display the size of the preparation, while black represents the ratio of ,1 in musculature preparations, because the size of preparations were larger than the frame of the image. After 5- HT application the active area markedly increased. (TIF) Figure S2 Regular occurrence of pacemaker [Ca2+]i oscillations after application of 5-HT. A) Ca2+ images acquired from a cell cluster preparation in control (a–c) and 5 min after 5-HT (10 mM) application (d, e). This preparation showed intermittent [Ca2+]i oscillations in control condition. B) Time course of pacemaker [Ca2+]i activity recorded in the square (x) indicated in A: control (left) and 5 min after application of 10 mM 5-HT (right). Dotted lines correspond to the times when images (a–e) were acquired. C– E) Bar graphs showing changes in the peak amplitude (C), frequency (D), and active area (E) of spontaneous pacemaker [Ca2+]i activity (Mean 6 S.D., n = 6). (TIF) Figure S3 An example of the effect of 2-Me-5-HT (10 mM) on field potentials. Field potential images in control (A) and during application of 2-Me-5-HT (B) are displayed at 200 ms intervals. Note, 2-Me-5-HT-enhanced ICC electrical activity, as seen during 5-HT application. (TIF) Figure S4 Possible underlying mechanisms for 5-HT-enhance- ment of gut pacemaker activity and contractility. It is thought that intracellular Ca2+ release channels, i.e. ryanodine receptors (RyR) and InsP3 receptors (IP3R) in ICC are periodically activated by the support of a Ca2+ influx pathway across the plasma membrane, although mechanisms underlying the coordinated actions of intracellular and plasma membrane ion channels are not yet known. 5-HT augments ICC [Ca2+]i oscillations presumably (1) by facilitating Ca2+ influx via 5-HT3 receptors, and (2) simultaneous activation of voltage-gated Ca2+-permeable insensitive to DHP Ca2+ antagonists (DHP(-)VGCC). In ICC, [Ca2+]i oscillations periodically activate Ca2+-activated ion channels in the plasma- membrane, i.e. Ca2+-activated Cl2 channels (ClCa) and/or Ca2+- References 1. Maeda H, Yamagata A, Nishikawa S, Yoshinaga K, Kobayashi S, et al. (1992) Rrequirement of c-kit for development of intestinal pacemaker system. Development 116: 369–375. 2. Faussone-Pellegrini MS, Thuneberg L (1999) Guide to the identification of interstitial cells of Cajal. Microsc Res Tech 47: 248–266. 3. Sanders KM, O¨rdo¨g T, Koh SD, Torihashi S, Ward SM (1999) Development and plasticity of interstitial cells of Cajal. Neurogastroenterol Motil 11: 311–338. 4. Huizinga JD (1999) Gastrointestinal peristalsis: joint action of enteric nerves, smooth muscle, and interstitial cells of Cajal. Microsc Res Tech 47: 239–247. 5. Camilleri M (2001) Enteric nervous system disorders: genetic and molecular insights for the neurogastroenterologist. Neurogastroenterol Motil 13: 277–295. 6. 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