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C Sternini

Publications and source records attributed to C Sternini.

At least 19 recordsLinked to original sources
PubMed2006

Role of carbon monoxide in electrically induced non-adrenergic, non-cholinergic relaxations in the guinea-pig isolated whole trachea.

BACKGROUND AND PURPOSE: Nitric oxide (NO) and vasoactive intestinal peptide (VIP) are considered transmitters of non-adrenergic, non-cholinergic (NANC) relaxations in guinea-pig trachea, whereas the role of carbon monoxide (CO) is unknown. This study was designed to assess the participation of CO, and to investigate the localization of haem oxygenase-2 (HO-2), the CO-producing enzyme, in tracheal neurons. EXPERIMENTAL APPROACH: NANC responses to electrical field stimulation (EFS) at 3 and 10 Hz were evaluated in epithelium-free whole tracheal segments as intraluminal pressure changes. Drugs used were: L-nitroarginine methyl ester (L-NAME, 100 microM) to inhibit NO synthase (NOS), alpha-chymotrypsin (2 U ml(-1)) to inactivate VIP, zinc protoporphyrin-IX (ZnPP-IX, 10 microM) to inhibit HO-2, and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 10 microM), a soluble guanylyl cyclase inhibitor. For immunohistochemistry, tissues were exposed to antibodies to PGP 9.5, a general neuronal marker, HO-2 and NOS, and processed with an indirect immunofluorescence method. KEY RESULTS: alpha-Chymotrypsin did not affect NANC relaxations. ODQ inhibited NANC responses by about 60%, a value similar to that obtained by combining L-NAME and ZnPP-IX. The combination of ODQ, L-NAME and ZnPP-IX reduced the responses by 90%. Subpopulations of HO-2 positive neurons containing NOS were detected in tracheal sections. CONCLUSIONS AND IMPLICATIONS: In the guinea-pig trachea, NANC inhibitory responses at 3 and 10 Hz use NO and CO as main transmitters. Their participation is revealed following inhibition of NOS, HO-2 and soluble guanylyl cyclase. The involvement of CO as a relaxing transmitter paves the way for novel therapeutic approaches in the treatment of airway obstruction.

Animals↗
PubMed2005

Effect of N-methyl-d-aspartate receptor blockade on neuronal plasticity and gastrointestinal transit delay induced by ischemia/reperfusion in rats.

Intestinal ischemia impairs gastrointestinal motility. The aims of this study were to investigate the effect of intestinal ischemia on gastrointestinal transit and on the expression of enteric transmitters in the rat, and whether the glutamate N-methyl-d-aspartate receptors influence these effects. Ischemia (1 h), induced by occluding the superior mesenteric artery, was followed by 0 or 24 h of reperfusion. Normal and sham-operated rats served as controls. Serosal blood flow was measured with laser Doppler flow meter. Gastrointestinal transit was measured as time of appearance of a marker in fecal pellets. Immunohistochemistry was used to evaluate the number of neurons immunoreactive for neuronal nitric oxide synthase (NOS) or vasoactive intestinal polypeptide and the density of substance P immunoreactive fibers in the myenteric plexus. The N-methyl-d-aspartate receptors antagonist, (+)-5-methyl-10,11-dihydro-5HT-[a,b] cyclohepten-5,10-imine (MK-801) (1 mg/kg i.v.) or the NOS inhibitor, N-nitro-l-arginine (10 mg/kg i.v.) was administered prior to ischemia. Serosal blood flow was decreased by 70% during ischemia, but it was not altered in sham-operated rats. Gastrointestinal transit was significantly prolonged in ischemic/reperfused rats compared with controls. There was a significant increase in the number of vasoactive intestinal polypeptide and neuronal nitric oxide synthase immunoreactive neurons, and a marked decrease of substance P immunoreactive fibers in ischemia followed by 24 h of reperfusion animals compared with controls. These alterations were not observed in ischemia without reperfusion. A significant delay of gastrointestinal transit and increase of vasoactive intestinal polypeptide neurons were also observed in sham-operated rats. The changes in transmitter expression and gastrointestinal transit in ischemic/reperfused rats were prevented by pre-treatment with the NOS inhibitor, N-nitro-l-arginine or the N-methyl-d-aspartate receptors antagonist, MK-801. This study suggests an involvement of the glutamatergic system and its interaction with nitric oxide in intestinal ischemia/reperfusion. Ischemia/reperfusion might induce local release of glutamate that activates N-methyl-d-aspartate receptors leading to increased production of nitric oxide and adaptive changes in enteric transmitters that might contribute to gastrointestinal dysmotility.

Analysis of Variance↗
PubMed2005

Identification of galanin receptor 1 on excitatory motor neurons in the guinea pig ileum.

Exogenously administered galanin inhibits cholinergic transmission to the longitudinal muscle and reduces peristaltic efficiency in the guinea pig ileum with a mechanism partially mediated by galanin receptor 1 (GAL-R1). We investigated the effect of exogenous galanin 1-16, which has high affinity for GAL-R1, on the ascending excitatory reflex of the circular muscle elicited by radial distension in isolated segments of guinea pig ileum. We used a three-compartment bath that allows dissecting the ascending pathway into the oral (site of excitatory motor neurons), intermediate (site of ascending interneurons) and caudal compartment (site of intrinsic primary afferent neurons). Galanin 1-16 (0.3-3 micromol L(-1)) applied to the oral compartment inhibited in a concentration-dependent manner the ascending excitatory reflex elicited by the wall distension in the caudal compartment. This effect was antagonized by the GAL-R1 antagonist, RWJ-57408 (1 and 10 micromol L(-1)). By contrast, galanin 1-16 was ineffective when added to the intermediate or caudal compartment up to 3 micromol L(-1). GAL-R1 immunoreactive neurons did not contain neuron-specific nuclear protein, a marker for intrinsic primary afferent neurons. These findings indicate that GAL-R1s are present on motor neurons responsible for the ascending excitatory reflex, but not on ascending interneurons and intrinsic primary afferent neurons.

Animals↗
PubMed2004

Abdominal surgery induces mu opioid receptor endocytosis in enteric neurons of the guinea-pig ileum.

Immunohistochemistry and confocal microscopy were used to investigate mu opioid receptor (muOR) internalization in enteric neurons of the guinea-pig ileum following abdominal surgery. The following surgical procedures were performed under halothane or isofluorane anesthesia: a) midline abdominal skin incision, b) laparotomy or c) laparotomy with intestinal manipulation. Gastrointestinal transit was evaluated by using a non-absorbable marker and measuring fecal pellet output. In neurons from normal and control (anesthesia alone) animals, muOR was predominantly at the cell surface. muOR endocytosis following skin incision was not significantly different from controls (21.2+/-3.5% vs. 13.7+/-2.1%, mean+/-S.E.M.), whereas it was significantly increased by laparotomy (46.5+/-6.1%; P<0.01 vs. controls) or laparotomy plus intestinal manipulation (40.5+/-6.1%; P<0.01 vs. controls) 30 min following surgery compared with controls. muOR endocytosis remained elevated at 4 h (38.6+/-1.2%; P<0.01 vs. controls), whereas it was similar to controls at 6 and 12 h (17.5+/-5.8% and 11.2+/-3.0%). muOR endocytosis occurred in cholinergic and nitrergic neurons. Gastrointestinal transit was significantly delayed by laparotomy or laparotomy plus intestinal manipulation (12.8+/-1.2 and 13.8+/-0.6 h vs. 7.0+/-0.5 in controls; P<0.01), but was not significantly changed by skin incision (8.2+/-0.6 h). The findings of the present study support the concept that the noxious stimulation caused by abdominal surgery induces release of endogenous opioids thus resulting in muOR endocytosis in neurochemically distinct enteric neurons. muOR internalization can serve as indirect evidence of opioid release and as a means to visualize neuronal pathways activated by opioids.

Abdomen↗
PubMed2004

Role of galanin receptor 1 in peristaltic activity in the guinea pig ileum.

Galanin effects are mediated by distinct receptors, galanin receptor 1 (GAL-R1), GAL-R2 and GAL-R3. Here, we analyzed 1) the role of GAL-R1 in cholinergic transmission and peristalsis in the guinea-pig ileum using longitudinal muscle-myenteric plexus preparations and intact segments of the ileum in organ bath, and 2) the distribution of GAL-R1 immunoreactivity in the myenteric plexus with immunohistochemistry and confocal microscopy. Galanin inhibited electrically stimulated contractions of longitudinal muscle-myenteric plexus preparations with a biphasic curve. Desensitization with 1 microM galanin suppressed the high potency phase of the curve, whereas the GAL-R1 antagonist, RWJ-57408 (1 microM), inhibited the low potency phase. Galanin (3 microM) reduced the longitudinal muscle contraction and the peak pressure, and decreased the compliance of the circular muscle. All these effects were antagonized by RWJ-57408 (1 or 10 microM). RWJ-57408 (10 microM) per se did not affect peristalsis parameters in normal conditions, nor when peristalsis efficiency was reduced by partial nicotinic transmission blockade with hexamethonium. In the myenteric plexus, GAL-R1 immunoreactivity was localized to neurons and to fibers projecting within the plexus and to the muscle. GAL-R1 was expressed mostly by cholinergic neurons and by some neurons containing vasoactive intestinal polypeptide or nitric oxide synthase. This study indicates that galanin inhibits cholinergic transmission to the longitudinal muscle via two separate receptors; GAL-R1 mediates the low potency phase. The reduced peristalsis efficiency could be explained by inhibition of the cholinergic drive, whereas the decreased compliance is probably due to inhibition of descending neurons and/or to the activation of an excitatory muscular receptor. Endogenous galanin does not appear to affect neuronal pathways subserving peristalsis in physiologic conditions via GAL-R1.

Acetylcholine↗
PubMed2004

Role of galanin receptor 1 in gastric motility in rat.

Galanin actions are mediated by distinct galanin receptors (GAL-R), GAL-R1, -R2 and -R3. We investigated the role of GAL-R1 in gastric motility and the expression of GAL-R1 in the rat stomach. In vivo, in urethane-anaesthetized rats, galanin (equipotent for all GAL-Rs) induced a short inhibition of gastric motility, followed by increase in tonic and phasic gastric motility; the latter was significantly reduced by the GAL-R1 antagonist, RWJ-57408. Galanin 1-16 (high affinity for GAL-R1 and -R2) induced a long-lasting decrease of intragastric pressure, which was not modified by RWJ-57408. In vitro, galanin and galanin 1-16 induced increase of intragastric pressure that was not affected by RWJ-57408. Tetrodotoxin (TTX) did not suppress the galanin excitatory effect, whereas the effect of galanin 1-16 on gastric contraction was increased by TTX- or N-nitro-L-arginine, an inhibitor of nitric oxide synthase. GAL-R1 immunoreactivity was localized to cholinergic and tachykinergic neurons and to neurons immunoreactive for nitric oxide synthase or vasoactive intestinal polypeptide. This study suggests that an extrinsic GAL-R1 pathway mediates the galanin excitatory action, whereas an extrinsic, non GAL-R1 pathway is likely to mediate the galanin inhibitory effect in vivo. GAL-R1 intrinsic neurons do not appear to play a major role in the control of gastric motility.

Animals↗
PubMed2004

The opioid system in the gastrointestinal tract.

Mu-, delta- and kappa-opioid receptors (ORs) mediate the effects of endogenous opioids and opiate drugs. Here we report (1) the distribution of muOR in the guinea-pig and human gastrointestinal tract in relation to endogenous ligands, to functionally distinct structures in the gut and to deltaOR and kappaOR; and (2) the ligand-induced muOR endocytosis in enteric neurones using in vitro and in vivo models. In the guinea pig, muOR immunoreactivity is confined mainly to the myenteric plexus. MuOR myenteric neurones are most numerous in the small intestine, followed by the stomach and the proximal colon. MuOR immunoreactive fibres are dense in the muscle layer and the deep muscular plexus, where they are in close association with interstitial cells of Cajal. This distribution closely matches the pattern of enkephalin. MuOR enteric neurones comprise functionally distinct populations of neurones of the ascending and descending pathways of the peristaltic reflex. In human gut, muOR immunoreactivity is localized to myenteric and submucosal neurones and to immune cells of the lamina propria. DeltaOR immunoreactivity is located in both plexuses where it is predominantly in varicose fibres in the plexuses, muscle and mucosa, whereas kappaOR immunoreactivity appears to be confined to the myenteric plexus and to bundles of fibres in the muscle. MuOR undergoes endocytosis in a concentration-dependent manner, in vitro and in vivo. Pronounced muOR endocytosis is observed in neurones from animals that underwent abdominal surgery that has been shown to induce delay in gastrointestinal transit. We can conclude that all three ORs are localized to the enteric nervous system with differences among species, and that muOR endocytosis can be utilized as a means to visualize enteric neurones activated by opioids and sites of opioid release.

Adult↗
PubMed2003

Ligand-induced mu opioid receptor endocytosis and recycling in enteric neurons.

Immunohistochemistry and confocal microscopy were used to investigate endocytosis and recycling of the native mu opioid receptor (muOR) in enteric neurons. Isolated segments of the guinea-pig ileum were exposed to increasing concentrations of muOR agonists at 4 degrees C to allow ligand binding and warming to 37 degrees C for 0 min (baseline) to 6 h in ligand-free medium to allow receptor internalization and recycling. The endogenous ligand, [Met]enkephalin, and [D-Ala(2),MePhe(4),Gly-ol(5)] enkephalin (DAMGO), an opioid analog, and the alkaloids, etorphine and fentanyl, induced rapid internalization of muOR immunoreactivity in enteric neurons, whereas morphine did not. muOR internalization was prevented by muOR antagonists. Basal levels of muOR immunoreactivity in the cytoplasm were 10.52+/-2.05%. DAMGO (1 nM-100 microM) induced a concentration-dependent increase of muOR immunofluorescence density in the cytoplasm to a maximum of 84.37+/-2.26%. Translocation of muOR immunoreactivity in the cytoplasm was detected at 2 min, reached the maximum at 15-30 min, remained at similar levels for 2 h, began decreasing at 4 h, and was at baseline values at 6 h. A second exposure to DAMGO (100 nM) following recovery of internalized muOR immunoreactivity at the cell surface induced a translocation of muOR immunoreactivity in the cytoplasm comparable to the one observed following the first exposure (46.89+/-3.11% versus 43.31+/-3.80%). muOR internalization was prevented by hyperosmolar sucrose, phenylarsine oxide or potassium depletion, which inhibit clathrin-mediated endocytosis. muOR recycling was prevented by pre-treatment with bafilomycin A1, an acidotropic agent that inhibits endosomal acidification, but not by the protein synthesis inhibitor, cycloheximide. This study shows that native muOR in enteric neurons undergoes ligand-selective endocytosis, which is primarily clathrin-mediated, and recycles following endosomal acidification. Following recycling, muOR is activated and internalized by DAMGO indicating that recycled receptors are functional.

Analgesics, Opioid↗
PubMed2001

Dynamin and Rab5a-dependent trafficking and signaling of the neurokinin 1 receptor.

Understanding the molecular mechanisms of agonist-induced trafficking of G-protein-coupled receptors is important because of the essential role of trafficking in signal transduction. We examined the role of the GTPases dynamin 1 and Rab5a in substance P (SP)-induced trafficking and signaling of the neurokinin 1 receptor (NK1R), an important mediator of pain, depression, and inflammation, by studying transfected cells and enteric neurons that naturally express the NK1R. In unstimulated cells, the NK1R colocalized with dynamin at the plasma membrane, and Rab5a was detected in endosomes. SP induced translocation of the receptor into endosomes containing Rab5a immediately beneath the plasma membrane and then in a perinuclear location. Expression of the dominant negative mutants dynamin 1 K44E and Rab5aS34N inhibited endocytosis of SP by 45 and 32%, respectively. Dynamin K44E caused membrane retention of the NK1R, whereas Rab5aS34N also impeded the translocation of the receptor from superficially located to perinuclear endosomes. Both dynamin K44E and Rab5aS34N strongly inhibited resensitization of SP-induced Ca(2+) mobilization by 60 and 85%, respectively, but had no effect on NK1R desensitization. Dynamin K44E but not Rab5aS34N markedly reduced SP-induced phosphorylation of extracellular signal regulated kinases 1 and 2. Thus, dynamin mediates the formation of endosomes containing the NK1R, and Rab5a mediates both endosomal formation and their translocation from a superficial to a perinuclear location. Dynamin and Rab5a-dependent trafficking is essential for NK1R resensitization but is not necessary for desensitization of signaling. Dynamin-dependent but not Rab5a-dependent trafficking is required for coupling of the NK1R to the mitogen-activated protein kinase cascade. These processes may regulate the nociceptive, depressive, and proinflammatory effects of SP.

Animals↗
PubMed2001

Expression of the human insulin gene in the gastric G cells of transgenic mice.

The goal of this study was to engineer gastrin-producing G cells of the gastric antrum to produce insulin. A pGas-Ins chimeric gene in which the gastrin promoter drives expression of the human insulin gene was constructed and was validated by transient transfection of GH4 and AGS cells. RT-PCR analysis and sequencing revealed three forms of differentially spliced insulin mRNA in GH4 cells transiently transfected by pGas-Ins. Gas-Ins transgenic mice were generated utilizing this chimeric gene. Northern blot analysis, in situ hybridization, and immunohistochemistry demonstrated expression of the human insulin gene specifically in antral G cells. Northern blot analysis demonstrated that the shortest of the insulin mRNA three forms is predominantly expressed in stomach tissue. RT-PCR analysis also showed expression of the transgene in colon, pancreas, and brain tissues that was undetectable by northern analysis. We conclude that gastrin promoter can be used for targeting expression of human insulin to antral G cells and that antral G cells can express human insulin. Further refining of the chimeric gene design is required to enhance expression.

Animals↗
PubMed2001

Tachykinin-dependent and -independent components of peristalsis in the guinea pig isolated distal colon.

BACKGROUND & AIMS: In the intestine, tachykinins regulate motility by participating in neuromuscular and neuro-neuronal transmission. The aim of this study was to test the hypothesis that colonic propulsion is regulated by an interplay between tachykinergic and cholinergic transmission. METHODS: Propulsion was elicited by intraluminal distention of a thin rubber balloon, which traveled from the oral to the anal end of guinea pig isolated distal colon segments. The overall contribution of endogenous tachykinins to colonic propulsion was examined by blocking NK1, NK2, and NK3 receptors simultaneously. RESULTS: NK2-receptor blockade by MEN 11420 inhibited propulsion, whereas blockade of NK(1) by SR 140333 or of NK3 receptors by SR 142801 had minor effects on motility. Blockade of muscarinic or nicotinic receptors by hyoscine or hexamethonium decelerated peristalsis up to propulsion arrest. In the presence of partial muscarinic receptor blockade, the NK1-receptor antagonist SR 140333 and the NK2-receptor antagonist MEN 11420 markedly inhibited propulsion. Propulsion was also inhibited by the NK3-receptor antagonist SR 142801 in the presence of partial nicotinic receptor blockade. The simultaneous administration of the 3 tachykinin antagonists inhibited propulsion by 50%. CONCLUSIONS: This study demonstrates the existence of an interplay between tachykinergic and cholinergic pathways during peristalsis and the importance of endogenous tachykinins acting at multiple receptor sites in the control of colonic propulsion.

Animals↗
PubMed2001

Monoclonal antibody to rat galanin: production, characterization, and in vivo immunoneutralization activity.

A monoclonal antibody (MAb) to galanin was prepared by cell fusion of myeloma Fox-NY and spleen cells from Robertsonian mice immunized with rat galanin. Hybridomas producing high-affinity antibodies were cloned in pristine-primed Balb/c mice. The antibody was purified by affinity chromatography and concentrated to 12 mg IgG/mL by dialysis. Immunoreactivity of the antibody was screened by radioimmunoassay. Ascites fluid contained approximately 10 mg/mL IgG that belong to the subclass of IgG2a as determined by enzyme-linked immunoadsorbent assay (ELISA). The titer of this IgG2a antibody entitled #G65G was 1:10,000 and the ID50 for rat galanin was 1000 fmol/mL as determined by liquid phase radioimmunoassay. Immunohistochemistry showed that this galanin MAb stains densed, beaded processes distributed to the enteric plexuses, where they appear to encircle neuronal cell bodies, to the muscle layer, where they are particularly abundant in the circular muscle layer and in the deep muscular layer, and to the mucosa. In vivo capacity of immunoneutralization by this antibody was tested in male Sprague-Dawley rats fasted for 24 h and anesthetized with urethane. Systemic injection of protein A purified galanin antibody (6 mg/rat) decreased by 70% of the inhibitory effect of intravenous galanin (2 nmol/kg/h i.v.) on gastric acid secretion induced by intracisternal TRH analog. These results show that galanin antibody #G65G is useful for in vivo immunoneutralization of galanin effects and is a valuable tool for immunohistochemical localization of galanin in gastrointestinal tissues.

Animals↗
PubMed2001

Receptors and transmission in the brain-gut axis: potential for novel therapies. III. Mu-opioid receptors in the enteric nervous system.

G protein-coupled receptors are cell surface signal-transducing proteins, which elicit a variety of biological functions by the activation of different intracellular effector systems. Many of these receptors, including the mu-opioid receptor (mu OR), have been localized in the gastrointestinal tract. mu OR is the target of opioids and alkaloids, potent analgesic drugs with high potential for abuse. mu OR is expressed by enteric neurons, and it undergoes ligand-selective endocytosis. It is of clinical importance because it mediates tolerance and other major side effects of opiate analgesics, including impairment of gastrointestinal propulsion. An important observation of mu OR is its differential trafficking and desensitization properties in response to individual agonists, which might have long-term physiological consequences and be involved in the development of opiate side effects. Receptor activation by agonists is the basis for signaling, and alterations of the mechanisms controlling cellular responses of G protein-coupled receptors to agonists might be the basis of several diseases, including gastrointestinal diseases. Therefore, understanding these basic cellular mechanisms is important for developing appropriate therapeutic agents.

Animals↗
PubMed2000

Neurotrophin-3 and neurotrophin receptor immunoreactivity in peptidergic enteric neurons.

In the rat small intestine, neurotrophin-3 immunoreactivity was identified in ganglion cells and in processes mostly innervating the mucosa and occasionally the muscle layer and vasculature. The vast majority of neurotrophin-3 immunoreactive neurons contained vasoactive intestinal polypeptide (VIP), but not substance P or related tachykinin (SP/TK). Neurotrophin receptors visualized by pan-trk immunoreactivity were found in numerous ganglion cells of both plexuses and in nerve processes in the intestinal wall. Pan-trk submucosal neurons contained VIP (36%) or SP/TK-IR (47%). Pan-trk myenteric neurons contained VIP-IR (57%) or SP/TK (27%). Our data suggest that neurotrophin-3 and neurotrophin receptors may be involved in the maintenance of enteric neuronal circuits, transmission and phenotypic expression.

Animals↗
PubMed2000

Role of agonist-dependent receptor internalization in the regulation of mu opioid receptors.

Organotypic cultures and ileal neuromuscular preparations were used to determine (i) whether endogenous release of opioids by electrical stimulation induces mu receptor endocytosis, and (ii) whether and under which conditions ligand-induced mu receptor endocytosis influences the responsiveness of neurons expressing native mu receptors. In longitudinal muscle-myenteric plexus preparations, electrical stimulation at 20 Hz induced a prominent endocytosis of mu receptors in enteric neurons, indicating endogenous release of opioids. A similar massive endocytosis was triggered by exogenous application of the mu receptor agonist, [D-Ala(2),MePhe(4), Gly-ol(5)] enkephalin, whereas exogenous application of morphine was ineffective. [D-Ala(2),MePhe(4),Gly-ol(5)] enkephalin and morphine induced a concentration-dependent inhibition of neurogenic cholinergic twitch contractions to electrical stimulation at 0.1 Hz. beta-Chlornaltrexamine shifted to the right the inhibitory curve of both agonists with a concentration-dependent reduction of the maximum agonist response, which is consistent with the existence of spare mu opioid receptors. Under these conditions, the induction of mu receptor endocytosis by exogenously applied [D-Ala(2), MePhe(4),Gly-ol(5)] enkephalin diminished the inhibitory effect of this agonist on twitch contractions and tritiated acetylcholine release. In contrast, there was no reduction of the inhibitory effect of morphine, which failed to induce mu receptor endocytosis, on neurogenic cholinergic response. These results provide the first evidence for the occurrence of mu receptor endocytosis in neurons by endogenously released opioids and show that agonist-dependent mu receptor endocytosis could serve as a mechanism to regulate mu opioid receptor responsiveness to ligand stimulation when the opioid receptor reserve is reduced.

Acetylcholine↗
PubMed2000

Etorphine increases the number of mu-opioid receptor-positive cells in the cerebral cortex.

Imunocytochemical techniques were used to determine whether agonist-induced activation of mu-opioid receptors alters the number and distribution of mu-opioid receptor-positive cells in the rat cerebral cortex. In untreated rats, mu-opioid receptor immunoreactivity was localized to neuronal perikarya and dendrites and to neuropilar punctate structures. mu-Opioid receptor-positive neurons were mostly in layers II and III and exhibited a bipolar or bitufted morphology. In rats treated with the mu-opioid receptor agonist etorphine (0.1mg/kg intraperitoneally) and perfused after different survival periods, there was an enhancement of immunostaining for mu-opioid receptors observed at 15min, reaching a maximum at 60min, and which returned to normal at 480min. Etorphine-induced effects included an increase in the intensity of cellular and neuropil staining; statistical analysis showed that the number of mu-opioid receptor-positive cells in etorphine-treated groups was significantly higher than in controls or saline-treated rats. In animals that received both etorphine and the mu-opioid receptor antagonist naloxone, the pattern of mu-opioid receptors immunoreactivity was similar to that of untreated animals. This study shows that the number of mu-opioid receptor-positive cells is significantly increased following etorphine treatment and suggests that agonist treatment may be exploited to increased immunostaining of mu-opioid receptors and also of other G-protein coupled receptors.

Animals↗
PubMed2000

Role of nitric oxide- and vasoactive intestinal polypeptide-containing neurones in human gastric fundus strip relaxations.

The morphological pattern and motor correlates of nitric oxide (NO) and vasoactive intestinal polypeptide (VIP) innervation in the human isolated gastric fundus was explored. By using the nicotinamide adenine dinucleotide phosphate hydrogen (NADPH)-diaphorase and specific rabbit polyclonal NO-synthase (NOS) and VIP antisera, NOS- and VIP-containing varicose nerve fibres were identified throughout the muscle layer or wrapping ganglion cell bodies of the myenteric plexus. NOS-immunoreactive (IR) neural cell bodies were more abundant than those positive for VIP-IR. The majority of myenteric neurones containing VIP coexpressed NADPH-diaphorase. Electrical stimulation of fundus strips caused frequency-dependent NANC relaxations. N(G)-nitro-L-arginine (L-NOARG: 300 microM) enhanced the basal tone, abolished relaxations to 0.3 - 3 Hz (5 s) and those to 1 Hz (5 min), markedly reduced ( approximately 50%) those elicited by 10 - 50 Hz, and unmasked or potentiated excitatory cholinergic responses at frequencies > or =1 Hz. L-NOARG-resistant relaxations were virtually abolished by VIP (100 nM) desensitization at all frequencies. Relaxations to graded low mechanical distension (< or =1 g) were insensitive to tetrodotoxin (TTX: 1 microM) and L-NOARG (300 microM), while those to higher distensions (2 g) were slightly inhibited by both agents to the same extent ( approximately 25%). In the human gastric fundus, NOS- and VIP immunoreactivities are colocalized in the majority of myenteric neurones. NO and VIP mediate electrically evoked relaxations: low frequency stimulation, irrespective of the duration, caused NO release only, whereas shortlasting stimulation at high frequencies induced NO and VIP release. Relaxations to graded mechanical distension were mostly due to passive viscoelastic properties, with a slight NO-mediated neurogenic component at 2 g distension. The difference between NO and VIP release suggests that in human fundus accommodation is initiated by NO. British Journal of Pharmacology (2000) 129, 12 - 20

Aged↗