Impaired intestinal barrier function, low-grade inflammation and altered neuronal control are reported in functional gastrointestinal disorders. However, the sequence of and causal relation between ...these events is unclear, necessitating a spontaneous animal model. The aim of this study was to describe the natural history of intestinal permeability, mucosal and neuromuscular inflammation and nitrergic motor neuron function during the lifetime of the BioBreeding (BB) rat.
Normoglycemic BB-diabetes prone (DP) and control rats were sacrificed at different ages and jejunum was harvested to characterize intestinal permeability, inflammation and neuromuscular function.
Both structural and functional evidence of increased intestinal permeability was found in young BB-DP rats from the age of 50 days. In older animals, starting in the mucosa from 70 days and in half of the animals also in the muscularis propria from 110 days, an inflammatory reaction, characterized by an influx of polymorphonuclear cells and higher myeloperoxidase activity, was observed. Finally, in animals older than 110 days, coinciding with a myenteric ganglionitis, a loss of nitrergic neurons and motor function was demonstrated.
In the BB-rat, mucosal inflammatory cell infiltration is preceded by intestinal barrier dysfunction and followed by myenteric ganglionitis and loss of nitrergic function. This sequence supports a primary role for impaired barrier function and provides an insightful model for the pathogenesis of functional gastrointestinal disorders.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Vagus nerve stimulation (VNS), most likely via enteric neurons, prevents postoperative ileus (POI) by reducing activation of alpha7 nicotinic receptor (α7nAChR) positive
macrophages (mMφ) and ...dampening surgery-induced intestinal inflammation. Here, we evaluated if 5-HT4 receptor (5-HT4R) agonist prucalopride can mimic this effect in mice and human.
Using Ca
imaging, the effect of electrical field stimulation (EFS) and prucalopride was evaluated in situ on mMφ activation evoked by ATP in jejunal
tissue. Next, preoperative and postoperative administration of prucalopride (1-5 mg/kg) was compared with that of preoperative VNS in a model of POI in wild-type and α7nAChR knockout mice. Finally, in a pilot study, patients undergoing a Whipple procedure were preoperatively treated with prucalopride (n=10), abdominal VNS (n=10) or sham/placebo (n=10) to evaluate the effect on intestinal inflammation and clinical recovery of POI.
EFS reduced the ATP-induced Ca
response of mMφ, an effect that was dampened by neurotoxins tetrodotoxin and ω-conotoxin and mimicked by prucalopride. In vivo, prucalopride administered before, but not after abdominal surgery reduced intestinal inflammation and prevented POI in wild-type, but not in α7nAChR knockout mice. In humans, preoperative administration of prucalopride, but not of VNS, decreased
and
expression in the
and improved clinical recovery.
Enteric neurons dampen mMφ activation, an effect mimicked by prucalopride. Preoperative, but not postoperative treatment with prucalopride prevents intestinal inflammation and shortens POI in both mice and human, indicating that preoperative administration of 5-HT4R agonists should be further evaluated as a treatment of POI.
NCT02425774.
Collagen fibre orientation in human bridging veins Kapeliotis, Markos; Gavrila Laic, Rebeca Alejandra; Peñas, Alvaro Jorge ...
Biomechanics and modeling in mechanobiology,
12/2020, Letnik:
19, Številka:
6
Journal Article
Recenzirano
Odprti dostop
Bridging veins (BVs) drain the blood from the cerebral cortex into dural sinuses. BVs have one end attached to the brain and the other to the superior sagittal sinus (SSS), which is attached to the ...skull. Relative movement between these two structures can cause BV to rupture producing acute subdural haematoma, a head injury with a mortality rate between 30 and 90%. A clear understanding of the BVs microstructure is required to increase the biofidelity of BV models when simulating head impacts. Twelve fresh BV samples draining in the superior sagittal sinus (SSS) from a single human cadaver were cut open along their length and placed on an inverted multiphoton microscope. To ensure that the BVs were aligned with the axial direction an in-house built, uniaxial tension set-up was used. Two scans were performed per sample. Before the first scan, a minor displacement was applied to align the tissue; then, a second scan was taken applying 50% strain. Each BV was scanned for a length of 5 mm starting from the drainage site into the SSS. Imaging was performed on a Zeiss LSM780 microscope with an 25
×
water immersion objective (NA 0.8), coupled to a tunable MaiTai DS (Spectraphysics) pulsed laser with the wavelength set at 850 nm. Second harmonic and fluorescence signals were captured in forward and backward direction on binary GaAsP (BiG) detectors and stored as four colour Z-stacks. Prior to the calculation of the local orientations, acquired Z-stacks were denoised and enhanced to highlight fibrillar structures from the background. Then, for each
Z
-plane of the stack, the ImageJ plugin OrientationJ was used to extract the local 2D orientations of the fibres based on structure tensors. Two kinds of collagen architectures were seen. The most common (8
/
12 samples) was single layered and had a uniform distribution of collagen. The less common (4
/
12 samples) had 2 layers and 7 to 34 times thicker collagen bundles on the outer layer. Fibre angle analysis showed that collagen was oriented mainly along the axial direction of the vessel. The von Mises fittings showed that in order to describe the fibre distribution 3 components were needed with mean angles
μ
at
-
0.35, 0.21,
-
0.02 rad or
-
20.2
∘
, 12.1
∘
,
-
1.2
∘
relative to the vessel’s axial direction which was also the horizontal scan direction.
The enteric nervous system (ENS) is a vital part of the autonomic nervous system that regulates many gastrointestinal functions, including motility and secretion. All neurons and glia of the ENS ...arise from neural crest-derived cells that migrate into the gastrointestinal tract during embryonic development. It has been known for many years that a subpopulation of the enteric neural crest-derived cells expresses pan-neuronal markers at early stages of ENS development. Recent studies have demonstrated that some enteric neurons exhibit electrical activity from as early as E11.5 in the mouse, with further maturation of activity during embryonic and postnatal development. This article discusses the maturation of electrophysiological and morphological properties of enteric neurons, the formation of synapses and synaptic activity, and the influence of neural activity on ENS development.
► Electrical activity occurs early during development of the enteric nervous system. ► Electrical activity influences enteric neuron differentiation. ► Maturation of neuronal properties continues in early postnatal ages during murine enteric development. ► Future studies are required to examine the interactions between genetic factors and neural activity.
Live imaging has become an essential tool to investigate the coordinated activity and output of cellular networks. Within the last decade, 2 Nobel prizes have been awarded to recognize innovations in ...the field of imaging: one for the discovery, use, and optimization of the green fluorescent protein (2008) and the second for the development of super-resolved fluorescence microscopy (2014). New advances in both optogenetics and microscopy now enable researchers to record and manipulate ac-tivity from specific populations of cells with better contrast and resolution, at higher speeds, and deeper into live tissues. In this review, we will discuss some of the recent developments in microscope technology and in the synthesis of fluorescent probes, both synthetic and genetically encoded. We focus on how live imaging of cellular physiology has progressed our under-standing of the control of gastrointestinal motility, and we discuss the hurdles to overcome in order to apply the novel tools in the field of neurogastroenterology and motility.
Acetylcholine-activating pentameric nicotinic receptors (nAChRs) are an essential mode of neurotransmission in the enteric nervous system (ENS). In this study, we examined the functional development ...of specific nAChR subtypes in myenteric neurons using Wnt1-Cre;R26R-GCaMP3 mice, where all enteric neurons and glia express the genetically encoded calcium indicator, GCaMP3. Transcripts encoding α3, α4, α7, β2, and β4 nAChR subunits were already expressed at low levels in the E11.5 gut and by E14.5 and, thereafter, α3 and β4 transcripts were the most abundant. The effect of specific nAChR subtype antagonists on evoked calcium activity in enteric neurons was investigated at different ages. Blockade of the α3β4 receptors reduced electrically and chemically evoked calcium responses at E12.5, E14.5, and P0. In addition to the α3β4 antagonist, antagonists to α3β2 and α4β2 also significantly reduced responses by P10-11 and in adult preparations. Therefore, there is an increase in the diversity of functional nAChRs during postnatal development. However, an α7 nAChR antagonist had no effect at any age. Furthermore, at E12.5 we found evidence for unconventional receptors that were responsive to the nAChR agonists 1-dimethyl-4-phenylpiperazinium and nicotine, but were insensitive to the general nicotinic blocker, hexamethonium. Migration, differentiation, and neuritogenesis assays did not reveal a role for nAChRs in these processes during embryonic development. In conclusion, there are significant changes in the contribution of different nAChR subunits to synaptic transmission during ENS development, even after birth. This is the first study to investigate the development of cholinergic transmission in the ENS.
The enteric nervous system (ENS) situated within the gastrointestinal tract comprises an intricate network of neurons and glia which together regulate intestinal function. The exact neuro-glial ...circuitry and the signaling molecules involved are yet to be fully elucidated. Vasoactive intestinal peptide (VIP) is one of the main neurotransmitters in the gut, and is important for regulating intestinal secretion and motility. However, the role of VIP and its VPAC receptors within the enteric circuitry is not well understood. We investigated this in the submucosal plexus of mouse jejunum using calcium (Ca
)-imaging. Local VIP application induced Ca
-transients primarily in neurons and these were inhibited by VPAC1- and VPAC2-antagonists (PG 99-269 and PG 99-465 respectively). These VIP-evoked neural Ca
-transients were also inhibited by tetrodotoxin (TTX), indicating that they were secondary to action potential generation. Surprisingly, VIP induced Ca
-transients in glia in the presence of the VPAC2 antagonist. Further, selective VPAC1 receptor activation with the agonist (K15, R16, L27VIP(1-7)/GRF(8-27)) predominantly evoked glial responses. However, VPAC1-immunoreactivity did not colocalize with the glial marker glial fibrillary acidic protein (GFAP). Rather, VPAC1 expression was found on cholinergic submucosal neurons and nerve fibers. This suggests that glial responses observed were secondary to neuronal activation. Trains of electrical stimuli were applied to fiber tracts to induce endogenous VIP release. Delayed glial responses were evoked when the VPAC2 antagonist was present. These findings support the presence of an intrinsic VIP/VPAC-initiated neuron-to-glia signaling pathway. VPAC1 agonist-evoked glial responses were inhibited by purinergic antagonists (PPADS and MRS2179), thus demonstrating the involvement of P2Y
receptors. Collectively, we showed that neurally-released VIP can activate neurons expressing VPAC1 and/or VPAC2 receptors to modulate purine-release onto glia. Selective VPAC1 activation evokes a glial response, whereas VPAC2 receptors may act to inhibit this response. Thus, we identified a component of an enteric neuron-glia circuit that is fine-tuned by endogenous VIP acting through VPAC1- and VPAC2-mediated pathways.
Coordination of gastrointestinal function relies on joint efforts of enteric neurons and glia, whose crosstalk is vital for the integration of their activity. To investigate the signaling mechanisms ...and to delineate the spatial aspects of enteric neuron‐to‐glia communication within enteric ganglia we developed a method to stimulate single enteric neurons while monitoring the activity of neighboring enteric glial cells. We combined cytosolic calcium uncaging of individual enteric neurons with calcium imaging of enteric glial cells expressing a genetically encoded calcium indicator and demonstrate that enteric neurons signal to enteric glial cells through pannexins using paracrine purinergic pathways. Sparse labeling of enteric neurons and high‐resolution analysis of the structural relation between neuronal cell bodies, varicose release sites and enteric glia uncovered that this form of neuron‐to‐glia communication is contained between the cell body of an enteric neuron and its surrounding enteric glial cells. Our results reveal the spatial and functional foundation of neuro‐glia units as an operational cellular assembly in the enteric nervous system.
Main Points
Enteric neurons release purines to communicate with enteric glial cells in their direct vicinity.
Nerve cell bodies are closely associated with glial cells to form functional neuro‐glia units within ganglia of the enteric nervous system.
Neurons of the enteric nervous system (ENS) arise from neural crest cells that migrate into and along the developing gastrointestinal tract. A subpopulation of these neural-crest derived cells ...express pan-neuronal markers early in development, shortly after they first enter the gut. However, it is unknown whether these early enteric "neurons" are electrically active. In this study we used live Ca(2+) imaging to examine the activity of enteric neurons from mice at embryonic day 11.5 (E11.5), E12.5, E15.5, and E18.5 that were dissociated and cultured overnight. PGP9.5-immunoreactive neurons from E11.5 gut cultures responded to electrical field stimulation with fast Ca(2+)(i) transients that were sensitive to TTX and ω-conotoxin GVIA, suggesting roles for voltage-gated Na(+) channels and N-type voltage-gated Ca(2+) channels. E11.5 neurons were also responsive to the nicotinic cholinergic agonist, dimethylphenylpiperazinium, and to ATP. In addition, spontaneous Ca(2+)(i) transients were present. Similar responses were observed in neurons from older embryonic gut. Whole-cell patch-clamp recordings performed on E12.5 enteric neurons after 2-10 h in culture revealed that these neurons fired both spontaneous and evoked action potentials. Together, our results show that enteric neurons exhibit mature forms of activity at early stages of ENS development. This is the first investigation to directly examine the presence of neural activity during enteric neuron development. Along with the spinal cord and hindbrain, the ENS appears to be one of the earliest parts of the nervous system to exhibit electrical activity.
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