Intact Drosophila Whole Brain Cellular Quantitation reveals Sexual Dimorphism

Establishing with precision the quantity and identity of the cell types of the brain is a prerequisite for a detailed compendium of gene and protein expression in the central nervous system. Currently however, strict quantitation of cell numbers has been achieved only for the nervous system of C.elegans. Here we describe the development of a synergistic pipeline of molecular genetic, imaging, and computational technologies designed to allow high-throughput, precise quantitation with cellular resolution of reporters of gene expression in intact whole tissues with complex cellular constitutions such as the brain. We have deployed the approach to determine with exactitude the number of functional neurons and glia in the entire intact Drosophila larval brain, revealing fewer neurons and many more glial cells than previously estimated. Moreover, we discover an unexpected divergence between the sexes at this juvenile developmental stage, with female brains having significantly more neurons than males. Topological analysis of our data establishes that this sexual dimorphism extends to deeper features of brain organisation. Our methodology enables robust and accurate quantification of the number and positioning of cells within intact organs, facilitating sophisticated analysis of cellular identity, diversity, and expression characteristics.