Marie Louise Grøndahl

Secretory pathways in Salmonella Typhimurium- induced fluid accumulation in the porcine small intestine

The involvement of 5-hydroxytryptamine (5-HT) and 5-HT3 receptors and prostaglandin E2 (PGE2) in Salmonella Typhimurium-induced fluid accumulation in the porcine small intestine was investigated. Salmonella Typhimurium (10(8) and 10(10) cfu) and cholera toxin (CT; 20 microg) were instilled for 8 and 11 h in ligated loops in the porcine jejunum and ileum. Fluid accumulation and concentrations of Na+, K+, Cl-, 5-HT and PGE2 in the fluid accumulated in the loops were measured. The fluid accumulation was also measured when Salmonella Typhimurium (10(10) cfu) and CT (20 microg) were instilled for 8 h in ligated loops in jejunum and ileum in pigs given subcutaneous injections of saline or the 5-HT3 receptor antagonist ondansetron (200 microg/kg). Salmonella Typhimurium (10(10) cfu) and CT both induced fluid accumulation in jejunum and ileum after 8 and 11 h. Both treatments also induced an increase in luminal release of 5-HT and PGE2. The accumulated fluid was iso-osmotic and hyperosmotic in CT- and Salmonella Typhimurium-treated loops, respectively. Ondansetron reduced the Typhimurium-induced fluid accumulation in both jejunum and ileum by c. 40%, while it failed to reduce the response to CT. These results demonstrate that 5-HT and PGE2 are released and 5-HT3 receptors activated in the secretory pathway of Typhimurium in the porcine small intestine.

Transport of neutral, cationic and anionic amino acids by systems B, bo, +, XAG, and ASC in swine small intestine.

Amino acid influx across the brush border membrane of the intact pig ileal epithelium was studied. It was examine whether in addition to system B, systems ASC and b(o,+) were involved in transport of bipolar amino acids. The kinetics of interactions between lysine and leucine demonstrates that system b(o,+) is present and accessible also to L-glutamine. D-aspartate (K(1/2) 0.3 mM) and L-glutamate (K(i) 0.5 mM) share a high affinity transporter with a maximum rate of 1.3 micromol cm(-2) h(-1), while only L-glutamate with a K(1/2) of 14.4 mM uses a low affinity transporter with a maximum rate of 2. 7 micromol cm(-2) h(-1), system ASC, against which serine has a K(i) of 1.6 mM. In the presence of 100 mM lysine, L-glutamine (A), leucine (B), and methionine (C) fulfilled the criteria of the ABC test for transport by one and the same transporter. However, serine inhibits not only transport of L-glutamate but also of glutamine (K(i) 0.5 mM), and L-glutamate inhibits part of the transport of glutamine. The test does, therefore, only indicate that the three bipolar amino acids have similar affinities for transport by systems B and ASC. Further study of the function of system B must be carried out under full inhibition by lysine and glutamate.

Effect of age on vasoactive intestinal polypeptide-induced short-circuit current in porcine jejunum

Vasoactive intestinal polypeptide is a transmitter at the neuroepithelial junction of the small intestine in cholera toxin-induced secretion. We investigated whether the secretory effect in vitro of vasoactive intestinal polypeptide in porcine jejunum was changed with age. Stripped tissue preparations from three age groups, neonatal (7-11 days), young (6-8 weeks) and adult (13-15 weeks) pigs, were mounted in Ussing chambers and short-circuited. Vasoactive intestinal polypeptide produced concentration dependent increases in short-circuit current in all three age groups with EC50 values (in nM) of 14.5 +/- 1.9, 16.2 +/- 2.0 and 147 +/- 0 in neonatal, young and adult pigs, respectively. The peak increases in short-circuit current in adult pigs were significantly decreased compared with the other two age groups. To evaluate the secretory capacity, theophylline was added to tissue preparations in which baseline short-circuit current again was established. Theophylline caused a significantly lesser increase in short-circuit current in adult pigs (25.4 +/- 2.0 microA.cm-2) than neonatal (57.1 +/- 3.6 microA.cm-2) and young pigs (63.1 +/- 2.9 microA.cm-2). In conclusion, vasoactive intestinal polypeptide showed a marked decrease in the secretory response with age in porcine jejunum, at least partly caused by a reduced secretory capacity of the enterocytes.

Effect of Mucosal Amino Acids on SCC and Na and Cl Fluxes in the Porcine Small Intestine

Most amino acids are, like glucose, co-transported with sodium and are thereby potential additives to oral rehydration solutions for the treatment of diarrhea. In this study the effects of mucosal amino acids on short-circuit current and Na and Cl fluxes in three segments of the porcine small intestine were studied. L-alanine, L-leucine, L-lysine, L-proline, L-phenylalanine and L-glutamine were added (chamber concentration of each amino acid was 20 mmol x l-1) to the mucosal side of stripped proximal, mid, and distal small intestine sheets mounted in Ussing chambers in glucose containing (16 mmol x l-1) bathing medium; electrical parameters and unidirectional fluxes were measured. The amino acids induced a significant elevation in mucosa to serosa flux and unaltered serosa to mucosa flux of Na in all three segments, resulting in significant elevation in net Na absorption in proximal (2.4 +/- 0.3 microEq x cm-2 x hr-1 to 3.5 +/- 0.3 microEq x cm-2 x hr-1, P < 0.05), mid (5.9 +/- 0.5 microEq x cm-2 x hr-1 to 8.1 +/- 0.7 microEq x cm-2 x hr-1, P < 0.05) and distal (5.9 +/- 0.6 microEq x cm-2 x hr-1 to 8.3 +/- 0.7 microEq x cm-2 x hr-1, P < 0.05) small intestine. The mucosa to serosa Cl flux was significantly elevated in all three segments (P < 0.01) by amino acids abolishing the net Cl secretion in proximal (2.5 +/- 0.4 microEq x cm-2 x hr-1 to 0.8 +/- 0.6 microEq x cm-2 x hr-1, P < 0.01) and mid (2.2 +/- 0.4 microEq x cm-2 x hr-1 to 0.1 +/- 0.9 microEq x cm-2 x hr-1, P < 0.01) small intestine. The changes in SCC were carried by the electrogen Na absorption. In conclusion, the amino acids, in addition to glucose, stimulate Na absorption by about 1.0 microEq x cm-2 x hr-1 in the proximal part, and 2.5 microEq x cm-2 x hr-1 in the mid and distal part of the small intestine.

beta-Amino acid transport in pig small intestine in vitro by a high-affinity, chloride-dependent carrier.

This study describes unidirectional influx of amino acids and D-glucose across the small intestinal brush-border membrane of fully weaned eight week old pigs. Influx is minimal in the duodenum and maximal in the distal and/or mid small intestine. Influx of beta-alanine, taurine and N-methyl-aminoisobutyric acid is chloride-dependent. The activation stoichiometry for taurine influx is 1.0 +/- 0.2 chloride/2.4 +/- 0.3 sodium/1 taurine. Influx of D-glucose, lysine, glycine and glutamate is chloride-independent. An ABC test demonstrates a common beta-amino acid carrier: (a) the apparent affinity constant K1/2Taurine is 44 +/- 13 microM (means +/- S.D.) and the inhibitory constant (KiTaurine) against beta-alanine influx is 41 +/- 5 microM (means +/- S.E.). (b) K1/2beta-alanine is 97 +/- 23 microM and Kibeta-alanine against taurine influx is 160 +/- 22 microM. (c) KiHypotaurine against taurine and beta-alanine influx is 43 +/- 4 (n = 7) and 22 +/- 5 microM (n = 7), respectively. In conclusion, a high affinity, low capacity, sodium- and chloride-dependent carrier of beta-amino acids is present in pig small intestine.

EFFECT OF ONDANSETRON ON SALMONELLA TYPHIMURIUM-INDUCED NET FLUID ACCUMULATION IN THE PIG JEJUNUM IN VIVO

Two major pathophysiological mechanisms explaining the diarrhoea induced by Salmonella typhimurium have been suggested to be: (a) invasion of the intestine by the bacteria, and (b) an enterotoxin resembling Vibrio cholerae toxin. Cholera toxin is a potent secretagogue in pig small intestine and induces secretion partly by activating 5-hydroxytryptamine receptors, following release of 5-hydroxytryptamine. Ondansetron is a selective 5-hydroxytryptamine-3 receptor antagonist, which reduces the cholera toxin-evoked fluid accumulation in pig jejunum. The aim of this study was to investigate the effect of ondansetron on Salmonella typhimurium-induced fluid accumulation in ligated loops of pig jejunum in vivo. 10(10) colony-forming units of the bacteria was injected into loops and incubated for 8 hr. 200 mg x kg-1 ondansetron given subcutaneously reduced the Salmonella typhimurium-induced fluid accumulation by about 40%. This results suggests the involvement of 5-hydroxytryptamine and 5-hydroxytryptamine-3 receptors in Salmonella typhimurium-induced diarrhoea.