Rett syndrome is a neurodevelopmental disorder caused by mutations in the methyl-CpG binding protein 2 gene (MECP2). Several neural systems are affected in Rett, resulting in an autonomic dysfunction, a movement disorder with characteristic loss of locomotor abilities and profound cognitive impairments. A deregulation of monoamines has been detected in the brain and cerebrospinal fluid of both Rett patients and a Rett syndrome murine model, the Mecp2 knock-out mouse. Our goal was to characterize the onset and progression of motor dysfunction in Mecp2^tm1.1Bird knock-out mice and the possible neurochemical alterations in different brain regions potentially playing a role in Rett-like pathophysiology, at two different time-points, at weaning (3 weeks old) and in young adults when overt symptoms are observed (8 weeks old). Our results revealed significant age- and region-dependent impairments in these modulatory neurotransmitter systems that correspond well with the motor phenotype observed in these mice. At 3 weeks of age, male Mecp2 knock-out mice exhibited ataxia and delayed motor initiation. At this stage, noradrenergic and serotonergic transmission was mainly altered in the prefrontal and motor cortices, whereas during disease progression the neurochemical changes were also observed in hippocampus and cerebellum. Our data suggest that the deregulation of norepinephrine and serotonin systems in brain regions that participate in motor control are involved in the pathophysiology of Rett syndrome motor phenotypes. Moreover, we highlight the contribution of cortical regions along with the brainstem to be in the origin of the pathology and the role of hippocampus and cerebellum in the progression of the disease rather than in its establishment.