In cultured microglial cells pro-inflammatory substances such as lipopolysaccharide and interferon-gamma induce outward-rectifying K+ channels (OR) exhibiting features of the Kv1.3 type. Here we studied the modulation of this channel by arachidonic acid (AA). Micromolar doses of AA (0.3-30 microM; ED50 1.55 microM) depressed OR currents at all the potentials tested. Such effect appeared in less than 30 s, reached the maximum in approximately 2 min, and partially reverted upon removal of AA. We then tested whether AA acted by mechanisms involving enzyme activation. The AA effect on OR remained unchanged in the presence of staurosporine (protein kinase C inhibitor), indomethacin (cyclooxygenase inhibitor), nordihydroguaiaretic acid (lipoxygenase inhibitor), and diphenylene iodonium (NADPH-oxidase inhibitor). The same effect was present in cell-free membrane patches in the outside-out configuration. In whole-cell recording AA induced the following changes in OR current: (a) decrease of OR current peak at all voltages tested; (b) increase of the activation rate and mild shift of the voltage-dependence of the activation; (c) acceleration of the inactivation rate with a concomitant appearance of a second and faster inactivation component; and (d) shift of the voltage-dependence of the inactivation curve. We also observed that upon AA application, the acceleration of activation and inactivation developed earlier than the second component of inactivation. We conclude that AA, by acting on both the channel protein and its lipid environment, causes a quick negative modulation of microglial OR current. We propose that a raised free AA may determine a loss of efficiency of OR in controlling the membrane potential and thus influence the functional reactivity of microglia to inflammatory stimuli.