The 5-hydroxytryptamine(4) (5-HT(4)) receptors have recently emerged as key modulators of learning, memory, and cognitive processes. In neurons, 5-hydroxytryptamine(4) receptors (5-HT(4)Rs) activate cAMP production and protein kinase A (PKA); however, nothing is known about their ability to activate another key signaling pathway involved in learning and memory: the extracellular signal-regulated kinase (ERK) pathway. Here, we show that 5-HT(4)R stimulation, in primary neurons, produced a potent but transient activation of the ERK pathway. Surprisingly, this activation was mostly PKA independent. Similarly, using pharmacological, genetic, and molecular tools, we observed that 5-HT(4)Rs in human embryonic kidney 293 cells, activated the ERK pathway in a G(s)/cAMP/PKA-independent manner. We also demonstrated that other classical G proteins (G(q)/G(i)/G(o)) and associated downstream messengers were not implicated in the 5-HT(4)R-activated ERK pathway. The 5-HT(4)R-mediated ERK activation seemed to be dependent on Src tyrosine kinase and yet totally independent of beta-arrestin. Immunocytofluorescence revealed that ERK activation by 5-HT(4)R was restrained to the plasma membrane, whereas p-Src colocalized with the receptor and carried on even after endocytosis. This phenomenon may result from a tight interaction between 5-HT(4)R and p-Src detected by coimmunoprecipitation. Finally, we confirmed that the main route by which 5-HT(4)Rs activate ERKs in neurons was Src dependent. Thus, in addition to classical cAMP/PKA signaling pathways, 5-HT(4)Rs may use ERK pathways to control memory process.