Submitted on January 26, 2005
Accepted on August 12, 2005
Subtractive hybridization unravels a role for the ion co-transporter NKCC1 in the murine intestinal pacemaker
Mira Wouters1, Ann De Laet2, Luc Ver Donck3, Eric Delpire4, Pierre-Paul van Bogaert5, Jean-Pierre Timmermans6, Alban de Kerchove d'Exaerde7, Karine Smans3, and Jean-Marie Vanderwinden7*
1 Laboratoire de Neurophysiology, Faculte de Medecine, Universite Libre de Bruxelles, Bruxelles, Belgium; Department of Gastro-intestinal Pharmacology, Johnson and Johnson, Pharmaceutical Research and Development, a Division of Janssen Pharmaceutica N.V., Beerse, Belgium
2 Laboratory of Cell Biology and Histology, University of Antwerp, Antwerp, Belgium; Laboratory of Electrobiology, University of Antwerp, Antwerp, Belgium
3 Department of Gastro-intestinal Pharmacology, Johnson and Johnson, Pharmaceutical Research and Development, a Division of Janssen Pharmaceutica N.V., Beerse, Belgium
4 Department of Anesthesiology, Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN, USA
5 Laboratory of Electrobiology, University of Antwerp, Antwerp, Belgium
6 Laboratory of Cell Biology and Histology, University of Antwerp, Antwerp, Belgium
7 Laboratoire de Neurophysiology, Faculte de Medecine, Universite Libre de Bruxelles, Bruxelles, Belgium
* To whom correspondence should be addressed. E-mail: jmvdwin{at}ulbe.ac.be.
In the small intestine, interstitial cells of Cajal (ICC) surrounding the myenteric plexus generate
the pacemaking slow waves that are essential for an efficient intestinal transit. The underlying
molecular mechanisms of the slow wave are poorly known. Our aim was to identify ICC-specific
genes and their function in the mouse jejunum.
Suppression subtractive hybridization using two independent ICC-deficient mouse models
identified 56 genes putatively downregulated in the muscularis propria compared to wild type
littermates. Differential expression was confirmed by real time quantitative PCR for the tyrosine
kinase receptor KIT, the established marker for ICC, and for the Na+-K+-2Cl- cotransporter
NKCC1. Immunoreactivity for NKCC1 was detected in myenteric ICC but not in the ICC
population located at the deep muscular plexus. NKCC1 was also expressed in enteric neurons
and mucosal crypts. Bumetanide, a NKCC1 inhibitor, reversibly affected the shape, amplitude
and frequency of the slow waves. Similar alterations were observed in NKCC1 knock out mice.
These data support the hypothesis that NKCC1 expressed in myenteric ICC is involved in the
mechanism of slow waves in the murine jejunum.
