Mécanismes moléculaires régulant la pathologie dendritique dans la rétine adulte lésée in vivo

Dendrites are major determinants of how retinal neurons integrate and process incoming information. In neurodegenerative disorders, such as glaucoma, dendritic alterations are usually accompanied by axonopathy. However, the mechanisms that regulate these pathological changes are poorly understood. The physiopathology of glaucoma, characterized by a progressive and irreversible degeneration of retinal ganglion cells (RGCs), is the leading cause of irreversible blindness worldwide. This desease is associated with a gradual thining of RGC axons and loss of RGC soma. Most studies on neuroprotection have focused on morphological and quantitative analysis of RGC bodies and axons. Yet, dendrites are critical for neurotransmission between retinal neurons and, as such, play a vital role in the functional properties of the retinal circuit. The main hypotheses of this thesis is that: i) axonal injury induces early dendrite pathology, and ii) the identification of signaling pathways underlying dendritic changes is critical to design neuroprotective strategies and restore neuronal function.  In the first study, we investigated the molecular mechanisms involved in the early dendritic changes that occur in RGCs induced by an acute axonal injury (axotomy) in vivo. Using adult transgenic mice carrying the yellow fluorescent protein (YFP) gene subjected to optic nerve axotomy, we demonstrate a marked retraction of RGC dendritic arbors and selective downregulation of the mammalian target of rapamycin (mTOR) activity prior to overt cell death. Axotomy triggered rapid up-regulation of the stress-induced protein Regulated in development and DNA damage response 2 (REDD2), a potent inhibitor of mTOR. Treatment with siRNA against REDD2 stimulated mTOR activity in axotomized RGCs and promoted a significant increase in the length and complexity of injured RGCs compared to retinas treated with control siRNA. Administration of rapamycin completely blocked the effect of siREDD2 on dendritic growth and branching thus confirming that this response occurred via mTOR stimulation. Whole-cell recordings demonstrated that REDD2 depletion leading to mTOR activation in RGCs restored their light response properties. Lastly, we show that REDD2-dependent mTOR activity extended RGC survival following axonal damage. Collectively, these results demonstrate that injury-induced stress leads to REDD2 upregulation, mTOR inhibition and dendrite pathology causing neuronal dysfunction and subsequent cell death. Recent data demonstrate a neuroinflammatory component in glaucoma characterized by upregulation of pro-inflammatory cytokines, most notably tumor necrosis factor α (TNFα) (Tezel, 2013, Soto and Howell, 2014). Therefore, in the second study of my thesis, we investigated the effect of exogenous TNFα on RGC dendritic structure in vivo and initiated the analysis of molecular signals involved in this response. Our results show that injection of recombinant TNFα into the vitreous chamber of YFP mice induced early retraction of the dendritic arbors of RGCs concomitant with a reduction of mTOR activity in these neurons. Taken together, the results shown in this thesis reveal a novel role for mTOR in the stability and maintenance of dendrites in adult retinal neurons. These studies also demonstrate the effect of early direct and indirect stressors, axotomy or TNFα respectively, on the pathology of dendrites and their effect on neuronal function.

Keyphrases: Dendrite, Mammalian target of rapamycin, Regulated in development and dna damage 2, Neuroprotection, Retinal ganglion cell, Axonal injury, Neurodegeneration, Tumor necrosis factor - alpha