Accuracy in assessment of colonic transit time with particles: how many markers should be used?

Background Colonic transit time (CTT) is often measured with particle methods in clinical practice, but few studies have evaluated the role of particle amounts for the results obtained. Methods Colonic transit time was studied in 28 subjects taking radiopaque particles for six consecutive days followed by an abdominal radiograph on day 7. Four distinguishable marker types were ingested simultaneously in an amount of 5, 10, 15, and 20 daily, respectively, and CTT calculated for each type as the number of retained markers divided by the daily intake. Reference values were based on 50 markers daily. Key Results Accuracy measured as median deviation of CTT from reference was for 20 markers day⁻¹ 0.08 days, for 15 markers day⁻¹ 0.10 days, for 10 markers day⁻¹ 0.12 days, and for 5 markers day⁻¹ 0.20 days. The CTT values obtained with 5 markers day⁻¹ deviated significantly more from the reference value than CTT values obtained with 10 markers day⁻¹ (P < 0.05) and with 15 and 20 markers day⁻¹ (P < 0.01). Colonic transit times obtained with 20, 15, or 10 markers day⁻¹ did not differ significantly (P > 0.1). Conclusions & Inferences Colonic transit time can be assessed with reasonable accuracy after repeated ingestion of particles. For clinical use, a daily amount of 10 or 12 markers is proposed for reporting CTT in days or hours, respectively. Doses below 10 daily yield a steeply increasing deviation from reference values.     

Effect of transdermal application of nicotine on colonic transit in healthy nonsmoking volunteers

The radio-opaque marker technique (ROMT) is a safe and noninvasive method to determine total colonic (TCTT) and segmental colonic transit times (SCTT). Previous results have shown that smoking volunteers had significantly longer TCTT than nonsmokers, but the underlying mechanism was not clear. We investigated the effect of transdermal nicotine application in two different doses in a nonblind randomized experiment involving three distinct phases. In phase 1 baseline transit times were determined with an abdominal X-ray after a 6-day period of marker ingestion and again after the following bowel movement to study the influence of a bowel movement just before the X-ray. TCTT was nearly twice as high before than after defaecation (42.6 h vs. 25.1 h, P < 0.05). The main acceleration was found in the rectosigmoid (RS) (18.6 h vs 7.1 h, P < 0.05) with no significant changes in right (RC) and left colon (LC). In phase 2 and 3 nicotine was applied in two doses of 17.5 mg day⁻¹ and 35 mg day⁻¹ in random order. Both doses resulted in a significant decrease of TCTT compared to the predefaecation baseline (42.6 h vs 32.2 h/28.2 h, respectively, P < 0.05). Again the main effect was located in the RS (18.6 h vs 9.9 h/7.6 h, P < 0.05). Short-term application nicotine results in a decrease of TCTT which is due to an accelerated transit in the RS.     

Involvement of peripheral and central motor and secretory components in the action of a prostaglandin E2 analogue on colonic transit in rats

The effects of central and peripheral administration of a prostaglandin E₂ analogue (enprostil) on colonic transit time, faecal dry matter and colonic myoelectrical activity, were examined in rats chronically fitted with a cannula in a lateral ventricle of the brain and a catheter inserted in the lumen of the proximal colon, or nichrome electrodes implanted on the proximal colon. In control studies, the mean retention time (MRT) of a marker ([⁵¹Cr] sodium chromate) administered into the proximal colon and determined in the faeces collected at hourly intervals was 7.3 ± 1.7 h, the frequency of colonic long spike bursts was 59.1 ± 9.1/h and faecal dry matter (DM) 65.4 ± 3.4%. Enprostil administered orally (350 μg/kg) decreased colonic MRT (4.9 ± 0.6 h) and faecal DM (50.1 ± 3.2%) and increased by 68% the colonic myoelectric index. Administered intracerebroventricularly enprostil (12 μg/kg) also accelerated colonic transit (MRT = 5.1 ± 1.6 h) and decreased faecal DM (48.7 ± 3.4%) but did not modify colonic spiking activity. Intraperitonal administration of the PG receptor antagonist SC-19220 (1 mg/kg) did not affect the effects of enprostil administered centrally on MRT and DM but blocked the action of oral enprostil on colonic transit time, DM and colonic spiking activity. Centrally administered, SC-19220 (100 μg/kg) antagonized the effects of oral enprostil on colonic transit and faecal DM but not on colonic spiking activity. It is concluded that in rats the enprostil-induced acceleration of colonic transit involves a peripheral action on colonic motility and a secretory component which is centrally mediated. Moreover these results show that colonic hypersecretion, but not hypermotility, can alone accelerate colonic transit.     

Inhibitory effect of oxytocin on accelerated colonic motility induced by water-avoidance stress in rats

Abstract  Recent studies have indicated that brain and gut activities are interrelated and exposure to several stressors, such as water-avoidance stress, stimulates the motor function of the gut through corticotropin-releasing factor (CRF)-signalling pathways in the brain. Central oxytocin is known to attenuate stress responses, including CRF expression in the brain. Here, we examined whether central oxytocin attenuated the acceleration of colonic motility induced by water-avoidance stress. A force transducer was attached to the distal colon of male rat, and the colonic motility and faecal pellet output were recorded while the rats were exposed to water-avoidance stress. Intracerebroventricular (i.c.v.) injections of oxytocin (5, 50 and 500 pmol) and the oxytocin receptor antagonist tocinoic acid (25 μg) were administered before exposure to water-avoidance stress, and the effect of oxytocin on colonic motor function was determined. Centrally administered oxytocin inhibited the accelerated colonic motility induced by water-avoidance stress. The effective dose ranged between 5 and 50 pmol on i.c.v. injection. Oxytocin also decreased the number of CRF-positive cells in the paraventricular nucleus and corticosterone release. The inhibitory effect of oxytocin on accelerated colonic motility was blocked by pretreatment with oxytocin receptor antagonist. Furthermore, centrally administered tocinoic acid enhanced the acceleration of colonic motility. These results suggested that endogenous central oxytocin may contribute to the regulation of colonic function and inhibit the brain CRF-signalling pathways targeting the gut, resulting in the inhibition of stress-induced colonic contractions.     

The effects of methane and hydrogen gases produced by enteric bacteria on ileal motility and colonic transit time

Background Gases produced by intestinal flora may modulate intestinal motor function in healthy individuals as well as those with functional bowel disease. Methane, produced by enteric bacteria in the human gut, is associated with slowed intestinal transit and constipation. The effects of hydrogen, another main gas produced by bacterial fermentation in the gut, on small bowel and colonic motor function remains unrecognized. Therefore, we set out to investigate whether intestinal gases including methane and hydrogen could influence the small bowel motility and colonic transit. Methods Guinea pig ileum was placed in the peristaltic bath with tension transducers attached to measure velocity and amplitude of peristaltic contraction before and after the infusion of control, hydrogen, and methane gases. Also, changes in the intraluminal pressures were monitored before and after the gas infusions. Key Results Methane decreased peristaltic velocity and increased contraction amplitude significantly of guinea pig ileum (P < 0.05). The AUC of intraluminal pressure was significantly increased with methane in guinea pig ileum (P < 0.05). In a second experiment, guinea pig colon was placed in the peristaltic bath to measure transit time before and after control, hydrogen, methane, and methane‐hydrogen mixture gas infusions. Hydrogen shortened colonic transit time by 47% in the proximal colon, and by 10% in the distal colon, when compared with baselines (P < 0.05). Conclusions & Inferences Methane delayed ileal peristaltic conduction velocity by augmenting contractility. Hydrogen shortened colonic transit, and that effect was more prominent in the proximal colon than distal colon.     

Effect of bowel cleansing on colonic transit in constipation due to slow transit or evacuation disorder

Colon transit time measurement with radio-opaque markers is a method of studying the passage of luminal contents throughout the colon. Overall colonic transit time (CTT), as well as segmental transit times [right (RTT), left (LTT) and rectosigmoid (RSTT)], can be calculated. We hypothesize that CTT is influenced by faecal impaction when the rectum is emptied infrequently. The aim of this study is to investigate the effect of bowel cleansing on colonic transit time in patients with chronic constipation. In 25 women (age 41 years; range 20-65 years) with constipation according to Thompson criteria, CTT measurement was performed in an unprepared situation and repeated after cleansing with 4 L of Klean-Prepreg. Ten healthy female volunteers (age 41 years; range 27-57 years) were used as controls. In constipated patients, CTT decreased from a median 70 h (range 10-130 h) to 48 h (5-94 h) in the cleansed state (P < 0.001). A shortening of transit time was found in all three segments. In 10 patients with slow transit (ST) (CTT > 86 h), CTT decreased from 110 h (range 94-130) to 86 (38-94) (P < 0.001). Five of the 10 patients with ST before bowel cleansing had a CTT below 86 h after cleansing. In female controls, uncleansed CTT and RSTT shortened from 39 h (23 to 62) and 17 h (8-29) to 29 h (17-48) and 10 h (0-20) after bowel cleansing (P=0.058 and P=0.046). Colonic intraluminal contents have a substantial effect on colonic transit. In female controls, bowel cleansing shortened rectosigmoid transit. Women with constipation had faster transit in the cleansed state, however, the distribution of markers was not altered. Despite the effect of bowel cleansing on CTT, it seems unnecessary to prepare the bowel in clinical practice because the differentiation of patients between slow transit constipation and outlet obstruction is not changed. However, because in an infrequent defecation pattern, the influence of faecal impaction is considerable, CTT should be applied with care for critical clinical decisions in the treatment of constipation.     

Glucagon like peptide-1 accelerates colonic transit via central CRF and peripheral vagal pathways in conscious rats

Glucagon like peptide-1 (7-36) (GLP-1), one of the gastrointestinal (GI) regulatory peptide, is known to act as a stress related brain neurotransmitter mediating GI function. Central administration of GLP-1 inhibits gastric emptying. However, little is known about the effect of central GLP-1 on colonic transit. Effects and mechanism of GLP-1 on colonic transit were investigated in conscious rats. Immediately after intracerebroventricular (icv)-injection of GLP-1, 51Cr was applied via the catheter positioned to the proximal colon. 90 min after 51Cr injection, rats were euthanized and the colon was removed and divided into 10 equal segments. The radioactivity of each segment was counted and the geometric center (GC) was calculated. Icv-injection of GLP-1 (0.3-3 nmol) dose-dependently accelerated colonic transit [(GC: 4.4+/-0.2 in controls, 7.8+/-0.5 in GLP-1 (3 nmol)]. In contrast, intraperitoneal (ip)-injection of GLP-1 (3 nmol) did not modify colonic transit. Icv-injection of GLP-1 (3 nmol)-induced acceleration of colonic transit was attenuated by vagotomy, atropine and hexamethonium, but not by guanethidine. Icv-injection of GLP-1 (3 nmol)-induced acceleration of colonic transit was abolished by corticotropin releasing factor (CRF) antagonist, astressin. Restraint stress-induced acceleration of colonic transit was abolished by a selective GLP-1 receptor antagonist, exendin. These results indicate that the endogenous GLP-1 is involved in mediating stress-induced alteration of colonic transit via a central CRF and peripheral cholinergic pathways in rats.     

Wireless pH‐motility capsule for colonic transit: prospective comparison with radiopaque markers in chronic constipation

Background Colon transit (CT) measurements are used in the management of significant constipation. The radiopaque marker (ROM) method provides limited information. Methods We proposed to validate wireless motility capsule (WMC), that measures pH, pressure and temperature, to ROM measurement of CT in patients with symptomatic constipation evaluated at multiple centers. Of 208 patients recruited, 158 eligible patients underwent simultaneous measurement of colonic transit time (CTT) using ROM (Metcalf method, cut off for delay >67 h), and WMC (cutoff for delay >59 h). The study was designed to demonstrate substantial equivalence, defined as diagnostic agreement >65% for patients who had normal or delayed ROM transit. Key Results Fifty‐nine of 157 patients had delayed ROM CT. Transit results by the two methods differed: ROM median 55.0 h [IQR 31.0–85.0] and WMC (43.5 h [21.7–70.3], P < 0.001. The positive percent agreement between WMC and ROM for delayed transit was ～80%; positive agreement in 47 by WMC/59 by ROM or 0.796 (95% CI = 0.67–0.98); agreement vs null hypothesis (65%) P = 0.01. The negative percent agreement (normal transit) was ～91%: 89 by WMC/98 by ROM or 0.908 (95% CI = 0.83–0.96); agreement vs null hypothesis (65%), P = 0.00001. Overall device agreement was 87%. There were significant correlations (P < 0.001) between ROM and WMC transit (CTT [r = 0.707] and between ROM and combined small and large bowel transit [r = 0.704]). There were no significant adverse events. Conclusions & Inferences The 87% overall agreement (positive and negative) validates WMC relative to ROM in differentiating slow vs normal CT in a multicenter clinical study of constipation.     

Long-term expression of corticotropin-releasing factor (CRF) in the paraventricular nucleus of the hypothalamus in response to an acute colonic inflammation

This study examined the long-term effects of an acute colitis on central corticotropin-releasing factor (CRF) expression. An increase in CRF mRNA in the paraventricular nucleus of the hypothalamus (PVN) and the central nucleus of the amygdala (CeA) was observed during active colitis, and the effect in the PVN was maintained following recovery. In summary, hypothalamic gene expression of CRF persists in the PVN despite resolution of an acute colitis.     

Influence of age,gender, hormonal status and smoking habits on colonic transit time

Abstract The factors that influence colonic transit time in healthy humans are not yet clearly defined. The aim of this study was therefore to determine (a) if there are differences in colonic transit time between men and women and (b) if age, female hormonal status or smoking habits are associated with alterations in these parameters. Colonic transit time was measured in 164 asymptomatic subjects (80 males, 84 females) by a radio-opaque marker technique with one single plain abdominal X-ray. Colonic transit time was significantly shorter in men than in women (30 ± 2 vs. 42 ± 3 h, P < 0.05). Colonic transit time in non-smoking males was significantly shorter compared with smoking males (26 ± 2 vs. 40 ± 5 h, P < 0.05). In females only height and menstrual cycle influenced colonic transit times. We conclude that gender and smoking habits should be considered when studying colonic transit time in health and disease.     

Local gamma-aminobutyric acid and glutamate circuit control of hypophyseotrophic corticotropin-releasing factor neuron activity in the paraventricular nucleus of the hypothalamus

Paraventricular corticotropin-releasing factor (CRF) neurons play a pivotal role in regulating neuroendocrine responses to stress. The mechanisms by which synaptic inputs control the activity of these neurons are not well understood. The present study was undertaken to determine the role of the intrinsic gamma-aminobutyric acid (GABA)- and glutamatergic neural circuits of the hypothalamic paraventricular nucleus (PVN) in the control of CRF neural activity. We show that in organotypic cultures of the PVN, blockade of the intrinsic GABAergic neurotransmission by the GABAA receptor antagonist bicuculline resulted in a significant increase in CRF secretion. The bicuculline-induced CRF secretory activity was abolished by the coadministration of the selective alpha-amino-3-hydroxy-5-methyl-4-isoxazoleprionic acid (AMPA)/kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). Electrical stimulation of the CRF cell division elicited glutamatergic extracellular field potentials that were dramatically enhanced by bicuculline and were suppressed by CNQX. These results show that the functional activity of CRF neurons in organotypic cultures of the PVN is under a tonic inhibitory influence of an intrinsic GABAergic circuit. Suppression of GABAergic transmission appears to have a permissive role for inducing an increased secretory activity of CRF neurons that is driven by an excitatory glutamatergic network via AMPA/kainate receptors.     

Neuropeptide Y and [Leu,Pro]neuropeptide Y potentiate potassium-induced noradrenaline release in the paraventricular nucleus of the aged rat

This microdialysis study investigated the effects of NPY and the Y₁ selective agonist [Leu³¹,Pro³⁴]NPY on basal and potassium-stimulated noradrenaline release in the PVN of 18-month-old anaesthetised male Sprague–Dawley rats. Microdialysate noradrenaline, DOPAC and HVA concentrations were measured by HPLC after i.c.v. administration of 2 nmol NPY, [Leu³¹,Pro³⁴]NPY or vehicle. [Leu³¹,Pro³⁴]NPY produced a significant 40% reduction in basal noradrenaline concentration (P<0.05). Aged rats had blunted noradrenaline responses to potassium stimulation, however stimulated noradrenaline release was similar in 18-month-old NPY-treated animals and 3-month-old saline treated age controls (2.8 and 3.2 times resting, respectively). [Leu³¹,Pro³⁴]NPY induced a significantly greater release of noradrenaline in response to KCl (5.0 times resting, P<0.05). Thus, in 18-month-old animals with reduced endogenous hypothalamic NPY content, administration of NPY or [Leu³¹,Pro³⁴]NPY increased potassium-induced noradrenaline release to levels seen in 3-month-old rats. This effect may be mediated by an NPY Y₁ receptor.     

Neuropeptide organization of the hypothalamic projection to the parabrachial nucleus in the rat

The hypothalamus is a major source of afferents to the parabrachial nucleus (PB), but the neurotransmitters in this pathway are largely unknown. In this study, we examine the neuropeptide immunoreactivities of neurons in the hypothalamus that project to the PB by using the combined retrograde fluorescence-immunofluorescence method. After injections of the fluorescent tracer fast blue into the PB, retrogradely labeled neurons were observed in the paraventricular, dorsomedial, ventromedial, median preoptic, and anteroventral periventricular hypothalamic nuclei; in the dorsal, retrochiasmatic, and lateral hypothalamic areas; and in the medial and lateral preoptic areas. Our results show that at least five distinct neuropeptide-immunoreactive cell populations in the hypothalamus project to the PB. In the perifornical lateral hypothalamus, many neurotensin (NT)-, corticotropin-releasing factor-, dynorphin (DYN)-, angiotensin II (AII)-, and galanin-like immunoreactive (-ir) neurons were retrogradely labeled. A cluster of retrogradely labeled neurons in the juxtacapsular lateral hypothalamus stained with an antiserum against alpha-melanocyte stimulating hormone (alpha MSH). Over 50% of the retrogradely labeled cells in the arcuate nucleus were adrenocorticotropin (ACTH)-or alpha MSH-ir. Many alpha MSH- and ACTH-ir, and a few DYN-, NT- and AII-ir neurons in the retrochiasmatic area were retrogradely labeled. Only small numbers of double-labeled neurons were found in the paraventricular nucleus, and, of these, enkephalin-ir and dynorphin-ir neurons were the most common. Somatostatin-ir cells in the hypothalamus were rarely double-labeled. The chemical coding of these hypothalamic projections to the PB may provide important clues to the functional organization of these descending pathways.     

Neuropeptide Y Inhibits the Migrating Myoelectric Complex and Delays Small Intestinal Transit in Rats

The effect of neuropeptide Y (NPY) on myoelectric activity of the small intestine was related to transit of a radioactive marker in fasted conscious rats. Myoelectric activity was recorded with bipolar electrodes implanted 5, 20, and 35 em distal to pylorus. A radioactive marker was administered luminally in the duodenum immediately after an activity front of a migrating myoelectric complex (MMC) had passed the first recording site. Intravenous infusion of NPY (50–100 pmol kg⁻¹ min ⁻¹) did not affect the MMC in the duodenum but interrupted its distal propagation and inhibited spiking in the jejunum. At higher doses, NPY (200–800 pmol kg⁻¹ min⁻¹) abolished the MMC at all recording sites. Effects of NPY on myoelectric activity were not prevented by guanethidine, phentolamine, propranolol, or naloxone. The effects of NPY on myoelectric activity corresponded to a dose-related slowing of transit of the marker. In controls, the peak of the marker was propagated ahead of the activity front over the three recording sites. NPY (100 pmol kg⁻¹ min⁻¹) slowed transit, with the peak of the marker proximal to the third electrode site. NPY (400 pmol kg⁻¹ min⁻¹) further delayed transit, with the peak of the marker proximal to the second electrode site. We conclude that the activity front of the MMC exerts a high propulsive capacity. NPY produced a dose-related nonadrenergic inhibition of the intestinal myoelectric activity, which may account for a delayed transit of intestinal contents.     

Microinjection of thyrotropin-releasing hormone in the paraventricular nucleus of the hypothalamus stimulates gastric contractility

Changes in gastric contractility following microinjection of thyrotropin-releasing hormone (TRH) into the paraventricular nucleus of the hypothalamus (PVN) were examined in fasted, urethane-anesthetized rats. Gastric contractility was measured with extraluminal force transducers and analysed by computer. Unilateral and bilateral PVN microinjections of TRH (0.5 and 1.0 μg) significantly increased the force index of gastric contractions from 0 to 60 min postinjection, when compared with animals microinjected with 0.1 μg TRH, 0.1% BSA or TRH (0.5 and 1.0 μg TRH) in sites adjacent to the PVN. The gastric force index was also significantly elevated from 61 to 120 min postinjection in rats receiving bilateral PVN microinjections of TRH (0.5 and 1.0 μg). Peak gastric responses occurred within 10–20 min postinjection and represented an approximately eight-fold increase over basal values. In the remaining groups, the force index was not significantly altered from preinjection values. The excitatory action of TRH (1.0 μg) on gastric contractility was completely abolished by subdiaphragmatic vagotomy. These results suggest that TRH acts within the PVN to stimulate gastric contractility via vagal-dependent pathways.     

Role of CRF receptor 1 in central CRF-induced stimulation of colonic propulsion in rats

The CRF receptor subtype mediating the colonic and gastric motor responses to central CRF was investigated in conscious rats. CRF (0.6 microg/rat) injected intracerebroventicularly (i.c.v.) or 1 h water avoidance stress stimulated defecation (pellet/60 min: 4.1+/-1.0 and 8.7+/-0.7 respectively vs. 0.3+/-0.3 in i.c.v. vehicle/no stress). The CRF receptor 1 (CRF-R1) antagonist, NBI-27914 (50-100 microg/rat) injected i.c.v., abolished the colonic response to i.c.v. CRF and dose-dependently reduced that induced by water avoidance stress. NBI-27914 (100 microg/rat) injected peripherally did not influence the defecatory response to stress. The peptide CRF-R1/R2 antagonist, astressin (10 microg/rat, i.c.v.) inhibited the colonic motor response to i.c.v. CRF and stress similarly as NBI-27914 injected i.c.v. at 100 microg/rat. Intracisternal (i.c.) injection of astressin (10 microg/rat) also completely prevented CRF (0.6 g, i.c.)-induced delayed gastric emptying while i.c. NBI-27914 (50 or 100 microg) had no effect. These results indicate a differential role of central CRF receptor subtypes in the colonic stimulatory and gastric inhibitory motor responses to central CRF and that the CRF component of stress-related activation of colonic expulsion is primarily mediated by CRF-R1.     

Anatomical specificity of noradrenergic inputs to the paraventricular and supraoptic nuclei of the rat hypothalamus

The distribution of neural inputs to the paraventricular (PVH) and supraoptic (SO) nuclei from the regions of the A1, the A2, and the A6 (locus coeruleus) noradrenergic cell groups was investigated by using a plant lectin, Phaseolus vulgaris leucoagglutinin (PHA-L), as an anterogradely transported tracer. An immunofluorescence double-labeling procedure was used to determine the extent to which individual anterogradely labeled fibers and terminals in the PVH and the SO also displayed immunoreactive dopamine-beta-hydroxylase (DBH), a marker for catecholaminergic neurons. The results may be summarized as follows: (1) Projections from the A1 region were found primarily, and in some experiments almost exclusively, in those parts of the magnocellular division of the PVH and the SO known to contain vasopressinergic neurons. (2) Projections from the A2 region were distributed primarily throughout the parvicellular division of the PVH and were most dense in the dorsal medial part, a region known to contain a prominent population of corticotropin-releasing factor (CRF)-immunoreactive neurons. In addition, a less-dense projection to the magnocellular division of the PVH and to the SO was consistently found. (3) Fibers originating from the locus coeruleus were distributed almost exclusively to the parvicellular division of the PVH, with the most prominent input localized to the periventricular zone, a part of the PVH known to contain dopamine-, somatostatin-, and thyrotropin-releasing-hormone-containing neurons. We found no evidence for a projection from A6 to the SO. (4) The majority of fibers originating from the A1, the A2 or the A6 regions contained DBH immunoreactivity, although an appreciable number did not. These results suggest that each of the three brainstem noradrenergic cell groups that contribute to the innervation of the PVH and/or the SO is in a position to modulate the activity of anatomically and chemically distinct groups of neurosecretory neurons.