Drosophila beta-Heavy-Spectrin is required in polarized ensheathing glia that form a diffusion-barrier around the neuropil

In the central nervous system (CNS), functional tasks are often allocated to distinct compartments. This is also evident in the insect CNS where synapses and dendrites are clustered in distinct neuropil regions. The neuropil is separated from neuronal cell bodies by ensheathing glia, which as we show using dye injection experiments forms an internal diffusion barrier. We find that ensheathing glial cells are polarized with a basolateral plasma membrane rich in phosphatidylinositol-(3,4,5)-triphosphate (PIP3) and the Na+/K+-ATPase Nervana2 (Nrv2) that abuts an extracellular matrix formed at neuropil-cortex interface. The apical plasma membrane is facing the neuropil and is rich in phosphatidylinositol-(4,5)-bisphosphate (PIP2) that is supported by a sub-membranous beta-Heavy-Spectrin cytoskeleton. beta-Heavy-spectrin mutant larvae affect ensheathing glial cell polarity with delocalized PIP2 and Nrv2 and exhibit an abnormal locomotion which is similarly shown by ensheathing glia ablated larvae. Thus, polarized glia compartmentalizes the brain and is essential for proper nervous system function.

RDL receptors

RDL receptors are invertebrate members of the Cys-loop family of ligand-gated ion channels. They are GABA (γ-aminobutyric acid)-activated chloride-selective receptors that are closely related to their vertebrate orthologues, the GABAA receptors, as well as other Cys-loop receptors such as the ionotropic glycine, nicotinic acetylcholine and 5-HT3 receptors. RDL receptors are widely expressed throughout the insect CNS (central nervous system) and are important in inhibitory neurotransmission. They are therefore a major insecticidal target site.

Targeting of an Expressed Insect Selective Neurotoxin by its Recombinant Baculovirus: Pharmacokinetic and Pharmacodynamic Aspects

Objectives: 1) Clarification of the mode of potentiation of an expressed insect selective neurotoxin (AaIT) by its recombinant baculovirus. 2) In vitro formation and/or modification of neuroactive polypeptides for the design of new improved recombinant baculoviruses. Results: 1) A combined utilization of bioassays, LM-cytochemistry, the highly resolutive EM immunogold and electrical recording from the CNS of baculovirus and AaIT - expressing – recombinant baculovirus infected larvae it has been shown that the recombinant virus potentiates the effect of the toxin. Potentiation is achieved through its continuous expression in the infected tracheal epithelia thus providing a: a) Local supply of freshly produced toxin in the vicinity of its traget sites; b) Translocation of the expressed toxin to the insect CNS. The latter exposes the recombinant toxin to new, critical, target sites which are inaccessible through the natural route of scorpion envenomation. 2) Subjecting a recombinant AaIT toxin to a newly designed system of random mutagenesis results in large numbers of new AaIT genes with amino acid substitutions. The new or modified toxin genes were inserted into a linear BmNPV expressed in silkworm cell culture and assayed on blowfly and silkworm larvae. Thus a system for mass formation and screening of neuroactive agents was developed. Contribution to agriculture: 1) Demonstration of the insecticidal mechanism, capacity and utility of the combination of neuroactive polypeptides and recombinant pathogens. 2) Development of a simple in vitro system for the formation and selection of new neuroactive polypeptides.

Neurogenesis in the insect enteric nervous system: generation of premigratory neurons from an epithelial placode

The enteric plexus (EP) is a major division of the enteric nervous system (ENS) in the moth Manduca sexta and contains a dispersed population of about 360 bipolar neurons, the EP cells. Previously we showed that embryonic EP cells achieve their mature distributions by extensive migration along the gut surface and then display position-specific phenotypes. We now demonstrate that the entire EP cell population is generated from an ectodermal placode that invaginates from the embryonic foregut. Individual EP cells become postmitotic just as they leave the epithelium, but their terminal differentiation is subsequently delayed until after their migratory dispersal. Clonal analysis by injection of lineage-tracing dyes has shown that the EP cell population is derived from a large number of placodal cells, each of which contributes a limited number of neurons to the ENS. Placodally derived clones produce neurons exclusively, while clones arising from cells adjacent to the placode are incorporated into the gut epithelium. These results indicate that neurogenesis in the insect ENS involves a developmental strategy that is distinct from that seen in the insect CNS and which resembles the generation of certain cell classes in the vertebrate nervous system.