We investigated the effects of 1-ethyl-2-benzimidazolinone (1-EBIO) on ion transport in the mouse jejunum through the use of the short-circuit (I _sc) current technique and the application of the patch-clamp technique to isolated jejunal crypts. In HCO₃ ^– Ringer's, 1-EBIO stimulated a dose-dependent (EC₅₀ 964 µmol/l), bumetanide-sensitive increase in I _sc consistent with stimulation of Cl^– secretion. In contrast, in Cl^–-free HCO₃ ^– Ringer's containing glucose, 1-EBIO (500 µmol/l) did not increase the phloridzin (100 µmol/l) sensitive I _sc, suggesting that electrogenic Na⁺ absorption was unaltered. Measurement of the membrane potential (V _m) with the perforated-patch technique indicated that in isolated crypts, 1-EBIO caused a reversible hyperpolarization of V _m and an increase in the change in V _m associated with step changes in bath K⁺, consistent with an increase in K⁺ conductance. In on-cell patch experiments with KCl Ringer's in the patch pipette and crypts bathed with NaCl Ringer's, 1-EBIO (500 µmol/l) increased the open probability (NP _o; 0.01±0.01 to 0.45±0.11, n=7) of an inwardly rectified intermediate conductance (g) channel. In inside-out patches with KCl Ringer's in the patch pipette and KCl Ringer's containing 100 nmol/l Ca²⁺ in the bath, the current-voltage relationship of the channel was inwardly rectified (g of 10 and 52 pS at –V _p of 100 and –100 mV, respectively) and reversed at 0 mV (n=5). Replacement of bath K⁺ with Na⁺ shifted the reversal potential toward the equilibrium potential for K⁺. In the presence of 1-EBIO, reducing the bath Ca²⁺ from 200 nmol/l to nominally Ca²⁺-free conditions decreased NP _o from 0.90±0.27 to 0.07±0.03 (n=3). We conclude that in the mouse jejunum, 1-EBIO does not stimulate electrogenic Na⁺ absorption. It does, however, stimulate secretion primarily through the activation of a basolateral, intermediate conductance Ca²⁺-sensitive K⁺ channel.