Reciprocal interactions between olfactory receptor axons and olfactory nerve glia cultured from the developing moth Manduca sexta

AbstractIn olfactory systems, neuron–glia interactions have been implicated in the growth and guidance of olfactory receptor axons. In the moth Manduca sexta, developing olfactory receptor axons encounter several types of glia as they grow into the brain. Antennal nerve glia are born in the periphery and enwrap bundles of olfactory receptor axons in the antennal nerve. Although their peripheral origin and relationship with axon bundles suggest that they share features with mammalian olfactory ensheathing cells, the developmental roles of antennal nerve glia remain elusive. When cocultured with antennal nerve glial cells, olfactory receptor growth cones readily advance along glial processes without displaying prolonged changes in morphology. In turn, olfactory receptor axons induce antennal nerve glial cells to form multicellular arrays through proliferation and process extension. In contrast to antennal nerve glia, centrally derived glial cells from the axon sorting zone and antennal lobe never form arrays in vitro, and growth-cone glial-cell encounters with these cells halt axon elongation and cause permanent elaborations in growth cone morphology. We propose that antennal nerve glia play roles similar to olfactory ensheathing cells in supporting axon elongation, yet differ in their capacity to influence axon guidance, sorting, and targeting, roles that could be played by central olfactory glia in Manduca

Activation of Glial FGFRs Is Essential in Glial Migration, Proliferation, and Survival and in Glia-Neuron Signaling during Olfactory System Development

Development of the adult olfactory system of the moth Manduca sexta depends on reciprocal interactions between olfactory receptor neuron (ORN) axons growing in from the periphery and centrally-derived glial cells. Early-arriving ORN axons induce a subset of glial cells to proliferate and migrate to form an axon-sorting zone, in which later-arriving ORN axons will change their axonal neighbors and change their direction of outgrowth in order to travel with like axons to their target areas in the olfactory (antennal) lobe. These newly fasciculated axon bundles will terminate in protoglomeruli, the formation of which induces other glial cells to migrate to surround them. Glial cells do not migrate unless ORN axons are present, axons fail to fasciculate and target correctly without sufficient glial cells, and protoglomeruli are not maintained without a glial surround. We have shown previously that Epidermal Growth Factor receptors and the IgCAMs Neuroglian and Fasciclin II play a role in the ORN responses to glial cells. In the present work, we present evidence for the importance of glial Fibroblast Growth Factor receptors in glial migration, proliferation, and survival in this developing pathway. We also report changes in growth patterns of ORN axons and of the dendrites of olfactory (antennal lobe) neurons following blockade of glial FGFR activation that suggest that glial FGFR activation is important in reciprocal communication between neurons and glial cells

Roles of Specific Membrane Lipid Domains in EGF Receptor Activation and Cell Adhesion Molecule Stabilization in a Developing Olfactory System

BACKGROUND:Reciprocal interactions between glial cells and olfactory receptor neurons (ORNs) cause ORN axons entering the brain to sort, to fasciculate into bundles destined for specific glomeruli, and to form stable protoglomeruli in the developing olfactory system of an experimentally advantageous animal species, the moth Manduca sexta. Epidermal growth factor receptors (EGFRs) and the cell adhesion molecules (IgCAMs) neuroglian and fasciclin II are known to be important players in these processes. METHODOLOGY/PRINCIPAL FINDINGS:We report in situ and cell-culture studies that suggest a role for glycosphingolipid-rich membrane subdomains in neuron-glia interactions. Disruption of these subdomains by the use of methyl-beta-cyclodextrin results in loss of EGFR activation, depletion of fasciclin II in ORN axons, and loss of neuroglian stabilization in the membrane. At the cellular level, disruption leads to aberrant ORN axon trajectories, small antennal lobes, abnormal arrays of olfactory glomerul, and loss of normal glial cell migration. CONCLUSIONS/SIGNIFICANCE:We propose that glycosphingolipid-rich membrane subdomains (possible membrane rafts or platforms) are essential for IgCAM-mediated EGFR activation and for anchoring of neuroglian to the cytoskeleton, both required for normal extension and sorting of ORN axons

Astrocytic glutamate transport regulates a Drosophila CNS synapse that lacks astrocyte ensheathment.

Anatomical, molecular, and physiological interactions between astrocytes and neuronal synapses regulate information processing in the brain. The fruit fly Drosophila melanogaster has become a valuable experimental system for genetic manipulation of the nervous system and has enormous potential for elucidating mechanisms that mediate neuron-glia interactions. Here, we show the first electrophysiological recordings from Drosophila astrocytes and characterize their spatial and physiological relationship with particular synapses. Astrocyte intrinsic properties were found to be strongly analogous to those of vertebrate astrocytes, including a passive current-voltage relationship, low membrane resistance, high capacitance, and dye-coupling to local astrocytes. Responses to optogenetic stimulation of glutamatergic premotor neurons were correlated directly with anatomy using serial electron microscopy reconstructions of homologous identified neurons and surrounding astrocytic processes. Robust bidirectional communication was present: neuronal activation triggered astrocytic glutamate transport via excitatory amino acid transporter 1 (Eaat1), and blocking Eaat1 extended glutamatergic interneuron-evoked inhibitory postsynaptic currents in motor neurons. The neuronal synapses were always located within 1 μm of an astrocytic process, but none were ensheathed by those processes. Thus, fly astrocytes can modulate fast synaptic transmission via neurotransmitter transport within these anatomical parameters. J. Comp. Neurol. 524:1979-1998, 2016. © 2016 Wiley Periodicals, Inc.This is the author accepted manuscript. The final version is available from Wiley via http://dx.doi.org/10.1002/cne.2401

The structure of the PanD/PanZ protein complex reveals negative feedback regulation of pantothenate biosynthesis by coenzyme A.

Coenzyme A (CoA) is an ubiquitous and essential cofactor, synthesized from the precursor pantothenate. Vitamin biosynthetic pathways are normally tightly regulated, including the pathway from pantothenate to CoA. However, no regulation of pantothenate biosynthesis has been identified. We have recently described an additional component in the pantothenate biosynthetic pathway, PanZ, which promotes the activation of the zymogen, PanD, to form aspartate ?-decarboxylase (ADC) in a CoA-dependent manner. Here we report the structure of PanZ in complex with PanD, which reveals the structural basis for the CoA dependence of this interaction and activation. In addition, we show that PanZ acts as a CoA-dependent inhibitor of ADC catalysis. This inhibitory effect can effectively regulate the biosynthetic pathway to pantothenate, and thereby also regulate CoA biosynthesis. This represents a previously unobserved mode of metabolic regulation whereby a cofactor-utilizing protein negatively regulates the biosynthesis of the same cofactor

Human gut Bacteroidetes can utilize yeast mannan through a selfish mechanism

Yeasts, which have been a component of the human diet for at least 7,000 years, possess an elaborate cell wall α-mannan. The influence of yeast mannan on the ecology of the human microbiota is unknown. Here we show that yeast α-mannan is a viable food source for the Gram-negative bacterium Bacteroides thetaiotaomicron, a dominant member of the microbiota. Detailed biochemical analysis and targeted gene disruption studies support a model whereby limited cleavage of α-mannan on the surface generates large oligosaccharides that are subsequently depolymerized to mannose by the action of periplasmic enzymes. Co-culturing studies showed that metabolism of yeast mannan by B. thetaiotaomicron presents a ‘selfish’ model for the catabolism of this difficult to breakdown polysaccharide. Genomic comparison with B. thetaiotaomicron in conjunction with cell culture studies show that a cohort of highly successful members of the microbiota has evolved to consume sterically-restricted yeast glycans, an adaptation that may reflect the incorporation of eukaryotic microorganisms into the human diet

MβCD treatment of cultured ORNs decreases MFas II labeling.

<p>A_1–4: In control conditions, a subset of ORN axons extending from explants are MFas II-positive, some strongly (arrowheads), some moderately (open arrowheads). Arrows indicate several unlabeled axons visible under brightfield optics. B_1–4: After 24-hr exposure to 1mM MβCD, more MFas II axons are moderately or only weakly labeled. C_1–4: At 2 mM MβCD, nearly all MFas II-positive axons are only faintly labeled. Rare axons that appear brightly labeled (C₃) were always less strongly labeled than those found in control or 1 mM dishes. No consistent changes were seen in axonal or growth cone morphology at the 1 mM dose; axon outgrowth was reduced at the 2 mg/ml dose.</p

Blocking activation of the FGFR blocks migration of neuropil glia.

<p><b>A,B:</b> Animals injected at stage 4 with DMSO or DMSO + PD173074 and examined at stage 7. <b>A:</b> Control animal injected with vehicle (DMSO) and labeled with the anti-pFGFR antibody (magenta) and Syto 13 (green) to show cell nuclei. Neuropil glial cells have migrated to surround glomeruli as in untreated animals. A more anterior view than those used in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0033828#pone-0033828-g002" target="_blank">Fig. 2</a> was chosen to better illustrate the intense labeling of NP glial processes surrounding glomeruli. <b>B:</b> Animal injected with DMSO containing 0.5 mg PD173074. Labeling for activated FGFRs on glial cells is absent, and most neuropil-associated (NP) glia have failed to migrate to surround glomeruli. Sorting zone (SZ) glia have migrated normally despite their lack of labeling for activated FGFRs. <b>C:</b> A western blot of control and PD173074-treated antennal-lobe tissue from which neuronal cell bodies had been removed demonstrates a nearly complete absence of labeling for pFGFRs for the PD173074-treated lobes. <b>D:</b> Another animal treated with 0.5 mg PD173074 (beginning at stage 3), dissected at stage 7, and labeled with Syto 13. At increased gain, some NP glial processes can be seen to have extended into the neuropil (arrowheads) despite the absence of cell-body migration. <b>E:</b> A stage-6 antennal lobe from an animal chronically deprived of ORN innervation on one side (antennal anlagen removed at stage 1.) Although lack of ORN innervation resulted in lack of glial migration, glial cells did exhibit activated FGFRs (magenta, arrowheads). <b>E′:</b> Enlarged section from boxed area of panel <b>E</b> better illustrates the labeling of glial processes. <b>E″:</b> Opposite lobe, which was not deprived of ORN input, appears to have the same intensity of pFGFR labeling. <b>F′,F″:</b> Individual cells of deafferented (<b>F′</b>) and control (<b>F″</b>) lobes both display colocalization of pFGFR and DNA labels. LG, MG = lateral and medial group of AL neuron cell bodies. Projection depths = 10 µm in <b>A, B, D, E, E′</b>. <b>F′, F″</b> are single optical sections (40× objective).</p