Sensory Calcitonin Gene-related Peptide Research Articles

Hydrogen Protons Modulate Perivascular Axo-axonal Interactions in the Middle Cerebral Artery of Rats.

Previous studies have demonstrated that nicotine can induce relaxation of the middle cerebral artery (MCA). However, whether this relaxation is associated with the activity of sensory calcitonin gene-related peptide (CGRP) nerves and whether this is modulated by hydrogen protons (H), facilitating the release of CGRP from sensory CGRPergic nerve terminals in the MCA, remains unclear. In this study, we examined the role of H in the modulation of neurogenic vasomotor responses in the rat-isolated endothelium-denuded MCA. Wire myography was used to measure vasoreactivity and indicated that nicotine-induced relaxation was sensitive to tetrodotoxin and lidocaine and drastically reduced levels of guanethidine (an adrenergic neuronal blocker), N-nitro-L-arginine (L-NNA), CGRP8-37, vasoactive intestinal polypeptide (VIP)6-28, capsaicin, capsazepine (a transient receptor potential vanilloid-1 inhibitor), and tetraethylammonium. However, this nicotine-induced relaxation was not sensitive to propranolol. Lowering the pH of the buffer solution with HCl caused pH-dependent vasorelaxation and deceased intracellular pH in the MCA rings, which was sensitive to L-NNA, CGRP8-37, VIP6-28, capsazepine, 4-aminopyridine (a voltage-gated potassium channel antagonist), and paxilline (a large conductance Ca-activated K channel antagonist). However, HCl-induced relaxation was not inhibited by glibenclamide (an ATP-sensitive K channel blocker). These results suggested that electrical and chemical activation of cerebral perivascular adrenergic nerves led to the release of H, which then facilitated the release of NO, VIP, and CGRP, resulting in vasorelaxation. Lowering the pH of the buffer solution caused potassium channels of vascular smooth muscle cells and perivascular nerves to open. In conclusion, our results demonstrated that H may act as a modulator on MCA perivascular nerves and/or smooth muscles.

Regeneration of adult rat sensory and motor neuron axons through chimeric peroneal nerve grafts containing donor Schwann cells engineered to express different neurotrophic factors

Large peripheral nerve (PN) defects require bridging substrates to restore tissue continuity and permit the regrowth of sensory and motor axons. We previously showed that cell-free PN segments repopulated ex vivo with Schwann cells (SCs) transduced with lentiviral vectors (LV) to express different growth factors (BDNF, CNTF or NT-3) supported the regeneration of axons across a 1 cm peroneal nerve defect (Godinho et al., 2013). Graft morphology, the number of regrown axons, the ratio of myelinated to unmyelinated axons, and hindlimb locomotor function differed depending on the growth factor engineered into SCs. Here we extend these observations, adding more LVs (expressing GDNF or NGF) and characterising regenerating sensory and motor neurons after injection of the retrograde tracer Fluorogold (FG) into peroneal nerve distal to grafts, 10 weeks after surgery. Counts were also made in rats with intact nerves and in animals receiving autografts, acellular grafts, or grafts containing LV-GFP transduced SCs. Counts and analysis of FG positive (+) DRG neurons were made from lumbar (L5) ganglia. Graft groups contained fewer labeled sensory neurons than non-operated controls, but this decrease was only significant in the LV-GDNF group. These grafts had a complex fascicular morphology that may have resulted in axon trapping. The proportion of FG+ sensory neurons immunopositive for calcitonin-gene related peptide (CGRP) varied between groups, there being a significantly higher percentage in autografts and most neurotrophic factor groups compared to the LV-CNTF, LV-GFP and acellular groups. Furthermore, the proportion of regenerating isolectin B4+ neurons was significantly greater in the LV-NT-3 group compared to other groups, including autografts and non-lesion controls. Immunohistochemical analysis of longitudinal graft sections revealed that all grafts contained a reduced number of choline acetyltransferase (ChAT) positive axons, but this decrease was significant only in the GDNF and NT-3 graft groups. We also assessed the number and phenotype of regrowing lumbar FG+ motor neurons in non-lesioned animals, and in rats with autografts, acellular grafts, or in grafts containing SCs expressing GFP, CNTF, NGF or NT-3. The overall number of FG+ motor neurons per section was similar in all groups; however in tissue immunostained for NeuN (expressed in α- but not γ-motor neurons) the proportion of NeuN negative FG+ neurons ranged from about 40–50% in all groups except the NT-3 group, where the percentage was 82%, significantly more than the SC-GFP group. Immunostaining for the vesicular glutamate transporter VGLUT-1 revealed occasional proprioceptive terminals in ‘contact’ with regenerating FG+ α-motor neurons in PN grafted animals, the acellular group having the lowest counts. In sum, while all graft types supported sensory and motor axon regrowth, there appeared to be axon trapping in SC-GDNF grafts, and data from the SC-NT-3 group revealed greater regeneration of sensory CGRP+ and IB4+ neurons, preferential regeneration of γ-motor neurons and perhaps partial restoration of monosynaptic sensorimotor relays.

Calcitonin gene-related peptide down-regulates bleomycin-induced pulmonary fibrosis.

We have found that eIF3a plays an important role in bleomycin-induced pulmonary fibrosis, and up-regulation of eIF3a induced by TGF-β1 is mediated via the ERK1/2 pathway. Whether ERK1/2 - eIF3a signal pathway is involved in calcitonin gene-related peptide (CGRP)-mediated pathogenesis of bleomycin-induced pulmonary fibrosis remains unknown. Pulmonary fibrosis was induced by intratracheal instillation of bleomycin (5 mg/kg) in rats. Primary pulmonary fibroblasts were cultured to investigate the proliferation by BrdU incorporation method and flow cytometry. Sensory CGRP depletion by capsaicin exacerbated bleomycin-induced pulmonary fibrosis in rats, as shown by a significant disturbed alveolar structure, marked thickening of the interalveolar septa and dense interstitial infiltration by inflammatory cells and fibroblasts, accompanied with increased expression of TGF-β1, eIF3a, phosphorylated ERK1/2, α-SMA, collagen I, and collagen III. Exogenous application of CGRP significantly inhibited TGF-β1-induced proliferation and differentiation of pulmonary fibroblasts concomitantly with decreased expression of eIF3a, phosphorylated ERK1/2, α-SMA, collagen I, and collagen III. These effects of CGRP were abolished in the presence of CGRP8-37. These results suggest that endogenous CGRP is related to the development of pulmonary fibrosis induced by bleomycin, and the inhibitory effect of CGRP on proliferation of lung fibroblasts involves the ERK1/2 - eIF3a signaling pathway.

Maintenance of bladder innervation in diabetes: A stereological study of streptozotocin-treated female rats

Neuropathy and cystopathy are two common conditions in patients with chronic diabetes. Despite obvious bladder sensory and motor nerve dysfunction in diabetes, no studies have selectively explored whether sensory or motor innervation is affected in the bladder. In the present study, we tested the hypothesis that loss of bladder sensory and motor fibers is responsible for bladder sensory and motor dysfunction. Parasympathetic and sensory innervation of the bladder dome and neck were examined using immunohistochemistry (IHC) and stereology in adult female rats 12weeks after induction of diabetes by streptozotocin. Naïve and age matched rats were evaluated as controls. Diabetic rats had mean blood glucose level of >400mg/dl, and bladder weights and thicknesses that were more than doubled compared to naïve rats. In naïve rats, parasympathetic vesicular acetylcholine transporter (VAT) and sensory calcitonin gene-related peptide (CGRP) immunopositive nerve fibers were located in bladder smooth muscle and were more densely distributed in the neck compared to the dome. Within the urothelial region, CGRP nerve fibers were densely distributed while VAT nerve fibers were sparsely distributed in the bladder neck and both were virtually absent in the bladder dome. Streptozotocin induced diabetes did not change the total nerve fiber length of either VAT or CGRP stained fibers in either the neck or dome. These studies indicate that hyperglycemia, induced by streptozotocin treatment, does not result in a loss of parasympathetic VAT or CGRP sensory nerve fibers, per se, but the doubling of bladder weight and mass does indicate a decrease in innervation density.

Inhibitory effect of calcitonin gene-related peptide on hypoxia-induced rat pulmonary artery smooth muscle cells proliferation: Role of ERK1/2 and p27

Calcitonin gene-related peptide (CGRP) inhibits angiotensin II-induced proliferation of aortic smooth muscle cells via inactivation of extracellular signal-regulated protein kinase 1/2 (ERK1/2). ERK1/2 is necessary for the degradation or down-regulation of the cell cycle inhibitor p27, and is also crucial in mediating proliferation of pulmonary artery smooth muscle cells (PASMCs). Whether ERK1/2/p27 signal pathway is involved in CGRP-mediated pathogenesis of pulmonary hypertension and vascular remodeling remains unknown. Pulmonary hypertension was induced by hypoxia in rats, and capsaicin (50mg/kg, s.c.) was used to deplete endogenous CGRP. Proliferation of cultured PASMCs was determined by BrdU incorporation method and flow cytometry. The expression/level of CGRP, p27, ERK1/2, c-fos and c-myc was analyzed by radioimmunoassay, immunohistochemistry, real-time PCR or Western blot. Sensory CGRP depletion by capsaicin exacerbated hypoxia-induced pulmonary hypertension in rats, as shown by an increase in right ventricle systolic pressure, mean pulmonary artery pressure and vascular hypertrophy, accompanied with decreased p27 expression and increased expression of phosphorylated ERK1/2, c-fos and c-myc. Exogenous application of CGRP significantly inhibited hypoxia-induced proliferation of PASMCs concomitantly with increased p27 expression and decreased expression of phosphorylated ERK1/2, c-fos and c-myc. These effects of CGRP were abolished in the presence of CGRP8–37. Knockdown of p27 also reversed the inhibitory effect of CGRP on proliferation of PASMCs and expression of c-fos and c-myc, but not on ERK1/2 phosphorylation. These results suggest that CGRP inhibits hypoxia-induced proliferation of PASMCs via ERK1/2/p27/c-fos/c-myc pathway. Down-regulation of CGRP may contribute to remodeling of pulmonary arteries in hypoxia-induced pulmonary hypertension.

Secretagogin is Expressed in Sensory CGRP Neurons and in Spinal Cord of Mouse and Complements other Calcium-Binding Proteins, with a Note on Rat and Human

BackgroundSecretagogin (Scgn), a member of the EF-hand calcium-binding protein (CaBP) superfamily, has recently been found in subsets of developing and adult neurons. Here, we have analyzed the expression of Scgn in dorsal root ganglia (DRGs) and trigeminal ganglia (TGs), and in spinal cord of mouse at the mRNA and protein levels, and in comparison to the well-known CaBPs, calbindin D-28k, parvalbumin and calretinin. Rat DRGs, TGs and spinal cord, as well as human DRGs and spinal cord were used to reveal phylogenetic variations.ResultsWe found Scgn mRNA expressed in mouse and human DRGs and in mouse ventral spinal cord. Our immunohistochemical data showed a complementary distribution of Scgn and the three CaBPs in mouse DRG neurons and spinal cord. Scgn was expressed in ~7% of all mouse DRG neuron profiles, mainly small ones and almost exclusively co-localized with calcitonin gene-related peptide (CGRP). This co-localization was also seen in human, but not in rat DRGs. Scgn could be detected in the mouse sciatic nerve and accumulated proximal to its constriction. In mouse spinal cord, Scgn-positive neuronal cell bodies and fibers were found in gray matter, especially in the dorsal horn, with particularly high concentrations of fibers in the superficial laminae, as well as in cell bodies in inner lamina II and in some other laminae. A dense Scgn-positive fiber network and some small cell bodies were also found in the superficial dorsal horn of humans. In the ventral horn, a small number of neurons were Scgn-positive in mouse but not rat, confirming mRNA distribution. Both in mouse and rat, a subset of TG neurons contained Scgn. Dorsal rhizotomy strongly reduced Scgn fiber staining in the dorsal horn. Peripheral axotomy did not clearly affect Scgn expression in DRGs, dorsal horn or ventral horn neurons in mouse.ConclusionsScgn is a CaBP expressed in a subpopulation of nociceptive DRG neurons and their processes in the dorsal horn of mouse, human and rat, the former two co-expressing CGRP, as well as in dorsal horn neurons in all three species. Functional implications of these findings include the cellular refinement of sensory information, in particular during the processing of pain.

Re-innervation pattern of the ‘neovagina’ created from the bladder flap in patients with Mayer-Rokitanski-Kistner-Hauser syndrome: An immunochemical study

Background. Cystovaginoplasty (CVP) is a method of vaginal reconstruction in women with Mayer-Rokitansky-Kistner-Hauser Syndrome (MRKHS). The neo-vagina allows normal sexual intercourses, but after CVP, the sexual life of patients with MRKHS does not differ significantly from normal females. Therefore, we decided to elucidate the pattern of sensory re-innervation of the bladder flap used for the surgery.Methods. Biopsies were taken from vaginal vestibule and urinary bladder during the CVP and 1 year later in four patients with MRKHS. The following neurotransmitters were studied calcitonin gene-related peptide (CGRP), galanin (GAL), vasoactive intestinal polypeptide (VIP) and pituitary adenylate cyclase-activating peptide (PACAP).Results. CGRP and PACAP nerve fibres were sparse under the urothelium and in submucosal layer of the neovagina, they were more numerous around blood vessels and in the vicinity of smooth muscles. This was similar to the pattern observed in the urinary bladder. VIP- and GAL-positive nerve fibres were most numerous in the submucosa around blood vessels and in smooth muscle bundles of neovagina. They were distinctly less numerous beneath the epithelium. This innervation pattern mimicked that seen in normal vagina and in vaginal vestibule of patients with MRKHS.Conclusions. Our findings demonstrate considerable nervous system plasticity in the bladder flap. Distribution of presumably sensory CGRP and PACAP immunoreactive nerve fibers was similar to the pattern observed within the intact bladder wall, and VIP or GAL immunoreactive nerve fibers (vasomotor functions) were distributed in a manner similar to that observed in the intact vaginal wall.

Quantification of Neuroepithelial Bodies and Their Innervation in Fawn-Hooded and Wistar Rat Lungs

The Fawn-Hooded rat (FHR), a model for primary pulmonary hypertension, shows an unexplained hypersensitivity to airway hypoxia. Because pulmonary neuroepithelial bodies (NEBs) appear to express a functional oxygen-sensing mechanism and an extensive sensory innervation, possible changes in this system should be taken into consideration. In the present study a comparative analysis of NEBs and their selective innervation was performed in FHRs and Wistar control rats. In both rat strains, the number of NEBs was estimated to be around 3,500, approximately 40% of which were innervated by vagal sensory calbindin D28k-immunoreactive (IR) nerve endings and approximately 50% by spinal sensory calcitonin gene-related peptide (CGRP)-IR nerve terminals. The number of intrinsic pulmonary nitrergic neurons and the percentage of pulmonary NEBs revealing a nitrergic innervation were highly significantly lower in FHRs. In both FHRs and Wistar rats, a remarkable morphologic interaction was observed between the intrinsic nitrergic and the CGRP-IR sensory population contacting NEBs. Our findings suggest a possible link between the hypersensitivity to airway hypoxia observed in FHRs and a reduced intrinsic pulmonary nitrergic innervation, possibly via the interaction with pulmonary NEBs and their spinal sensory CGRP-IR innervation.

Capsaicin-sensitive nerve fibers: a potential extra-ACTH mechanism participating in adrenal regeneration in rats.

Pituitary-derived factors, including ACTH, have been widely implicated in initiating adrenal regeneration. However, recent work has demonstrated that adrenal regeneration is also modulated by adrenal nerves that extensively reinnervate the regenerating adrenal. Moreover, transection of the splanchnic nerve removes sensory calcitonin gene-related peptide (CGRP) and preganglionic sympathetic vesicular acetylcholine transporter (VAChT)-positive fibers from the regenerating gland and delays regeneration. However, it is not known whether the splanchnic nerve effects on adrenal regeneration are mediated by the CGRP-positive or VAChT-positive innervation. The present studies use the drug capsaicin, a neurotoxin selective for a subset of primary afferent neurons, to specifically remove CGRP-positive fibers from the adrenal gland and assess subsequent effects on the recovery of adrenal mass and function after surgical enucleation. Male, Sprague-Dawley rats were anesthetized and treated with capsaicin (vs. vehicle) periaxonally to the thoracic splanchnic nerve (33 mM, 15 minutes) or systemically (30-100 mg/kg for 4 days, s.c.). After 7-12 days of recovery, rats received right adrenalectomy and left adrenal enucleation. At 14 and 21 days postenucleation, prestress and poststress plasma and adrenals glands were collected; adrenals were weighed and fixed for immunolabeling of CGRP-positive nerve fibers. Periaxonal capsaicin treatment decreased adrenal CGRP content prior to surgical enucleation; however, reinnervation by CGRP-positive fibers was not prevented and regeneration was not affected. Systemic capsaicin treatment attenuated the reinnervation by CGRP-positive fibers and increased the rate, but not extent, of adrenal regeneration. These results support the hypothesis that adrenal innervation represents an extra-ACTH mechanism to modulate the rate of adrenal regeneration.

Do sensory calcitonin gene-related peptide nerve fibres in the rat pelvic plexus supply autonomic neurons projecting to the uterus and cervix?

Sensory nerve fibres containing calcitonin gene-related peptide (CGRP) innervate neurons of the paracervical ganglion (PCG) in the female rat pelvic plexus. We have combined retrograde tracing with immunocytochemistry to investigate whether CGRP-immunoreactive (-IR) fibres supply neurons targeting the genital tract. Of the total neurons projecting to either the uterine horns or the cervix, 38 and 41% received CGRP-IR innervation, respectively. All these neurons displayed choline acetyltransferase-IR, thus are cholinergic. They were found throughout the PCG and other pelvic plexus ganglia, namely accessory ganglia (AG) and hypogastric plexus (HP). Pelvic nerve section showed that afferent fibres in these nerves provided most of the CGRP-IR fibres supplying uterine- or cervical-related neurons in the PCG/AG, none in HP. It is suggested that such sensory-motor network may provide a local pathway for reflex control of genital tract activity, acting through cholinergic nerve projections.

Calcitonin gene-related peptide gene expression in collagen-induced arthritis is differentially regulated in primary afferents and motoneurons: influence of glucocorticoids

Calcitonin gene-related peptide is involved in peripheral and spinal mechanisms of inflammatory pain. In this paper, we used collagen II-induced arthritis in the rat as a model to investigate the influence of chronic arthritic pain on calcitonin gene-related peptide gene expression in sensory and motor pathways. Additionally, we examined the effect of the glucocorticoid drug budesonide on arthritis-induced changes of calcitonin gene-related peptide expression and constitutive calcitonin gene-related peptide expression. Thirteen days after the immunization with native rat collagen type II rats developed a progressive and chronic polyarthritis which was scored with respect to the degree of swelling and/or redness of the paw and ankle joints. Budesonide significantly attenuated the extent of arthritis. Changes in calcitonin gene-related peptide expression were evaluated by semiquantitative in situ hybridization and immunocytochemistry on day 21 post-immunization. In sensory neurons of dorsal root ganglia of arthritic rats, a significant increase in calcitonin gene-related peptide messenger RNA and protein levels was seen. These increases were completely blocked by budesonide. Also in dorsal root ganglia of non-arthritic rats, budesonide had an effect, with reduced calcitonin gene-related peptide messenger RNA levels below constitutive concentrations. Image analysis of calcitonin gene-related peptide immunoreactivity revealed that changes in calcitonin gene-related peptide expression were due to alterations in calcitonin gene-related peptide expression levels rather than to de novo synthesis or changes in the numbers of calcitonin gene-related peptide expressing neurons. In spinal motoneurons of arthritic rats, marked decreases in calcitonin gene-related peptide messenger RNA and protein levels were measured. These reductions were attenuated by budesonide. The changes in calcitonin gene-related peptide expression in motoneurons correlated with the severity of arthritis in the ipsilateral hind paw. Budesonide had no effects on calcitonin gene-related peptide messenger RNA levels in motoneurons of non-arthritic rats. The opposite regulation of calcitonin gene-related peptide gene expression in primary sensory and spinal somatomotor pathways in collagen-induced arthritis suggests that calcitonin gene-related peptide plays a specific role in both chronic inflammatory pain and arthritis-induced motor dysfunction. The sensitivity of constitutive and inflammation-induced sensory calcitonin gene-related peptide expression to budesonide treatment may indicate that the beneficial effects of steroid treatment in inflammation is partly mediated by down-regulation of calcitonin gene-related peptide in sensory neurons involved in neurogenic inflammation.

5-Hydroxytryptamine 4 receptor agonists initiate the peristaltic reflex in human, rat, and guinea pig intestine

Background & Aims: The peristaltic reflex induced by mucosal stimuli is mediated by intrinsic sensory calcitonin gene-related peptide (CGRP) neurons activated by 5-hydroxytryptamine (5-HT) released from enterochromaffin cells. The involvement of 5-HT 4 receptors was examined with selective 5-HT 4 agonists. Methods: Compartmented intestinal segments were used to measure neurotransmitter release and the mechanical components of the reflex. Results: In human jejunal and rat and guinea pig colonic segments, addition of the 5-HT 4 agonist HTF 919 elicited release of CGRP only into the compartment where the 5-HT 4 agonist was added; vasoactive intestinal peptide (VIP) was released only into the compartment where descending relaxation was measured, and substance P (SP) was released only into the compartment where ascending contraction was measured. The CGRP antagonist hCGRP8-37 inhibited both mechanical responses by 75%–80%. Release of CGRP, VIP, and SP as well as ascending and descending responses were inhibited by selective 5-HT 4 but not by selective 5-HT 3 antagonists. Similar results were obtained with a different 5-HT 4 agonist, R093877. However, HTF 919 was 10–30 times more potent (median effective concentration, ~10 nmol/L for peptide release and 5 nmol/L for mechanical responses) than R093877. Conclusions: Selective 5-HT 4 agonists applied to the mucosa in nanomolar concentrations trigger the peristaltic reflex in human, rat, and guinea pig intestine. GASTROENTEROLOGY 1998;115:370-380

Sensory CGRP depletion by capsaicin exacerbates hypoxia-induced pulmonary hypertension in rats

Pulmonary hypertension is a debilitating disease that occurs among infants and adults. One of many etiologies is airway hypoxia. We previously demonstrated a role of endogenous calcitonin gene-related peptide (CGRP), a potent vasodilator, in ameliorating the pulmonary vascular pressor response to chronic hypoxia and related changes in the lungs and heart. This study evaluates the role of endogenous sensory CGRP in hypoxic pulmonary hypertension and examines the intrinsic neural microcircuitry. Rats were pretreated with capsaicin i.p. to deplete pulmonary sensory C-fiber stores of CGRP and substance P and placed in hypobaric hypoxia (10% O 2, 16 days) or normoxia together with sham controls. Hypoxia increased pulmonary artery pressure, right-ventricular weight, arterial medial thickness, elasticized capillaries, endothelial cell density, lung water and hematocrit in control rats. Capsaicin augmented pulmonary artery pressure and right-ventricular hypertrophy in hypoxia, and medial thickness and endothelial cell density both in normoxia and hypoxia. Because of the limited effects on these parameters by substance P and other capsaicin-sensitive lung agents, our results suggest that sensory CGRP deficit severely exacerbates pathological signs of hypoxic pulmonary hypertension. A neural microcircuitry consistent with an axon reflex pathway is outlined histochemically. We conclude that endogenous CGRP modulates pulmonary vascular tone in hypoxic pulmonary hypertension which requires intact primary sensory fibers.

Postnatal development of the autonomic and sensory innervation of the musculature in the rat urinary bladder

The postnatal development of the innervation of the muscle layer in the rat urinary bladder was analysed in whole mount preparations using immunohistochemistry against protein gene-product 9.5 (PGP; general neuronal marker), growth-associated protein 43 (GAP), dopamine β-hydroxylase (DBH), neuropeptide Y (NPY), vasoactive intestinal polypeptide (VIP), calcitonin gene-related peptide (CGRP) and substance P (SP). Immunoreactive nerve fibres for all markers were already present at birth. The density of PGP- and GAP-positive nerve fibres was similar and remained constant throughout the postnatal development. The rank order of densities for the other markers relative to PGP was NPY (129–189%)>CGRP (20–63%)>SP (7–23%)>DBH (7–12%)>VIP (2–11%). While the density of presumably efferent VIP- and DBH-positive fibres did not change postnatally, NPY-positive fibres reached adult density at the fifth postnatal day. Sensory CGRP- and SP-positive nerve fibres approached adult levels at the end of the second week, shortly before the micturition reflex was completely developed. The data suggest that a sufficient relative density of sensory and certain efferent elements might be a prerequisite for the development of the mature micturition reflex.

Distinct 5-HT receptors mediate the peristaltic reflex induced by mucosal stimuli in human and guinea pig intestine

BACKGROUND & AIMS: The role of 5-hydroxytryptamine (5-HT) in regulating the peristaltic reflex in humans is unknown. The neural pathways subserving peristalsis induced by mucosal stimulation were characterized in human jejunum and guinea pig colon. METHODS: Compartmented flat-sheet preparations that enable measurement of 5-HT and sensory transmitter release into one compartment and mechanical responses in adjacent compartments were used. RESULTS: Mucosal stimuli (2-8 brush strokes) caused concomitant release of 5-HT and calcitonin gene-related peptide (CGRP) into the compartment where stimulation was applied in both species; in contrast, muscle stretch caused release of CGRP only. CGRP release as well as ascending contraction and descending relaxation of circular muscle induced by mucosal stimulation were inhibited by a selective 5-HT4 antagonist in human jejunum and by selective 5-HT4 and 5-HT3 antagonists in guinea pig colon. The effects of the 5-HT3 and 5-HT4 antagonists in guinea pig colon were additive. A selective 5-HT1P antagonist mimicked the effect of the 5-HT4 antagonist. The CGRP antagonist human CGRP8-37 inhibited ascending and descending responses in both species. CONCLUSIONS: 5-HT released by mucosal stimulation initiates a peristaltic reflex by activating 5- HT4/5-HT1P receptors on sensory CGRP neurons in human intestine and 5- HT4/5-HT1P and 5-HT3 receptors in guinea pig colon. (Gastroenterology 1996 Nov;111(5):1281-90)

5-HT released by mucosal stimuli initiates peristalsis by activating 5-HT4/5-HT1p receptors on sensory CGRP neurons.

The intestinal peristaltic reflex can be elicited by mucosal stimulation or circular muscle stretch. Muscle stretch activates extrinsic, whereas mucosal stimulation activates intrinsic calcitonin gene-related peptide (CGRP)-containing sensory neurons. The present study examined the role of 5-hydroxytryptamine (5-HT) in sensory transmission. A three-compartment preparation of rat colon was used that enables separate measurement of sensory transmitters and modulators. Mucosal stimuli (2-8 brush strokes) caused concurrent increase in 5-HT and CGRP release in proportion to the intensity of stimulation. Release of both 5-HT and CGRP occurred exclusively into the central compartment where the stimuli were applied. Exogenous 5-HT caused a concentration-dependent release of CGRP. Release of CGRP induced by exogenous 5-HT or mucosal stimulation was inhibited by selective 5-HT4 and 5-HT1p antagonists but was not affected by 5-HT1A, 5-HT2, and 5-HT3 antagonists. Ascending contraction and descending relaxation of circular muscle measured in the peripheral orad and caudad compartments, respectively, were also selectively inhibited by 5-HT4 and 5-HT1p antagonists added to the central but not peripheral compartments. In contrast, muscle stretch elicited CGRP but not 5-HT release; the ascending contraction and descending relaxation components of the peristaltic reflex induced by muscle stretch were not affected by 5-HT antagonists. We conclude that 5-HT released by mucosal stimulation initiates the peristaltic reflex by activating 5-HT4/5-HT1p receptors on sensory CGRP-containing neurons.

Neonatal capsaicin-treatment in mice: effects on pancreatic peptidergic nerves and 2-deoxy- d-glucose-induced insulin and glucagon secretion

It is not known whether sensory nerves are involved in the insulin, glucagon or glucose responses to autonomic nerve activation induced by 2-deoxy- d-glucose (2-DG). We therefore treated mice neonatally with capsaicin which permanently destroys sensory afferent nerve fibers. Immunohistochemistry of the pancreas at 13–14 weeks of age revealed a substantial reduction of calcitonin gene-related peptide (CGRP)-immunoreactive nerves and a partial reduction of substance P-immunoreactive nerves. In contrast, no effect was observed on galanin-immunoreactive nerves. At age 10–12 weeks, the mice were injected intravenously with 2-DG (500 mg/kg). In controls, 2-DG stimulated insulin and glucagon secretion and induced hyperglycemia ( P < 0.01). Capsaicin treatment partially reduced the glucose and glucagon responses to 2-DG ( P < 0.01). In contrast, the insulin response to 2-DG was not affected by capsaicin. It is concluded that the mouse pancreas contains capsaicin-sensitive sensory CGRP- and substance P-immunoreactive nerve fibers, whereas the galanin-immunoreactive nerve fibers are not sensitive to capsaicin. Furthermore, capsaicin-sensitive sensory nerve fibers are partially involved in 2-DG-induced glucagon secretion and hyperglycemia, whereas sensory nerves are not involved in 2-DG-induced insulin secretion.

Calcitonin gene-related peptide and substance P in afferents to the upper gastrointestinal tract in the rat

Combined immunohistochemistry and retrograde tracing was used to investigate the origin of the sensory calcitonin gene-related peptide (CGRP) innervation of the rat stomach. Up to 85% of spinal gastric afferents (T 6-L 1) contained CGRP immunoreactivity compared with <6% of vagal gastric afferents. By comparison substance P immunoreactivity occurred in about 50% of spinal gastric afferents and less than 2% of vagal afferents. The vagal afferents to the oesophagus were 14 and 26% substance P- and CGRP-immunoreactive respectively. The results suggest an important role for CGRP in gastric spinal afferents.