The molecular mechanisms that mediate experience-based changes in the function of the cerebral cortex, particularly in the adult animal, are poorly understood. Here we show using in vivo voltage-sensitive dye imaging, that whisker trimming leads to depression of whiskerevoked sensory responses in primary, secondary and associative somatosensory cortical regions. Given the importance of cholinergic neurotransmission in cognitive and sensory functions, we examined whether ␣4-containing (␣4*) nicotinic acetylcholine receptors (nAChRs) mediate cortical depression. Using knock-in mice that express YFP-tagged ␣4 nAChRs subunits, we show that whisker trimming selectively increased the number ␣4*-YFP nAChRs in layer 4 of deprived barrel columns within 24 h, which persisted until whiskers regrew. Confocal and electron microscopy revealed that these receptors were preferentially increased on the cell bodies of GABAergic neurons. To directly link these receptors with functional cortical depression, we show that depression could be induced in normal mice by topical application or micro-injection of ␣4* nAChR agonist in the somatosensory cortex. Furthermore, cortical depression could be blocked after whisker trimming with chronic infusions of an ␣4* nAChR antagonist. Collectively, these results uncover a new role for ␣4* nAChRs in regulating rapid changes in the functional responsiveness of the adult somatosensory cortex.