Glial cells mediate target layer selection of retinal axons in the developing visual system of Drosophila.

In the fly visual system, each class of photoreceptor neurons (R cells) projects to a different synaptic layer in the brain. R1-R6 axons terminate in the lamina, while R7 and R8 axons pass through the lamina and stop in the medulla. As R cell axons enter the lamina, they encounter both glial cells and neurons. The cellular requirement for R1-R6 targeting was determined using loss-of-function mutations affecting different cell types in the lamina. nonstop (encoding a ubiquitin-specific protease) is required for glial cell development and hedgehog for neuronal development. Removal of glial cells but not neurons disrupts R1-R6 targeting. We propose that glial cells provide the initial stop signal promoting growth cone termination in the lamina. These findings uncover a novel function for neuron-glial interactions in regulating target specificity.

Quantitative morphological analysis of embryonic cockroach (Periplaneta americana) brain neurons developing in vitro.

Neurons dissociated from the brain of embryonic cockroaches (Periplaneta americana) can be maintained in culture for several weeks. The survival as well as the progressive organization of the neurons into a complex network was studied during a 5-week period under different culture conditions. About 10% of the dissociated cells adhered to the culture dish. This figure remained constant throughout the culture. The cell diameter ranged from 10 to 20 microns and did not change significantly over time in culture. Whereas only a few cells exhibited neurites at the start of the culture, the number of cells exhibiting neurites increased to reach about 99% after 2 weeks. The different cells were then connected to each other, forming a network, which became more and more complex. The number of cells per cluster as well as the length and the diameter of the "connectives" that linked the different clusters were found to increase with time. The morphology of individual neurons within the network was visualized after intracellular injection of biocytin. Labeling with antibodies raised against serotonin or GABA indicated that neurons were able to differentiate and to acquire specific neurotransmitter fates. The serotonergic phenotype was found to appear progressively throughout the culture, in parallel with the formation of the network. Cell density, addition of fetal calf serum, and ecdysone were shown to influence the development of the network.

Retinal axon target selection in Drosophila is regulated by a receptor protein tyrosine phosphatase.

Different Drosophila photoreceptors (R cells) connect to neurons in different optic lobe layers. R1-R6 axons project to the lamina; R7 and R8 axons project to separate layers of the medulla. We show a receptor tyrosine phosphatase, PTP69D, is required for lamina target specificity. In Ptp69D mutants, R1-R6 project through the lamina, terminating in the medulla. Genetic mosaics, transgene rescue, and immunolocalization indicate PTP69D functions in R1-R6 growth cones. PTP69D overexpression in R7 and R8 does not respecify their connections, suggesting PTP69D acts in combination with other factors to determine target specificity. Structure-function analysis indicates the extracellular fibronectin type III domains and intracellular phosphatase activity are required for targeting. We propose PTP69D promotes R1-R6 targeting in response to extracellular signals by dephosphorylating substrate(s) in R1-R6 growth cones.

Polyglutamine-expanded human huntingtin transgenes induce degeneration of Drosophila photoreceptor neurons.

Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder. Disease alleles contain a trinucleotide repeat expansion of variable length, which encodes polyglutamine tracts near the amino terminus of the HD protein, huntingtin. Polyglutamine-expanded huntingtin, but not normal huntingtin, forms nuclear inclusions. We describe a Drosophila model for HD. Amino-terminal fragments of human huntingtin containing tracts of 2, 75, and 120 glutamine residues were expressed in photoreceptor neurons in the compound eye. As in human neurons, polyglutamine-expanded huntingtin induced neuronal degeneration. The age of onset and severity of neuronal degeneration correlated with repeat length, and nuclear localization of huntingtin presaged neuronal degeneration. In contrast to other cell death paradigms in Drosophila, coexpression of the viral antiapoptotic protein, P35, did not rescue the cell death phenotype induced by polyglutamine-expanded huntingtin.

Influence of receptor axons on the formation of olfactory glomeruli in a hemimetabolous insect, the cockroach Periplaneta americana.

The embryonic development of the hemimetabolous insect Periplaneta americana requires approximately 31 days. Deafferentation experiments were used to investigate the role of ingrowing receptor axons during embryogenesis, specifically their influence 1) on the subdivision of the antennal lobe neuropil into glomeruli, 2) on the morphology and number of glial cells, and 3) on the arborization pattern of central neurons. The flagellum of one antenna was removed from embryos at different developmental stages starting with day 10. Subsequently, they were raised in culture until a total age of 26 days. At day 10, the deutocerebrum has received only a very small number (ca. 0.4%) of antennal receptor axons; deafferentation at this stage allowed us to deprive the deutocerebrum of approximately 99% of its normal antennal input. Deafferentation has marked effects on the organization of the antennal lobe neuropil. The deafferented lobe is reduced in volume compared to the control side; the characteristic glomeruli are missing. During normal development glomeruli are formed between day 19 and 22, first in dorsal and then in ventral antennal lobe regions. By removing the antenna before day 20, their formation is disturbed in all parts of the antennal lobe. If deafferentation is performed after stage 20, glomeruli persist in dorsal regions, but are missing in ventral regions. On day 24 or later, glomeruli in both dorsal and ventral regions are unaffected by deafferentation. Glial cells continue to extend fine processes into the neuropil in the absence of ingrowing receptor axons. The number of glial cells is reduced compared to control lobes. Multiglomerular local interneurons and other gamma-amino butyric acid-immunoreactive neurons, as well as projection neurons, fail to develop glomerular arborization patterns in antennal lobes deprived of sensory axons.

Drosophila photoreceptor axon guidance and targeting requires the dreadlocks SH2/SH3 adapter protein.

SUMMARY: Mutations in the Drosophila gene dreadlocks (dock) disrupt photoreceptor cell (R cell) axon guidance and targeting. Genetic mosaic analysis and cell-type-specific expression of dock transgenes demonstrate dock is required in R cells for proper innervation. Dock protein contains one SH2 and three SH3 domains, implicating it in tyrosine kinase signaling, and is highly related to the human proto-oncogene Nck. Dock expression is detected in R cell growth cones in the target region. We propose Dock transmits signals in the growth cone in response to guidance and targeting cues. These findings provide an important step for dissection of signaling pathways regulating growth cone motility.

Embryonic development of the antennal lobes of a hemimetabolous insect, the cockroach Periplaneta americana: light and electron microscopic observations.

In the hemimetabolous insect Periplaneta americana, the adult-like organization of the primary olfactory centers, the antennal lobes, is established during the approximately 31 days of embryogenesis. This report describes the temporal sequence of developmental events as viewed in the light and electron microscope by means of histological stains and by DiI labeling of antennal receptor axons with subsequent photoconversion. Glomeruli, characteristic differentiations of the antennal lobe neuropil, are first observed on day 19; their development, which is not synchronous in the various parts of the antennal lobe, lasts until about day 22. From day 10 on, glial cells begin to form a narrow boundary layer between the soma cortex and the central neuropil. They exhibit a lengthening of their processes in parallel with the formation of glomeruli. Marked proliferation or migration of these glial cells into the neuropil between glomeruli has not been observed. Antennal receptor axons could be labeled from stage 15 on. They terminate in an elongated growth cone with numerous filopodia. From day 18 on, some of these become bent or show an initial bifurcation. From day 22 on, the first afferent axons develop an adult-like arborization pattern. Synaptic contacts between receptor axons and unidentified neurons were observed as early as stages 16 and 19, in which the axons still have a growth cone-like form. In stage 27, in which the fibers have adult-like arborizations, many output contacts and few input contacts were found.

Serotonin-immunoreactive neurons in the antennal lobes of the American cockroach Periplaneta americana: light- and electron-microscopic observations.

A large deutocerebral serotonin-immunoreactive neuron arborizes profusely in the glomeruli of the antennal lobes, and also sends neurites into the lateral lobe and the calyces of the mushroom bodies in the ipsilateral protocerebrum. Electron micrographs of the glomerular neuropil show that the main synapses of the serotonin-immunoreactive arborizations are output contacts with unidentified neuron profiles. Only a few synaptic input contacts with serotonin-labeled fibers were observed.