Although the p38 mitogen-activated protein kinases are active in many neuronal populations in the peripheral and central nervous systems, little is known about the physiological functions of p38 in postmitotic neurons. We report that p38 activity determines in vitro and in vivo the switch from noradrenergic to cholinergic neurotransmission that occurs in sympathetic neurons on exposure to the neuropoietic cytokines CNTF and LIF. This transdifferentiation serves as a model for the plastic mechanisms that enable mature neurons to change some of their central functions without passing through the cell cycle. We demonstrate that in postmitotic neurons, p38 and STAT pathways are concurrently activated by neuropoietic cytokine treatment for at least 12 h overlapping with changes in neurotransmitter marker gene expression. Inhibition of p38 blocks the upregulation of the nuclear matrix protein Satb2 and of cholinergic markers by CNTF without affecting STAT3 phosphorylation. Conversely, overexpression of p38 alpha or beta in the absence of cytokines stimulates cholinergic marker expression. The neurotransmitter switch in vitro is impaired in neurons isolated from p38 beta(-/-) mice. Consistent with these in vitro results, a substantial loss of cells expressing cholinergic properties is observed in vivo in the stellate ganglion of mature mice deficient in the p38 beta isoform.