show Abstracthide AbstractNeuronal activity is temperature sensitive and affects important traits, such as behavior. Yet we know next to nothing about the evolutionary response of neuronal phenotypes in new temperature environments. Here we use long-term experimental evolution of Drosophila simulans populations exposed to novel temperature regimes. For the first time, we demonstrate a direct causal relationship between thermal selective pressure and the evolution of neuronally expressed molecular and behavioral phenotypes. Several essential neuronal genes evolve lower expression at high temperatures and higher expression at low temperatures, with dopaminergic neurons standing out by displaying the most consistent expression change across independent replicates. We functionally validate the link between evolved gene expression and behavioral changes by pharmacological intervention in the experimentally evolved D. simulans populations as well as by genetically triggered expression changes of key genes in D. melanogaster. Since natural temperature clines confirm our results for Drosophila and Culex populations, we conclude neuronal dopamine evolution is a key factor for temperature adaptation.