We investigated the effects of different protein kinase C (PKC) activators on Na+ currents using the conventional whole-cell and the inside-out macropatch voltage-clamp techniques in mouse neuroblastoma cells (N1E-115). Two different categories of PKC activators were investigated: the cis-unsaturated fatty acids (CUFAs): oleic (cis-9-octadecenoic), linoleic (cis-9-12-octadecadienoic), and linolenic acid (cis-9-12-15-octadecatrienoic), and, the diacylglycerol (DAG) derivative 1-2-dioctanoyl-sn-glycerol (DOG). These substances caused the following alterations on Na+ currents: (i) Na+ currents were attenuated as a function of voltage. While DOG attenuated both inward and outward Na+ currents in a monotonic and continuous voltage-dependent manner, CUFAs preferentially attenuated inward currents; (ii) the steady-state activation curve of Na+ currents shifted to more depolarized voltages; (iii) opposite to the activation curve, the steady-state inactivation curve of Na+ channels (h curve) shifted to more hyperpolarized voltages; (iv) the time course of inactivation development was accelerated by PKC activators, while the recovery from inactivation was not affected; (v) substances that inhibit different metabolic pathways (PKC activation, cyclooxygenase, lipooxygenase, and P-450 pathways) did not prevent the effects of PKC activators on Na+ currents. One fully saturated fatty acid (octadecanoic acid), a trans-unsaturated fatty acid (trans-9-octadecenoic), and different phorbol esters did not affect Na+ currents; (vi) effects of different PKC activators on Na+ currents were completely reversible. These observations suggest that PKC activators might interact with Na+ channels directly. These direct effects must be taken into consideration in evaluating the overall effect of PKC activation on Na+ channels. Moreover, it is likely that this direct interaction could account, at least in part, for the diversity of effects of PKC activators on Na+ channels.