Neurons must function for decades of life, but how these non-dividing cells are preserved is poorly understood. Using mouse serotonin (5-HT) neurons as a model, we report an adult-stage transcriptional program specialized to ensure the preservation of neuronal connectivity. We uncover a switch in Lmx1b and Pet1 transcription factor function from controlling embryonic axonal growth to sustaining a transcriptomic signature of 5-HT connectivity comprising functionally diverse synaptic and axonal genes. Adult-stage deficiency of Lmx1b and Pet1 causes slowly progressing degeneration of 5-HT synapses and axons, increased susceptibility of 5-HT axons to neurotoxic injury, and abnormal stress responses. Axon degeneration occurs in a die back pattern and is accompanied by accumulation of α-synuclein and amyloid precursor protein in spheroids and mitochondrial fragmentation without cell body loss. Our findings suggest that neuronal connectivity is transcriptionally protected by maintenance of connectivity transcriptomes; progressive decay of such transcriptomes may contribute to age-related diseases of brain circuitry. Display omitted •Lmx1b and Pet1 sustain an adult-stage connectivity transcriptome in 5-HT neurons•Adult-stage loss of Lmx1b and Pet1 causes degeneration of 5-HT synapses and axons•APP and α-syn accumulate in spheroids and mitochondria fragment as fibers degenerate•Connectivity decay is accompanied by physiological and behavioral alterations Kitt et al. show that adult serotonergic connectivity is protected from expansive degeneration by an Lmx1b/Pet1-regulated transcriptional program. Adult-stage deficiency of these transcription factors causes progressive breakdown of 5-HT synapses and axons accompanied by accumulation of α-synuclein and amyloid precursor protein in spheroids and mitochondrial fragmentation without cell body loss.