Abstract
Neuroregeneration is a dynamic process synergizing the functional outcomes of multiple signaling circuits. Channelrhodopsin-based optogenetics shows the feasibility of stimulating neural repair but does not pin down specific signaling cascades. Here, we utilized optogenetic systems, optoRaf and optoAKT, to delineate the contribution of the ERK and AKT signaling pathways to neuroregeneration in live Drosophila larvae. We showed that optoRaf or optoAKT activation not only enhanced axon regeneration in both regeneration-competent and -incompetent sensory neurons in the peripheral nervous system but also allowed temporal tuning and proper guidance of axon regrowth. Furthermore, optoRaf and optoAKT differ in their signaling kinetics during regeneration, showing a gated versus graded response, respectively. Importantly in the central nervous system, their activation promotes axon regrowth and functional recovery of the thermonociceptive behavior. We conclude that non-neuronal optogenetics targets damaged neurons and signaling subcircuits, providing a novel strategy in the intervention of neural damage with improved precision.
Highlights
Using a thermonociception based behavioral recovery assay, we found that optoRaf and optoAKT activation led to effective axon regeneration as well as functional recovery after central nervous system (CNS) injury
We focused on the axons of C4da neurons, which project into the ventral nerve cord 275 (VNC) and form a ladder-like structure
Neurotrophins are known to activate Trk receptors and trigger the Ras/MEK/ERK, AKT, and phospholipase C (PLC) pathways which are involved in cell survival, neural differentiation, axon and dendrite growth and sensation (Bibel and Barde 2000; Huang and Reichardt 2001; Chao 2003; Cheng et al 2011; Joo et al 2014)
Summary
Inadequate neuroregeneration remains a major roadblock towards functional recovery after nervous system damage such as stroke, spinal cord injury (SCI), and multiple sclerosis.Extracellular factors from oligodendrocyte, astroglial, and fibroblastic sources restrict axon regrowth (Liu et al 2006; Yiu and He 2006; Liu et al 2011; Lu et al 2014; Schwab and Strittmatter 2014) but eliminating these molecules only allows limited sprouting (Sun and He2010), suggesting a down-regulation of the intrinsic regenerative program in injured neurons (Sun and He 2010; He and Jin 2016). The neurotrophic signaling pathway, which regulates neurogenesis during embryonic development, represents an important intrinsic regenerative machinery (Ramer et al 2000). Elimination of the PTEN phosphatase, an endogenous brake for neurotrophic signaling, yields axonal regeneration (Park et al 2008). An important feature of the neurotrophin signaling pathway is that the functional outcome depends on signaling kinetics (Marshall 1995) and subcellular localization (Watson et al 2001). Neural regeneration from damaged neurons is synergistically regulated by multiple signaling circuits in space and time. Pharmacological and genetic approaches do not provide sufficient spatial and temporal resolutions in the modulation of signaling outcomes in terminally differentiated neurons in vivo. The functional link between signaling kinetics and functional recovery of damaged neurons remains unclear. The emerging non-neuronal optogenetic technology uses light to control protein-protein interaction and enables light-mediated signaling modulation in live cells and multicellular organisms
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