Otilonium bromide inhibits muscle contractions via L-type calcium channels in the rat colon M. T. MARTIN, L. HOVE-MADSEN & M. JIMENEZ Department of Cell Biology, Physiology and Immunology, Universitat Auto ´ noma de Barcelona (UAB), Barcelona, Spain Abstract The aim of this study is to evaluate in vitro the effect of otilonium bromide (OB) on the mechanical and electrical activities of the rat colonic smooth muscle using muscle bath, microelectrodes and patch-clamp techniques. Otilonium bromide dose dependently inhibited the spontaneous activity (logIC 50 ± SE: )5.31 ± 0.05). This effect was not modified by TTX (10 )6 mol L )1 ). Cyclic depolariza- tions were abolished by OB (10 )4 mol L )1 ). Electrical field stimulation induced inhibitory junction poten- tials (IJPs) followed by a depolarization with super- imposed spikes causing a contraction. In the presence of OB (10 )4 mol L )1 ) IJPs were recorded, but spikes and contractions were abolished. Otilonium bromide (3 · 10 )6 mol L )1 ) inhibited inward current obtained in isolated cells (amphotericin perforated patch technique). The otilonium-sensitive current ampli- tude was maximal (75pA) around 0 mV. The effect of different doses of OB was tested by depolarizing cells from )70 mV to 0 mV. OB dose dependently inhib- ited the inward current with an EC 50 of 885 nmol L )1 . Abolishment of the otilonium-sensitive current by 3 · 10 )6 mol L )1 nifedipine confirmed that it was an L-type Ca 2+ current. Our results show that OB inhibits the spontaneous and triggered muscular contractions. This effect is produced by the inhibition of muscular action potentials carried by L-type cal- cium current, confirming the spasmolytic properties of OB. Keywords electrophysiology, muscle bath, otilonium bromide, patch-clamp, rat colon. INTRODUCTION Otilonium bromide (OB) is a spasmolytic agent widely used for the treatment of gastrointestinal diseases such as irritable bowel syndrome. 1,2 OB reduces abdominal pain and discomfort, 3 and enhances sensory thresholds to recto-sigmoideal distention. 4 Several pharmacologi- cal properties have been attributed to OB such as calcium channel blocker, muscarinic and tachykinin (NK2) receptor antagonist. 5,6 Previous studies have reported that OB has antimuscarinic properties and also binds the L-type calcium channel in the rat colon. 7 In rat chromaffin cells, OB acts as an L-type calcium channel blocker. 8 On the other hand, this substance has a poor systemic absorption, but it penetrates and accumulates in the walls of the large intestine of the rat colon. 9 All these characteristics suggest that OB has excellent properties as spasmolytic drug that can be used in a wide variety of diseases including the irritable bowel syndrome. The rat is a well-characterized model to study colonic motility. We have recently reported that the rat colon has two types of contractions: low fre- quency with high amplitude contractions and high frequency with low amplitude contractions. High amplitude contractions probably participate in pro- pulsive motility, whereas low amplitude contractions are local movements that contribute to mixing and water absorption. Both types of contractions are originated by spontaneous cyclic depolarizations and slow wave activity. 10,11 High amplitude contractions can be recorded in vitro and in vivo and have been associated with Ôgiant contractionsÕ that can occur in humans. 12–14 In the rat colon, inhibitory neurotrans- mission is mediated by both nitric oxide and ATP, although other neurotransmitters such as PACAP can also be involved. 10,15 ATP or PACAP might be related to the fast component of the inhibitory junction potential (IJP) which is apamin-sensitive, whereas nitric oxide participates in the sustained component of the IJP. Address for correspondence Marcel Jimenez, Department of Physiology, Veterinary Faculty, Universitat Auto ` noma de Barcelona 08193, Bellaterra, Barcelona, Spain. Tel: 34 935811898; fax: 34 935812006; e-mail: marcel.jimenez@uab.es Received: 31 October 2003 Accepted for publication: 28 January 2004 Neurogastroenterol Motil (2004) 16, 175–183 Ó 2004 Blackwell Publishing Ltd 175