Numerous in vivo studies have demonstrated that psychostimulant drugs such as amphetamine and cocaine can induce the expression of the immediate early gene c-fos in striatal neurons via the activation of D1 dopamine receptors. NMDA receptor activation is also known to induce c-fos in the striatum. In the present study we have used a primary striatal neuronal culture preparation to examine the mechanisms whereby these stimuli lead to changes in gene expression. Direct application of NMDA to striatal cells in culture caused a rapid increase in the expression of c-fos as well as an increase in the phosphorylation of the transcription factor CRE binding protein (CREB). This was prevented by NMDA receptor antagonists, and required extracellular calcium, but did not involve L-type calcium channels. The induction of c-fos and CREB phosphorylation following NMDA were unaffected by inhibition of protein kinase C; tyrosine kinases or nitric oxide synthase. However, the response to NMDA was blocked by KN62, a selective inhibitor of calcium/calmodulin-dependent protein kinase. Application of the D1 agonist SKF 38393, or direct stimulation of adenylyl cyclase with forskolin, also resulted in the phosphorylation of CREB and the induction of c-fos in striatal neurons. These effects were blocked by the protein kinase A inhibitor H89. These observations are consistent with the hypothesis that calcium/calmodulin-dependent phosphorylation of CREB induced by NMDA, or cAMP-dependent phosphorylation of CREB induced by D1 agonists, underlie the induction of c-fos seen following activation of these receptors in striatal neurons.