Quantitative measurement of mRNA coding for the receptors controlling acidsecretion in the ovine fundus and antrum by using RT-PCR

Background and Aims: Gastric acid secretion is stimulated by the action of gastrin, histamine and acetylcholine on their respective receptors. To dete rmine the regulation of synthesis of these receptors during different gastr ic secretory states a competitive RT-PCR method for quantitating the mRNA f or these receptors was developed. Methods: Partial cDNA clones (400-500 base pairs (bp)) for the ovine gastri n, histamine (H-2) and acetylcholine (M-3) receptors were isolated and sequ enced. These cDNA constructs were modified by the inclusion of approximatel y 100 bp of unrelated sequence within the plasmids. cDNA was synthesized fr om a mixture of known amounts of RNA transcribed from the modified plasmids and from total RNA extracted from sheep stomach. Proportional coamplificat ion of mixed cDNA was demonstrated using common primer sets. Results: All three receptors were more highly expressed in the fundus than the antrum. The concentration of cholecystokinin-B/gastrin receptor mRNA wa s 75-fold higher in the fundus than in the antrum, and the concentration of both histamine and acetylcholine receptor mRNA were fivefold higher in the fundus than in the antrum. Infusion of gastrin caused a significant increa se in fundic histamine mRNA receptor expression, but not in the expression of the gastrin or muscarinic receptors. Conclusions: No significant differences were observed in the levels of rece ptor mRNA between normal adult and fetal animals despite markedly reduced g astric secretion in the fetus, suggesting that gastric receptor gene expres sion is not the rate-limiting factor in determining gastric acid secretion in the neonatal animal. (C) 2000 Blackwell Science Asia Pty Ltd.