The neuroanatomical ultrastructure and function of a biological ring attractor

Neural representations of head direction (HD) have been discovered in many species. Theoretical work has proposed that the dynamics associated with these representations are generated, maintained, and updated by recurrent network structures called ring attractors. We evaluated this theorized structure-function relationship by performing electron microscopy-based circuit reconstruction and RNA profiling of identified cell types in the HD system of Drosophila melanogaster. We identified motifs that have been hypothesized to maintain the HD representation in darkness, update it when the animal turns, and tether it to visual cues. Functional studies provided support for the proposed roles of individual excitatory or inhibitory circuit elements in shaping activity. We also discovered recurrent connections between neuronal arbors with mixed pre- and post-synaptic specializations. Our results confirm that the Drosophila HD network contains the core components of a ring attractor while also revealing unpredicted structural features that might enhance the network's computational power. Overall design: Three different types of sequencing protocols were performed: bulk sequencing for the EPGs, ?7s, PEN2s, and PEGs, low-cell sequencing for the PEN2s and R4d neurons, and single cell sequencing for the PEN1 and PEN2 lines. Two biological replicates were run for each genetic line for the bulk sequencing as well as for whole brain samples, which served as a control. Two genetic lines were run for the EPGs and ?7s while one line was run for the PEGs. For the low-cell sequencing, 14 replicates were run for the PEN2s, 9 for the R4d neurons, and 8 for the whole brain material. One genetic line was used for each. The optic lobes were removed from the brain for the low-cell controls, but not for the bulk sequencing. The former is referred to as the central brain (CB) while the latter is referred to as whole brain (WB). For the single-cell sequencing, one genetic line was used for the PEN1s and one was used for the PEN2s. Six 96 well plates were run for the PEN1s and five 96 well plates were run for the PEN2s. Each plate featured two wells with ERCC RNA and two well with whole brain RNA to act as controls.