Currents through anion channels in the plasma membrane of Lilium longiflorum pollen grain protoplasts were studied under conditions of symmetrical anionic concentrations by means of patch-clamp whole-cell configuration. With Cl(-)-based intra-and extracellular solutions, three outward-rectifying anion conductances, I(Cl1), I(Cl2) and I(Cl3), were identified. These three activities were discriminated by differential rundown behaviour and sensitivity to 5-nitro-2-(phenylpropylamino)-benzoate (NPPB), which could not be attributed to one or more channel types. All shared strong outward rectification, activated instantaneously and displayed a slow time-dependent activation for positive potentials. All showed modulation by intracellular calcium ([Ca(2+)](in)), increasing intensity from 6.04 nM up to 0.5 mM (I(Cl1)), or reaching a maximum value with 8.50 mu M (I(Cl2) and I(Cl3)). After rundown, the anionic currents measured using NO(3)(-)-based solutions were indistinguishable, indicating that the permeabilities of the channels for Cl(-) and NO(3)(-) are similar. Additionally, unitary anionic currents were measured from outside-out excised patches, confirming the presence of individual anionic channels. This study shows for the first time the presence of a large anionic conductance across the membrane of pollen protoplasts, resulting from the presence of Ca(2+)-regulated channels. A similar conductance was also found in germinated pollen. We hypothesize that these putative channels may be responsible for the large anionic fluxes previously detected by means of self-referencing vibrating probes.