GABAA receptor subunit messenger RNA expression in the enteric nervous system of the rat: implications for functional diversity of enteric GABAA receptors
Section snippets
Experimental procedures
All tissues used in this study were obtained from male Sprague–Dawley rats (125–175 g; Charles River, Canada) killed by decapitation according to accepted standards of the University of Ottawa and National Research Council animal care commitees. Each animal was subjected to a laparotomy and the small intestine exposed for resection of the ileum. Segments of the proximal ileum (6 cm in length) were removed into an oxygenated Krebs' buffer, maintained at 37°C and pH 7.2. The individual segments
Reverse transcription–polymerase chain reaction
GABA subunit mRNAs were detected using subunit-specific primers in RNA isolated from homogenates of gut wall free of mucosa. After reverse transcription (RT) and PCR amplification of the cDNA, bands of the predicted size were observed in ethidium bromide-stained agarose gels. This assay consistently detected α1, α3, α5, β2, β3, γ1 and γ3 subunit mRNAs (Fig. 1). However, α4 and α6 message were detected at threshold levels in two out of six samples. This was not thought to be the result of
Discussion
In order to determine how the enteric GABAergic system is integrated in the neural control of gut function we have undertaken the first comprehensive study of the subunit expression profile of GABAA receptors within the nerve networks of the intestine. This information provides a framework for further functional and pharmacological studies since we have found a distinctive and heterogeneous distribution of GABAA receptor subunit mRNAs. In contrast to the CNS, α3, γ1 and γ3 subunit mRNAs were
Acknowledgements
This work was supported by the Medical Research Council of Canada; National Research Council of Canada; Canadian Association of Gastroenterology Studentship to R.S.
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