Signaling through nicotinic acetylcholine (nACh) receptors underlies a diverse array of behaviors. In order for appropriate signaling to occur via nACh receptors, it is necessary for the genes encoding the receptor subunits to be expressed in a highly regulated temporal and spatial manner. Here we report a transgenic mouse approach to characterize the transcriptional regulation of the gene encoding the nACh receptor beta4 subunit. nACh receptors containing this subunit play critical roles in both the central and peripheral nervous systems. We demonstrate that a 2.3-kilobase pair fragment of the beta4 5'-flanking region is capable of directing reporter gene expression in transgenic animals. Importantly, the transcriptional activity of the promoter region is cell-type-specific and developmentally regulated and overlaps to a great extent with endogenous beta4 mRNA expression. These data indicate that the 2.3-kilobase pair fragment contains transcriptional regulatory elements critical for appropriate beta4 subunit gene expression.