The 5-HT 3 receptors are serotonin-gated ion channels that physically couple with purinergic P2X2 receptors to trigger a functional cross-inhibition leading to reciprocal channel occlusion. Although this functional receptor–receptor coupling seems to serve a modula-tory role on both channels, this might not be its main physiological purpose. Using primary cultures of rat hippocampal neurons as a quantitative model of polarized targeting, we show here a novel function for this interaction. In this model, 5-HT 3A receptors did not exhibit by themselves the capability of distal targeting in dendrites and axons but required the presence of P2X2R for their proper subcellular localization. 5-HT 3A R distal targeting occurred with a delayed time course and exhibited a neuron phenotype dependency. In the subpopulation of neurons expressing endogenous P2X2R, 5-HT 3A R distal neuritic localization correlated with P2X2R expression and could be selectively inhibited by P2X2R RNA interference. Cotransfection of both receptors revealed a specific colocalization, cotraffick-ing in common surface clusters, and the axonal rerouting of 5-HT 3A R. The physical association between the two receptors was dependent on the second intracellular loop of the 5-HT 3A subunit, but not on the P2X2R C-terminal tail that triggers the functional cross-inhibition with the 5-HT 3A R. Together, these data establish that 5-HT 3A R distal targeting in axons and dendrites primarily depends on P2X2R expression. Because several P2XR have now been shown to functionally interact with several other members of the 4-TMD family of receptor channels, we propose to reconsider the real functional role for this receptor family, as trafficking partner proteins dynamically involved in other receptors targeting.