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International Journal of Molecular... Mar 2023Interleukin 1β (IL1β) is a pro-inflammatory cytokine that may play a crucial role in enteric neuroinflammation in type 1 diabetes. Therefore, our goal is to evaluate...
Interleukin 1β (IL1β) is a pro-inflammatory cytokine that may play a crucial role in enteric neuroinflammation in type 1 diabetes. Therefore, our goal is to evaluate the effects of chronic hyperglycemia and insulin treatment on IL1β immunoreactivity in myenteric neurons and their different subpopulations along the duodenum-ileum-colon axis. Fluorescent immunohistochemistry was used to count IL1β expressing neurons as well as the neuronal nitric oxide synthase (nNOS)- and calcitonin gene-related peptide (CGRP)-immunoreactive myenteric neurons within this group. Tissue IL1β level was measured by ELISA in muscle/myenteric plexus-containing homogenates. IL1β mRNA was detected by RNAscope in different intestinal layers. The proportion of IL1β-immunoreactive myenteric neurons was significantly higher in the colon than in the small intestine of controls. In diabetics, this proportion significantly increased in all gut segments, which was prevented by insulin treatment. The proportion of IL1β-nNOS-immunoreactive neurons only increased in the diabetic colon, while the proportion of IL1β-CGRP-immunoreactive neurons only increased in the diabetic ileum. Elevated IL1β levels were also confirmed in tissue homogenates. IL1β mRNA induction was detected in the myenteric ganglia, smooth muscle and intestinal mucosa of diabetics. These findings support that diabetes-related IL1β induction is specific for the different myenteric neuronal subpopulations, which may contribute to diabetic motility disturbances.
Topics: Rats; Animals; Diabetes Mellitus, Type 1; Calcitonin Gene-Related Peptide; Interleukin-1beta; Diabetes Mellitus, Experimental; Neurons; Nitric Oxide Synthase Type I; RNA, Messenger; Insulins
PubMed: 36982878
DOI: 10.3390/ijms24065804 -
Communications Biology Mar 2023Major advances have been achieved in imaging technologies but most methodological approaches currently used to study the enteric neuronal functions rely on exogenous...
Major advances have been achieved in imaging technologies but most methodological approaches currently used to study the enteric neuronal functions rely on exogenous contrast dyes that can interfere with cellular functions or survival. In the present paper, we investigated whether full-field optical coherence tomography (FFOCT), could be used to visualize and analyze the cells of the enteric nervous system. Experimental work on whole-mount preparations of unfixed mouse colons showed that FFOCT enables the visualization of the myenteric plexus network whereas dynamic FFOCT enables to visualize and identify in situ individual cells in the myenteric ganglia. Analyzes also showed that dynamic FFOCT signal could be modified by external stimuli such veratridine or changes in osmolarity. These data suggest that dynamic FFOCT could be of great interest to detect changes in the functions of enteric neurons and glia in normal and disease conditions.
Topics: Animals; Mice; Tomography, Optical Coherence; Enteric Nervous System; Neuroglia; Neurons; Osmolar Concentration
PubMed: 36864093
DOI: 10.1038/s42003-023-04593-9 -
Cell Reports Mar 2023The enteric nervous system (ENS) consists of glial cells (EGCs) and neurons derived from neural crest precursors. EGCs retain capacity for large-scale neurogenesis in...
The enteric nervous system (ENS) consists of glial cells (EGCs) and neurons derived from neural crest precursors. EGCs retain capacity for large-scale neurogenesis in culture, and in vivo lineage tracing has identified neurons derived from glial cells in response to inflammation. We thus hypothesize that EGCs possess a chromatin structure poised for neurogenesis. We use single-cell multiome sequencing to simultaneously assess transcription and chromatin accessibility in EGCs undergoing spontaneous neurogenesis in culture, as well as small intestine myenteric plexus EGCs. Cultured EGCs maintain open chromatin at genomic loci accessible in neurons, and neurogenesis from EGCs involves dynamic chromatin rearrangements with a net decrease in accessible chromatin. A subset of in vivo EGCs, highly enriched within the myenteric ganglia and that persist into adulthood, have a gene expression program and chromatin state consistent with neurogenic potential. These results clarify the mechanisms underlying EGC potential for neuronal fate transition.
Topics: Single-Cell Analysis; Neuroglia; Neurogenesis; Chromatin; Chromatin Assembly and Disassembly; RNA; Ganglia; Multiomics; Male; Female; Animals; Mice; Enteric Nervous System; Single-Cell Gene Expression Analysis; Cell Culture Techniques; Intestine, Small; Weaning
PubMed: 36857184
DOI: 10.1016/j.celrep.2023.112194 -
Frontiers in Neuroanatomy 2022The enteric nervous system (ENS), sometimes referred to as a "second brain" is a quasi-autonomous nervous system, made up of interconnected plexuses organized in a...
The enteric nervous system (ENS), sometimes referred to as a "second brain" is a quasi-autonomous nervous system, made up of interconnected plexuses organized in a mesh-like network lining the gastrointestinal tract. Originally described as an actor in the regulation of digestion, bowel contraction, and intestinal secretion, the implications of the ENS in various central neuropathologies has recently been demonstrated. However, with a few exceptions, the morphology and pathologic alterations of the ENS have mostly been studied on thin sections of the intestinal wall or, alternatively, in dissected explants. Precious information on the three-dimensional (3-D) architecture and connectivity is hence lost. Here, we propose the fast, label-free 3-D imaging of the ENS, based on intrinsic signals. We used a custom, fast tissue-clearing protocol based on a high refractive-index aqueous solution to increase the imaging depth and allow us the detection of faint signals and we characterized the autofluorescence (AF) from the various cellular and sub-cellular components of the ENS. Validation by immunofluorescence and spectral recordings complete this groundwork. Then, we demonstrate the rapid acquisition of detailed 3-D image stacks from unlabeled mouse ileum and colon, across the whole intestinal wall and including both the myenteric and submucosal enteric nervous plexuses using a new spinning-disk two-photon (2P) microscope. The combination of fast clearing (less than 15 min for 73% transparency), AF detection and rapid volume imaging [less than 1 min for the acquisition of a z-stack of 100 planes (150*150 μm) at sub-300-nm spatial resolution] opens up the possibility for new applications in fundamental and clinical research.
PubMed: 36844894
DOI: 10.3389/fnana.2022.1070062 -
Medicina (Kaunas, Lithuania) Feb 2023: Diabetic gastroenteropathy (DG) is a common complication of diabetes mellitus type 2. Interstitial cells are non-neural cells of mesenchymal origin inserted between...
: Diabetic gastroenteropathy (DG) is a common complication of diabetes mellitus type 2. Interstitial cells are non-neural cells of mesenchymal origin inserted between nerve elements and smooth muscle cells, necessary for normal function and peristaltic contractions in the gastrointestinal (GI) tract. There are at least two types of interstitial cells within the GI muscle layer-interstitial cells of Cajal (ICC) and interstitial platelet-derived growth factor receptor α-positive cells (IPC). The mechanism of diabetic gastroenteropathy is unclear, and interstitial cells disorders caused by metabolic changes in diabetes mellitus (DM) could explain the symptoms of DG (slow intestinal transit, constipation, fecal incontinence). The aim of this study was to identify PDGFRα and c-kit immunoreactive cells in the colon of rats with streptozotocin-nicotinamide-induced diabetes mellitus type 2, as well as to determine their distribution in relation to smooth muscle cells and enteric nerve structures. : Male Wistar rats were used, and diabetes type 2 was induced by an intraperitoneal injection of streptozotocin, immediately after intraperitoneal application of nicotinamide. The colon specimens were exposed to PDGFRα and anti-c-kit antibodies to investigate interstitial cells; enteric neurons and smooth muscle cells were immunohistochemically labeled with NF-M and desmin antibodies. : Significant loss of the intramuscular ICC, myenteric ICC, and loss of their connection in intramuscular linear arrays and around the ganglion of the myenteric plexus were observed with no changes in nerve fiber distribution in the colon of rats with diabetes mellitus type 2. IPC were rarely present within the colon muscle layer with densely distributed PDGFRα+ cells in the colon mucosa and submucosa of both experimental groups. In summary, a decrease in intramuscular ICC, discontinuities and breakdown of contacts between myenteric ICC without changes in IPC and nerve fibers distribution were observed in the colon of streptozotocin/nicotinamide-induced diabetes type 2 rats.
Topics: Rats; Male; Animals; Interstitial Cells of Cajal; Receptor, Platelet-Derived Growth Factor alpha; Streptozocin; Rats, Wistar; Diabetes Mellitus, Type 2; Colon; Gastrointestinal Diseases
PubMed: 36837509
DOI: 10.3390/medicina59020308 -
International Journal of Molecular... Feb 2023Aquaporin 1 (AQP1) is one of thirteen known mammalian aquaporins. Its main function is the transport of water across cell membranes. Lately, a role of AQP has been...
Aquaporin 1 (AQP1) is one of thirteen known mammalian aquaporins. Its main function is the transport of water across cell membranes. Lately, a role of AQP has been attributed to other physiological and pathological functions including cell migration and peripheral pain perception. AQP1 has been found in several parts of the enteric nervous system, e.g., in the rat ileum and in the ovine duodenum. Its function in the intestine appears to be multifaceted and is still not completely understood. The aim of the study was to analyze the distribution and localization of AQP1 in the entire intestinal tract of mice. AQP1 expression was correlated with the hypoxic expression profile of the various intestinal segments, intestinal wall thickness and edema, as well as other aspects of colon function including the ability of mice to concentrate stools and their microbiome composition. AQP1 was found in a specific pattern in the serosa, the mucosa, and the enteric nervous system throughout the gastrointestinal tract. The highest amount of AQP1 in the gastrointestinal tract was found in the small intestine. AQP1 expression correlated with the expression profiles of hypoxia-dependent proteins such as HIF-1α and PGK1. Loss of AQP1 through knockout of AQP1 in these mice led to a reduced amount of bacteroidetes and firmicutes but an increased amount of the rest of the phyla, especially deferribacteres, proteobacteria, and verrucomicrobia. Although AQP-KO mice retained gastrointestinal function, distinct changes regarding the anatomy of the intestinal wall including intestinal wall thickness and edema were observed. Loss of AQP1 might interfere with the ability of the mice to concentrate their stool and it is associated with a significantly different composition of the of the bacterial stool microbiome.
Topics: Animals; Mice; Rats; Aquaporin 1; Aquaporins; Colon; Duodenum; Edema; Hypoxia; Mammals; Mice, Knockout; Sheep; Gastrointestinal Tract
PubMed: 36835026
DOI: 10.3390/ijms24043616 -
International Journal of Environmental... Feb 2023A particularly pressing problem is determining consumer-safe doses of potentially health- and life-threatening substances, such as acrylamide. The aim of the study was...
BACKGROUND
A particularly pressing problem is determining consumer-safe doses of potentially health- and life-threatening substances, such as acrylamide. The aim of the study was to determine how acrylamide affects the pituitary adenylate cyclase-activating polypeptide (PACAP)-immunoreactive intramural neurons in the small intestine of sexually immature gilts.
METHODS
The study was conducted on 15 sexually immature Danish gilts receiving for 28 days empty gelatin capsules or acrylamide in low (0.5 µg/kg of body weight (b.w.)/day) and high (5 µg/kg b.w./day) doses. After euthanasia, intestinal sections were stained using the double immunofluorescence staining procedure.
RESULTS
Studies have shown that oral administration of acrylamide in both doses induced a response of intramural neurons expressed as an increase in the population of PACAP-immunoreactive neurons in the small intestine. In the duodenum, only in the myenteric plexus (MP) was an increase in the number of PACAP-immunoreactive (IR) neurons observed in both experimental groups, while in the outer submucous plexus (OSP) and inner submucous plexus (ISP), an increase was noted only in the high-dose group. In the jejunum, both doses of acrylamide led to an increase in the population of PACAP-IR neurons in each enteric plexus (MP, OSP, ISP), while in the ileum, only supplementation with the higher dose of acrylamide increased the number of PACAP-IR enteric neurons in the MP, OSP, and ISP.
CONCLUSIONS
The obtained results suggest the participation of PACAP in acrylamide-induced plasticity of enteric neurons, which may be an important line of defence from the harmful action of acrylamide on the small intestines.
Topics: Swine; Animals; Female; Pituitary Adenylate Cyclase-Activating Polypeptide; Acrylamide; Intestine, Small; Neurons; Sus scrofa
PubMed: 36833970
DOI: 10.3390/ijerph20043272 -
International Journal of Pharmaceutics Mar 2023The addiction induced by the misuse of opioids, is not only a public health emergency but also a social and economic welfare. The main therapy is based on opioid...
The addiction induced by the misuse of opioids, is not only a public health emergency but also a social and economic welfare. The main therapy is based on opioid antagonists. Oral and injectable naltrexone administration is the most widely used, presenting some inconveniences: poor patient adherence to the oral daily dosing schedule, cases of hepatitis and clinically significant liver dysfunction. This study proposes the in vitro e in vivo evaluation of anti-opioid properties of naloxone loaded-poly(lactic-co-glycolic) acid microparticles (NX-MP). In previous studies, NX-MP showed in vitro sustained naloxone release for one week at least. Our results demonstrate the in vitro efficacy of the NX-MP antagonizing for 7 days the morphine effect in SH-SY5Y cells and myenteric plexus-longitudinal muscle preparations isolated from guinea-pig ileum. The in vivo evaluation of the NX-MP was carried out in mice testing the antagonism of the antinociceptive effect of morphine. Results showed that subcutaneous administration of NX-MP blocked the morphine effect. The results of this work suggest that the subcutaneous administration of NX-MP enhances naloxone therapeutic efficacy as non-addictive medication and could be a promising alternative to naltrexone. Furthermore, the dose of NX-MP can be adapted to the patient necessities. It would be an interesting advantage to treat opioid-addiction.
Topics: Humans; Mice; Animals; Guinea Pigs; Naloxone; Morphine; Analgesics, Opioid; Naltrexone; Neuroblastoma; Narcotic Antagonists
PubMed: 36822337
DOI: 10.1016/j.ijpharm.2023.122766 -
Autonomic Neuroscience : Basic &... May 2023Substance-P (SP) is a commonly used marker of nociceptive afferent axons, and it plays an important role in a variety of physiological functions including the regulation...
Substance-P (SP) is a commonly used marker of nociceptive afferent axons, and it plays an important role in a variety of physiological functions including the regulation of motility, gut secretion, and vascular flow. Previously, we found that SP-immunoreactive (SP-IR) axons densely innervated the pyloric antrum of the flat-mount of the mouse whole stomach muscular layer. However, the regional distribution and morphology of SP-IR axons in the submucosa and mucosa were not well documented. In this study, the mouse antrum-pylorus-duodenum (APD) were transversely and longitudinally sectioned. A Zeiss M2 imager was used to scan the serial sections of each APD (each section montage consisted of 50-100 all-in-focus maximal projection images). To determine the detailed structures of SP-IR axons and terminals, we used the confocal microscope to scan the regions of interest. We found that 1) SP-IR axons innervated the muscular, submucosal, and mucosal layers. 2) In the muscular layer, SP-IR varicose axons densely innervated the muscles and formed varicose terminals which encircled myenteric neurons. 3) In the submucosa, SP-IR axons innervated blood vessels and submucosal ganglia and formed a network in Brunner's glands. 4) In the mucosa, SP-IR axons innervated the muscularis mucosae. Some SP-IR axons entered the lamina propria. 5) The muscular layer of the antrum and duodenum showed a higher SP-IR axon density than the pyloric sphincter. 6) SP-IR axons were from extrinsic and intrinsic origins. This work provided a comprehensive view of the distribution and morphology of SP-IR axons in the APD at single cell/axon/varicosity scale. This data will be used to create a 3D scaffold of the SP-IR axon innervation of the APD.
Topics: Mice; Animals; Pylorus; Substance P; Axons; Duodenum; Neurons
PubMed: 36804650
DOI: 10.1016/j.autneu.2023.103074