Dopamine regulates colonic glial cell-derived neurotrophic factor secretion through cholinergic dependent and independent pathways.

BACKGROUND AND PURPOSE: Glial cell-derived neurotrophic factor (GDNF) maintains gut homeostasis. Dopamine promotes GDNF release in astrocytes. We investigated the regulation by dopamine of colonic GDNF secretion. EXPERIMENTAL APPROACH: D1 receptor knockout (D1 R-/- ) mice, adeno-associated viral 9-short hairpin RNA carrying D2 receptor (AAV9-shD2 R)-treated mice, 6-hydroxydopamine treated (6-OHDA) rats and primary enteric glial cells (EGCs) culture were used. Incubation fluid from colonic submucosal plexus and longitudinal muscle myenteric plexus were collected for GDNF and ACh measurements. KEY RESULTS: D2 receptor-immunoreactivity (IR), but not D1 receptor-IR, was observed on EGCs. Both D1 receptor-IR and D2 receptor-IR were co-localized on cholinergic neurons. Low concentrations of dopamine induced colonic GDNF secretion in a concentration-dependent manner, which was mimicked by the D1 receptor agonist SKF38393, inhibited by TTX and atropine and eliminated in D1 R-/- mice. SKF38393-induced colonic ACh release was absent in D1 R-/- mice. High concentrations of dopamine suppressed colonic GDNF secretion, which was mimicked by the D2 receptor agonist quinpirole, and absent in AAV-shD2 R-treated mice. Quinpirole decreased GDNF secretion by reducing intracellular Ca2+ levels in primary cultured EGCs. Carbachol ( ACh analogue) promoted the release of GDNF. Quinpirole inhibited colonic ACh release, which was eliminated in the AAV9-shD2 R-treated mice. 6-OHDA treated rats with low ACh and high dopamine content showed decreased GDNF content and increased mucosal permeability in the colon. CONCLUSION AND IMPLICATIONS: Low concentrations of dopamine promote colonic GDNF secretion via D1 receptors on cholinergic neurons, whereas high concentrations of dopamine inhibit GDNF secretion via D2 receptors on EGCs and/or cholinergic neurons.

Regional changes in brain apparent diffusion coefficient in fetuses with complex congenital heart disease and normal pregnancy assessed using diffusion-weighted imaging.

Objectives: To explore changes in brain apparent diffusion coefficient (ADC) in normal fetuses and fetuses with complex congenital heart disease (CHD) during the second and early third trimesters. Methods: This single-center prospective study was conducted from May 2019 through October 2021. We measured and compared the mean ADC values between 23 fetuses with CHD and 27 gestational age (GA)-matched controls using covariance analyses. ADC density plots and histograms were used to compare brain characteristics. False-discovery rates (FDR, α = 0.05) correction was used for multiple testing. Results: The mean ADC in the frontal white matter, temporal white matter, parietal white matter, occipital white matter, cerebellar hemisphere, central area of the centrum semiovale, basal ganglia region, thalamus, and pons were not significantly different (all p > 0.05). Based on histogram analysis, there were no significant differences between the controls and fetuses with CHD after FDR correction. However, the ADC density plots showed significant heterogeneity between the controls and fetuses with CHD. Conclusion: The mean ADC values and ADC histogram analysis did not differ between the CHD and normal groups. The ADC density plots may provide supplementary information and improve the sensitivity for detecting early brain changes in fetuses with CHD.

[Distribution Characteristics, Ecological Risks, and Source Identification of Heavy Metals in Cultivated Land Under Non-grain Production].

The purpose of this study was to explore the distribution characteristics and potential ecological risks of soil heavy metal pollution of cultivated land under non-grain production. Taking a typical area around Hangzhou Bay as an example, 254 topsoil samples (0-20 cm) of cultivated land were collected, and the content of eight soil heavy metals in four different cultivated land use types, including grain, seedlings, vegetables, and fruits, was analyzed. The ecological risk was assessed by the Nemerow pollution index and the potential ecological risk index, and the PMF model was used to identify the source of soil heavy metals in the study area. The results showed that the average contents of As, Cr, Cd, Cu, Hg, Ni, and Zn were all higher than the soil background value, except for Pb, but were lower than the national risk control standard for soil contamination of agricultural land. Non-grain production had a significant impact on the accumulation of heavy metals in soil. The content of heavy metals in nurseries and orchards was relatively high, followed by vegetable fields, and the lowest in grain fields. The Nemerow index showed that the cultivated land in the study area was in a light pollution level as a whole, and the single-factor pollution risks of Hg, Cd, and As were relatively high. The potential ecological risk levels of heavy metals in different cultivated land use types were:nurseries>orchards>vegetable fields>grain fields. The PMF results showed that the main sources of soil heavy metals in the study area were mixed sources of industrial emissions (36.8%), natural parent material sources (28.4%), atmospheric deposition sources (21.4%), and agricultural activity sources (13.4%). In conclusion, the increase in the application of chemical fertilizers and pesticides was the direct reason for the increase in soil heavy metal content caused by non-grain production of the cultivated land, whereas the industrial and mining emissions and atmospheric deposition accelerated the increase in soil heavy metal content in the study area.

Comparison of a deep learning-accelerated T2-weighted turbo spin echo sequence and its conventional counterpart for female pelvic MRI: reduced acquisition times and improved image quality.

OBJECTIVES: To investigate the feasibility of a deep learning-accelerated T2-weighted turbo spin echo (TSE) sequence (T2DL) applied to female pelvic MRI, using standard T2-weighted TSE (T2S) as reference. METHODS: In total, 24 volunteers and 48 consecutive patients with benign uterine diseases were enrolled. Patients in the menstrual phase were excluded. T2S and T2DL sequences in three planes were performed for each participant. Quantitative image evaluation was conducted by calculating the signal-to-noise ratio (SNR) and contrast-to-noise ratio (CNR). Image geometric distortion was evaluated by measuring the diameters in all three directions of the uterus and lesions. Qualitative image evaluation including overall image quality, artifacts, boundary sharpness of the uterine zonal layers, and lesion conspicuity were assessed by three radiologists using a 5-point Likert scale, with 5 indicating the best quality. Comparative analyses were conducted for the two sequences. RESULTS: T2DL resulted in a 62.7% timing reduction (1:54 min for T2DL and 5:06 min for T2S in axial, sagittal, and coronal imaging, respectively). Compared to T2S, T2DL had significantly higher SNR (p ≤ 0.001) and CNR (p ≤ 0.007), and without geometric distortion (p = 0.925-0.981). Inter-observer agreement regarding qualitative evaluation was excellent (Kendall's W > 0.75). T2DL provided superior image quality (all p < 0.001), boundary sharpness of the uterine zonal layers (all p < 0.001), lesion conspicuity (p = 0.002, p < 0.001, and p = 0.021), and fewer artifacts (all p < 0.001) in sagittal, axial, and coronal imaging. CONCLUSIONS: Compared with standard TSE, deep learning-accelerated T2-weighted TSE is feasible to reduce acquisition time of female pelvic MRI with significant improvement of image quality.

[Dual regulation of Qizhiweitong particles on gastric motility in the model rats with gastroparesis].

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.

[Methods for the evaluation of intestinal mucosal permeability].

The intestinal mucosal barrier (IMB), which consists of mechanical barrier, chemical barrier, biological barrier and immune barrier, plays an important role in the maintenance of intestinal epithelium integrity and defense against invasion of bacteria, endotoxins and foreign antigens. Impaired IMB, characterized by increased intestinal mucosal permeability (IMP) and decreased transmembrane resistance (TR), has been implicated in the pathogenesis of various digestive, urinary, circulatory, neurological and metabolic dysfunctions. Electrophysiological recording of TR in the ex vivo intestinal tissues or cultured epithelial cell monolayers, or biochemical quantification of transepithelial movement of orally-administered molecular probes or specific endogenous protein molecules has frequently been used in the evaluation of IMB. In this paper, the composition and function of IMB will be summarized, with emphasis on the evaluation methods of IMP.

Impaired nitrergic relaxation in pyloric sphincter of the 6-OHDA Parkinson's disease rat.

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.

Activation of dopamine D2 receptor promotes pepsinogen secretion by suppressing somatostatin release from the mouse gastric mucosa.

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.

Activation of α7nAChR Protects Against Gastric Inflammation and Dysmotility in Parkinson's Disease Rats.

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.

Feasibility study of simultaneous multislice diffusion kurtosis imaging with different acceleration factors in the liver.

BACKGROUND: Simultaneous multislice diffusion-weighted imaging (SMS-DWI) has been used to reduce image acquisition time. The purpose of this study was to investigate the feasibility of diffusion kurtosis imaging (DKI) based on the SMS technique in the liver and the influence of this method compared with that of conventional DWI sequences on image quality and DKI-derived quantitative parameters. METHODS: Forty volunteers underwent SMS-DWI sequences with acceleration factors of 2 and 3 (SMS2-DWI, SMS3-DWI) and conventional DWI (C-DWI) of the liver with three b-values (50, 800, 2000 s/mm2) in a 3T system. Qualitative image quality parameters and quantitative measurements of the signal-to-noise ratio (SNR), mean kurtosis (MK), mean apparent diffusivity (MD) and apparent diffusion coefficient (ADC) for the liver were compared between the three sequences. RESULTS: The scan times of C-DWI, SMS2-DWI, and SMS3-DWI were 4 min 11 s, 2 min 2 s, and 1 min 34 s, respectively. For all image quality parameters, there were no significant differences observed between C-DWI and SMS2-DWI (all p > 0.05) in the images with b-values of 800 and 2000 s/mm2. C-DWI and SMS2-DWI exhibited better scores than SMS3-DWI (all p < 0.01) in the images with b-values of 2000 s/mm2. In the images with b-values of 800 s/mm2, C-DWI and SMS2-DWI exhibited better scores than SMS3-DWI for artefacts and overall image quality (all p < 0.01), and C-DWI exhibited better scores than SMS3-DWI for the visibility of intrahepatic vessels (p < 0.001). There were no significant differences in the sharpness of the right lobe edge (p = 0.144), conspicuity of the left lobe (p = 0.370) or visibility of intrahepatic vessels (p = 0.109) between SMS2-DWI and SMS3-DWI. There were no significant differences in the sharpness of the right lobe edge (p = 0.066) or conspicuity of the left lobe (p = 0.131) between C-DWI and SMS3-DWI. For the b-value of 800 s/mm2, there were no statistically significant differences between SMS2-DWI and C-DWI (p = 1.000) or between SMS2-DWI and SMS3-DWI (p = 0.059), whereas SMS3-DWI had a significantly lower SNR than C-DWI (p = 0.024). For the DKI-derived parameters (MK and MD) and ADC values, there were no significant differences between the three sequences (MK, p = 0.606; MD, p = 0.831; ADC, p = 0.264). CONCLUSIONS: SMS-DWI with an acceleration factor of 2 is feasible for the liver, resulting in considerable reductions in scan time while maintaining similar image quality, comparable DKI parameters and ADC values compared with those of C-DWI.

Reduced acetylcholine and elevated muscarinic receptor 2 in duodenal mucosa contribute to the impairment of mucus secretion in 6-hydroxydopamine-induced Parkinson's disease rats.

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.

Scan Time Reduction in Intravoxel Incoherent Motion Diffusion-Weighted Imaging and Diffusion Kurtosis Imaging of the Abdominal Organs: Using a Simultaneous Multislice Technique With Different Acceleration Factors.

OBJECTIVE: To investigate the feasibility of quantitative intravoxel incoherent motion (IVIM) and diffusion kurtosis imaging (DKI) analyses in the upper abdominal organs by simultaneous multislice diffusion-weighted imaging (SMS-DWI). SUBJECTS AND METHODS: In this prospective study, a total of 32 participants underwent conventional DWI (C-DWI) and SMS-DWI sequences with acceleration factors of 2 and 3 (SMS2-DWI and SMS3-DWI, respectively) in the upper abdomen with multiple b-values (0, 10, 20, 50, 80, 100, 150, 200, 500, 800, 1000, 1500, and 2000 seconds/mm2) on a 3 T system (MAGNETOM Prisma; Siemens Healthcare, Erlangen, Germany). Image quality and quantitatively measurements of apparent diffusion coefficient (ADC), true diffusion coefficient (D), pseudodiffusion coefficient (D*), perfusion fraction (f), mean kurtosis (MK), and mean apparent diffusivity (MD) for the liver, pancreas, kidney cortex and medulla, spleen, and erector spine muscle were compared between the 3 sequences. RESULTS: The acquisition times for C-DWI, SMS2-DWI, and SMS3-DWI were 10 minutes 57 seconds, 5 minutes 9 seconds, and 3 minutes 54 seconds. For image quality parameters, C-DWI and SMS2-DWI yielded better results than SMS3-DWI (P < 0.05). SMS2-DWI had equivalent IVIM and DKI parameters compared with that of C-DWI (P > 0.05). No statistically significant differences in the ADC, D, f, and MD values between the 3 sequences (P > 0.05) were observed. The D* and MK values of the liver (P = 0.005 and P = 0.012) and pancreas (P = 0.019) between SMS3-DWI and C-DWI were significantly different. CONCLUSIONS: SMS2-DWI can substantially reduce the scan time while maintaining equivalent IVIM and DKI parameters in the abdominal organs compared with C-DWI.

Pancreatic acinar cells utilize tyrosine to synthesize L-dihydroxyphenylalanine.

The pancreatic ß cells can synthesize dopamine by taking L-dihydroxyphenylalanine, but whether pancreatic acinar cells synthesize dopamine has not been confirmed. By means of immunofluorescence, the tyrosine hydroxylase -immunoreactivity and aromatic amino acid decarboxylase (AADC)- immunoreactivity were respectively observed in pancreatic acinar cells and islet ß cells. Treatment with L-dihydroxyphenylalanine, not tyrosine, caused the production of dopamine in the incubation of INS-1 cells (rat islet ß cell line) and primary isolated islets, which was blocked by AADC inhibitor NSD-1015. However, only L-dihydroxyphenylalanine, but not dopamine, was detected when AR42J cells (rat pancreatic acinar cell line) were treated with tyrosine, which was blocked by tyrosine hydroxylase inhibitor AMPT. Dopamine was detected in the coculture of INS-1 cells with AR42J cells after treatment with tyrosine. In an in vivo study, pancreatic juice contained high levels of L-dihydroxyphenylalanine and dopamine. Both L-dihydroxyphenylalanine and dopamine accompanied with pancreatic enzymes and insulin in the pancreatic juice were all significantly increased after intraperitoneal injection of bethanechol chloride and their increases were all blocked by atropine. Inhibiting TH with AMPT blocked bethanechol chloride-induced increases in L-dihydroxyphenylalanine and dopamine, while inhibiting AADC with NSD-1015 only blocked the dopamine increase. Bilateral subdiaphragmatic vagotomy of rats leads to significant decreases of L-dihydroxyphenylalanine and dopamine in pancreatic juice. These results suggested that pancreatic acinar cells could utilize tyrosine to synthesize L-dihydroxyphenylalanine, not dopamine. Islet ß cells only used L-dihydroxyphenylalanine, not tyrosine, to synthesize dopamine. Both L-dihydroxyphenylalanine and dopamine were respectively released into the pancreatic duct, which was regulated by the vagal cholinergic pathway. The present study provides important evidences for the source of L-dihydroxyphenylalanine and dopamine in the pancreas.

Enhanced Contractive Tension and Upregulated Muscarinic Receptor 2/3 in Colorectum Contribute to Constipation in 6-Hydroxydopamine-Induced Parkinson's Disease Rats.

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.

[Source, metabolism and function of dopamine in digestive tract].

Dopamine (DA), as a catecholamine neurotransmitter widely distributed in the central nervous system and the peripheral tissues, has attracted a lot of attention. Especially in recent years, DA has been found to regulate the function of the immune system, and the involvement of DA in the intestinal mucosal inflammation-related diseases has become a hot research topic. The digestive tract is an important source of peripheral DA, and DA is not only produced in the enteric nervous system and gastrointestinal epithelium, but also produced by intestinal microorganisms. In addition to the synthetases of DA, the DA contents in body tissues are also affected by the two kinds of metabolic enzymes, monoamine oxidase (MAO) and catechol-O-methyltransferase (COMT). This article reviewed the sources, metabolism, and functions of DA in digestive tract, especially focusing on the distribution and function of MAO and COMT, the enzymes degrading DA.

Na+-K+-2Cl- cotransporter 2 located in the human and murine gastric mucosa is involved in secretagogue-induced gastric acid secretion and is downregulated in lipopolysaccharide-treated mice.

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.

Gastric smooth muscle cells manifest an abnormal phenotype in Parkinson's disease rats with gastric dysmotility.

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.

Source of dopamine in gastric juice and luminal dopamine-induced duodenal bicarbonate secretion via apical dopamine D2 receptors.

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.

Downregulated Dopamine Receptor 2 and Upregulated Corticotrophin Releasing Hormone in the Paraventricular Nucleus Are Correlated With Decreased Glucose Tolerance in Rats With Bilateral Substantia Nigra Lesions.

Patients with Parkinson's disease (PD) have a high prevalence of glucose metabolism abnormalities. However, the mechanism underlying these symptoms remains unclear. The hypothalamic-pituitary-adrenal (HPA) axis is the major neuroendocrine axis that regulates homeostasis in mammals, including glucose metabolism. Corticotrophin releasing hormone (CRH), which is synthesized in the paraventricular nucleus (PVN) of the hypothalamus, plays an important role in the regulation of blood glucose levels via the HPA axis. Our previous studies have reported that PVN neurons express numerous dopamine receptors (DRs) and accept direct projections from the substantia nigra (SN). We hypothesize that damage to dopaminergic neurons in the SN might influence the blood glucose level through the HPA system. Rats with bilateral SN lesions induced by 6-hydroxydopamine (6-OHDA) (referred to as 6-OHDA rats) were used to investigate alterations in the levels of blood glucose, CRH, and factors related to the HPA axis and to explore possible mechanisms. Blood glucose levels were detected at different time points after the glucose solution was intraperitoneally administered. CRH and DRs in the PVN were evaluated by immunofluorescence and western blot analysis. Adrenocorticotropic hormone (ACTH) in the pituitary and plasma corticosterone (CORT) was evaluated by radioimmunoassay (RIA). The results showed that 6-OHDA rats exhibited significantly decreased tyrosine hydroxylase (TH) in the SN and decreased glucose tolerance at 6 weeks, but not at 4 weeks. In the PVN, dopamine receptor 2 (D2) was expressed on CRH-positive neurons, and D2-positive neurons were surrounded by TH-positive fibers. Additionally, the expression of CRH was upregulated, whereas the expression of D2 and TH were downregulated in 6-OHDA rats compared with control rats. In D2 knock-out mice, the significantly enhanced expression of CRH and reduced expression of D2 were detected in the PVN. Furthermore, RIA revealed increased ACTH in the pituitary and elevated CORT in the blood. In summary, the present study suggests that the dopaminergic neurons in the SN are involved in the regulation of body glucose metabolism through CRH neurons that express D2 in the hypothalamic PVN. SN lesions decrease glucose tolerance mainly by downregulating D2 and upregulating CRH in the PVN through the HPA neuroendocrine system.