Modulation of heme oxygenase/carbon monoxide system affects the inhibitory neurotransmission involved in gastrointestinal motility of streptozotocin-treated diabetic rats.

Alterations in gastrointestinal motility of diabetic patients have been linked to degenerative changes induced by glucose abnormalities in the peripheral nervous system. The heme oxygenase/carbon monoxide (HO/CO) signalling represents one of the non-adrenergic/non-cholinergic (NANC) neurotransmission pathways involved in regulation of physiological peristalsis. To investigate the role of HO/CO system in intestinal motility under diabetic conditions, the response to electrical field stimulation (EFS) and western blot analysis of HO/CO pathway components were studied on duodenum longitudinal smooth muscle strips isolated from streptozotocin (STZ)-treated diabetic rats (65 mg kg(-1), i.p.) and respective controls (CTRL), 6 weeks after the onset of diabetes. When compared to CTRL, the ability of CO releasing molecule (CORM-3) (100-400 micromol L(-1)) to enhance NANC relaxation was significantly impaired in STZ-treated rats (P < 0.05). Conversely, in vitro incubation with the HO inhibitor ZnPPIX (10 micromol L(-1), 60 min) significantly reduced EFS-induced relaxation in CTRL (P < 0.05), but not in STZ-treated rats. Interestingly, the ability of ZnPPIX to inhibit EFS-induced relaxation was partially restored in STZ-treated rats co-administered in vivo with the HO-1 inducer cobalt protoporphyrin IX (CoPPIX) (0.5 mg per 100 g body weight weekly). Expression of inducible HO-1 protein was increased in homogenates from STZ-treated rats (vs CTRL, P < 0.01), and further increased in STZ-treated rats receiving CoPPIX (P < 0.05). Taken together, our data underline the essential role of HO/CO system in regulation of inhibitory NANC neurotransmission in the duodenum and suggest that dysregulation of HO/CO activity may represent one mechanism by which gastrointestinal motility is altered in diabetes.

Indomethacin-induced ileitis is associated with tensiometric, vascular and oxidative changes in the experimental rat model.

BACKGROUND: Indomethacin-induced enteritis is a model of inflammatory bowel disease. MATERIALS AND METHODS: To further characterize this model, rats received two injections of indomethacin (7.5 mg kg(-1)) 24 h apart and histological damage of intestinal mucosa, oxidative stress, alterations of intestinal motility and mesenteric vascular bed (MVB) reactivity were investigated after 5 days. RESULTS: The results show that indomethacin caused several histological and functional changes at the ileal level. In particular, response to carbachol as well as the nonadrenergic-noncholinergic inhibitory response to electrical field stimulation (EFS) was lower in the treated than control rats. Moreover, nitric oxide (NO)-component of the inhibitory response was higher in the treated than control rats. Mesenteric vessels preparations from the treated rats showed increased noradrenaline (NA)-induced perfusion pressure, whereas relaxant responses to acetylcholine, although not significantly reduced in the treated rats, had a higher nitrergic component. This finding suggests that vascular dysfunction may contribute to chronic inflammation. Indomethacin injection also determined acute and severe oxidative stress in ileum mucosa. CONCLUSIONS: In conclusion, our study contributes to further characterize the rat model of indomethacin-induced enteritis and suggests that it is suitable for drug screening in rats, as this model can be obtained in a very short period and is simple and reproducible.

Functional alterations of mesenteric vascular bed, vas deferens and intestinal tracts in a rat hindlimb unloading model of microgravity.

1. Prolonged bed rest or exposure to microgravity may cause several alterations in autonomic nervous system response (ANSR). 2. Hindlimb unloading (HU) rats were used as an animal model of simulated microgravity to investigate ANSR changes. The experiments were carried out to investigate the effects of simulated microgravity on the autonomic nervous response of the perfused mesenteric vascular bed (MVB), vas deferens and the colon and duodenum from 2-week HU rats. 3. In MVB preparations of HU rats, the frequency-dependent increases in perfusion pressure with perivascular nerve stimulation (PNS; 8-40 Hz) were inhibited, whereas the noradrenaline (NA) concentration-dependent (1-100 microM) perfusion pressure increases were potentiated. The latter most probably reflected up-regulation of alpha-adrenergic receptor function. Relaxant responses of NA-precontracted MVB to PNS (4-30 Hz) or isoprenaline were not different between control and HU preparations, while vasodilation induced by the endothelial agonist ACh was reduced. 4. Transmural stimulation (2-40 Hz) induced frequency-dependent twitches of the vas deferens which were reduced in vas deferens of HU rats, while the sensitivity to NA-induced contraction was significantly increased. 5. In the gastroenteric system of HU rat, direct contractile responses to carbachol or tachykinin as well as relaxant or contractile responses to nervous stimulation appeared unchanged both in the proximal colon rings and in duodenal longitudinal strips. 6. In conclusion, HU treatment affects peripheral tissues in which the main contractile mediators are the adrenergic ones such as resistance vessels and vas deferens, probably by reducing the release of neuromediator. This study validates NA signalling impairment as a widespread process in microgravity, which may most dramatically result in the clinical phenotype of orthostatic intolerance.

Colonic smooth muscle responses in patients with diverticular disease of the colon: effect of the NK2 receptor antagonist SR48968.

BACKGROUND: Little is known about the pathophysiology of diverticular disease. AIM: To compare passive and active stress and the response to carbachol of colonic smooth muscle specimens from patients with diverticular disease and patients with colon cancer. The effect of the NK2 receptor antagonist, SR48968, on electrically evoked contractions of circular muscle was also investigated. PATIENTS: Sigmoid colon segments were obtained from 16 patients (51-83 years) undergoing elective sigmoid resection for diverticular disease and 39 patients (50-88 years) undergoing left hemicolectomy for non-obstructive sigmoid colon cancer. METHODS: Isometric tension was measured on circular or longitudinal taenial muscle. Strips were stretched gradually to Lo (length allowing the development of optimal active tension with carbachol) and were also exposed to increasing carbachol concentrations. The effects of atropine, tetrodotoxin and SR48968 on electrically evoked (supramaximal strength, 0.3 ms, 0.1-10 Hz) contractions of circular strips from 8 patients with diverticular disease and 19 patients with colon cancer were also studied. RESULTS: Both passive and active stress in circular muscle strips obtained from patients with diverticular disease was higher than in patients with colon cancer (P < 0.05). Electrically evoked contractions were significantly reduced by atropine in all preparations and were virtually suppressed by combined SR48968 and atropine. Tetrodotoxin suppressed electrically evoked contractions only in patients with colon cancer, whereas a tetrodotoxin-resistant component was identified in patients with diverticular disease. CONCLUSIONS: The changes in both passive and active stress in specimens from patients with diverticular disease may reflect circular smooth muscle dysfunction. Acetylcholine and tachykinins are the main excitatory neurotransmitters mediating electrically evoked contractions in human sigmoid colon circular muscle.

Effects of in vivo treatment with interleukins 1beta and 6 on rat mesenteric vascular bed reactivity.

1. Inflammatory bowel disease (IBD) is a condition that involves proinflammatory cytokines such as interleukins 1beta and 6 (ILs). In this disease, it has been shown that an abnormal microcirculatory system is implicated. 2. Therefore, the effects of in vivo treatment for three days with interleukins 1beta and 6 were investigated on rat isolated mesenteric vascular bed (MVB). 3. A significant concentration-dependent increase in vascular response to noradrenaline (NA) was found, with a significant difference in Emax between control (93.01 +/- 16.78 mmHg) and treated preparations (137.91 +/- 5.20 mmHg). Endothelin-1(ET-1) induced a significantly greater increase of perfusion pressure in treated rats in comparison with control rats at the highest concentration used (0.1 microm). 4. The concentration-dependent decrease of perfusion pressure induced by acetylcholine (ACh) in MVB precontracted with NA was significantly reduced in specimens from treated rats in comparison with control rats, with a significant difference in Emax between control and treated preparations. 5. Perivascular nerve stimulation (PNS) evoked contractions with no difference between treatments. Similarly, no difference in relaxant effect was found after PNS in specimens precontracted with NA, in the presence of guanethidine. 6. These findings indicate that the precocious inflammation acts only at postsynaptic level, facilitating vascular contraction. These data seem to support the hypothesis that vascular dysfunction caused by overproduction of ILs may contribute, among other immunological factors, to vasculitis in IBD that leads to intestinal ischaemia through vasoconstriction.

Interleukins 1 beta and 6 induce functional alteration of rat colonic motility: an in vitro study.

BACKGROUND: In rodents, interleukins administration induces intestinal changes similar to those found in inflammatory bowel disease. We investigated the effects of in vivo subchronic treatment with IL-1 beta and IL-6 on rat colonic mucosa and circular smooth muscle. MATERIALS AND METHOD: We evaluated transmucosal electrical parameters (Ussing chambers) and early changes of in vitro direct contractility induced by carbachol and tachykinins. Alterations in excitatory and inhibitory neurotransmission were studied with electrical field stimulation (EFS). RESULTS: Treatment with interleukins induces inflammation proved by fever, early signs of colonic histological damage and changes in mucosal ion transport. Concentration response-curve to carbachol was significantly lower in treated rats (P<0.02) with significant difference in Emax between control (1.67+/-0.17 g) and treated preparations (1.20+/-0.13 g) (P<0.05). Concentration response-curve to NK2 agonist was significantly lower in the treated rats (P<0.005) with a significant difference in Emax between the control (0.26+/-0.04 g) and treated preparations (0.12+/-0.02 g) (P<0.02). None of the drugs used induces changes in EC50. The contractile reflex response to electrically induced distension was significantly higher in the treated rats and more reduced after administration of atropine. Adding NK2 receptor antagonist resulted in a further reduction being observed in the treated and control rats (P=NS). Relaxation by EFS on cholinergic tone was not different between treatments, although pretreatment with L-NNA resulted in greater relaxation in the treated (-21.7%) than in the control rats (-14.8%). CONCLUSION: Early inflammation induced by a subchronic treatment with ILs causes changes in mucosal ionic transport parameters, a reduction in the direct contractile response, and an alteration in the neurotransmission (by an enhancing cholinergic component) that may affect the physiological pattern of colonic motility and the sensory reflex.

Tauroursodeoxycholic acid reduces damaging effects of taurodeoxycholic acid on fundus gastric mucosa.

We investigated the effects of tauroursodeoxycholic acid (TUDCA) to assess whether this acid may also have "protective" effects similar to those found with ursodeoxycholic acid (UDCA). We used a well-known amphibian model of gastric mucosa, and studied the effects of taurodeoxycholic acid (TDCA) on electrical transepithelial parameters, acid secretion and histology in absence or in presence of TUDCA. Mucosal exposure to TDCA, after stimulation with histamine, caused a reduction in transepithelial potential difference (V(t)) and transepithelial resistance (R(t)) and a decrease in acid secretion while mucosal exposure to TUDCA did not cause a significant change in the electrical parameters. Moreover, TDCA primarily affected the neck cells, while TUDCA affected only oxyntic cells, causing a similar degree of injury to that observed in controls. Mucosal exposure to TUDCA plus TDCA caused a reduction in short circuit current (I(sc)) and R(t), whereas acid secretion did not change. These results suggest that: (1) TUDCA reduces the damaging effects of TDCA on fundus gastric mucosa; (2) TUDCA may play an important role in the treatment of gastritis associated with bile reflux.

Effect of three nonpeptide cholecystokinin antagonists on human isolated gallbladder.

Cholecystokinin is the most important stimulant of postprandial gallbladder contraction, and a regulator of gallbladder fasting tone. The aim of this study was to evaluate the effect of dexloxiglumide on isolated human gallbladder contraction induced by cholecystokinin-octapeptide and to compare this effect to that of lorglumide and amiglumide, two glutaramic acid analogs of dexloxiglumide. The negative logarithms of the antagonist dissociation constant (pK(B)) values were 7.00 +/- 0.14, 6.95 +/- 0.11, and 6.71 +/- 0.10 for lorglumide, dexloxiglumide, and amiglumide, respectively. Dexloxiglumide produced a concentration-dependent rightward shift of the cholecystokinin-octapeptide curve, without affecting its maximal response. A similar effect was obtained both with lorglumide and amiglumide. Moreover, the slopes for the three antagonists did not differ significantly from unity. These data show that the three molecules have a potent antagonistic effect, of a competitive nature, on gallbladder cholecystokinin type 1 receptors. It may be concluded that dexloxiglumide, lorglumide, and amiglumide exhibit a promising therapeutic profile for biliary colic and other gastrointestinal disorders in which CCK1 receptors play important physiological roles.

Effect of somatostatin on human gallbladder motility: an in vitro study.

In vivo studies have demonstrated that somatostatin induces human gallbladder relaxation. To determine whether this polypeptide acts directly on the gallbladder muscle, its effect on strips of human gallbladder was studied in vitro. Strips of gallbladder were set up isometrically in an organ bath containing oxygenated Krebs' solution. Dose-response curves to cholecystokinin-octapeptide and carbachol were first established. The ability of somatostatin to cause relaxation under basal conditions and during 50% maximal stimulation by cholecystokinin-octapeptide (7.2 x 10(-8) M) and carbachol (3.5 x 10(-6) M) was assessed in 32 strips at 4.3 x 10(-6) M concentration which mimics the plasma concentrations found in patients with somatostatinoma and in 12 additional strips at 4.3 x 10(-8) M concentration. Somatostatin action on the intrinsic innervation by using electrical field stimulation (EFS) (200 mA 5 msec in duration, 30 Hz; 400 mA, 1 msec in duration, 10 Hz) was also evaluated in 39 strips. Somatostatin had no effect on the basal or carbachol-generated tensions. On the contrary, somatostatin (4.3 x 10(-6) M) reduced cholecystokinin-octapeptide-generated tensions by 8% (P < 0.001) and reduced EFS-generated tensions at 30 Hz by 7.7% (P < 0.01) and those at 10 Hz by 41.2% (P < 0.01). All responses to cholecystokinin-octapeptide and carbachol were abolished by dibutyryl-guanosine 3', 5'-cyclic monophosphate (5 x 10(-3) M) and atropine (10(-5) M), respectively (P < 0.0002 and P < 0.0002). All responses to electrical field stimulation were reduced or abolished by tetrodotoxin (2 x 10(-6) M) (P < 0.001 and P < 0.0001, respectively). Our findings show that somatostatin exerts its inhibitory action on the response to cholecystokinin-octapeptide and on the intrinsic innervation of the gallbladder smooth muscle. The probable neurotransmitter is the acetylcholine.

Motor responsiveness of proximal and distal human colonic muscle layers to carbachol and neurotensin.

A regional heterogeneity of the responsiveness to neurohumoral agents has been demonstrated in proximal and distal colon. The aim of this study was to compare the motor responsiveness of circular and longitudinal muscles from ascending and sigmoid colon to carbachol and neurotensin. Ascending colon circular muscle was more sensitive to carbachol than sigmoid colon circular muscle (P < 0.05). Moreover, the potency for the carbachol-stimulated contraction was greater in ascending colon circular than longitudinal muscle (P < 0.05). The potency for carbachol and neurotensin stimulations was similar in longitudinal and circular muscles of both sections, respectively. However, the ascending colon circular muscle had a greater potency to neurotensin than longitudinal muscle (P < 0.03). Longitudinal muscle was more sensitive to the effects of neurotensin in sigmoid than ascending colon (P < 0.01). On a molar basis, neurotensin was approximately fivefold more potent than carbachol in producing similar contractions. These data suggest that these agents stimulate human colon smooth muscle according to region and type of muscle layer.

Gallbladder motility in vitro in men with gallstones following Billroth II gastric resection.

Gastric surgery induces an increased incidence of gallstones. To investigate the changes in gallbladder kinetics after gastric resection, 20 male patients were studied: ten patients undergoing cholecystectomy for gallstones developed after Billroth II gastric resection and ten patients undergoing cholecystectomy for cholelithiasis without previous abdominal surgery. Longitudinal strips from the gallbladder wall were suspended in an organ bath and the isometric tension recorded. Dose-response curves to cholecystokinin-octapeptide and carbachol were obtained. Half the maximal response to cholecysto-kinin-octapeptide was 0.50 +/- 0.11 x 10(-7) M in the first group and 1.36 +/- 0.37 x 10(-7) M in the second group (P < 0.05). The ED50 to carbachol was 24.33 +/- 2.69 x 10(-7) M in the gastrectomy group and 40.39 +/- 5.01 x 10(-7) M in the control group (P < 0.01). There was no significant difference in the maximal contractile response either to cholecystokinin-octa-peptide or carbachol in the two groups. Our study shows an increased gallbladder sensitivity to cholecystokinin-octapeptide and carbachol in patients with gallstones developed after Billroth II gastric resection.