The transport properties of Mongolian gerbil middle ear epithelial cells grown in primary culture were studied. These cells formed polarized monolayers that exhibited domes on nonporous supports. On porous supports, monolayers developed an apical-negative transepithelial electric potential difference (VT = -37.2 +/- 2.7 mV) and a transepithelial resistance (RT = 519 +/- 56 omega.cm2). The short-circuit current equivalent (Ieq) was 62.4 +/- 6.2 microA/cm2 (mean +/- SE, n = 15). Na+ and Cl- accumulated in the basal bath and generated a basolateral hyperosmolarity that drove a net water flow. Amiloride (10 microM), when added to the apical but not to the basal bath, induced a 23.4 +/- 1.5 mV and 44.1 +/- 1.3 microA/cm2 decrease of VT and Ieq, respectively, while RT increased by 403 +/- 69 omega.cm2 (P less than 0.001, n = 15). Exposure of the monolayers to a low-Cl- solution (30 mM) enhanced the transepithelial potential, possibly by means of a Cl- secretion through apical Cl- channels. Isoproterenol (10(-4) M basolateral) increased intracellular adenosine 3',5'-cyclic monophosphate (cAMP) content (concentration of half-maximal response = 2.5 x 10(-7) M) and decreased VT, RT, and Ieq. The isoproterenol-induced fall of VT occurred even in the presence of low-Cl-solutions. This suggested an increase of the paracellular pathway conductance, although there was no significant modification of the mannitol permeability.(ABSTRACT TRUNCATED AT 250 WORDS)