Rett syndrome is characterized by the postnatal loss of neurophysiological function and regression of childhood development. Because the syndrome is X linked, and males with MECP2 mutations generally do not survive birth, the study of this syndrome has been complicated by the fact that, in the female brain, a portion of neurons express wild-type MECP2, and another portion of neurons express a non-functional allele of MECP2. Here, we present an approach that enables transcriptional profiling of individual neurons and direct comparison of neurons that express functional MECP2 with those that have diminished MECP2 function. With this novel profiling approach, we found that mutant neurons from the brains of patients with Rett syndrome show patterns of defects in expression of synaptic and metabolic genes. A similar analysis of rat brain lacking MECP2 expression yielded similar patterns, suggesting that rat is a suitable in vivo model of Rett syndrome. These analyses also identified DNA damage and senescence transcriptional signatures specifically in MECP2-null neurons, suggesting a possible trigger of dysfunction in Rett syndrome. Together, these data highlight potentially defective molecular, physiological and metabolic pathways in brain neurons of patients with Rett syndrome.