The monocular deprivation model of amblyopia is characterized by a reduction in cortical responses to stimulation of the deprived eye. Although the effects of monocular deprivation on the primary visual cortex have been well characterized physiologically and anatomically, the molecular mechanisms underlying ocular dominance plasticity remain unknown. Previous studies have indicated that the transcription factor adenosine cAMP/Ca(2+) response element-binding protein (CREB) is activated during monocular deprivation. However, it remains unknown whether CREB function is required for the loss of cortical responses to the deprived eye. To address this issue, we used the herpes simplex virus (HSV) to express a dominant negative form of CREB (HSV-mCREB) containing a single point mutation that prevents its activation. Quantitative single-unit electrophysiology showed that cortical expression of this mutated form of CREB during monocular deprivation prevented the loss of responses to the deprived eye. This effect was specific and not related to viral toxicity, because overexpression of functional CREB or expression of beta-galactosidase using HSV injections did not prevent the ocular dominance shift during monocular deprivation. Additional evidence for specificity was provided by the finding that blockade of ocular dominance plasticity was reversible; animals treated with HSV-mCREB recovered ocular dominance plasticity when mCREB expression declined. Moreover, this effect did not result from a suppression of sensory responses caused by the viral infection because neurons in infected cortex responded normally to visual stimulation. These findings demonstrate that CREB function is essential for ocular dominance plasticity.