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The pig is an important translational model for studying intestinal physiology and disorders for its many homologies with humans, including the organization of the enteric nervous system (ENS), the major regulator of gastrointestinal functions. This study focused on the quantification and neurochemical characterization of substance P (SP) neurons in the pig ascending (AC) and descending colon (DC) in wholemount preparations of the inner submucosal plexus (ISP), outer submucosal plexus (OSP), and myenteric plexus (MP). We used antibodies for the pan-neuronal marker HuCD, and choline acetyltransferase (ChAT) and neuronal nitric oxide synthase (nNOS), markers for excitatory and inhibitory transmitters, for multiple labeling immunofluorescence and high-resolution confocal microscopy. The highest density of SP immunoreactive (IR) neurons was in the ISP (222/mm2 in the AC, 166/mm2 in the DC), where they make up about a third of HuCD-IR neurons, compared to the OSP and MP (19-22% and 13-17%, respectively, P < 0.001-0.0001). HuCD/SP/ChAT-IR neurons (up to 23%) were overall more abundant than HuCD/SP/nNOS-IR neurons (< 10%). Most SP-IR neurons contained ChAT-IR (62-85%), whereas 18-38% contained nNOS-IR with the highest peak in the OSP. A subpopulation of SP-IR neurons contains both ChAT- and nNOS-IR with the highest peak in the OSP and ISP of DC (33-36%) and the lowest in the ISP of AC (< 10%, P < 0.001). SP-IR varicose fibers were abundant in the ganglia. This study shows that SP-IR neurons are functionally distinct with variable proportions in different plexuses in the AC and DC reflecting diverse functions of specific colonic regions.
Assuntos
Plexo Mientérico , Plexo Submucoso , Humanos , Suínos , Animais , Substância P , Neurônios , Colo , Colina O-AcetiltransferaseRESUMO
The enteric nervous system (ENS) controls gastrointestinal functions. In large mammals' intestine, it comprises an inner (ISP) and outer (OSP) submucous plexus and a myenteric plexus (MP). This study quantifies enteric neurons in the ISP, OSP, and MP of the pig ascending (AC) and descending colon (DC) using the HuC/D, choline acetyltransferase (ChAT), and neuronal nitric oxide synthase (nNOS) neuronal markers in whole mount preparations with multiple labeling immunofluorescence. We established that the ISP contains the highest number of HuC/D neurons/mm2, which were more abundant in AC vs. DC, followed by OSP and MP with similar density in AC and DC. In the ISP, the density of ChAT immunoreactive (IR) neurons was very similar in AC and DC (31% and 35%), nNOS-IR neurons were less abundant in AC than DC (15% vs. 42%, P < 0.001), and ChAT/nNOS-IR neurons were 5% and 10%, respectively. In the OSP, 39-44% of neurons were ChAT-IR in AC and DC, while 45% and 38% were nNOS-IR and 10-12% were ChAT/nNOS-IR (AC vs. DC P < 0.05). In the MP, ChAT-IR neurons were 44% in AC and 54% in DC (P < 0.05), nNOS-IR neurons were 50% in both, and ChAT/nNOS-IR neurons were 12 and 18%, respectively. The ENS architecture with multilayered submucosal plexuses and the distribution of functionally distinct groups of neurons in the pig colon are similar to humans, supporting the suitability of the pig as a model and providing the platform for investigating the mechanisms underlying human colonic diseases.
Assuntos
Colina O-Acetiltransferase/imunologia , Colo/inervação , Sistema Nervoso Entérico/citologia , Plexo Mientérico/citologia , Neurônios/enzimologia , Óxido Nítrico Sintase/imunologia , Plexo Submucoso/citologia , Animais , Contagem de Células , Masculino , Suínos , Porco MiniaturaRESUMO
OBJECTIVES: Proteases are key mediators of pain and altered enteric neuronal signalling, although the types and sources of these important intestinal mediators are unknown. We hypothesised that intestinal epithelium is a major source of trypsin-like activity in patients with IBS and this activity signals to primary afferent and enteric nerves and induces visceral hypersensitivity. DESIGN: Trypsin-like activity was determined in tissues from patients with IBS and in supernatants of Caco-2 cells stimulated or not. These supernatants were also applied to cultures of primary afferents. mRNA isoforms of trypsin (PRSS1, 2 and 3) were detected by reverse transcription-PCR, and trypsin-3 protein expression was studied by western blot analysis and immunohistochemistry. Electrophysiological recordings and Ca2+ imaging in response to trypsin-3 were performed in mouse primary afferent and in human submucosal neurons, respectively. Visceromotor response to colorectal distension was recorded in mice administered intracolonically with trypsin-3. RESULTS: We showed that stimulated intestinal epithelial cells released trypsin-like activity specifically from the basolateral side. This activity was able to activate sensory neurons. In colons of patients with IBS, increased trypsin-like activity was associated with the epithelium. We identified that trypsin-3 was the only form of trypsin upregulated in stimulated intestinal epithelial cells and in tissues from patients with IBS. Trypsin-3 was able to signal to human submucosal enteric neurons and mouse sensory neurons, and to induce visceral hypersensitivity in vivo, all by a protease-activated receptor-2-dependent mechanism. CONCLUSIONS: In IBS, the intestinal epithelium produces and releases the active protease trypsin-3, which is able to signal to enteric neurons and to induce visceral hypersensitivity.
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Células Epiteliais/enzimologia , Mucosa Intestinal/enzimologia , Síndrome do Intestino Irritável/enzimologia , Síndrome do Intestino Irritável/genética , Tripsina/genética , Tripsina/metabolismo , Animais , Células CACO-2 , Estudos de Casos e Controles , Colo/enzimologia , Colo/inervação , Meios de Cultivo Condicionados/farmacologia , Dipeptídeos/farmacologia , Sistema Nervoso Entérico/citologia , Sistema Nervoso Entérico/diagnóstico por imagem , Sistema Nervoso Entérico/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Feminino , Gânglios Espinais/citologia , Humanos , Hipersensibilidade/enzimologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/patologia , Isoxazóis/farmacologia , Lipopolissacarídeos/farmacologia , Masculino , Camundongos , Microscopia Confocal , Neurônios Aferentes/efeitos dos fármacos , Neurônios Aferentes/fisiologia , Permeabilidade/efeitos dos fármacos , RNA Mensageiro/análise , Ratos , Receptor PAR-2/antagonistas & inibidores , Receptor PAR-2/metabolismo , Tripsina/farmacologia , Tripsinogênio/genética , Regulação para CimaRESUMO
The immediate and long-term effects of exposure to early life stress (ELS) have been documented in humans and animal models. Even relatively brief periods of stress during the first 10 days of life in rodents can impact later behavioral regulation and the vulnerability to develop adult pathologies, in particular an impairment of cognitive functions and neurogenesis, but also modified social, emotional, and conditioned fear responses. The development of preclinical models of ELS exposure allows the examination of mechanisms and testing of therapeutic approaches that are not possible in humans. Here, we describe limited bedding and nesting (LBN) procedures, with models that produce altered maternal behavior ranging from fragmentation of care to maltreatment of infants. The purpose of this paper is to discuss important issues related to the implementation of this chronic ELS procedure and to describe some of the most prominent endpoints and consequences, focusing on areas of convergence between laboratories. Effects on the hypothalamic-pituitary adrenal (HPA) axis, gut axis and metabolism are presented in addition to changes in cognitive and emotional functions. Interestingly, recent data have suggested a strong sex difference in some of the reported consequences of the LBN paradigm, with females being more resilient in general than males. As both the chronic and intermittent variants of the LBN procedure have profound consequences on the offspring with minimal external intervention from the investigator, this model is advantageous ecologically and has a large translational potential. In addition to the direct effect of ELS on neurodevelopmental outcomes, exposure to adverse early environments can also have intergenerational impacts on mental health and function in subsequent generation offspring. Thus, advancing our understanding of the effect of ELS on brain and behavioral development is of critical concern for the health and wellbeing of both the current population, and for generations to come.
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Maus-Tratos Infantis , Cognição , Emoções , Comportamento Materno , Comportamento de Nidação , Estresse Psicológico/psicologia , Tecido Adiposo Branco/metabolismo , Animais , Animais Recém-Nascidos , Roupas de Cama, Mesa e Banho , Comportamento Animal , Epigênese Genética , Feminino , Humanos , Sistema Hipotálamo-Hipofisário/metabolismo , Recém-Nascido , Masculino , Modelos Animais , Neurogênese , Sistema Hipófise-Suprarrenal/metabolismo , Reprodutibilidade dos Testes , Resiliência Psicológica , Roedores , Fatores Sexuais , Estresse Psicológico/metabolismoRESUMO
BACKGROUND & AIMS: Diet has major effects on the intestinal microbiota, but the exact mechanisms that alter complex microbial communities have been difficult to elucidate. In addition to the direct influence that diet exerts on microbes, changes in microbiota composition and function can alter host functions such as gastrointestinal (GI) transit time, which in turn can further affect the microbiota. METHODS: We investigated the relationships among diet, GI motility, and the intestinal microbiota using mice that are germ-free (GF) or humanized (ex-GF mice colonized with human fecal microbiota). RESULTS: Analysis of gut motility revealed that humanized mice fed a standard polysaccharide-rich diet had faster GI transit and increased colonic contractility compared with GF mice. Humanized mice with faster transit due to administration of polyethylene glycol or a nonfermentable cellulose-based diet had similar changes in gut microbiota composition, indicating that diet can modify GI transit, which then affects the composition of the microbial community. However, altered transit in mice fed a diet of fermentable fructooligosaccharide indicates that diet can change gut microbial function, which can affect GI transit. CONCLUSIONS: Based on studies in humanized mice, diet can affect GI transit through microbiota-dependent or microbiota-independent pathways, depending on the type of dietary change. The effect of the microbiota on transit largely depends on the amount and type (fermentable vs nonfermentable) of polysaccharides present in the diet. These results have implications for disorders that affect GI transit and gut microbial communities, including irritable bowel syndrome and inflammatory bowel disease.
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Bactérias/genética , DNA Bacteriano/análise , Dieta , Metabolismo Energético , Trato Gastrointestinal/microbiologia , Trânsito Gastrointestinal/fisiologia , Vida Livre de Germes , Metagenoma , Animais , Trato Gastrointestinal/metabolismo , CamundongosRESUMO
Thyrotropin-releasing hormone (TRH) acts centrally to exert pleiotropic actions independently from its endocrine function, including antinociceptive effects against somatic pain in rodents. Whether exogenous or endogenous activation of TRH signaling in the brain modulates visceral pain is unknown. Adult male Sprague-Dawley rats received an intracerebroventricular (ICV) injection of the stable TRH analog, RX-77368 (10, 30 and 100â¯ng/rat) or saline (5⯵l) or were semi-restrained and exposed to cold (4°C) for 45â¯min. The visceromotor response (VMR) to graded phasic colorectal distensions (CRD) was monitored using non-invasive intracolonic pressure manometry. Naloxone (1â¯mg/kg) was injected subcutaneously 10â¯min before ICV RX-77368 or saline. Fecal pellet output was monitored for 1â¯h after ICV injection. RX-77368 ICV (10, 30 and 100â¯ng/rat) reduced significantly the VMR by 56.7%, 67.1% and 81.1% at 40â¯mmHg and by 30.3%, 58.9% and 87.4% at 60â¯mmHg respectively vs ICV saline. Naloxone reduced RX-77368 (30 and 100â¯ng, ICV) analgesic response by 51% and 28% at 40â¯mmHg and by 30% and 33% at 60â¯mmHg respectively, but had no effect per se. The visceral analgesia was mimicked by the acute exposure to cold. At the doses of 30 and 100â¯ng, ICV RX-77368 induced defecation within 30â¯min. These data established the antinociceptive action of RX-77368 injected ICV in a model of visceral pain induced by colonic distension through recruitment of both opioid and non-opioid dependent mechanisms.
Assuntos
Neoplasias Colorretais , Ácido Pirrolidonocarboxílico/análogos & derivados , Hormônio Liberador de Tireotropina/análogos & derivados , Dor Visceral , Ratos , Masculino , Animais , Ratos Sprague-Dawley , Dor Visceral/tratamento farmacológico , Analgésicos/farmacologia , Naloxona/farmacologiaRESUMO
Helicobacter pylori is a highly prevalent human gastric pathogen that causes gastritis, ulcer disease, and gastric cancer. It is not yet fully understood how H. pylori injures the gastric epithelium. The Na,K-ATPase, an essential transporter found in virtually all mammalian cells, has been shown to be important for maintaining the barrier function of lung and kidney epithelia. H. pylori decreases levels of Na,K-ATPase in the plasma membrane of gastric epithelial cells, and the aim of this study was to demonstrate that this reduction led to gastric injury by impairing the epithelial barrier. Similar to H. pylori infection, the inhibition of Na,K-ATPase with ouabain decreased transepithelial electrical resistance and increased paracellular permeability in cell monolayers of human gastric cultured cells, 2D human gastric organoids, and gastric epithelium isolated from gerbils. Similar effects were caused by a partial shRNA silencing of Na,K-ATPase in human gastric organoids. Both H. pylori infection and ouabain exposure disrupted organization of adherens junctions in human gastric epithelia as demonstrated by E-cadherin immunofluorescence. Functional and structural impairment of epithelial integrity with a decrease in Na,K-ATPase amount or activity provides evidence that the H. pylori-induced downregulation of Na,K-ATPase plays a role in the complex mechanism of gastric disease induced by the bacteria.
Assuntos
Mucosa Gástrica , Infecções por Helicobacter , Helicobacter pylori , Ouabaína , ATPase Trocadora de Sódio-Potássio , ATPase Trocadora de Sódio-Potássio/metabolismo , ATPase Trocadora de Sódio-Potássio/genética , Humanos , Animais , Ouabaína/farmacologia , Infecções por Helicobacter/metabolismo , Infecções por Helicobacter/microbiologia , Infecções por Helicobacter/patologia , Mucosa Gástrica/metabolismo , Mucosa Gástrica/microbiologia , Mucosa Gástrica/patologia , Mucosa Gástrica/efeitos dos fármacos , Gerbillinae , Membrana Celular/metabolismo , Membrana Celular/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/microbiologia , Células Epiteliais/efeitos dos fármacos , Organoides/metabolismo , Organoides/microbiologiaRESUMO
Introduction: The central and peripheral nervous systems provide cholinergic innervation in the colon. The ability to assess their neuroanatomical distinctions is still a challenge. The pig is regarded as a relevant translational model due to the close similarity of its enteric nervous system (ENS) with that of human. Opioid-induced constipation is one of the most common side effects of opioid therapy. Methods: We developed an approach to differentiate the central and peripheral cholinergic innervation of the pig colon using double immunolabeling with a novel mouse anti-human peripheral type of choline acetyltransferase (hpChAT) antibody combined with a rabbit anti-common type of ChAT (cChAT) antibody, a reliable marker of cholinergic neurons in the central nervous system. We examined their spatial configurations in 3D images of the ENS generated from CLARITY-cleared colonic segments. The density was quantitated computationally using Imaris 9.7. We assessed changes in the distal colon induced by daily oral treatment for 4 weeks with the µ opioid receptor agonist, loperamide (0.4 or 3 mg/kg). Results: The double labeling showed strong cChAT immunoreactive (ir) fibers in the cervical vagus nerve and neuronal somata and fibers in the ventral horn of the sacral (S2) cord while hpChAT immunoreactivity was visualized only in the ENS but not in the vagus or sacral neural structures indicating the selectivity of these two antibodies. In the colonic myenteric plexus, dense hpChAT-ir neurons and fibers and varicose cChAT-ir fibers surrounding hpChAT-ir neurons were simultaneously visualized in 3D. The density of cChAT-ir varicose fibers in the outer submucosal plexus of both males and females were higher in the transverse and distal colon than in the proximal colon and in the myenteric plexus compared to the outer submucosal plexus and there was no cChAT innervation in the inner submucosal plexus. The density of hpChAT in the ENS showed no segmental or plexus differences in both sexes. Loperamide at the highest dose significantly decreased the density hpChAT-ir fibers + somata in the myenteric plexus of the distal colon. Discussion: These data showed the distinct density of central cholinergic innervation between myenteric and submucosal plexuses among colonic segments and the localization of cChAT-ir fibers around peripheral hpChAT neurons in 3D. The reduction of cholinergic myenteric innervation by chronic opiate treatment points to target altered prokinetic cholinergic pathway to counteract opiate constipation.
RESUMO
The porcine gut is increasingly regarded as a useful translational model. The enteric nervous system in the colon coordinates diverse functions. However, knowledge of the molecular profiling of porcine enteric nerve system and its similarity to that of human is still lacking. We identified the distinct transcriptional programs associated with functional characteristics between inner submucosal and myenteric ganglia in porcine proximal and distal colon using bulk RNA and single-cell RNA sequencing. Comparative transcriptomics of myenteric ganglia in corresponding colonic regions of pig and human revealed highly conserved programs in porcine proximal and distal colon, which explained >96% of their transcriptomic responses to vagal nerve stimulation, suggesting that porcine proximal and distal colon could serve as predictors in translational studies. The conserved programs specific for inflammatory modulation were displayed in pigs with vagal nerve stimulation. This study provides a valuable transcriptomic resource for understanding of human colonic functions and neuromodulation using porcine model.
Assuntos
Sistema Nervoso Entérico , Transcriptoma , Humanos , Animais , Suínos , Colo/inervaçãoRESUMO
Monitoring of colon activity is currently limited to tethered systems like anorectal manometry. These systems have significant drawbacks, but fundamentally limit the observation time of colon activity, reducing the likelihood of detecting specific clinical events. While significant technological advancement has been directed to mobile sensor capsules, this work describes the development and feasibility of a stationary sensor for describing the coordinated activity between neighboring segments of the colon. Unlike wireless capsules, this device remains in position and measures propagating pressure waves and impedances between colon segments to describe activity and motility. This low-power, flexible, wireless sensor-the colon monitor to capture activity (ColoMOCA) was validated in situ and in vivo over seven days of implantation. The ColoMOCA diameter was similar to common endoscopes to allow for minimally invasive diagnostic placement. The ColoMOCA included two pressure sensors, and three impedance-sensing electrodes arranged to describe the differential pressures and motility between adjacent colon segments. To prevent damage after placement in the colon, the ColoMOCA was fabricated with a flexible polyimide circuit board and a silicone rubber housing. The resulting device was highly flexible and suitable for surgical attachment to the colon wall. In vivo testing performed in eleven animals demonstrated suitability of both short term (less than 3 hours) and 7-day implantations. Data collected wirelessly from animal experiments demonstrated the ColoMOCA described colon activity similarly to wired catheters and allowed untethered, conscious monitoring of organ behavior.
Assuntos
Colo , Próteses e Implantes , Animais , Eletrodos , Impedância Elétrica , CatéteresRESUMO
BACKGROUND & AIMS: Corticotropin-releasing factor receptor-1 (CRF(1)) mediates the stress-induced colonic motor activity. Less is known about the role of CRF(2) in the colonic response to stress. METHODS: We studied colonic contractile activity in rats and CRF(2)-/-, CRF-overexpressing, and wild-type mice using still manometry; we analyzed defecation induced by acute partial-restraint stress (PRS), and/or intraperitoneal injection of CRF ligands. In rats, we monitored activation of the colonic longitudinal muscle myenteric plexus (LMMP) neurons and localization of CRF(1) and CRF(2) using immunohistochemical and immunoblot analyses. We measured phosphorylation of extracellular signal-regulated kinase 1/2 by CRF ligands in primary cultures of LMMP neurons (PC-LMMPn) and cyclic adenosine monophosphate (cAMP) production in human embryonic kidney-293 cells transfected with CRF(1) and/or CRF(2). RESULTS: In rats, a selective agonist of CRF(2) (urocortin 2) reduced CRF-induced defecation (>50%), colonic contractile activity, and Fos expression in the colonic LMMP. A selective antagonist of CRF(2) (astressin(2)-B) increased these responses. Urocortin 2 reduced PRS-induced colonic contractile activity in wild-type and CRF-overexpressing mice, whereas disruption of CRF(2) increased PRS-induced colonic contractile activity and CRF-induced defecation. CRF(2) colocalized with CRF(1) and neuronal nitric oxide synthase in the rat colon, LMMP, and PC-LMMPn. CRF-induced phosphorylation of extracellular signal-regulated kinase in PC-LMMPn; this was inhibited or increased by a selective antagonist of CRF(1) (NBI35965) or astressin(2)-B, respectively. The half maximal effective concentration, EC(50), for the CRF-induced cAMP response was 8.6 nmol/L in human embryonic kidney-293 cells that express only CRF(1); this response was suppressed 10-fold in cells that express CRF(1) and CRF(2). CONCLUSIONS: In colon tissues of rodents, CRF(2) activation inhibits CRF(1) signaling in myenteric neurons and the stress-induced colonic motor responses. Disruption of CRF(2) function impairs colonic coping responses to stress.
Assuntos
Colo/fisiopatologia , Motilidade Gastrointestinal/fisiologia , Plexo Mientérico/metabolismo , Receptores de Hormônio Liberador da Corticotropina/metabolismo , Estresse Fisiológico , Doença Aguda , Animais , Colo/metabolismo , Colo/patologia , Modelos Animais de Doenças , Feminino , Motilidade Gastrointestinal/efeitos dos fármacos , Humanos , Imuno-Histoquímica , Injeções Intraperitoneais , Masculino , Camundongos , Camundongos Endogâmicos , Plexo Mientérico/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Receptores de Hormônio Liberador da Corticotropina/efeitos dos fármacos , Urocortinas/administração & dosagemRESUMO
Electrical stimulation has been demonstrated as an alternative approach to alleviate intractable colonic motor disorders, whose effectiveness can be evaluated through colonic motility assessment. Various methods have been proposed to monitor the colonic motility and while each has contributed towards better understanding of colon motility, a significant limitation has been the spatial and temporal low-resolution colon motility data acquisition and analysis. This paper presents the study of employing bio-impedance characterization to monitor colonic motor activity. Direct distal colon stimulation was undertaken in anesthetized pigs to validate the bio-impedance scheme simultaneous with luminal manometry monitoring. The results indicated that the significant decreases of bio-impedance corresponded to strong colonic contraction in response to the electrical stimulation in the distal colon. The magnitude/power of the dominant frequencies of phasic colonic contractions identified at baseline (in the range 2-3 cycles per minute (cpm)) were increased after the stimulation. In addition, positive correlations have been found between bio-impedance and manometry. The proposed bio-impedance-based method can be a viable candidate for monitoring colonic motor pattern with high spatial and temporal resolution. The presented technique can be integrated into a closed-loop therapeutic device in order to optimize its stimulation protocol in real-time.
Assuntos
Doenças do Colo , Motilidade Gastrointestinal , Animais , Colo/fisiologia , Impedância Elétrica , Motilidade Gastrointestinal/fisiologia , Manometria/métodos , SuínosRESUMO
Peptide CRF antagonists injected peripherally alleviate stress-induced visceral hypersensitivity (SIVH) to colorectal distension (CRD) in rodents. Here we further evaluated the dose and time-dependent inhibitory activity of several long-acting peptide CRF receptor antagonists related to astressin on SIVH, focusing on astressin C (AstC), which previously showed high efficacy on stress-related alterations of HPA axis and gut secretomotor functions. Male and female Sprague-Dawley rats pretreated subcutaneously (SC) with AstC were injected intraperitoneally (IP) with CRF 15 min later. The visceromotor responses (VMR) to graded phasic CRD (10, 20, 40 and 60 mmHg) were monitored at basal, 15 min and up to 1-8 days after pretreatment. Two other astressin analogs, hexanoyl-astressin D (Hex-AstD) and [CαMeVal19,32]-AstC, were also tested. The response to IP CRF was sex-dependent with female rats requiring a higher dose to exhibit visceral hyperalgesia. Pretreatment with AstC (30-1000 µg/kg) resulted in a dose-related inhibition of IP CRF-induced SIVH and diarrhea in both sexes. The highest dose prevented SIVH and diarrhea up to 5-7 days after a single SC injection and was lost on day 7 (females) and day 8 (males) but reinstated after a second injection of AstC on day 8 or 9 respectively. [CαMeVal19,32]-AstC and Hex-AstD (1000 µg/kg in males) also prevented SIVH. These data show the potent long-lasting anti-hyperalgesic effect of AstC in an acute model of SIVH in both male and female rats. This highlights the potential of long-acting peripheral CRF antagonists to treat stress-sensitive irritable bowel syndrome.
Assuntos
Dor Visceral , Animais , Hormônio Liberador da Corticotropina/metabolismo , Diarreia , Feminino , Hiperalgesia/tratamento farmacológico , Sistema Hipotálamo-Hipofisário/metabolismo , Masculino , Fragmentos de Peptídeos , Sistema Hipófise-Suprarrenal/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Hormônio Liberador da Corticotropina/metabolismo , Estresse Psicológico/tratamento farmacológico , Dor Visceral/tratamento farmacológicoRESUMO
Peptide YY (PYY) antisecretory effect on intestinal epithelia is well established, whereas less is known about its actions to influence colonic motility in conscious animals. We characterized changes in basal function and stimulated colonic motor function induced by PYY-related peptides in conscious mice. PYY(3-36), PYY, and neuropeptide Y (NPY) (8 nmol/kg) injected intraperitoneally inhibited fecal pellet output (FPO) per hour during novel environment stress by 90%, 63%, and 57%, respectively, whereas the Y(1)-preferring agonists, [Pro(34)]PYY and [Leu(31),Pro(34)]NPY, had no effect. Corticotrophin-releasing factor 2 receptor antagonist did not alter PYY(3-36) inhibitory action. PYY and PYY(3-36) significantly reduced restraint-stimulated defecation, and PYY(3-36) inhibited high-amplitude distal colonic contractions in restrained conscious mice for 1 h, by intraluminal pressure with the use of a microtransducer. PYY suppression of intraperitoneal 5-hydroxytryptophan induced FPO and diarrhea was blocked by the Y(2) antagonist, BIIE0246, injected intraperitoneally and mimicked by PYY(3-36), but not [Leu(31),Pro(34)]NPY. PYY(3-36) also inhibited bethanechol-stimulated FPO and diarrhea. PYY(3-36) inhibited basal FPO during nocturnal feeding period and light phase in fasted/refed mice for 2-3 h, whereas the reduction of food intake lasted for only 1 h. PYY(3-36) delayed gastric emptying after fasting-refeeding by 48% and distal colonic transit time by 104%, whereas [Leu(31),Pro(34)]NPY had no effect. In the proximal and distal colon, higher Y(2) mRNA expression was detected in the mucosa than in muscle layers, and Y(2) immunoreactivity was located in nerve terminals around myenteric neurons. These data established that PYY/PYY(3-36) potently inhibits basal and stress/serotonin/cholinergic-stimulated propulsive colonic motor function in conscious mice, likely via Y(2) receptors.
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Colo/fisiologia , Diarreia/fisiopatologia , Motilidade Gastrointestinal/fisiologia , Peptídeo YY/metabolismo , Receptores de Neuropeptídeo Y/metabolismo , 5-Hidroxitriptofano/metabolismo , 5-Hidroxitriptofano/farmacologia , Doença Aguda , Animais , Arginina/análogos & derivados , Arginina/farmacologia , Benzazepinas/farmacologia , Betanecol/farmacologia , Colo/efeitos dos fármacos , Colo/inervação , Estado de Consciência , Diarreia/induzido quimicamente , Diarreia/tratamento farmacológico , Ingestão de Alimentos/efeitos dos fármacos , Ingestão de Alimentos/fisiologia , Vias Eferentes/efeitos dos fármacos , Vias Eferentes/fisiologia , Esvaziamento Gástrico/efeitos dos fármacos , Esvaziamento Gástrico/fisiologia , Motilidade Gastrointestinal/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Plexo Mientérico/efeitos dos fármacos , Plexo Mientérico/fisiologia , Neuropeptídeo Y/análogos & derivados , Neuropeptídeo Y/antagonistas & inibidores , Neuropeptídeo Y/metabolismo , Neuropeptídeo Y/farmacologia , Parassimpatomiméticos/farmacologia , Fragmentos de Peptídeos , Peptídeo YY/farmacologia , Restrição Física , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/fisiologiaRESUMO
The interaction between gut inflammatory processes and stress is gaining increasing recognition. Corticotropin-releasing factor (CRF)-receptor activation in the brain is well established as a key signaling pathway initiating the various components of the stress response including in the viscera. In addition, a local CRF signaling system has been recently established in the gut. This review summarize the present knowledge on mechanisms through which both brain and gut CRF receptors modulate intestinal inflammatory processes and its relevance towards increased inflammatory bowel disease (IBD) activity and post-infectious irritable bowel syndrome (IBS) susceptibility induced by stress.
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Inflamação/patologia , Doenças Inflamatórias Intestinais/patologia , Síndrome do Intestino Irritável/patologia , Receptores de Hormônio Liberador da Corticotropina/fisiologia , Estresse Psicológico/patologia , Animais , Humanos , Transdução de Sinais/fisiologiaRESUMO
Visceral pain modulation by chronic stress in mice has been little studied. Electromyography (EMG) recording of abdominal muscle contractions, as a proxy to the visceromotor response (VMR), requires electrode implantation and post-surgical single housing (SH) which could affect the VMR to stress. To test this hypothesis, male mice had electrode implantation surgery (S) plus SH, or no surgery and were group housed (NS-GH) or single housed (NS-SH) and exposed to either water avoidance stress (WAS, 1 h/day) or left undisturbed in their home cages for 10 days. The VMR to phasic ascending colorectal distension (CRD) was assessed before (basal) and 24 h after 10 days of WAS or no stress using a surgery-free method of intraluminal colonic pressure (ICP) recording (solid-state manometry). WAS heightened significantly the VMR to CRD at 30, 45, and 60 mmHg in S-SH vs. NS-GH, but not compared to NS-SH conscious mice. Compared to basal CRD, WAS increased VMR at 60 mmHg in the S-SH group and decreased it at 30-60 mmHg in NS-GH mice, while having no effect in NS-SH mice. The average defecation during the hour of repeated WAS over 10 days was 1.9 and 2.4 fold greater in S-SH vs. NS-GH and NS-SH mice, respectively. These data indicate that the combination of S-SH required for VMR monitoring with EMG is an important component of repeated WAS-induced post-stress visceral hypersensitivity and defecation in mice.
Assuntos
Músculos Abdominais/fisiologia , Isolamento Social , Estresse Psicológico/fisiopatologia , Músculos Abdominais/efeitos dos fármacos , Músculos Abdominais/cirurgia , Animais , Atropina/farmacologia , Buprenorfina/farmacologia , Colo/fisiologia , Defecação , Dilatação Patológica/fisiopatologia , Eletrodos Implantados/efeitos adversos , Eletromiografia , Hiperalgesia/psicologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Dor , Limiar da Dor/fisiologia , Vísceras , ÁguaRESUMO
BACKGROUND: Knowledge on optimal electrical stimulation (ES) modalities and region-specific functional effects of colonic neuromodulation is lacking. We aimed to map the regional colonic motility in response to ES of (a) the colonic tissue and (b) celiac branch of the abdominal vagus nerve (CBVN) in an anesthetized porcine model. METHODS: In male Yucatan pigs, direct ES (10 Hz, 2 ms, 15 mA) of proximal (pC), transverse (tC), or distal (dC) colon was done using planar flexible multi-electrode array panels and CBVN ES (2 Hz, 0.3-4 ms, 5 mA) using pulse train (PT), continuous (10 min), or square-wave (SW) modalities, with or without afferent nerve block (200 Hz, 0.1 ms, 2 mA). The regional luminal manometric changes were quantified as area under the curve of contractions (AUC) and luminal pressure maps generated. Contractions frequency power spectral analysis was performed. Contraction propagation was assessed using video animation of motility changes. KEY RESULTS: Direct colon ES caused visible local circular (pC, tC) or longitudinal (dC) muscle contractions and increased luminal pressure AUC in pC, tC, and dC (143.0 ± 40.7%, 135.8 ± 59.7%, and 142.0 ± 62%, respectively). The colon displayed prominent phasic pressure frequencies ranging from 1 to 12 cpm. Direct pC and tC ES increased the dominant contraction frequency band (1-6 cpm) power locally. Pulse train CBVN ES (2 Hz, 4 ms, 5 mA) triggered pancolonic contractions, reduced by concurrent afferent block. Colon contractions propagated both orally and aborally in short distances. CONCLUSION AND INFERENCES: In anesthetized pigs, the dominant contraction frequency band is 1-6 cpm. Direct colonic ES causes primarily local contractions. The CBVN ES-induced pancolonic contractions involve central neural network.
Assuntos
Colo/inervação , Estimulação Elétrica/métodos , Motilidade Gastrointestinal/fisiologia , Nervo Vago , Animais , Colo/fisiologia , Manometria , Sus scrofa , SuínosRESUMO
Corticotropin-releasing factor (CRF) 1 receptor (CRF(1)) activation in the brain is a core pathway orchestrating the stress response. Anatomical data also support the existence of CRF signaling components within the colon. We investigated the colonic response to intraperitoneal (ip) injection of cortagine, a newly developed selective CRF(1) peptide agonist. Colonic motor function and visceral motor response (VMR) were monitored by using a modified miniaturized pressure transducer catheter in adult conscious male Sprague-Dawley rats and C57Bl/6 mice. Colonic permeability was monitored by the Evans blue method and myenteric neurons activation by Fos immunohistochemistry. Compared with vehicle, cortagine (10 microg/kg ip) significantly decreased the distal colonic transit time by 45% without affecting gastric transit, increased distal and transverse colonic contractility by 35.6 and 66.2%, respectively, and induced a 7.1-fold increase in defecation and watery diarrhea in 50% of rats during the first hour postinjection whereas intracerebroventricular (icv) cortagine (3 microg/rat) had lesser effects. Intraperitoneal (ip) cortagine also increased colonic permeability, activated proximal and distal colonic myenteric neurons, and induced visceral hypersensitivity to a second set of phasic colorectal distention (CRD). The CRF antagonist astressin (10 mug/kg ip) abolished ip cortagine-induced hyperalgesia whereas injected icv it had no effect. In mice, cortagine (30 microg/kg ip) stimulated defecation by 7.8-fold, induced 60% incidence of diarrhea, and increased VMR to CRD. Stresslike colonic alterations induced by ip cortagine in rats and mice through restricted activation of peripheral CRF(1) receptors support a role for peripheral CRF(1) signaling as the local arm of the colonic response to stress.
Assuntos
Colo/inervação , Hormônio Liberador da Corticotropina/toxicidade , Motilidade Gastrointestinal/efeitos dos fármacos , Hiperalgesia/induzido quimicamente , Plexo Mientérico/efeitos dos fármacos , Receptores de Hormônio Liberador da Corticotropina/agonistas , Proteínas Recombinantes de Fusão/toxicidade , Estresse Psicológico/fisiopatologia , Animais , Hormônio Liberador da Corticotropina/administração & dosagem , Defecação/efeitos dos fármacos , Diarreia/induzido quimicamente , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Trânsito Gastrointestinal/efeitos dos fármacos , Hiperalgesia/metabolismo , Hiperalgesia/fisiopatologia , Hiperalgesia/prevenção & controle , Injeções Intraperitoneais , Injeções Intraventriculares , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Plexo Mientérico/metabolismo , Plexo Mientérico/fisiopatologia , Vias Neurais/efeitos dos fármacos , Vias Neurais/metabolismo , Vias Neurais/fisiopatologia , Medição da Dor , Fragmentos de Peptídeos/administração & dosagem , Permeabilidade , Estimulação Física , Pressão , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de Hormônio Liberador da Corticotropina/metabolismo , Proteínas Recombinantes de Fusão/administração & dosagem , Estresse Psicológico/metabolismoRESUMO
BACKGROUND: Water avoidance stress (WAS) induces a naloxone-independent visceral analgesia in male rats under non-invasive conditions of monitoring. The objective of the study was to examine the role of brain CRF signaling in acute stress-induced visceral analgesia (SIVA). METHODS: Adult male Sprague-Dawley rats were chronically implanted with an intracerebroventricular (ICV) cannula. The visceromotor response (VMR) to graded phasic colorectal distension (CRD: 10, 20, 40, 60 mm Hg, 20 seconds, 4 minutes intervals) was monitored using manometry. The VMR to a first CRD (baseline) was recorded 5 minutes after an ICV saline injection, followed 1 hour later by ICV injection of either CRF (30, 100, or 300 ng and 1, 3, or 5 µg/rat) or saline and a second CRD, 5 minutes later. Receptor antagonists against CRF1 /CRF2 (astressin-B, 30 µg/rat), CRF2 (astressin2 -B, 10 µg/rat), oxytocin (tocinoic acid, 20 µg/rat), or vehicle were injected ICV 5 minutes before CRF (300 ng/rat, ICV) or 15 minutes before WAS (1 hour). KEY RESULTS: ICV CRF (100 and 300 ng) reduced the VMR to CRD at 60 mm Hg by -36.6% ± 6.8% and -48.7% ± 11.7%, respectively, vs baseline (P < 0.001), while other doses had no effect and IP CRF (10 µg/kg) induced visceral hyperalgesia. Astressin-B and tocinoic acid injected ICV induced hyperalgesia and prevented the analgesic effect of ICV CRF (300 ng/rat) and WAS, while astressin2 -B only blocked WAS-induced SIVA. CONCLUSIONS & INFERENCES: These data support a role for brain CRF signaling via CRF2 in SIVA in a model of WAS and CRD likely mediated by the activation of brain oxytocin pathway.