Heat stable enterotoxin (STa) from E.coli reduces net fluid absorption from rat proximal jejunum by interruption of sodium ion uptake. Stimulation by STa of epithelial cell chloride ion secretion could also contribute through associated fluid secretion. However, secretion is never detected in vivo with STa when a perfusate recovery method is used (Lucas et al., 2001). Experiments were therefore done under circumstances where sodium ion uptake and hence fluid uptake was minimised in the expectation that any underlying STa stimulated secretory process would become evident. Fluid uptake was investigated in vivo in anaesthetised (70 mg /kg i.p Sagatal) Wistar rats in recirculated perfused loops using recovered luminal volume to assess net fluid absorption. Loops of twenty five cm of proximal jejunum were perfused with buffers in which sodium ion was replaced by choline ion or by mannitol, with and without STa. In addition, some buffers contained 0.1 mM ethyl-iso-propyl-amiloride (EIPA) to inhibit any residual sodium ion uptake arising from nominally zero sodium ion perfusates. At the end of the experiment, the animals were humanely killed. Fluid absorption (mean +/-s.e (n)) from 150 mM saline was 56.1 +/- 10.0(6)µl/cm/hr v 28.6 +/- 4.5(7) µl/cm/hr in the presence of STa (p<0.05). Replacement of sodium ion by choline ion caused negative net fluid absorption of 13.3 +/- 4.0(7) µl/cm/hr, further reduced (p<0.05) to 30.4 +/- 5.5(6) µl/cm/hr when STa was added. When mannitol substituted for sodium chloride, negative fluid absorption was 11.9 +/- 4.9 (6) µl/cm/hr and increased to 32.4 +/-8.2(5) µl/cm/hr by STa. To determine whether the smaller effect of STa in the absence of sodium ion was dependent still on residual sodium ion uptake, the experiments were repeated with the NHE3 inhibitor EIPA additionally present. In the zero sodium ion perfusates with 01.uM EIPA negative fluid movement was 17.4 +/-5.7(6) µl/cm/hr and 16.6 +/-3.4(6) µl/cm/hr when STa was present. These experiments indicate that when fluid uptake is inhibited by inhibiting sodium ion uptake, STa does not have the expected further effect on fluid movement through additional secretion. These observations make unlikely the hypothesised epithelial cell chloride ion secretion, proposed as the basis of enterotoxin action (Field et al.,1978).