The role of Ca2+ in insect olfactory transduction was studied in the moth Spodoptera littoralis. Single sensillum recordings were made to investigate in vivo the role of sensillar Ca2+ on the electrophysiological properties of sex pheromone responsive olfactory receptor neurons (ORNs). Lowering the sensillar Ca2+ concentration to 2 x 10(-8) M increased ORN spontaneous firing activity and induced long bursts of action potentials (APs) superimposed on spontaneous negative deflections of the transepithelial potential. We inferred that Ca2+ stabilizes the membrane potential of ORNs, keeping the spontaneous firing activity at a low and regular level. Neither the amplitude and kinetics of the rising phase of sensillar potentials (SPs) recorded in response to pheromone stimuli nor the AP generation during stimulation depended on the extracellular Ca2+ concentration. Thus, extracellular Ca2+ is not absolutely necessary for ORN response. Partial inhibition of responses with a calmodulin antagonist, W-7, also indicates that intracellular Ca2+ contributes to the ORN response and suggests that Ca2+ release from internal stores is involved. In 2 x 10(-8) M Ca2+, the repolarization of the SP was delayed when compared with higher Ca2+ concentrations. Therefore, in contrast to depolarization, ORN repolarization depends on extracellular Ca2+. Ca2+-gated K+ channels identified from cultured ORNs with whole-cell recordings are good candidates to mediate ORN repolarization.