Gasotransmitters do not prevent changes in transepithelial ion transport induced by hypoxia followed by reoxygenation

Abstract

Objectives: How gaseous signalling molecules affect ion transport processes contributing to the physiological functions of the gastrointestinal tract under hypoxic conditions still needs to be clarified. The objective of the present study was to characterize the impact of gaseous signalling molecules on parameters of colonic ion transport during a hypoxia/reoxygenation cycle and the remaining secretory capacity of the epithelium after such a cycle.

Methods: Short-circuit current (I_sc) and tissue conductance (G_t) recordings in Ussing chamber experiments were performed on rat colon samples using CORM-2 (putative CO donor; 35 and 350 µM), sodium nitroprusside (NO donor; 100 µM), NaHS (fast H₂S donor; 10 - 1,000 µM), GYY 4137 (slow H₂S donor; 50 µM) and Angeli's salt (HNO donor; 100 µM) as donors for gasotransmitters. Inhibition of endogenous synthesis of H₂S was operated by inhibitors of cystathionin-γ-lyase, i.e. dl-propargylglycine (1 mM) or β-cyano-l-alanine (5 mM), and the inhibitor of cystathionine-β-synthase, amino-oxyacetate (5 mM).

Results: The fast gasotransmitter donors NaHS, sodium nitroprusside and Angeli's salt, administered 5 min before the onset of hypoxia, induced an increase in I_sc. The response to the subsequently applied hypoxia was characterized by a decrease in I_sc, which tended to be reduced only in the presence of the lowest concentration of NaHS (10 µM) tested. Reoxygenation resulted in a slow increase in I_sc, which was unaffected by all donors or inhibitors tested. The stable acetylcholine derivative carbachol (50 µM) was administered at the end of each hypoxia/reoxygenation cycle to test the secretory capacity of the epithelium. Pretreatment of the tissue with the putative CO donor CORM-2 suppressed the secretory response induced by carbachol. The same was observed when cystathionin-γ-lyase and cystathionin-γ-synthase were inhibited simultaneously. Under both conditions, G_t drastically increased suggesting an impaired tissue integrity.

Conclusions: The present results demonstrate that none of the exogenous gasotransmitter releasing drugs significantly ameliorated the changes in epithelial ion transport during the hypoxia/reoxygenation cycle ex vivo. In contrast, the putative CO donor CORM-2 exerted a toxic effect on the epithelium. The endogenous production of H₂S, however, seems to have a protective effect on the mucosal integrity and the epithelial transport functions, which - when inhibited - leads to a loss of the secretory ability of the mucosa. This observation together with the trend for improvement observed with a low concentration of the H₂S donor NaHS suggests a moderate protective role of low concentrations of H₂S under hypoxic conditions.

Keywords: colon; electrolyte transport; gaseous signalling molecules; hypoxia and reoxygenation.