Search

Key points Chloroquine (CQ) stimulates itch nerves and causes intense scratching in mice by activating the G-protein coupled receptor (GPCR) MrgprA3; it is not known how stimulation of MrgprA3 (or other GPCRs) leads to activation of the itch nerve terminals in the skin, but previous studies have found that transient receptor potential A1 (TRPA1) gene deletion blocks CQ-induced scratching., In the present study we used a novel dorsal skin-nerve preparation to evaluate mechanisms underlying CQ- and histamine-induced action potential discharge in itch nerve terminals., We found that CQ activation of the nerves requires the beta3 isoform of phospholipase C, but TRPA1 or other TRP channel are not required., Evidence is provided for a role for calcium-activated chloride channels such as TMEM16a in GPCR-activation of itch nerve terminals., The mechanism by which TRP channels participate in pruritogen-induced scratching may involve sites of action other than the primary afferent terminals., Abstract Chloroquine (CQ) and histamine are pruritogens commonly used to study itch in the mouse. A novel skin-nerve preparation was used to evaluate chloroquine (CQ)- and histamine-induced activation of afferent nerves in the dorsal thoracic skin of the mouse. All CQ sensitive nerves were C-fibres, and were also sensitive to histamine. The response to CQ, but not histamine, was largely absent in mrgpr-cluster Δ−/− mice, supporting the hypothesis that CQ evokes itch largely via stimulation of MrgprA3 receptors. The CQ-induced action potential discharge was largely absent in phospholipase Cβ3 knockout animals. The CQ and histamine responses were not influenced by removal of TRPA1, TRPV1, TRPC3 or TRPC6, nor by the TRP channel blocker Ruthenium Red. The bouts of scratching in response to CQ were not different between wild-type and TRPA1-deficient mice. A selective inhibitor of the calcium-activated chloride channel TMEM16A, N-((4-methoxy)-2-naphthyl)-5-nitroanthranilic acid (MONNA), inhibited CQ-induced action potential discharge at itch nerve terminals and bouts of scratching by about 50%. Although TRPA1 and TRPV1 channels may be involved in the scratching responses to intradermal pruritogens, this is unlikely to be due to an effect at the nerve terminals, where chloride channels may play a more important role. [ABSTRACT FROM AUTHOR]

Background Gastroesophageal reflux can cause high acidity in the esophagus and trigger heartburn and pain. However, because of the esophageal mucosal barrier, the acidity at the nerve terminals of pain-mediating C-fibers in esophageal mucosa is predicted to be substantially lower. We hypothesized that the esophageal dorsal root ganglia ( DRG) C-fibers are activated by mild acid (compared to acidic reflux), and express receptors and ion channels highly sensitive to acid. Methods Extracellular single unit recordings of activity originating in esophageal DRG C-fiber nerve terminals were performed in the innervated esophagus preparation ex vivo. Acid was delivered in a manner that bypassed the esophageal mucosal barrier. The expression of mRNA for selected receptors in esophagus-specific DRG neurons was evaluated using single cell RT- PCR. Key Results Mild acid ( pH = 6.5-5.5) activated esophageal DRG C-fibers in a pH-dependent manner. The response to mild acid at pH = 6 was not affected by the TRPV1 selective antagonist iodo-resiniferatoxin. The majority (70-95%) of esophageal DRG C-fiber neurons ( TRPV1-positive) expressed mRNA for acid sensing ion channels ( ASIC1a, ASIC1b, ASIC2b, and/or ASIC3), two-pore-domain (K2P) potassium channel TASK1, and the proton-sensing G-protein coupled receptor OGR1. Other evaluated targets ( PKD2L1, TRPV4, TASK3, TALK1, G2A, GPR4, and TDAG8) were expressed rarely. Conclusions & Inferences Guinea pig esophageal DRG C-fibers are activated by mild acid via a TRPV1-independent mechanism, and express mRNA for several receptors and ion channels highly sensitive to acid. The high acid sensitivity of esophageal C-fibers may contribute to heartburn and pain in conditions of reduced mucosal barrier function. [ABSTRACT FROM AUTHOR]

Background The mouse is an invaluable model for mechanistic studies of esophageal nerves, but the afferent innervation of the mouse esophagus is incompletely understood. Vagal afferent neurons are derived from two embryonic sources: neural crest and epibranchial placodes. We hypothesized that both neural crest and placodes contribute to the TRPV1-positive (potentially nociceptive) vagal innervation of the mouse esophagus. Methods Vagal jugular/nodose ganglion (JNG) and spinal dorsal root ganglia (DRG) neurons were retrogradely labeled from the cervical esophagus. Single cell RT-PCR was performed on the labeled neurons. Key Results In the Wnt1Cre/R26R mice expressing a reporter in the neural crest-derived cells we found that both the neural crest- and the placodes-derived vagal JNG neurons innervate the mouse esophagus. In the wild-type mouse the esophageal vagal JNG TRPV1-positive neurons segregated into two subsets: putative neural crest-derived purinergic receptor P2X2-negative/preprotachykinin-A (PPT-A)-positive subset and putative placodes-derived P2X2-positive/PPTA-negative subset. These subsets also segregated by the expression of TrkA and GFRα3 in the putative neural crest-derived subset, and TrkB in the putative placodes-derived subset. The TRPV1-positive esophageal DRG neurons had the phenotype similar to the vagal putative neural crest-derived subset. Conclusions & Inferences The TRPV1-positive (potentially nociceptive) vagal afferent neurons innervating the mouse esophagus originate from both neural crest and placodes. The expression profile of the receptors for neurotrophic factors is similar between the neural crest-derived vagal and spinal nociceptors, but distinct from the vagal placodes-derived nociceptors. [ABSTRACT FROM AUTHOR]

Background The TRPA1 receptor is directly activated by a wide range of chemicals including many endogenous molecules relevant for esophageal pathophysiology. We addressed the hypothesis that the TRPA1 agonists differentially activate esophageal nociceptive subtypes depending on their embryological source (neural crest or epibranchial placodes). Methods Single cell RT-PCR and whole cell patch clamp recordings were performed on the vagal neurons retrogradely labeled from the guinea pig esophagus. Extracellular recordings were made in the isolated innervated esophagus preparation ex vivo. Key Results Single cell RT-PCR revealed that the majority of the nodose (placodes-derived) and jugular (neural crest-derived) TRPV1-positive esophageal nociceptors express TRPA1. Single fiber recording showed that the TRPA1 agonists allyl-isothiocyanate (AITC) and cinnamaldehyde were effective in inducing robust action potential discharge in the nerve terminals of nodose nociceptors, but had far less effect in jugular nociceptors (approximately fivefold less). Higher efficacy of the TRPA1 agonists to activate nodose nociceptors was confirmed in the isolated esophagus-labeled vagal neurons in the whole cell patch clamp studies. Similarly to neural crest-derived vagal jugular nociceptors, the spinal DRG nociceptors that are also neural crest-derived were only modestly activated by allyl-isothiocyanate. Conclusions & Inferences We conclude that the TRPA1 agonists are substantially more effective activators of the placodes-derived than the neural crest-derived esophageal nociceptors. Our data predict that in esophageal diseases the presence of endogenous TRPA1 activators will be preferentially signaled by the vagal nodose nociceptors. [ABSTRACT FROM AUTHOR]

Clinical studies implicate adenosine acting on esophageal nociceptive pathways in the pathogenesis of noncardiac chest pain originating from the esophagus. However, the effect of adenosine on esophageal afferent nerve subtypes is incompletely understood. We addressed the hypothesis that adenosine selectively activates esophageal nociceptors. Whole cell perforated patch-clamp recordings and single-cell RT-PCR analysis were performed on the primary afferent neurons retrogradely labeled from the esophagus in the guinea pig. Extracellular recordings were made from the isolated innervated esophagus. In patch-clamp studies, adenosine evoked activation (inward current) in a majority of putative nociceptive (capsaicin-sensitive) vagal nodose, vagal jugular, and spinal dorsal root ganglia (DRG) neurons innervating the esophagus. Single-cell RT-PCR analysis indicated that the majority of the putative nociceptive (transient receptor potential V1-positive) neurons innervating the esophagus express the adenosine receptors. The neural crest-derived (spinal DRG and vagal jugular) esophageal nociceptors expressed predominantly the adenosine A1 receptor while the placodes-derived vagal nodose nociceptors expressed the adenosine A1 and/or A2A receptors. Consistent with the studies in the cell bodies, adenosine evoked activation (overt action potential discharge) in esophageal nociceptive nerve terminals. Furthermore, the neural crest-derived jugular nociceptors were activated by the selective A1 receptor agonist CCPA, and the placodes-derived nodose nociceptors were activated by CCPA and/or the selective adenosine A2A receptor CGS-21680. In contrast to esophageal nociceptors, adenosine failed to stimulate the vagal esophageal low-threshold (tension) mechanosensors. We conclude that adenosine selectively activates esophageal nociceptors. Our data indicate that the esophageal neural crest-derived nociceptors can be activated via the adenosine A1 receptor while the placodes-derived esophageal nociceptors can be activated via A1 and/or A2A receptors. Direct activation of esophageal nociceptors via adenosine receptors may contribute to the symptoms in esophageal diseases. [ABSTRACT FROM AUTHOR]

Abstract: Vagal afferent nerves are essential for optimal neural regulation of visceral organs, but are not often considered important for their defense. However, there are well-defined subsets of vagal afferent nerves that have activation properties indicative of specialization to detect potentially harmful stimuli (nociceptors). This is clearly exemplified by the vagal bronchopulmonary C-fibers that are quiescent in healthy lungs but are readily activated by noxious chemicals and inflammatory molecules. Vagal afferent nerves with similar activation properties have been also identified in the esophagus and probably exist in other visceral tissues. In addition, these putative vagal nociceptors often initiate defensive reflexes, can be sensitized, and have the capacity to induce central sensitization. This set of properties is a characteristic of nociceptors in somatic tissues. [Copyright &y& Elsevier]

The afferent neurons innervating the oesophagus originate from two embryonic sources: neurons located in vagal nodose ganglia originate from embryonic placodes and neurons located in vagal jugular and spinal dorsal root ganglia (DRG) originate from the neural crest. Here, we address the hypothesis that 5-hydroxytryptamine (5-HT) differentially stimulates afferent nerve subtypes in the oesophagus. Extracellular recordings of single unit activity originating from nerve terminals were made in the isolated innervated guinea-pig oesophagus. Whole cell patch clamp recordings (35 °C) were made from the primary afferent neurons retrogradely labelled from the oesophagus. 5-Hydroxytryptamine (10 μmol L−1) activated vagal nodose C-fibres (70%) in the oesophagus but failed to activate overtly vagal jugular nerve fibres and oesophagus-specific spinal DRG neurons. The response to 5-HT in nodose C-fibre nerve terminals was mimicked by the selective 5-HT3 receptor agonist 2-methyl-5-HT (10 μmol L−1) and nearly abolished by the 5-HT3 receptor antagonists ondansetron (10 μmol L−1) and Y-25130 (10 μmol L−1). In patch clamp studies, 2-methyl-5-HT (10 μmol L−1) activated a proportion of isolated oesophagus-specific nodose capsaicin-sensitive neurons (putative cell bodies of nodose C-fibres). We conclude that the responsiveness to 5-HT discriminates placode-derived (vagal nodose) C-fibres from the neural crest-derived (vagal jugular and spinal DRG) afferent nerves in the oesophagus. The response to 5-HT in nodose C-fibres is mediated by the 5-HT3 receptor in their neuronal membrane. [ABSTRACT FROM AUTHOR]

Clinical studies indicate that adeno- sine contributes to esophageal mechanical hypersensitivity in some patients with pain originating in the esophagus. We have previously reported that the esophageal vagal nodose C fibers express the aden- osine A2A receptor. Here we addressed the hypothesis that stimulation of the adenosine A2A receptor induces mechanical sensitization of esophageal C fibers by a mechanism involving transient receptor potential A1 (TRPA1). Extracellular single fiber recordings of activity originating in C-fiber terminals were made in the ex vivo vagally innervated guinea pig esophagus. The adenosine A2A receptor-selec- tive agonist CGS21680 induced robust, reversible sensitization of the response to esophageal distention (10 - 60 mmHg) in a concentration- dependent fashion (1-100 nM). At the half-maximally effective con- centration (EC50: =3 nM), CGS21680 induced an approximately twofold increase in the mechanical response without causing an overt activation. This sensitization was abolished by the selective A2A antagonist SCH58261. The adenylyl cyclase activator forskolin mim- icked while the nonselective protein kinase inhibitor H89 inhibited mechanical sensitization by CGS21680. CGS21680 did not enhance the response to the purinergic P2X receptor agonist cx,[3-methylene- ATP, indicating that CGS21680 does not nonspecifically sensitize to all stimuli. Mechanical sensitization by CGS21680 was abolished by pretreatment with two structurally different TRPA1 antagonists AP18 and HC030031. Single cell RT-PCR and whole cell patch-clamp studies in isolated esophagus-specific nodose neurons revealed the expression of TRPA1 in A2A-positive C-fiber neurons and demon- strated that CGS21682 potentiated TRPA1 currents evoked by allyl- isothiocyanate. We conclude that stimulation of the adenosine A2A receptor induces mechanical sensitization of nodose C fibers by a mechanism sensitive to TRPA1 antagonists indicating the involve- ment of TRPA1. [ABSTRACT FROM AUTHOR]

Background: Our aim was to summarize our experience with the diagnosis and surgical treatment of solid pseudopapillary neoplasm (SPN) of the pancreas to provide a reference for the management of this rare condition. Methods: We collected and analyzed retrospective data on the clinical presentation, laboratory investigations, radiologic imaging, pathology and operative details of patients with SPN of the pancreas diagnosed between February 2001 and December 2009. Results: In all, 23 of 24 patients were women, and the mean age of all patients was 31 years. The most common clinical presentation was vague abdominal pain. Abdominal imaging showed solid or solid cystic masses in the pancreas, mostly in the tail or head of the gland. All patients were treated surgically. There were no postoperative deaths. After follow-up ranging from 4 to 109 months (median 68 mo), 20 of 22 patients who underwent curative resection were alive with no evidence of disease recurrence. Of the 2 patients with R1 resections, 1 died 42 months after surgery, whereas the other underwent a second operation and was alive after 36 months' follow-up. Conclusion: Solid pseudopapillary neoplasm of the pancreas is a relatively indolent tumour. The initial diagnosis of SPN of the pancreas is suggested by radiologic imaging findings but should be considered in the context of clinical and histopathologic characteristics. We advocate for complete surgical resection once SPN is diagnosed. [ABSTRACT FROM AUTHOR]

Vagal afferent fibres innervating thoracic structures such as the respiratory tract and oesophagus are diverse, comprising several subtypes of functionally distinct C-fibres and A-fibres. Both morphological and functional studies of these nerve subtypes would be advanced by selective, effective and long-term transduction of vagal afferent neurons with viral vectors. Here we addressed the hypothesis that vagal sensory neurons can be transduced with adeno-associated virus (AAV) vectors in vivo, in a manner that would be useful for morphological assessment of nerve terminals, using enhanced green fluorescent protein (eGFP), as well as for the selective knock-down of specific genes of interest in a tissue-selective manner. We found that a direct microinjection of AAV vectors into the vagal nodose ganglia in vivo leads to selective, effective and long-lasting transduction of the vast majority of primary sensory vagal neurons without transduction of parasympathetic efferent neurons. The transduction of vagal neurons by pseudoserotype AAV2/8 vectors in vivo is sufficiently efficient such that it can be used to functionally silence TRPV1 gene expression using short hairpin RNA (shRNA). The eGFP encoded by AAV vectors is robustly transported to both the central and peripheral terminals of transduced vagal afferent neurons allowing for bright imaging of the nerve endings in living tissues and suitable for structure-function studies of vagal afferent nerve endings. Finally, the AAV2/8 vectors are efficiently taken up by the vagal nerve terminals in the visceral tissue and retrogradely transported to the cell body, allowing for tissue-specific transduction. Vagal sensory neurons play a central role in communication between visceral organs and the central nervous system. Research into the physiology of these neurons would be advanced by techniques that could deliver molecular biological tools allowing for the manipulation of selected genes. We investigated whether adeno-associated virus (AAV) vectors can be used to such ends. We found that certain AAV vectors can deliver the gene for green fluorescent protein (eGFP) resulting in robust production of eGFP that is then transported peripherally to the nerve terminals in the visceral organs of innervation and centrally to the brain, where their morphology can readily be visualized. We also found that AAV vectors can effectively deliver shRNA allowing for efficient silencing of genes of interest in the vagal sensory nerves. A combination of AAV transfection with available techniques will probably advance understanding of the structure and function of vagal sensory nerves. [ABSTRACT FROM AUTHOR]

Key points: The action potential initiation in the nerve terminals and its subsequent conduction along the axons of afferent nerves are not necessarily dependent on the same voltage‐gated sodium channel (NaV1) subunits. The action potential initiation in jugular C‐fibres within airway tissues is not blocked by TTX; nonetheless, conduction of action potentials along the vagal axons of these nerves is often dependent on TTX‐sensitive channels. This is not the case for nodose airway Aδ‐fibres and C‐fibres, where both action potential initiation and conduction is abolished by TTX or selective NaV1.7 blockers. The difference between the initiation of action potentials within the airways vs. conduction along the axons should be considered when developing NaV1 blocking drugs for topical application to the respiratory tract. Abstract: The action potential (AP) initiation in the nerve terminals and its subsequent AP conduction along the axons do not necessarily depend on the same subtypes of voltage‐gated sodium channels (NaV1s). We evaluated the role of TTX‐sensitive and TTX‐resistant NaV1s in vagal afferent nociceptor nerves derived from jugular and nodose ganglia innervating the respiratory system. Single cell RT‐PCR was performed on vagal afferent neurons retrogradely labelled from the guinea pig trachea. Almost all of the jugular neurons expressed the TTX‐sensitive channel NaV1.7 along with TTX‐resistant NaV1.8 and NaV1.9. Tracheal nodose neurons also expressed NaV1.7 but, less frequently, NaV1.8 and NaV1.9. NaV1.6 were expressed in ∼40% of the jugular and 25% of nodose tracheal neurons. Other NaV1 α subunits were only rarely expressed. Single fibre recordings were made from the vagal nodose and jugular nerve fibres innervating the trachea or lung in the isolated perfused vagally‐innervated preparations that allowed for selective drug delivery to the nerve terminal compartment (AP initiation) or to the desheathed vagus nerve (AP conduction). AP initiation in jugular C‐fibres was unaffected by TTX, although it was inhibited by NaV1.8 blocker (PF‐01247324) and abolished by combination of TTX and PF‐01247324. However, AP conduction in the majority of jugular C‐fibres was abolished by TTX. By contrast, both AP initiation and conduction in nodose nociceptors was abolished by TTX or selective NaV1.7 blockers. Distinction between the effect of a drug with respect to inhibiting AP in the nerve terminals within the airways vs. at conduction sites along the vagus nerve is relevant to therapeutic strategies involving inhaled NaV1 blocking drugs. [ABSTRACT FROM AUTHOR]

Background: Our aim was to summarize our experience with the diagnosis and surgical treatment of solid pseudopapillary neoplasm (SPN) of the pancreas to provide a reference for the management of this rare condition.Methods: We collected and analyzed retrospective data on the clinical presentation, laboratory investigations, radiologic imaging, pathology and operative details of patients with SPN of the pancreas diagnosed between February 2001 and December 2009.Results: In all, 23 of 24 patients were women, and the mean age of all patients was 31 years. The most common clinical presentation was vague abdominal pain. Abdominal imaging showed solid or solid cystic masses in the pancreas, mostly in the tail or head of the gland. All patients were treated surgically. There were no postoperative deaths. After follow-up ranging from 4 to 109 months (median 68 mo), 20 of 22 patients who underwent curative resection were alive with no evidence of disease recurrence. Of the 2 patients with R1 resections, 1 died 42 months after surgery, whereas the other underwent a second operation and was alive after 36 months' follow-up.Conclusion: Solid pseudopapillary neoplasm of the pancreas is a relatively indolent tumour. The initial diagnosis of SPN of the pancreas is suggested by radiologic imaging findings but should be considered in the context of clinical and histopathologic characteristics. We advocate for complete surgical resection once SPN is diagnosed. [ABSTRACT FROM AUTHOR]

The eight chlorines of C60Cl8 with two pairs of fused pentagons are successfully substituted by sterically encumbering groups through the Friedel–Crafts reaction to get C60(C6H4Me)4Ph4, which provides a facile way for multiple modification of special fullerenes. [ABSTRACT FROM AUTHOR]

A preduodenal position of the portal vein (PDPV) is a very rare congenital anomaly; even rarer is its association with a preduodenal position of the common bile duct (PDCBD). To the seven cases of PDCBD mentioned in the literature, we add this particularly rare case which is associated with multiple abnormalities such as situs inversus totalis, intestinal malrotation, short pancreas, bilobed spleen, accessory spleen, and abnormal ramification of the celiac axis, superior mesenteric artery and renal arteries. Besides describing and illustrating this case, we also discuss the anatomy and embryology of these structures and briefly review the patterns of previously reported cases that we found. We performed an immunohistochemical examination of the pancreas to demonstrate the ventro-dorsal pancreas in our case. For the explanation of the embryology of the PDCBD, the ventro-dorsal pancreas and PDPV malformation, we emphasized the reverse rotation of the ventral pancreas and duodenum. [ABSTRACT FROM AUTHOR]

We dissected 50 head halves of 25 Japanese cadavers (10 males, 15 females) to investigate the innervations of the levator veli palatini (LVP) and superior constrictor pharyngis. The branches supplying the LVP were classified into the following three types according to their origins: supplying branches that originated from the pharyngeal branch of the glossopharyngeal nerve (type I, four sides, 8%), branches that originated from a communicating branch between the pharyngeal branches of the glossopharyngeal and vagus nerves (type II, 36 sides, 72%), and those that originated from the pharyngeal branch of the vagus nerve (type III, 10 sides, 20%). In previous studies, supplying branches of type I were seldom described. Regarding the innervation of the superior constrictor, some variations were observed, and we consider it likely that there is a close relationship between these variations and the type of innervation of the LVP. [ABSTRACT FROM AUTHOR]

The eight chlorines of C60Cl8 with two pairs of fused pentagons are successfully substituted by sterically encumbering groups through the Friedel-Crafts reaction to get the title compound, which provides a facile way for multiple modification of special fullerenes. [ABSTRACT FROM AUTHOR]