVC might be indirectly innervated by the SN through the hypothalamus and/or the LC. These data provide important morphological insights into the potential mechanism underlying the gastroparesis observed in PD patients.
Asunto(s)
Bulbo Raquídeo/citología , Neuronas/citología , Sustancia Negra/citología , Animales , Encéfalo/citología , Colina O-Acetiltransferasa/metabolismo , Masculino , Red Nerviosa/citología , Vías Nerviosas/citología , Técnicas de Trazados de Vías Neuroanatómicas , Neuronas/enzimología , Ratas , Ratas Sprague-Dawley , Tirosina 3-Monooxigenasa/metabolismoRESUMEN
To study movement disorder in Parkinson's disease (PD), an animal model of PD can be created by injecting lipopolysaccharide (LPS) into the substantia nigra of rats. In addition to body movement disorders, patients with PD often experience gastrointestinal (GI) dysfunction, such as gastroparesis. However, the underlying mechanism of these disorders remains unclear. The dorsal motor nucleus of vagus (DMV) is a well-known visceral nucleus that regulates GI function. The present study investigated alterations in DMV neurons and gastric motility in rats with LPS-induced PD (LPS-PD rats). Gastric motility was recorded using a strain gauge force transducer in vivo. The distributions of tyrosine hydroxylase (TH)- and choline acetyltransferase (ChAT)-positive neurons in the DMV were determined using immunofluorescence and confocal laser microscopy. Our results indicated that in LPS-PD rats, the number of neurons in the substantia nigra, including neurons with TH immunoreactivity, was markedly reduced, although glial cell proliferation was clearly observed. However, enhanced TH immunoreactivity and decreased ChAT immunoreactivity were found in the DMV. Furthermore, weakened gastric motility was recorded in anesthetized LPS-PD rats. In conclusion, rats with LPS-induced PD exhibited gastric dysmotility with an alteration in DMV neurons. This PD model may be used to study autonomic nervous system disorders that are often observed in patients with early-stage PD.
Asunto(s)
Colina O-Acetiltransferasa/metabolismo , Modelos Animales de Enfermedad , Motilidad Gastrointestinal/fisiología , Enfermedad de Parkinson Secundaria/enzimología , Tirosina 3-Monooxigenasa/metabolismo , Nervio Vago/enzimología , Animales , Colina O-Acetiltransferasa/análisis , Ingestión de Alimentos/fisiología , Lipopolisacáridos/toxicidad , Masculino , Neuronas/química , Neuronas/enzimología , Enfermedad de Parkinson Secundaria/inducido químicamente , Enfermedad de Parkinson Secundaria/fisiopatología , Ratas , Ratas Sprague-Dawley , Tirosina 3-Monooxigenasa/análisis , Nervio Vago/químicaRESUMEN
The dorsal motor nucleus of vagus (DMV) plays an important role in the regulation of gastrointestinal function. Dopamine (DA) exerts potent neuromodulatory effects on the motoneurons in the DMV via dopamine receptors (DRs). However, the distribution of DRs and their neurochemical phenotypes in the DMV are unclear. In the present study, the distribution of DRs D1- and D2-immunoreactive (IR) neurons and their neurochemical phenotypes in the DMV of rats were investigated using a double-labeling immunofluorescence technique combined with confocal microscopy. The results indicated that a considerable quantity of D1 and D2 was expressed throughout the DMV. A large amount of choline acetyltransferase (ChAT)-IR and a few tyrosine hydroxylase (TH)-IR neurons were observed in the DMV. Nearly all of the neurons were also D1-IR and D2-IR. In conclusion, the present study demonstrates the wide distribution of D1 and D2 in the cholinergic and catecholaminergic neurons in the DMV of rats. The DRs might play an important role in the regulation of DA on the activity of cholinergic and catecholaminergic neurons in the DMV.
Asunto(s)
Neuronas/química , Neuronas/metabolismo , Receptores de Dopamina D1/biosíntesis , Receptores de Dopamina D2/biosíntesis , Nervio Vago/química , Nervio Vago/metabolismo , Animales , Inmunoquímica , Masculino , Ratas , Ratas Sprague-Dawley , Receptores de Dopamina D1/análisis , Receptores de Dopamina D2/análisisRESUMEN
Dopamine (DA) is synthesized in gastrointestinal epithelial cells and performs important regulatory effects on the duodenal mucosa. However, the underlying mechanism remains largely unknown. The present study investigated the effect of DA on the duodenal epithelial ion transport in rats by means of short-circuit current (ISC), real-time pH titration, enzyme-linked immunosorbent assay, and immunohistochemistry. The results indicate that basolateral, but not apical, application of DA induced a concentration-dependent ISC downward deflection with an apparent IC50 of 5.34 µmol/L. Basolateral application of dopaminergic receptor D1 (D1) antagonist, SCH-23390, inhibited DA-induced change in ISC (â³ISC) in a dose-dependent manner. D1 agonist, SKF38393, mimicked the effect of DA on the ISC. The clear immunoreactivity of D1 subtype D5 (D1b) was at the both apical and basolatoral sides of Brunner's glands and intestinal crypts. Basolateral pretreatment with adenylate cyclase inhibitor, MDL12330A, significantly inhibited DA- and forskolin-induced â³ISC. DA and SKF38393 increased the level of intracellular cyclic adenosine monophosphate (cAMP) from 1.55 ± 0.11 to 2.07 ± 0.11 and 5.91 ± 0.25 pmol/L·mg(-1), respectively. Furthermore, the serosal DA-induced â³ISC was remarkably inhibited by apical administration of K(+) channel blockers, Ba(2+) and tetraethylammonium, but not by Cl(-) channel blockers. Serosal DA and D1 agonist did not affect duodenal HCO3(-) secretion. In conclusion, the present results demonstrate that serosal DA is able to promote rat duodenal epithelial K(+) secretion, not HCO3(-) secretion through D1-mediated and cAMP-dependent pathway. The study provides a new insight in the modulation of DA on the ion transport of duodenal epithelia in rats.
Asunto(s)
Dopamina/metabolismo , Duodeno/metabolismo , Mucosa Intestinal/metabolismo , Receptores de Dopamina D1/fisiología , 2,3,4,5-Tetrahidro-7,8-dihidroxi-1-fenil-1H-3-benzazepina/farmacología , Animales , Benzazepinas/farmacología , Bicarbonatos/metabolismo , AMP Cíclico/metabolismo , Duodeno/efectos de los fármacos , Conductividad Eléctrica , Iminas/farmacología , Mucosa Intestinal/efectos de los fármacos , Transporte Iónico/efectos de los fármacos , Masculino , Técnicas de Placa-Clamp , Potasio/metabolismo , Ratas , Ratas Sprague-Dawley , Receptores de Dopamina D1/agonistas , Receptores de Dopamina D1/antagonistas & inhibidoresRESUMEN
Biology sequence comparison is a fundamental task in computational biology. According to the hydropathy profile of amino acids, a protein sequence is taken as a string with three letters. Three curves of the new protein sequence were defined to describe the protein sequence. A new method to analyze the similarity/dissimilarity of protein sequence was proposed based on the conditional probability of the protein sequence. Finally, the protein sequences of ND6 (NADH dehydrogenase subunit 6) protein of eight species were taken as an example to illustrate the new approach. The results demonstrated that the method is convenient and efficient.