Temporal and Notch identity determine layer targeting and synapse location of medulla neurons.

How specification mechanisms that generate neural diversity translate into specific neuronal targeting, connectivity, and function in the adult brain is not understood. In the medulla region of the optic lobe, neural progenitors generate different neurons in a fixed order by sequentially expressing a series of temporal transcription factors as they age. Then, Notch signaling in intermediate progenitors further diversifies neuronal progeny. By establishing the birth order of medulla neurons, we found that their temporal identity correlates with the depth of neuropil targeting in the adult brain, for both local interneurons and projection neurons. We show that this temporal identity-dependent targeting of projection neurons unfolds early in development and is genetically determined. By leveraging the Electron Microscopy reconstruction of the adult fly brain, we determined the synapse location of medulla neurons in the different optic lobe neuropils and find that it is significantly associated with both their temporal identity and Notch status. Moreover, we show that all the putative medulla neurons with the same predicted function share similar neuropil synapse location, indicating that ensembles of neuropil layers encode specific visual functions. In conclusion, we show that temporal identity and Notch status of medulla neurons can predict their neuropil synapse location and visual function, linking their developmental patterning with their specific connectivity and functional features in the adult brain.