Optimizing Drosophila olfactory learning with a semi-automated training device.

Citation:

Murakami S, Dan C, Zagaeski B, Maeyama Y, Kunes S, Tabata T. Optimizing Drosophila olfactory learning with a semi-automated training device. Journal of neuroscience methods. 2010;188 (2) :195-204.

Date Published:

2010 May 15

Abstract:

Drosophila olfactory aversive conditioning has served as a powerful model system with which to elucidate the molecular and neuronal mechanisms underlying memory formation. In the typical protocol, flies are exposed to a constant odor stream while receiving a pulsed electric shock in the conditioning tube of a manual apparatus. We have devised a simple, low-cost semi-automated conditioning apparatus that computationally controls the delivery of odor and shock. A semiconductor-based odor sensor is employed to monitor the change of odor concentration in the training tube. The system thus allows electric shocks to be precisely matched with odor concentration in the training tube. We found that short-term memory performance was improved with a pulsed odor flow protocol, in which odor is presented in short pulses, each paired with electric shock, rather than as a constant flow. The effect of pulsed odor flow might be ascribed to the phenomenon of 'conditioned approach', where approach toward an odor is induced when the electric shock is presented before odor pulse ends. Our data shows that the system is applicable to the study of olfactory memory formation and to the examination of conditioning parameters at a level of detail not practical with a manual apparatus.

Notes:

n/a

Recent Publications

Daniele JR, Baqri RM, Kunes S. Analysis of axonal trafficking via a novel live-imaging technique reveals distinct hedgehog transport kinetics. Biol Open. 2017;6 (5) :714-721.Abstract

The Drosophila melanogaster (Dmel) eye is an ideal model to study development, intracellular signaling, behavior, and neurodegenerative disease. Interestingly, dynamic data are not commonly employed to investigate eye-specific disease models. Using axonal transport of the morphogen Hedgehog (Hh), which is integral to Dmel eye-brain development and implicated in stem cell maintenance and neoplastic disease, we demonstrate the ability to comprehensively quantify and characterize its trafficking in various neuron types and a neurodegeneration model in live early third-instar larval Drosophila We find that neuronal Hh, whose kinetics have not been reported previously, favors fast anterograde transport and varies in speed and flux with respect to axonal position. This suggests distinct trafficking pathways along the axon. Lastly, we report abnormal transport of Hh in an accepted model of photoreceptor neurodegeneration. As a technical complement to existing eye-specific disease models, we demonstrate the ability to directly visualize transport in real time in intact and live animals and track secreted cargoes from the axon to their release points. Particle dynamics can now be precisely calculated and we posit that this method could be conveniently applied to characterizing disease pathogenesis and genetic screening in other established models of neurodegeneration.

Daniele JR, Chu T, Kunes S. A novel proteolytic event controls Hedgehog intracellular sorting and distribution to receptive fields. Biol Open. 2017;6 (5) :540-550.Abstract

The patterning activity of a morphogen depends on secretion and dispersal mechanisms that shape its distribution to the cells of a receptive field. In the case of the protein Hedgehog (Hh), these mechanisms of secretion and transmission remain unclear. In the developing Drosophila visual system, Hh is partitioned for release at opposite poles of photoreceptor neurons. Release into the retina regulates the progression of eye development; axon transport and release at axon termini trigger the development of postsynaptic neurons in the brain. Here we show that this binary targeting decision is controlled by a C-terminal proteolysis. Hh with an intact C-terminus undergoes axonal transport, whereas a C-terminal proteolysis enables Hh to remain in the retina, creating a balance between eye and brain development. Thus, we define a novel mechanism for the apical/basal targeting of this developmentally important protein and posit that similar post-translational regulation could underlie the polarity of related ligands.

Baqri RM, Pietron AV, Gokhale RH, Turner BA, Kaguni LS, Shingleton AW, Kunes S, Miller KE. Mitochondrial chaperone TRAP1 activates the mitochondrial UPR and extends healthspan in Drosophila. Mech Ageing Dev. 2014;141-142 :35-45.Abstract

The molecular mechanisms influencing healthspan are unclear but mitochondrial function, resistance to oxidative stress and proteostasis are recurring themes. Tumor necrosis factor Receptor Associated Protein 1 (TRAP1), the mitochondrial analog of Hsp75, regulates levels of reactive oxygen species in vitro and is found expressed at higher levels in tumor cells where it is thought to play a pro-survival role. While TRAP1-directed compartmentalized protein folding is a promising target for cancer therapy, its role at the organismal level is unclear. Here we report that overexpression of TRAP1 in Drosophila extends healthspan by enhancing stress resistance, locomotor activity and fertility while depletion of TRAP1 has the opposite effect, with little effect on lifespan under both conditions. In addition, modulating TRAP1 expression promotes the nuclear translocation of homeobox protein Dve and increases expression of genes associated with the mitochondrial unfolded protein response (UPR(mt)), indicating an activation of this proteostasis pathway. Notably, independent genetic knockdown of components of the UPR(mt) pathway dampen the enhanced stress resistance observed in TRAP1 overexpression flies. Together these studies suggest that TRAP1 regulates healthspan, potentially through activation of the UPR(mt).

Song E, de Bivort B, Dan C, Kunes S. Determinants of the Drosophila odorant receptor pattern. Dev Cell. 2012;22 (2) :363-76.Abstract

In most olfactory systems studied to date, neurons that express the same odorant receptor (Or) gene are scattered across sensory epithelia, intermingled with neurons that express different Or genes. In Drosophila, olfactory sensilla that express the same Or gene are dispersed on the antenna and the maxillary palp. Here we show that Or identity is specified in a spatially stereotyped pattern by the cell-autonomous activity of the transcriptional regulators Engrailed and Dachshund. Olfactory sensilla then become highly motile and disperse beneath the epidermis. Thus, positional information and cell motility underlie the dispersed patterns of Drosophila Or gene expression.

Dearborn RE, Dai Y, Reed B, Karian T, Gray J, Kunes S. Reph, a regulator of Eph receptor expression in the Drosophila melanogaster optic lobe. PLoS One. 2012;7 (5) :e37303.Abstract

Receptors of the Eph family of tyrosine kinases and their Ephrin ligands are involved in developmental processes as diverse as angiogenesis, axon guidance and cell migration. However, our understanding of the Eph signaling pathway is incomplete, and could benefit from an analysis by genetic methods. To this end, we performed a genetic modifier screen for mutations that affect Eph signaling in Drosophila melanogaster. Several dozen loci were identified on the basis of their suppression or enhancement of an eye defect induced by the ectopic expression of Ephrin during development; many of these mutant loci were found to disrupt visual system development. One modifier locus, reph (regulator of eph expression), was characterized in molecular detail and found to encode a putative nuclear protein that interacts genetically with Eph signaling pathway mutations. Reph is an autonomous regulator of Eph receptor expression, required for the graded expression of Eph protein and the establishment of an optic lobe axonal topographic map. These results reveal a novel component of the regulatory pathway controlling expression of eph and identify reph as a novel factor in the developing visual system.

Song E, Kunes S. Determinants of the Drosophila Odorant Receptor Pattern. Developmental Cell. 2012;22 (2) :363-376.

Kunes Laboratory

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