CALCIUM-DEPENDENT INTESTINAL CHLORIDE SECRETION BY VIBRIO-PARAHAEMOLYTICUS THERMOSTABLE DIRECT HEMOLYSIN IN A RABBIT MODEL

Background & Aims: Vibrio parahaemolyticus is a major agent of seafood gastroenteritis that induces intestinal secretion in the rabbit throu gh its thermostable direct hemolysin. The aim of this study was to cha racterize the enterotoxicity of purified hemolysin in vitro. Methods: Rabbit ileum was mounted in Ussing chambers, and changes in potential difference and short-circuit current were monitored after addition of hemolysin. Intracellular calcium concentrations in the nontumoral rat crypt-derived cell line IEC-6 were measured using microspectrofluorome try. Results: In Ussing chamber experiments, mucosal toxin addition up to 50 hemolytic units per milliliter induced a proportional increase of the electrical parameters in normal but not Cl--free Ringer's solut ion. The response to the toxin was not additive to that of calcium ion ophore A23187 and was eliminated by preloading the tissue with 1-2-bis (o-aminophenoxy)ethane N,N,N',N'-tetraacetic acid (BAPTA), a calcium b uffer. In IEC-6 cells, a 10-fold increase in intracellular calcium lev el was found after addition of hemolysin. Such an increase was totally quenched by BAPTA. Finally, preincubation with trisialoganglioside GT 1b, but not monosialoganglioside GM1, eliminated toxin-induced increas es in potential difference and short-circuit current. Conclusions: The se data support the hypothesis that the thermostable direct hemolysin induces intestinal chloride secretion using GT1b as a putative recepto r and Ca2+ as a second messenger.