Cox, D. N. | bioRxiv Science

bioRxiv2018

Injury-induced cold sensitization in Drosophila larvae involves behavioral shifts that require the TRP channels Pkd2 and Brv1.

Nociceptive sensitization involves an increase in responsiveness of pain sensing neurons to sensory stimuli, typically through the lowering of their nociceptive threshold. Nociceptive sensitization is common following tissue damage, inflammation, and disease and serves to protect the affected area while it heals. Organisms can become sensitized to a range of noxious and innocuous stimuli, including thermal stimuli. The basic mechanisms underlying sensitization to warm or painfully hot stimuli have begun to be elucidated, however, sensitization to cold is not well understood. Here, we develop a Drosophila assay to study cold sensitization after UV-induced epidermal damage in larvae. Larvae respond to acute cold stimuli with a set of unique behaviors that include a contraction of the head and tail (CT) or a raising of the head and tail into a U-Shape (US). Under baseline, non-injured conditions larvae primarily produce a CT response to an acute cold (10 {degrees}C) stimulus, however, we show that cold-evoked responses shift following tissue damage: CT responses decrease, US responses increase and some larvae exhibit a lateral body roll (BR) that is typically only observed in response to high temperature and noxious mechanical stimuli. At the cellular level, class III neurons are required for the decrease in CT, chordotonal neurons are required for the increase in US, and chordotonal and class IV neurons are required for the appearance of BR responses after UV. At the molecular level, we found that the transient receptor potential (TRP) channels Polycystic kidney disease gene 2 (Pkd2) and brivido-1 (brv1) are required for these behavioral shifts. Our Drosophila model will enable a sophisticated molecular genetic dissection of genes and circuits involved in cold nociceptive sensitization.

neuroscience↗

bioRxiv2017

Formin3 regulates dendritic architecture via microtubule stabilization and is required for somatosensory nociceptive behavior

The acquisition, maintenance and modulation of dendritic architecture are critical to neuronal form, plasticity and function. Morphologically, dendritic shape impacts functional connectivity and is largely mediated by organization and dynamics of cytoskeletal fibers that provide the underlying scaffold and tracks for intracellular trafficking. Identifying molecular factors that regulate dendritic cytoskeletal architecture is therefore important in understanding mechanistic links between cytoskeletal organization and neuronal function. In a neurogenomic-driven genetic screen of cytoskeletal regulatory molecules, we identified Formin3 (Form3) as a critical regulator of cytoskeletal architecture in Drosophila nociceptive sensory neurons. Form3 is a member of the conserved Formin family of multi-functional cytoskeletal regulators and time course analyses reveal Form3 is cell-autonomously required for maintenance of complex dendritic arbors. Cytoskeletal imaging demonstrates form3 mutants exhibit a specific destabilization of the dendritic microtubule (MT) cytoskeleton, together with defective dendritic trafficking of mitochondria, satellite Golgi and the TRPA channel Painless. Biochemical studies reveal Form3 directly interacts with MTs via FH1-FH2 domains and promotes MT stabilization via acetylation. Neurologically, mutations in human Inverted Formin 2 (INF2; ortholog of form3) have been causally linked to Charcot-Marie-Tooth (CMT) disease. CMT sensory neuropathies lead to impaired peripheral sensitivity. Defects in form3 function in nociceptive neurons results in a severe impairment in noxious heat evoked behaviors. Expression of the INF2 FH1-FH2 domains rescues form3 defects in MT stabilization and nocifensive behavior revealing conserved functions in regulating the cytoskeleton and sensory behavior thereby providing novel mechanistic insights into potential etiologies of CMT sensory neuropathies.\n\nSignificance StatementMechanisms governing cytoskeletal architecture are critical in regulating neural function as aberrations are linked to a broad spectrum of neurological and neurocognitive disorders. Formins are important cytoskeletal regulators however their mechanistic roles in neuronal architecture are poorly understood. We demonstrate mutations in Drosophila formin3 lead to progressive destabilization of the dendritic microtubule cytoskeleton resulting in severely reduced arborization coupled to impaired organelle and ion channel trafficking, as well as nociceptive sensitivity. INF2 mutations are implicated in CMT sensory neuropathies, and INF2 expression can rescue microtubule and nociceptive behavioral defects in form3 mutants. While CMT sensory neuropathies have been linked to defects in axonal development and myelination, our studies connect dendritic cytoskeletal defects with peripheral insensitivity suggesting possible alternative etiological bases.

neuroscience↗