title: Analyzing Synaptic Modulation of Drosophila melanogaster Photoreceptors after Exposure to Prolonged Light.
abstract: The nervous system has the remarkable ability to adapt and respond to various stimuli. This neural adjustment is largely achieved through plasticity at the synaptic level. The Active Zone (AZ) is the region at the presynaptic membrane that mediates neurotransmitter release and is composed of a dense collection of scaffold proteins. AZs of Drosophila melanogaster (Drosophila) photoreceptors undergo molecular remodeling after prolonged exposure to natural ambient light. Thus the level of neuronal activity can rearrange the molecular composition of the AZ and contribute to the regulation of the functional output. Starting from the light exposure set-up preparation to the immunohistochemistry, this protocol details how to quantify the number, the spatial distribution, and the delocalization level of synaptic molecules at AZs in Drosophila photoreceptors. Using image analysis software, clusters of the GFP-fused AZ component Bruchpilot were identified for each R8 photoreceptor (R8) axon terminal. Detected Bruchpilot spots were automatically assigned to individual R8 axons. To calculate the distribution of spot frequency along the axon, we implemented a customized software plugin. Each axon's start-point and end-point were manually defined and the position of each Bruchpilot spot was projected onto the connecting line between start and end-point. Besides the number of Bruchpilot clusters, we also quantified the delocalization level of Bruchpilot-GFP within the clusters. These measurements reflect in detail the spatially resolved synaptic dynamics in a single neuron under different environmental conditions to stimuli.
authors.affiliation: Department of Neuroscience of Disease, Center for Transdisciplinary Research, Niigata University; Brain Research Institute, Niigata University; Dendrite Differentiation, German Center for Neurodegenerative Diseases (DZNE); atsushi.sugie@bri.niigata-u.ac.jp.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Sugie
authors.affiliation: Image and Data Analysis Facility, German Center for Neurodegenerative Diseases (DZNE); christoph.moehl@dzne.de.
authors.forename: Christoph
authors.initials: C
authors.lastname: Möhl
authors.affiliation: Graduate School of Life Science and Technology, Tokyo Institute of Technology (Titech).
authors.forename: Satoko
authors.initials: S
authors.lastname: Hakeda-Suzuki
authors.affiliation: Department of Neuroscience of Disease, Center for Transdisciplinary Research, Niigata University; Brain Research Institute, Niigata University.
authors.forename: Hideaki
authors.initials: H
authors.lastname: Matsui
authors.affiliation: Graduate School of Life Science and Technology, Tokyo Institute of Technology (Titech).
authors.forename: Takashi
authors.initials: T
authors.lastname: Suzuki
authors.affiliation: Dendrite Differentiation, German Center for Neurodegenerative Diseases (DZNE).
authors.forename: Gaia
authors.initials: G
authors.lastname: Tavosanis
mesh_terms: D000818:Animals; D029721:Drosophila Proteins / Q000378:metabolism; D004331:Drosophila melanogaster / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008027:Light; D049408:Luminescent Agents / Q000378:metabolism; D017956:Photoreceptor Cells, Invertebrate / Q000378:metabolism / Q000528:radiation effects*; D017729:Presynaptic Terminals; D011485:Protein Binding; D021381:Protein Transport; D013569:Synapses / Q000378:metabolism; D009435:Synaptic Transmission / Q000502:physiology*
chemical_list: D029721:Drosophila Proteins; D049408:Luminescent Agents; D049452:Green Fluorescent Proteins