Production and action of local mediators in the rat gastric mucosa

Abstract

This study concerns the production and action of the local mediators nitric oxide (NO) and prostaglandin E₂ (PGE₂) in the rat gastric mucosa. The major objectives were: (i) to determine which mucosal cell type(s) contained NO synthase activity, (ii) to establish the functional role(s) of NO in the gastric mucosa and (iii) to investigate regulation of gastric PGE₂ production.
Gastric mucosal cells were isolated by pronase digestion coupled with intermittent calcium chelation and were separated by either density-gradient centrifugation or by counterflow elutriation. The distribution of Ca²⁺ -dependent NO synthase activity, measured via the conversion of [¹⁴C]-L-arginine to [¹⁴C]-L- citrulline, paralleled the distribution of mucous cells in elutriated fractions. Pre-treatment of rats with lipopolysaccharide caused the induction of Ca²⁺ -independent NO synthase in the elutriator fractions enriched with mucous cells.
Incubation of isolated cells with the NO donor isosorbide dinitrate (ISDN) produced a concentration-dependent increase in the guanosine 3',-5'-cyclic monophosphate (cGMP) content which was accompanied by a concentration-dependent increase in release of immunoreactive mucin. Intragastric administration of ISDN of dibutyryl cGMP in vivo increased the thickness of the mucus layer overlying the gastric mucosa.
The NO donor S-nitroso-N-acetylpenicillamine (SNAP) produced a concentration-dependent inhibition (IC₅₀ 247 μM) of histamine-stimulated aminopyrine accumulation, a measure of secretory activity, in cell suspensions containing > 80% parietal cells. SNAP increased the cGMP content of the suspension but did not decrease cellular viability, glucose oxidation or adenosine 3',5'-cyclic monophosphate content. The inhibitory effect of SNAP was observed in permeabilised cells stimulated with ATP and was stereospecifically blocked by preincubation with Rp-8-bromoguanosine 3'-5'-monophosphorothioate, which inhibits activation of cGMP-dependent protein kinase.
Stimulation of PGE₂ release by bradykinin in a low density cell fraction, enriched
with parietal cells and devoid of vascular endothelial cells and macrophages, involved a bradykinin B₁ receptor.
In summary, NO synthase activity is probably present in gastric mucous epithelial
cells. NO may promote mucus secretion by elevation of cGMP. NO donors inhibit acid secretion at a specific site and their action may involve cGMP. The bradykinin B₁ receptor is involved with PGE₂ production in the gastric mucosa.

Date of Award	Mar 1994
Original language	English
Supervisor	Peter J Hanson (Supervisor) & B.J.R. Whittle (Supervisor)