The neural stem cells that give rise to the neural lineages of the brain can generate their progeny directly or through transit amplifying intermediate neural progenitor cells (INPs). The INP-producing neural stem cells in Drosophila are called type II neuroblasts, and their neural progeny innervate the central complex, a prominent integrative brain center. Here we use genetic lineage tracing and clonal analysis to show that the INPs of these type II neuroblast lineages give rise to glial cells as well as neurons during postembryonic brain development. Our data indicate that two main types of INP lineages are generated, namely mixed neuronal/glial lineages and neuronal lineages. Genetic loss-of-function and gain-of-function experiments show that the gcm gene is necessary and sufficient for gliogenesis in these lineages. The INP-derived glial cells, like the INP-derived neuronal cells, make major contributions to the central complex. In postembryonic development, these INP-derived glial cells surround the entire developing central complex neuropile, and once the major compartments of the central complex are formed, they also delimit each of these compartments. During this process, the number of these glial cells in the central complex is increased markedly through local proliferation based on glial cell mitosis. Taken together, these findings uncover a novel and complex form of neurogliogenesis in Drosophila involving transit amplifying intermediate progenitors. Moreover, they indicate that type II neuroblasts are remarkably multipotent neural stem cells that can generate both the neuronal and the glial progeny that make major contributions to one and the same complex brain structure.

BackgroundThe central complex is a multimodal information-processing center in the insect brain composed of thousands of neurons representing more than 50 neural types arranged in a stereotyped modular neuroarchitecture. In Drosophila, the development of the central complex begins in the larval stages when immature structures termed primordia are formed. However, the identity and origin of the neurons that form these primordia and, hence, the fate of these neurons during subsequent metamorphosis and in the adult brain, are unknown.ResultsHere, we used two pointed-Gal4 lines to identify the neural cells that form the primordium of the fan-shaped body, a major component of the Drosophila central complex. We found that these early-born primordium neurons are generated by four identified type II neuroblasts that amplify neurogenesis through intermediate progenitors, and we demonstrate that these neurons generate the fan-shaped body primordium during larval development in a highly specific manner. Moreover, we characterize the extensive growth and differentiation that these early-born primordium neurons undergo during metamorphosis in pupal stages and show that these neurons persist in the adult central complex, where they manifest layer-specific innervation of the mature fan-shaped body.ConclusionsTaken together, these findings indicate that early-born neurons from type II neuroblast lineages have dual roles in the development of a complex brain neuropile. During larval stages they contribute to the formation of a specific central complex primordium; during subsequent pupal development they undergo extensive growth and differentiation and integrate into the modular circuitry of the adult brain central complex.

Graphical AbstractHighlights d Many early larval neurons lack a branched neurite tree and synapses (SU neurons) d Axons of SU neurons form bundles associated with the fiber tracts of most lineages d SU neurons of lineages DM1-DM4 pioneer the neural architecture of the central complex d SU neurons differentiate into the pontine neurons of the adult central complex SUMMARYSerial electron microscopic analysis shows that the Drosophila brain at hatching possesses a large fraction of developmentally arrested neurons with a small soma, heterochromatin-rich nucleus, and unbranched axon lacking synapses. We digitally reconstructed all 802 ''small undifferentiated'' (SU) neurons and assigned them to the known brain lineages. By establishing the coordinates and reconstructing trajectories of the SU neuron tracts, we provide a framework of landmarks for the ongoing analyses of the L1 brain circuitry. To address the later fate of SU neurons, we focused on the 54 SU neurons belonging to the DM1-DM4 lineages, which generate all columnar neurons of the central complex.Regarding their topologically ordered projection pattern, these neurons form an embryonic nucleus of the fan-shaped body (''FB pioneers''). Fan-shaped body pioneers survive into the adult stage, where they develop into a specific class of bi-columnar elements, the pontine neurons. Later born, unicolumnar DM1-DM4 neurons fasciculate with the fan-shaped body pioneers. Selective ablation of the fan-shaped body pioneers altered the architecture of the larval fan-shaped body primordium but did not result in gross abnormalities of the trajectories of unicolumnar neurons, indicating that axonal pathfinding of the two systems may be controlled independently.Our comprehensive spatial and developmental analysis of the SU neurons adds to our understanding of the establishment of neuronal circuitry.

The neurons and glial cells of the Drosophila brain are generated by neural stem cell-like progenitors during two developmental phases, one short embryonic phase and one more prolonged postembryonic phase. Like the bulk of the adult-specific neurons, most of glial cells found in the adult central brain are generated postembryonically. Five of the neural stem cell-like progenitors that give rise to glial cells during postembryonic brain development have been identified as type II neuroglioblasts that generate neural and glial progeny through transient amplifying INPs. Here we identify DL1 as a novel multipotent neuroglial progenitor in the central brain and show that this type II neuroblast not only gives rise to neurons that innervate the central complex but also to glial cells that contribute exclusively to the optic lobe. Immediately following their generation in the central brain during the second half of larval development, these DL1 lineage-derived glia migrate into the developing optic lobe, where they differentiate into three identified types of optic lobe glial cells, inner chiasm glia, outer chiasm glia and cortex glia. Taken together, these findings reveal an unexpected central brain origin of optic lobe glial cells and central complex interneurons from one and the same type II neuroglioblast.