Robo2 regulates synaptic oxytocin content by affecting actin dynamics
- Weizmann Institute of Science, Israel
Abstract
The regulation of neuropeptide level at the site of release is essential for proper neurophysiological functions. We focused on a prominent neuropeptide, oxytocin (OXT) in the zebrafish as an in vivo model to visualize and quantify OXT content at the resolution of a single synapse. We found that OXT-loaded synapses were enriched with polymerized actin. Perturbation of actin filaments by either cytochalasin-D or conditional Cofilin expression resulted in decreased synaptic OXT levels. Genetic loss of robo2 or slit3 displayed decreased synaptic OXT content and robo2 mutants displayed reduced mobility of the actin probe Lifeact-EGFP in OXT synapses.Using a novel transgenic reporter allowing real-time monitoring of OXT-loaded vesicles, we showed that robo2 mutants display slower rate of vesicles accumulation. OXT-specific expression of dominant-negative Cdc42, which is a key regulator of actin dynamics and a downstream effector of Robo2, led to a dose-dependent increase in OXT content in WT, and a dampened effect in robo2 mutants. Our results link Slit3-Robo2-Cdc42, which controls local actin dynamics, with the maintenance of synaptic neuropeptide levels.
Data availability
All source data files and codes were uploaded. Source data as summary tables is provided for all graphs and plots shown (Fig. 1E; 2B-E and G-J, 3E-H and J,K,M,N, 4D-F, 5O,R, 6I, 7C,D). R codes used for data import and analysis is provided.
Article and author information
Author details
Funding
Israel Science Foundation (1511/16)
- Savani Anbalagan
- Janna Blechman
- Michael Gliksberg
- Ludmila Gordon
- Gil Levkowitz
Israel Science Foundation (2137/16)
- Savani Anbalagan
- Janna Blechman
- Michael Gliksberg
- Ludmila Gordon
- Gil Levkowitz
Minerva Foundation ((Minerva Stiftung))
- Savani Anbalagan
- Janna Blechman
- Michael Gliksberg
- Ludmila Gordon
- Gil Levkowitz
United States-Israel Binational Science Foundation (2017325)
- Michael Gliksberg
- Gil Levkowitz
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: Experiments involving zebrafish were approved by the Weizmann Institute'sInstitutional Animal Care and Use Committee (protocol #27220516)
Copyright
© 2019, Anbalagan et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Robo2 regulates synaptic oxytocin content by affecting actin dynamics
eLife 8:e45650.
https://doi.org/10.7554/eLife.45650
Further reading
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- Developmental Biology
The evolutionarily conserved Hippo (Hpo) pathway has been shown to impact early development and tumorigenesis by governing cell proliferation and apoptosis. However, its post-developmental roles are relatively unexplored. Here, we demonstrate its roles in post-mitotic cells by showing that defective Hpo signaling accelerates age-associated structural and functional decline of neurons in Caenorhabditis elegans. Loss of wts-1/LATS, the core kinase of the Hpo pathway, resulted in premature deformation of touch neurons and impaired touch responses in a yap-1/YAP-dependent manner, the downstream transcriptional co-activator of LATS. Decreased movement as well as microtubule destabilization by treatment with colchicine or disruption of microtubule-stabilizing genes alleviated the neuronal deformation of wts-1 mutants. Colchicine exerted neuroprotective effects even during normal aging. In addition, the deficiency of a microtubule-severing enzyme spas-1 also led to precocious structural deformation. These results consistently suggest that hyper-stabilized microtubules in both wts-1-deficient neurons and normally aged neurons are detrimental to the maintenance of neuronal structural integrity. In summary, Hpo pathway governs the structural and functional maintenance of differentiated neurons by modulating microtubule stability, raising the possibility that the microtubule stability of fully developed neurons could be a promising target to delay neuronal aging. Our study provides potential therapeutic approaches to combat age- or disease-related neurodegeneration.
