What is new in radiology and pathology of motility disorders in children?

Seminars in Pediatric Surgery (2010) 19, 81-85

What is new in radiology and pathology of motility disorders in children? Jonathan R. Sutcliffe, MD, FRCS,a Sebastian King, MB, BS, BSc, MD, PhD,b John M. Hutson, MD, AO,c Bridget Southwell, BSc(Hon), MSc, PhD, AGAFd From the aDepartment of Paediatric Surgery, Leeds General Infirmary, Leeds, United Kingdom; b Department of General Surgery, Royal Children’s Hospital and Department of Paediatrics, University of Melbourne, Surgical Research Laboratory and Murdoch Children’s Research Institute, Victoria, Australia; c Department of Urology, Royal Children’s Hospital and Department of Paediatrics, University of Melbourne, Surgical Research Laboratory and Murdoch Children’s Research Institute, Victoria, Australia; and the d Surgical Research Laboratory, Murdoch Children’s Research Institute, Victoria, Australia. KEYWORDS Transit studies; Scintigraphy; Rectal ultrasound; Manometry; Neurotransmitters; Interstitial cells of Cajal

Disorders affecting colorectal motility lead to significant morbidity in children with surgical conditions. Etiology is frequently unknown, which in turn makes treatment empiric and compromises outcome. A thorough understanding of the normal mechanisms of control and the ability to recognize and manage defects is an important goal for clinicians. This article reviews recent advances made in the investigation of children with colorectal motility disorders, including the role of transit studies (marker studies and scintigraphy), options for assessing anatomy (ultrasound, contrast enema, and sectional imaging) and the use of manometry, both anorectal and colonic. Current concepts in microscopic evaluation are outlined. © 2010 Elsevier Inc. All rights reserved.

Despite recent advances, the management of children with colorectal disorders continues to provide significant challenges in terms of functional outcome. It is now more than 100 years since the discovery of Hirschsprung disease (HD), and almost 50 years since the development of a standard surgical treatment. Even after refinement of these techniques, including the introduction of laparoscopy, there still remain a significant percentage of children with functional problems of colonic evacuation after surgery for HD. For children with anorectal malformations (ARM) there have been few major advances since the introduction of Alberto Peña’s revolutionary operation more than 25 years ago. Again, laparoscopic surgery has been introduced facilAddress reprint requests and correspondence: Jonathan R. Sutcliffe, MD, FRCS, Department of Paediatric Surgery, Leeds General Infirmary, Great George Street, Leeds, LS1 3EX, United Kingdom. E-mail address: [email protected].

1055-8586/$ -see front matter © 2010 Elsevier Inc. All rights reserved. doi:10.1053/j.sempedsurg.2009.11.014

itating the management of rectourethral and rectovesical fistulae, but there remain many children with ongoing abnormalities of defecation. Apart from HD and ARM, there has been increasing surgical involvement in children with chronic idiopathic constipation since the introduction of the Malone appendicostomy for administration of antegrade colonic enemas. Coupled with this, there has been a realization that chronic idiopathic constipation is a heterogeneous disorder. Those referred for a surgical opinion are often the worst affected and have treatment-resistant symptoms. Once HD has been excluded, it is unusual for an organic cause to then be recognized. In many cases, the failure to provide a diagnosis represents our failure to understand the normal mechanisms of gastrointestinal motility yet patients are labeled as “functional.” It should be remembered that the complete absence of ganglion cells seen in HD is a relatively crude anomaly

82 in histologic terms and it seems intuitive that more subtle, but as yet unrecognized, abnormalities are likely to exist in symptomatic patients. More sophisticated histologic investigations may lead to the identification of the underlying cause, which will in turn lead to better-targeted therapies. Similarly, reliable methods to examine transit are needed to distinguish those children with primary colonic motility disorders from those with abnormal pelvic floor function. As with any investigation requested in a clinical setting, the aim must be to either influence management or help provide a prognosis. Symptoms such as constipation and soiling are seen in children with colorectal pathology and carry a significant long-term burden for children and families. This review concentrates on new advances in medical imaging, manometry, and histology, which it is hoped will allow more rational management for children with colorectal motility disorders.

Transit studies Transit studies are of use in treatment-resistant chronic idiopathic constipation to determine the level of hold-up. Objective confirmation of a motility disorder may in itself be helpful both to the family and to the treating clinician. For others, determination of the site of hold-up may be used to guide therapy. There are a minority of patients for whom symptoms are considered potentially fictitious and for whom transit studies may be useful, as demonstration of normal transit would influence future management. Normal patterns of motility in the colon are highly variable, and studies have aimed to establish the normal times for transit through the various regions of the bowel. The contents of a meal take 2 to 4 hours to pass from the pylorus to the cecum, and a further 12 to 72 hours to reach the rectum. Two basic techniques have been used to evaluate gastrointestinal motility: transit of sitz (plastic) markers, which are viewed by standard abdominal x-ray, or radioisotope tracer, viewed by gamma camera (scintigraphy). Sitz markers have been used for much longer and are widely available. Their use was first described in 1969, and they have subsequently been in common use to gain information about colorectal function in patients with constipation. Markers are ingested with a meal and abdominal x-rays are obtained to count the number of markers in different segments of the colon at defined times after ingestion. The method is simple, and clearly identifies patients with hold-up in the proximal colon from those with dysfunctional evacuation. Plastic marker studies are considered the gold standard for transit studies. There are 2 main methods for plastic marker studies. Pellets can be ingested in a single dose, often given inside a capsule, and abdominal images obtained every 24 hours until all the markers have passed. The test is simple but incurs repeated exposure to radiation, and is time-consuming and inconvenient for the family. In the second method,

Seminars in Pediatric Surgery, Vol 19, No 2, May 2010 markers of different shapes are ingested on 3 consecutive days and a single abdominal x-ray is obtained on day 4. Segmental transit is calculated from the relative positions of the different markers. Various refinements have been made to the interpretation of plastic marker studies, including drawing of areas of interest to more clearly define the different segments of the colon. However, there is the potential for observer error in drawing of these regions, although this error is inherent in all marker studies. Other contentious areas include whether bowel preparation should be carried out prior to the test (results suggesting that bowel preparation has a more significant effect in outlet obstruction than in slow colonic transit) and reproducibility (there seems to be a significantly wide variation in total colonic transit). Total and segmental transit times in the colon of children have been summarized by Wagener et al.1 Transit studies using radioisotopes involve ingestion of the isotope followed by multiple images taken during 2 to 5 days. As images are captured on a gamma camera there is no increase in radiation, as the radiation burden is the same irrespective of the number of times the patient is imaged. The total amount of radiation exposure is adjusted to be similar to 2 abdominal x-rays. Areas of interest over the colon can be drawn, which then allows for quantitation. This method is relatively new and is only available in a few centers.2 Before it can be put into widespread use it will be necessary to estimate reproducibility and intrasubject variability. Isotopes can be given in either liquid or solid form, although the liquid preparations overcome delayed gastric emptying caused by solids. Analysis of transit may be performed using the geometrical center (the weighted average of radioactivity over specific regions of the bowel) to determine the median point of radioactivity for each time point.3 There is consensus that images at 4 to 6 hours after ingestion, as well as 24 and 48 hours give a good summary of colonic transit with acceptable specificity and high sensitivity for detecting motility disorders.4 In Melbourne we have used radioactive isotope transit studies to characterize colonic transit times in 64 children with chronic idiopathic constipation. Intraobserver variability was found to be low (kappa ⫽ 0.79).5 Visual analysis demonstrated that the transit studies could be separated into 3 clear groups: those with normal transit, those with anorectal hold-up, and those with slow colonic transit. In addition, the patients with slow colonic transit could be separated into 3 subgroups, a severe group with hold-up in the proximal small bowel even before the radioactive tracer reached the cecum, a moderately severe group where the tracer only reached the splenic flexure by 48 hours, and a further group where the tracer reached the anorectum within a reasonable time, but then failed to clear from the more proximal colon. It remains to be determined whether these subgroups represent 3 separate entities, or merely examples of different levels of severity of slow-transit constipation (STC).

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83

Anatomical studies

Manometry

A distended rectum or colon indicates significant abnormality. This may be a primary pathology or secondary to a more distal hold-up. Nevertheless, a distended segment is likely to have impaired motility and its identification is therefore useful in directing treatment. Ultrasonography has been introduced for identification of rectal distension by some research groups.6 Measuring the rectal diameter through a partially filled bladder enables an estimate to be made of the degree of rectal distension. Comparison of symptomatic children and controls revealed significant differences and importantly established control data for a pediatric population. Because there is no radiation exposure, repeated evaluation at different time points may be performed making this of potential use in the follow-up of patients. Other centers have used contrast enemas to evaluate the degree of colorectal distension. A water-soluble contrast is used to demonstrate the presence or otherwise of distension and its proximal extent. A further advantage of this technique is the potentially therapeutic effect of instilling watersoluble contrast into a loaded colon, which helps empty the colon. Evaluation of the degree of colonic distension will help to guide the dose of laxative recommended.7,8 In the adult literature, magnetic resonance imaging has been used to examine not only pelvic anatomy9,10 but also the movement of the pelvic floor during defecation. Normal data have been difficult to interpret even for adult subjects11 and the additional consideration of the need for a general anesthetic for the majority of children makes this less easy to perform and obviates the ability to image voluntary defecation. Nevertheless, this modality may contribute to our understanding of the normal sequence of activity within the pelvic floor during defecation in a way not achievable by other methods and may influence our attempts to reconstruct perineal and pelvic anatomy in, for example, children with ARM. The fact that several different modalities are in use is a reflection of the piecemeal way in which the current approach to these problems has developed. Each investigation has its own advantages and disadvantages, and provides an answer to a different set of questions; transit studies will indicate the site of hold-up, ultrasound will give information about rectal anatomy in a way that can then be repeated without radiation exposure, whereas a contrast enema will demonstrate more proximal anatomy and help to clear the colon, albeit at the cost of some additional radiation exposure. Magnetic resonance will produce more detailed anatomical data, but requires a general anesthetic in many circumstances, which also limits the yield in terms of functional information. Normal data for children in particular are incomplete. Local expertise will further influence the choice of investigation. Progress will best be achieved when standardized protocols and normative data are available for each investigation as this will allow comparison between groups.

Anorectal manometry may be used in clinical practice to differentiate organic causes from behavioral causes of constipation. It is typically used as a technique to establish pelvic floor dysfunction in adult patients, though it is occasionally used in the pediatric population. Both reduced rectal sensation12 and increased rectal compliance have been found to be common in children with chronic constipation.13 The clinical relevance of these findings remains unclear. Anorectal manometry has also been used with the intent of excluding HD. However, as it often requires sedation or general anesthesia, it may have a limited role in the investigation of idiopathic constipation. Rectal biopsy remains the gold standard investigation for the exclusion of HD. Colonic manometry has been used to investigate motility patterns for almost 50 years. While most studies remain in the realm of research, a small number of centers use colonic manometry to guide clinical decisions. In most centers the manometry catheter is placed retrogradely at colonoscopy, which then allows for up to 6 hours of colonic monitoring to be performed before the catheter is removed. Short-duration colonic manometry has been used to differentiate colonic from anorectal dysmotility in a large cohort (n ⫽ 375) of children from 3 centers in the United States.14 The clinical significance of these findings is unclear; children who demonstrated an absence of appropriate colonic responses underwent colonic diversion, but the response was variable. Nevertheless, good correlation between colonic manometry and the flow of intraluminal contents has been demonstrated in adult volunteers using scintigraphy.15 Colonoscopic placement in a child requires general anesthesia, bowel preparation and gas insufflation all of which may affect results. An alternative to colonoscopic placement is the passage of the manometry catheter through a pre-existing appendicostomy. This then allows continuous 24-hour manometry and measurement of responses to physiological stimuli. This technique has demonstrated significant abnormalities in colonic motility in children with STC (n ⫽ 18).16 Abnormalities included a low frequency of antegrade propagating sequences (indicators of motility) in both the right and left colon, and an attenuation of the normally rapid increase in activity in response both to meals and waking. It was thought that loss of the postprandial response may indicate a defect in neurohormonal control of gut motility. Interestingly, most subjects with STC had a normal frequency of high-amplitude propagating sequences (thought to initiate mass colonic movement).

Histologic investigation Identification of a relevant defect in the control of motility would provide the opportunity to direct treatment specifically. For the last half-century the only common histologic analysis done in patients with colorectal disorders was a

84 rectal biopsy, which was carried out as a diagnostic test for HD. The presence of large nerve bundles associated with the absence of ganglion cells in the submucosa is demonstrated using a combination of histochemical staining for the enzyme acetylcholinesterase and H&E. Rectal biopsy has become such a routine test that when it shows no evidence of HD it is often misinterpreted as being “normal.” The complexity of the enteric nervous system and difficulty in obtaining reliable control tissue in children has made the search for more subtle enteric nervous system defects in human disease difficult. Defects may be quantitative (eg, reductions in components of the ENS) or qualitative (eg, loss of function). Once an abnormality has been identified, it is frequently difficult to establish whether the changes are primary or secondary to some other cause.17 Another important consideration has been difficulty in accessing colorectal tissue from anywhere other than the rectum. It is fortuitous in diagnostic terms that HD always affects the distal rectum as this is the only part of the hindgut readily available to biopsy. For some patients, however, the site of abnormality may be more proximal. Laparoscopic approaches have therefore been developed to allow biopsy.18 Due in part to these problems controversy has arisen with some diagnoses. Intestinal neuronal dysplasia (IND) was first described in 1971 by Meier-Ruge.19 In IND, the primary histologic abnormality was initially defined as the presence of enlarged ganglia in the submucosal plexus, characterized by more than 7 nerve cells.20 IND as an entity remains controversial. The diagnosis is difficult to make histologically, requiring examination of at least 30 sections.21 A variety of histologic techniques have been used, leading to a lack of consistency in the literature and significant interobserver variation between histologists.22 Because doubt arose regarding the existence of IND as a diagnosis, the recommended method of biopsy and diagnostic criteria themselves have changed.22 It is unclear how a diagnosis of IND should influence management. If it exists as described, the natural history is unknown and histologic changes may improve with time.23 Indeed, even the current diagnostic criteria are consistent with appearances in the bowel of asymptomatic children.24 Several types of treatment have been employed, from conservative management with laxatives, to sphincter myectomy, stoma formation, and partial or total colectomy, but no clear management strategy has emerged.25 Despite the problems encountered with the diagnosis of IND, the concept of a physiological explanation for dysmotility is an attractive one given the significant number of patients with severe idiopathic constipation,26 the number of HD patients who clearly have motility abnormalities after the aganglionic segment has been resected, or indeed the patients with ARM who have ongoing symptoms despite anatomical correction. For some of these patients, although the histologic appearances may not readily achieve the criteria required to make the formal diagnosis of IND, intes-

Seminars in Pediatric Surgery, Vol 19, No 2, May 2010 tinal neurons are nevertheless dysplastic. When recently discussed by a panel of experts,26 there was general agreement that IND did exist, although not necessarily with the current diagnostic criteria, nor necessarily as a primary abnormality. Patients with motility disorders may have other causes for their symptoms; immunohistochemical studies have reported changes in nerves containing nitric oxide synthase,27 vasoactive intestinal peptide,27 substance P,28 choline acetyl transferase,29 and serotonin30 in adult chronic idiopathic constipation. Our group reported low immunoreactivity of substance P in STC children.31,32 Other neuronal subgroups may be affected and require further study. Muscle defects,33,34 and loss of interstitial cells of Cajal (ICC) have been reported in adult STC,35,36 while autonomic nervous system defects have been reported in upper gastrointestinal tract disorders37 and colonic disorders38 and the central release of corticotrophin-releasing hormone has been shown to influence colonic motility.39 Children with STC lack a postprandial increase in colonic motor activity,40 suggesting that changes in postprandial hormones might be lacking. One interesting component of the control mechanism for normal motility is a group of non-neuronal cells, ICC. Although these cells were first identified in 1893, the identification of c-Kit as an ICC marker by Maeda41 allowed examination using immunohistochemistry and obviated the need for more complex and expensive techniques. This opened the door for a swathe of studies. Subsequent investigation in animal models has indicated important roles of ICC in pacemaking, as intermediaries between neurons and smooth muscle cells and in mechanoreception. Blockage of receptors involved in this system results in a lethal paralytic ileus in mice. In human subjects, ICC abnormalities are reported in various functional smooth muscle disorders. HD, ARM, and STC in adults have all been associated with ICC abnormalities. It remains unclear whether these abnormalities are primary or secondary, and the limited availability of control tissue makes confident identification of abnormal distribution difficult. Nevertheless, interest in this group of cells remains because ICC demonstrate plasticity; in animal models, a loss of ICC and associated function is seen with dilatation. Because partial restoration of distribution and function occur with resolution of dilatation, these cells may become therapeutic targets and further investigation is warranted.

Discussion The long-term sequelae of colorectal pathology may be particularly troublesome to patients and attempts to correct them are likely to be more successful with an accurate understanding of their pathophysiology rather than with empiric treatment. Our understanding of the normal mechanisms of control of intestinal function remain incomplete but nevertheless techniques are being developed, which

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have a potentially important role in a clinical setting in the objective assessment of transit, motility, anatomy, and histology. Clearly, for any individual, the clinical question will determine which investigation is most suitable, but in practice local expertise may influence this. Consistent methodology between centers for each investigation and reliable data from normal subjects are likely to further improve our ability to manage this difficult group of patients.

References 1. Wagener S, Shankar K, Turnock R, et al. Colonic transit time—what is normal? J Pediatr Surg 2004;39:166-9. 2. Maurer AH, Parkman HP. Update on gastrointestinal scintigraphy. Semin Nucl Med 2006;36:110-18. 3. Notghi A, Hutchinson R, Kumar D, et al. Use of geometric center and parametric images in scintigraphic colonic transit studies. Gastroenterology 1994;107:1270-7. 4. Southwell B, Clarke M, Sutcliffe J, et al. Colonic transit studies: normal values for adults and children with comparison of radiological and scintigraphic methods. Pediatr Surg Int 2009;25:559-72. 5. Sutcliffe J, King S, Hutson J, et al. Gastrointestinal transit in children with chronic idiopathic constipation. Pediatr Surg Int 2009;25:465-72. 6. Singh S, Gibbons N, Vincent M, et al. Use of pelvic ultrasound in the diagnosis of megarectum in children with constipation. J Pediatr Surg 2005;40:1941-4. 7. Levitt MA, Peña A. Surgery and constipation: when, how, yes, or no? J Pediatr Gastroenterol Nutr 2005;41:S58-60. 8. Levitt M, Martin C, Falcone R, et al. Transanal rectosigmoid resection for severe intractable idiopathic constipation. J Pediatr Surg 2009;44: 1285-91. 9. Fletcher J, Busse R, Riederer S, et al. Magnetic resonance imaging of anatomic and dynamic defects of the pelvic floor in defecatory disorders. Am J Gastroenterol 2003;98:399-411. 10. Stoker J, Bartram CI, Halligan S. Imaging of the posterior pelvic floor. Eur Radiol 2002;12:779-88. 11. Goh V, Halligan S, Kaplan G, et al. Dynamic MR imaging of the pelvic floor in asymptomatic subjects. AJR Am J Roentgenol 2000; 174:661-6. 12. Meunier P, Marechal JM, de Beaujeu MJ. Rectoanal pressures and rectal sensitivity studies in chronic childhood constipation. Gastroenterology 1979;77:330-6. 13. Voskuijl W, van Ginkel R, Benninga M, et al. New insight into rectal function in pediatric defecation disorders: disturbed rectal compliance is an essential mechanism in pediatric constipation. J Pediatr 2006; 148:62-7. 14. Villarreal J, Sood M, Zangen T, et al. Colonic diversion for intractable constipation in children: colonic manometry helps guide clinical decisions. J Pediatr Gastroenterol Nutr 2001;33:588-91. 15. Dinning PG, Szczesniak MM, Cook IJ. Determinants of postprandial flow across the human ileocaecal junction: a combined manometric and scintigraphic study. Neurogastroenterol Motil 2008;20:1119-26. 16. King S, Catto-Smith A, Stanton M, et al. 24-Hour colonic manometry in pediatric slow transit constipation shows significant reductions in antegrade propagation. Am J Gastroenterol 2008;103:2083-91. 17. Chitkara DK, Di Lorenzo C. From the bench to the ‘crib’-side: implications of scientific advances to paediatric neurogastroenterology and motility. Neurogastroenterol Motil 2006;18:251-62. 18. King SK, Sutcliffe JR, Hutson JM. Laparoscopic seromuscular colonic biopsies: a surgeon’s experience. J Pediatr Surg 2005;40:381-4. 19. Meier-Ruge W. Uber ein erkrankungsbild des colon mit hirschprungsymptomatik. Verh Dtsch Ges Pathol 1971;55:506-9.

85 20. Meier-Ruge W, Bronnimann P, Gambazzi F, et al. Histopathological criteria for intestinal neuronal dysplasia of the submucosal plexus (type B). Virchows Arch 1995;426:549-56. 21. Meier-Ruge WA, Longo-Bauer CH. Morphometric determination of the methodological criteria for the diagnosis of intestinal neuronal dysplasia (IND B). Pathol Res Pract 1997;193:465-9. 22. Koletzko S, Gambazzi F, Faus-Kessler T, et al. Rectal biopsy for diagnosis of intestinal neuronal dysplasia in children: a prospective multicentre study on interobserver variation and clinical outcome. Gut 1999;44:853-61. 23. Munakata K, Morita K, Okabe I, et al. Clinical and histologic studies of neuronal intestinal dysplasia. J Pediatr Surg 1985;20:231-5. 24. Coerdt W, Michel J, Rippin G, et al. Quantitative morphometric analysis of the submucous plexus in age-related control groups. Virchows Arch 2004;444:239-46. 25. Csury L, Pena A. Intestinal neuronal dysplasia. Myth or reality? Pediatr Surg Int 1995;10:441-6. 26. Martucciello G, Pini Prato A, Puri P, et al. Controversies concerning diagnostic guidelines for anomalies of the enteric nervous system: a report from the fourth International symposium on Hirschsprung’s disease and related neurocristopathies. J Pediatr Surg 2005;40:1527-31. 27. Cortesini C, Cianchi F, Infantino A, et al. Nitric oxide synthase and VIP distribution in enteric nervous system in idiopathic chronic constipation. Dig Dis Sci 1995;40:2450-5. 28. Porter A, Wattchow D, Hunter A, et al. Abnormalities of nerve fibers in the circular muscle of patients with slow transit constipation. Int J Colorectal Dis 1998;13:208-16. 29. Wattchow D, Brookes S, Murphy E, et al. Regional variation in the neurochemical coding of the myenteric plexus of the human colon and changes in patients with slow transit constipation. Neurogastroenterol Motil 2008;20:1298-305. 30. Lincoln J, Crowe R, Kamm M, et al. Serotonin and 5-hydroxyindoleacetic acid are increased in the sigmoid colon in severe idiopathic constipation. Gastroenterology 1990;98:1219-25. 31. Hutson JM, Chow CW, Borg J. Intractable constipation with a decrease in substance P-immunoreactive fibres: is it a variant of intestinal neuronal dysplasia? J Pediatr Surg 1996;31:580-3. 32. Hutson J, Chow C, Hurley M, et al. Deficiency of substance Pimmunoreactive nerve fibres in children with intractable constipation: a form of intestinal neuronal dysplasia. J Paediatr Child Health 1997; 33:187-9. 33. Slater BJ, Varma JS, Gillespie JI. Abnormalities in the contractile properties of colonic smooth muscle in idiopathic slow transit constipation. Br J Surg 1997;84:181-4. 34. Wedel T, Van Eys G, Waltregny D, et al. Novel smooth muscle markers reveal abnormalities of the intestinal musculature in severe colorectal motility disorders. Neurogastroenterol Motil 2006;18:526-38. 35. He C, Burgart L, Wang L, et al. Decreased interstitial cell of Cajal volume in patients with slow-transit constipation. Gastroenterology 2000;118:14-21. 36. Lyford G, He C, Soffer E, et al. Pan-colonic decrease in interstitial cells of Cajal in patients with slow transit constipation. Gut 2002;51: 496-501. 37. Chelimsky G, Boyle J, Tusing L, et al. Autonomic abnormalities in children with functional abdominal pain: coincidence or etiology? J Pediatr Gastroenterol Nutr 2001;33:47-53. 38. Altomare D, Portincasa P, Rinaldi M, et al. Slow-transit constipation: solitary symptom of a systemic gastrointestinal disease. Dis Colon Rectum 1999;42:231-40. 39. Martinez V, Tache Y. Role of CRF receptor 1 in central CRF-induced stimulation of colonic propulsion in rats. Brain Res 2001;893:29-35. 40. Stanton M, Hutson J, Simpson D, et al. Colonic manometry via appendicostomy shows reduced frequency, amplitude, and length of propagating sequences in children with slow-transit constipation. J Pediatr Surg 2005;40:1138-45. 41. Maeda H, Yamagata A, Nishikawa S, et al. Requirement of c-kit for development of intestinal pacemaker system. Development 1992;116: 369-75.