Caffeine affects early in vivo murine brain development by accelerating the evagination of the primitive neuroepithelium into telencephalic vesicles. In this model, caffeine induces the expression of the regulatory subunit alpha of protein kinase A (PKA RI alpha) and of Sonic hedgehog (Shh). The understanding of the molecular mechanisms linking caffeine and neural gene expression would benefit from a reproducible in vitro model. Accordingly, the present study aimed to determine whether caffeine modulated the expression of these genes in primary neuronal and astroglial cultures derived from developing murine neocortex. Using real-time PCR, the results showed that caffeine induced robust overexpression of Shh mRNA in both cell types without significantly modifying PKA RI alpha gene expression.