Alterations in spontaneous contractions in vitro after repeated inflammation of rat distal colon

Carol Bossone1, Jeanette M. Hosseini1, Victor Piñeiro-Carrero2, and Terez Shea-Donohue1,3

Departments of 1 Physiology, 2 Pediatrics, and 3 Medicine, Uniformed Services University of the Health Sciences, Bethesda, Maryland 20814-4799


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
MATERIAL AND METHODS
RESULTS
DISCUSSION
REFERENCES

In inflammatory bowel disease, smooth muscle function reportedly varies with disease duration. The aim of these studies was to determine changes in the control of spontaneous contractions in a model of experimental colitis that included reinflammation of the healed area. The amplitude and frequency of spontaneous contractions in circular smooth muscle were determined after intrarectal administration of trinitrobenzenesulfonic acid in rat distal colon. With the use of a novel paradigm, rats were studied 4 h (acute) or 28 days (healed) after the initial inflammation. At 28 days, rats were studied 4 h after a second inflammation (reinflamed) of the colon. Colitis induced transient increases in the amplitude of spontaneous contractions coincident with a loss of nitric oxide synthase activity. The frequency of contractions was controlled by constitutive nitric oxide in controls. Frequency was increased in healed and reinflamed colon and was associated with a shift in the dominance of neural constitutive nitric oxide synthase control to that of inducible nitric oxide synthase (iNOS). The initial colitis induced a remodeling of the neural control of spontaneous contractions reflecting changes in their regulation by constitutive nitric oxide synthase and iNOS.

inflammation; colitis; rat; nitric oxide; smooth muscle; enteric nerves


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
MATERIAL AND METHODS
RESULTS
DISCUSSION
REFERENCES

ULCERATIVE COLITIS and Crohn's disease are two chronic idiopathic diseases collectively known as inflammatory bowel disease (IBD). These disorders are characterized by prominent intestinal inflammation. Crohn's disease may affect any part of the gastrointestinal tract, while the inflammation in ulcerative colitis is restricted to the colon. The etiologies of these disorders are unclear, but prominent clinical features are diarrhea and abdominal cramping (17, 18). IBD is further characterized by periods of quiescence during which spontaneous relapse or acute reinflammation may occur. Early in the disease, the mucosa may be indistinguishable from that in healthy subjects during the inactive stage. Exacerbation of the disease occurs during the cycles of remission and relapse, and, in more advanced disease, the mucosa may be atrophic with a persistent diffuse inflammation. After healing of the affected area, clinical symptoms often reappear during acute relapse.

A major limitation in understanding the etiology of IBD is that few animals spontaneously develop colitis. Research into the pathogenesis of IBD has made use of several established animal models, particularly the hapten 2,4,6-trinitrobenzenesulfonic acid (TNBS) in ethanol (8, 15, 31, 34). This agent produces an acute inflammation that progresses over several weeks to a chronic stage that is morphologically similar to Crohn's disease. Typically, mucosal injury and polymorphonuclear cell infiltration are observed within 2 h after exposure to TNBS-ethanol (34). The lymphocytic infiltration characteristic of chronic inflammation is evident at 48 h and evolves over several weeks (15, 34). However, experimental models of inflammation have not specifically investigated the changes that may occur after acute relapse that arguably may have more relevance to clinical disease. To address this issue, we have modified an established animal model to evaluate changes in the response of smooth muscle to an initial inflammation of the distal colon followed by a period of recovery before reinflammation of the healed area.

It is well recognized that inflammation is associated with alterations in mucosal and motor function in both small intestine and colon (8, 11, 12, 14, 16-18, 21, 22, 25, 26, 28, 30). In addition, there is evidence suggesting that colonic motility varies with disease activity (8, 11). As inflammation progresses, there are changes in the profile of inflammatory/immune cells and associated mediators that may directly or indirectly affect smooth muscle contractility. Early clinical studies report both hypermotility (26) and hypomotility (11) in ulcerative colitis patients. Moreover, ulcerative colitis patients exhibit changes in the frequency of spontaneous contractions that vary with the duration of their disease (11). There are few studies of spontaneous contractions in the colon in experimental models of inflammation. However, in acetic acid-induced ileitis in dogs, the number of spontaneous phasic contractions was reduced in the first week, and this correlated well with the decreased contractility of smooth muscle strips in vitro (21).

Colonic circular smooth muscle exhibits spontaneous phasic contractions that are subservient to the slow wave frequency. Both excitatory and inhibitory neurotransmitters are important in the regulation of colonic smooth muscle contraction. Previous studies in our laboratory suggested a role for inhibitory neurotransmitters in controlling smooth muscle contraction (8, 11). Nitric oxide (NO) has been identified as an important inhibitory neurotransmitter or neuromodulator in the enteric nervous system (1, 2, 9, 10, 32). Morphological studies of canine colon suggest that NO is also a critical intermediary in the communication between interstitial cells of Cajal (ICC), enteric inhibitory nerves, and smooth muscle in the generation of spontaneous contractions (12). Recently, impaired nitric oxide synthase (NOS) activity in nerves was implicated in the reduced ability of smooth muscle to relax in colitis induced by dextran sulfate sodium in rats (14). Thus the aim of the proposed studies was to determine the changes in the nitrergic control of spontaneous contractions in an animal model of experimental colitis in the following three different stages: acute inflammation, healed mucosa, and acute reinflammation of the healed area.


    MATERIAL AND METHODS
TOP
ABSTRACT
INTRODUCTION
MATERIAL AND METHODS
RESULTS
DISCUSSION
REFERENCES

Induction of Inflammation

Experiments and animal care were conducted in compliance with guidelines outlined by the Guide for the Care and Use of Laboratory Animals (Institute of Animal Research, National Research Council). Male Sprague-Dawley rats (200-400 g; Taconic Farms) were assigned at random to one of the following four treatment groups: control, acute, healed, or reinflamed. After an overnight fast (water ad libitum), rats were anesthetized with 8 mg/kg xylazine (Miles, Shawnee Mission, KS) in combination with 40 mg/kg ketamine (Fort Dodge Laboratories, Fort Dodge, IA) intramuscularly. The control group received 1 ml saline intrarectally, whereas acute, healed, and reinflamed animals were given 100 mg/kg TNBS (Sigma, St. Louis, MO) in a 50% ethanol solution intrarectally. Four hours later, control and acute animals were reanesthetized for surgical removal of a 4- to 5-cm section of distal colon (midtransverse to distal portion). Healed and reinflamed animals were allowed to recover and were monitored for 4 wk. After four weeks, rats were reanesthetized and reinjected intrarectally with either 1 ml saline (healed) or 100 mg/ml TNBS in 50% ethanol (reinflamed), and tissue was harvested 4 h later as performed previously for control and acute animals. All rats were killed with an overdose of pentobarbital sodium (10 mg/kg).

Histology

Full-thickness sections of distal colon from each animal were fixed in 10% neutral buffered formalin, embedded in paraffin, sectioned, and stained with either hematoxylin and eosin (H & E) or Giemsa. With the use of an ocular micrometer (Olympus; Olympus Optical, Tokyo, Japan), the thicknesses of muscularis externa, circular muscularis externa, muscularis mucosae, and submucosa were measured on H & E-stained sections with a ×20 objective on a light microscope. At least three measurements were taken, and values were averaged. An investigator who was unaware of the treatment given graded the sections. A score of 1-5 was determined by giving one point for each of the following features: 1) mucosal sloughing, 2) vasodilatation, 3) submucosal edema, 4) hemorrhage, and 5) increased thickness of the muscle layer. A minimum of seven slides was evaluated in each treatment group. The number and distribution of inflammatory cells were evaluated in three to eight well-oriented areas in Giemsa-stained sections. In addition, mucosa-free sections of distal colon from each group were fixed in 4% paraformaldehyde in PBS for visualizing NADPH diaphorase as an index of NOS activity using the method of Shuttleworth et al. (23). For each section, the number of stained neurons was counted, and the section was photographed by an investigator blinded to the treatment group.

In Vitro Smooth Muscle Contractions

With the use of a double-blade scalpel, 2-mm-wide mucosa-free segments of the distal colon were cut, mounted in organ baths in their circular axis, and maintained in an oxygenated (95% O2-5% CO2) Krebs solution at 37°C throughout the experiment. Krebs solution contained (in mM) 118.5 NaCl, 4.75 KCl, 2.54 CaCl2, 1.19 MgSO4, 25 NaHCO3, 1.19 NaH2PO4, and 11.0 dextrose. The length of maximal active tension (Lo) was determined using ACh (10-4 M). All experiments were performed at Lo.

Muscle strips were allowed to equilibrate for 20-30 min before addition of the ganglionic blocker hexamethonium (100 µM), the NOS inhibitor NG-nitro-L-arginine (L-NNA, 10 µM), the inducible NOS (iNOS) inhibitor aminoguanidine (10 µM), or the sodium channel blocker TTX (1 µM). Aminoguanidine is reported to be a selective, but incomplete, inhibitor of iNOS at this concentration (33). TTX was prepared in a citrate buffer (50 mM citric acid and 48 mM NaH2PO4) and chilled. All chemicals were obtained from Sigma. All antagonists were added to the muscle baths 30 min before measurements. Fresh oxygenated Krebs solution was changed in the baths every 10 min, and the drugs were added back after each washing so that the tissue was exposed to the drug for a total of 30 min. Hexamethonium and aminoguanidine were dissolved in distilled water and chilled. L-NNA was dissolved in distilled water and maintained at 37°C. Vehicle-treated strips received the same volume and identical administration times of the appropriate dissolving solvent for each antagonist.

Data Analysis

The amplitude (mN/cm2), frequency (no. of contractions/10 min), and duration (s) of the spontaneous contractions were measured (model 79E polygraph; Grass, Quincy, MA) after the addition of vehicle, L-NNA, hexamethonium, TTX, or aminoguanidine. Tension was determined using the method of Percy et al. (16) and was defined as
Force<IT>/</IT>area<IT>=</IT>grams tension

<IT>×9.8 </IT>m<IT>/</IT>s<SUP><IT>2</IT></SUP><IT>/</IT>[(gram wet wt<IT>×0.726</IT>)<IT>/</IT>(<IT>1.05×L<SUB>o</SUB></IT>)]
where mass was corrected for water content by multiplying the wet weight by 1.056 (the muscle density). The mass of muscle was corrected for the circular muscle component of the muscularis externa by multiplying by 0.726 (the measured percent of circular muscle). Four strips of muscle were obtained from each animal, and values for each parameter (e.g., hexamethonium) represent the means of strips taken from six different rats. Each strip was incubated with only one antagonist (e.g., hexamethonium). Measurements from each strip were averaged, and data are expressed as means ± SE. There were no differences among individual responses in vehicle alone, so only the water vehicle is presented (Figs. 1-7 and Tables 1 and 2). Differences in morphometric parameters among groups were analyzed by one-way ANOVA followed by the t-test using the Bonferoni correction (Prism Graphpad, Philadelphia, PA). Differences among groups in concentration-response curves were analyzed by multiple ANOVA using the Systat 5.2 program (Systat) followed by post hoc analysis for differences between individual means. For analysis of differences in NADPH diaphorase staining, unlabeled photomicrographs were arranged in order of staining intensity and were analyzed using the Kruskal-Wallis test, a nonparametric method of evaluating more than two groups. A representative section from each group was selected for inclusion in this report.


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Fig. 1.   Representative photomicrographs of Giemsa-stained sections (5 µm) of the rat distal colon 4 h after intrarectal administration of saline (A) or 2,4,6-trinitrobenzene sulfonic acid (TNBS; B) or 4 wk and 4 h after initial intrarectal administration of TNBS followed by saline (C) or TNBS (D).



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Fig. 2.   Representative photomicrographs of NADPH diaphorase-stained sections (5 µm) of the rat distal colon 4 h after intrarectal administration of saline (A) or TNBS (B) or 4 wk and 4 h after initial intrarectal administration of TNBS followed by saline (C) or TNBS (D).



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Fig. 3.   Changes in the amplitude of spontaneous phasic contractions 4 h after intrarectal administration of saline (control) or TNBS (acute) vs. 4 wk and 4 h after initial TNBS followed by saline-treated healed tissue (healed) or TNBS-induced reinflammation (reinflamed) *P < 0.05 vs. control. tau P < 0.05 vs. healed.



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Fig. 4.   Examples of mechanical activity in circular smooth muscle taken from rat distal colon 4 h after intrarectal administration of saline (A) or TNBS (B) or 4 wk and 4 h after initial TNBS followed by saline (C) or TNBS reinflammation (D) in distal colon.



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Fig. 5.   Changes in the amplitude of spontaneous phasic contractions in the presence of vehicle (VEH), hexamethonium (HEX), TTX , NG-nitro-L-arginine (L-NNA), or aminoguanidine (AG) for control (4 h postsaline), acute (4 h post-TNBS), healed (4 wk post-TNBS + 4 h postsaline), or reinflamed (4 wk post-TNBS + 4 h post-TNBS) tissue. Tissue was incubated with antagonists for 30 min before measurements. *P < 0.05 and **P < 0.01 vs. control. phi P < 0.05 vs. respective vehicle within group.



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Fig. 6.   Changes in the frequency of spontaneous phasic contractions 4 h after intrarectal administration of saline (control) or TNBS (acute) vs. 4 wk and 4 h after initial TNBS followed by saline (healed) or TNBS-induced reinflammation (reinflamed). *P < 0.05 vs. control.



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Fig. 7.   Changes in the frequency of spontaneous phasic contractions in the presence of vehicle, hexamethonium, TTX, L-NNA, or aminoguanidine for control (4 h postsaline), acute (4 h post-TNBS), healed (4 wk post-TNBS + 4 h postsaline), or reinflamed (4 wk post-TNBS + 4 h) tissue. Tissue was incubated with antagonists for 30 min before measurements. *P < 0.05 vs. control. phi P < 0.05 vs. respective vehicle within group.


                              
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Table 1.   Changes in colonic smooth muscle morphology (muscle thickness), PMN count, and mucosal injury scoring during acute colitis followed by healing and reinflammation of the affected area


                              
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Table 2.   Duration of spontaneous contractions (seconds) during inflammation


    RESULTS
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ABSTRACT
INTRODUCTION
MATERIAL AND METHODS
RESULTS
DISCUSSION
REFERENCES

Histology

Histological findings are presented in Table 1, and representative photomicrographs of H & E-stained sections from all four treatment groups are shown in Fig. 1, A-D. The microscopic appearance of the colonic epithelium after the first acute inflammation showed significant neutrophilic infiltration in the crypts and mucosal injury featuring vascular congestion, submucosal edema, and mucosal sloughing. The muscularis propria was unchanged at this time. After 4 wk, the mucosa was healed but could be distinguished from controls by more prominent goblet cells, distorted crypts, and the presence of neutrophils and lymphoid infiltrates. During acute reinflammation, there was significantly greater tissue damage and neutrophilic infiltration compared with acute inflammation as well as the presence of lymphocytic-type infiltrates that were not observed in the acute group. The intensity of NADPH diaphorase staining of myenteric plexus and enteric nerves was dramatically less in acute and reinflamed tissue than in controls. The intensity of staining in healed tissue was less than in controls but was still present. There were no differences in the number of neurons present (control, 11.5 ± 1.5; acute, 8.0 ± 2.1; healed 12.33 ± 1.6; reinflamed, 11.5 ± 1.5), only in the intensity of the staining. Representative examples of NOS-stained photomicrographs are shown in Fig. 2.

Spontaneous Smooth Muscle Contractions

Amplitude. The colonic smooth muscle from the saline-treated group exhibited spontaneous contractions that were nifedipine sensitive. The amplitude of these contractions was increased significantly by acute inflammation, returned to control values in the healed group, and was elevated significantly again upon reinflammation (Figs. 3 and 4, A-D). To further explore the inflammation-induced alterations in spontaneous contractions, amplitude was compared in the presence and absence of the nonspecific NOS inhibitor L-NNA, the selective inhibitor of iNOS aminoguanidine (27), the sodium channel blocker TTX, or the ganglionic blocker hexamethonium (Fig. 5, A-D). In both control and healed tissue, the amplitude was elevated by L-NNA and TTX, suggesting that the amplitude of spontaneous contractions is subject to tonic inhibitory control by nitrergic and other nerves. In contrast, contractile amplitude was unaltered by hexamethonium and aminoguanidine, showing a lack of control by ganglionic neurons or inducible NO in controls. In acute inflammation, the significantly enhanced amplitude was not elevated further by any treatment, indicating a loss of nitrergic and other neural control. The effect of the various antagonists on spontaneous contractions in reinflamed tissue was nearly identical to that in the acute group; however, amplitude was elevated further by TTX, indicating a loss of nitergic but not other neural control in the reinflamed group. Thus, during the initial inflammation, there is a loss of both nitrergic and other neural control of amplitude that was restored in the healed group. In contrast, in response to a second inflammation of this healed area, only the nitrergic neural control was lost.

Duration. In controls, the duration of spontaneous contractions was unchanged in the presence of any antagonist tested (Table 2). Compared with controls, the duration of contractions was lengthened significantly by acute inflammation [Fig. 3 (amplitude) and Fig. 4, A vs. B (duration)] and was prolonged further in the presence of TTX (Table 2). Duration returned to control values in the healed stage, but, unlike control, duration was extended in the presence of L-NNA, TTX, or hexamethonium. Compared with controls, the duration of spontaneous contractions was enhanced again upon reinflammation and was lengthened further by TTX and L-NNA.

Frequency. When compared with controls, acute inflammation had no effect on the frequency of spontaneous contractions (Fig. 6 and Fig. 4, A vs. B). In contrast, there was a marked increase in frequency in both the healed and reinflamed groups (Fig. 6 and Fig. 4, C and D). In controls, frequency was significantly elevated only in the presence of L-NNA (Fig. 7, A-D), indicating a tonic inhibitory control by nitrergic nerves. In contrast, in the acute group, frequency was insensitive to any antagonist, showing a loss of this nitrergic influence. In the healed group, frequency was reduced significantly in the presence of hexamethonium and aminoguanidine but not TTX or L-NNA, while in the reinflamed group, frequency was reduced to control values by all antagonists tested. These data indicate that the initial inflammation induced a shift in control of the frequency of spontaneous contractions from nitrergic nerves to a pathway that involves myenteric neurons and inducible NO.


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
MATERIAL AND METHODS
RESULTS
DISCUSSION
REFERENCES

IBD patients generally experience periods of active disease followed by resolution of the mucosal injury and healing. This may be followed by a period of inactivity or quiescence that is interrupted by an acute relapse, the most frequent symptom of which is diarrhea. Repetition of this cycle often leads to exacerbation of the disease. In the present study, contractility of distal colonic circular smooth muscle was assessed in groups of rats 4 h after TNBS-induced inflammation of the distal colon, 4 wk after the initial inflammation during a period of quiescence, or 4 h after reinflammation of the affected area. The microscopic appearance of the distal colon taken from the acute and reinflamed groups featured mucosal damage, submucosal edema, and vasocongestion and/or hemorrhage. However, the initial acute inflammation had a significant neutrophil infiltration, whereas inflammatory cells in the reinflamed group were neutrophils as well as lymphocytes and monocytes. These lymphoid aggregates are a feature of more chronic disease in this model (6, 15, 34). More importantly, there were alterations in the neural control of both the amplitude and frequency of spontaneous contractions consistent with an inflammation-induced remodeling of enteric nerves.

Abnormal gut motility is a common feature of IBD (3, 17, 18, 26, 28, 30). Previous studies showed that inflammation alters circular smooth muscle contractile properties and responses to neurotransmitters or nerve stimulation (5, 15, 24). In healthy colon, circular smooth muscle exhibits nifedipine-sensitive rhythmic spontaneous contractions in vivo and in vitro that are electrically coupled to slow waves (9, 10). NO plays an important role in the regulation of the amplitude of contractions in the colon by controlling the inherently excitable smooth muscle syncytium (9, 10, 13, 32). In addition, there are reportedly fewer NOS-containing neurons in the distal than in the proximal colon, an observation that is consistent with its ability to generate large-amplitude propulsive contractions (29). In the present study, the amplitude of spontaneous contractions was elevated in healthy controls by either TTX or the nonspecific NOS inhibitor L-NNA. The lack of an effect on amplitude of contractions of the iNOS inhibitor aminoguanidine supports previous reports that phasic contractile activity is suppressed continuously by the release of constitutive NO from nerves (9, 29). The mechanical data and immunohistochemical localization of NADPH diaphorase staining in nerves and neurons in the rat colonic muscularis externa in the present study are consistent with earlier findings in rodents (2, 4, 14), dogs (10, 22), and humans (9, 24). The amplitude of spontaneous contractions was unaffected by hexamethonium, indicating that the tonic inhibitory neural control is independent of nicotinic ganglionic neurotransmission.

Acute inflammation of the colon significantly elevated the amplitude and duration of spontaneous contractions but had no effect on the frequency. Previous reports showed that smooth muscle contractility is decreased at 3-7 days after induction of colitis (6, 11, 15). Frequency was also unaffected in the first week after induction of colitis (12). Long-duration slow waves were associated with high-amplitude spontaneous contractions in healthy canine colon (10), suggesting that the duration and amplitude of phasic contractions in this study are electrically coupled to the duration of slow waves. The increased amplitude of contractions in the presence of L-NNA or TTX observed in control tissue was absent in acute inflammation, suggesting a loss of neural NO control. This is concurrent with the reduction in NADPH diaphorase staining in the distal colon of these rats, indicating a likely reduction in constitutive NOS (cNOS) activity. The specificity of NADPH diaphorase staining for neuronal NOS (nNOS) has been confirmed by others showing that NOS activity and NADPH diaphorase copurify to homogeneity and that both activities could be immunoprecipitated with an antibody recognizing neuronal NADPH diaphorase (7). Recent studies demonstrated that the impaired ability to relax smooth muscle was associated with a reduction in nNOS immunoreactivity (14), comparable to the reduction in NADPH diaphorase staining observed in the present study. Similarly, they also observed reduced nNOS without a reduction in the number of myenteric neurons, indicating that the effects on contractility are not due to a general decrement in the population of neurons (14). These data suggest that loss of NO control in acute inflammation in the present study leads to an elevation in contractile amplitude. This may be linked electrically to an overall depolarization of the resting membrane potential and/or an increase in the frequency of long-duration slow waves.

An alternative explanation for the increased contraction could be explained by changes in excitatory neurotransmitters such as ACh. We showed previously that acute inflammation increased the responses to ACh and substance P, an effect attributed to a loss of neural inhibition rather than an increase in excitation (8). In addition, these studies showed that colitis suppressed the neural inhibition derived, in part, from ACh acting at nicotinic receptors in the myenteric plexus (8). Thus it is likely that reduced inhibitory input resulting from a decrease in NOS activity in enteric nerves underlies the enhanced contractile amplitude observed in the present study.

Regulation of spontaneous contractile amplitude in healed tissue was nearly identical to that in controls, and the increased amplitude in response to reinflammation of the affected area was similar to that observed in acute colitis. Thus it appears that both the initial inflammation and recurrence of active disease induce a transient increase in contractile amplitude and duration. This is coincident with a loss of neural cNOS activity as evidenced by the reduction in NADPH diaphorase staining in the nerves in this tissue rather than to a decrease in the number of neurons.

In contrast to amplitude, the frequency of spontaneous contractions was not altered by the initial inflammation. Instead, frequency was elevated significantly at 4 wk after TNBS in both the healed and reinflamed colon, indicating that the initial colitis caused long-lasting alterations in the frequency of phasic contractions. In vivo studies in human colon have shown that the slow wave rhythm may be elevated in patients with ulcerative colitis (11, 26). NO is considered to be an important factor in the control of pacemaker activity. The results of the current study support a role for iNOS in the inflammation-induced alterations in slow wave frequency. Inflammation is known to enhance iNOS activity, resulting in an overproduction of NO. The origin of this NO may be from a number of sources, including macrophages, epithelial cells, or the smooth muscle cells themselves (19, 24). In the present study, the rate of spontaneous contractions in control animals was elevated only in the presence of L-NNA, suggesting a role for NO in the control of basal frequency. These data support previous reports implicating NO in the regulation of ICC function (20). In contrast, in healed and reinflamed tissue, the increased frequency was reduced by aminoguanidine but not by L-NNA. Thus the initial inflammation induced a remodeling of the control of contractile frequency from cNOS-derived NO to iNOS-derived NO. The ability of hexamethonium to reduce frequency in healed and reinflamed tissue indicated the acquisition of a ganglionic regulation of frequency in colitis.

In conclusion, we have modified an experimental model of inflammation to establish a novel approach to the effects of colitis on smooth muscle function. The morphological appearance of healed and reinflamed colon has similarities to patients with quiescent and active IBD. In vitro studies of smooth muscle demonstrate that the smooth muscle activity after an initial inflammation differs dramatically from the response after reinflammation of the healed area. Both acute and reinflammation colitis induce a transient increase in the amplitude of spontaneous phasic contractions that can be attributed to a loss of inhibitory nitrergic control. In addition, the initial inflammation induces a long-lasting alteration in the frequency of spontaneous contractions that suggests a remodeling of the interactions between smooth muscle and nerves. It is proposed that changes in cNOS affect contractile amplitude, although there is a shift in the control of frequency from cNOS to iNOS.


    ACKNOWLEDGEMENTS

The opinions contained herein are the private ones of the authors and are not to be construed as official policy or reflecting the views of the Department of Defense.


    FOOTNOTES

These studies were supported by a grant from the Crohn's and Colitis Foundation of America.

Address for reprint requests and other correspondence: T. Shea-Donohue, Dept. of Medicine, Uniformed Services Univ. of the Health Sciences, 4301 Jones Bridge Rd., Bethesda, MD 20814-4799 (E-mail: tshea{at}usuhs.mil).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Received 17 May 2000; accepted in final form 1 December 2000.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
MATERIAL AND METHODS
RESULTS
DISCUSSION
REFERENCES

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Am J Physiol Gastrointest Liver Physiol 280(5):G949-G957