Autophagy is critical for neuron survival. In neurons, loss of autophagy leads to the formation of protein aggregates and neurodegeneration, both in vitro and in vivo after genetic disruption. Conversely, increasing autophagy ameliorates toxicity in a variety of neurodegenerative models. Thus, stimulation of autophagy has been proposed as a therapeutic strategy for neurodegenerative diseases characterized by the accumulation of unfolded proteins. mTOR is the canonical regulator of autophagy in non-neuronal cells, but our lab has shown that autophagy in primary neurons is not effectively regulated by mTOR. Drugs which induce mTOR- independent autophagy have been identified by our lab and others and could provide starting points for potential therapeutics in neurons. Although these drugs act on a variety of targets, a common activity that many share is that they reduce cytosolic calcium levels. We have novel data showing that this mechanism may also be responsible for mTOR-independent autophagy in neurons. Mimicking low cytosolic calcium in neurons using the calmodulin inhibitors fluphenazine and trifluoperazine leads to robust induction of mTOR- independent autophagy. However, the potential downstream mediator of cytosolic Ca2+-mediated autophagy is unknown. We have identified calcineurin, the only calcium sensitive phosphatase, as a potential regulator of neuronal autophagy downstream of low cytosolic calcium. Inhibition of calcineurin by two structurally distinct compounds, Cyclosporin A (CsA) and FK506, induced autophagy in a dose-dependent manner in primary neurons. Autophagy induction by CsA and FK506 is intriguing in light of previous data showing them to be protective in several neurodegenerative models. Our previous work has shown the neuroprotective role of autophagy, but it is unknown if the protective effects of CsA and FK506 are the result of autophagy induction. We hypothesize that calcineurin integrates high cytosolic Ca2+ levels to negatively regulate neuronal autophagy induction and that the protective effects of CsA/FK506 are mediated through autophagy induction by the inhibition of calcineurin.
In Aim 1 we will determine whether calcineurin is sufficient to induce autophagy using more specific genetic means, and we will determine if neuronal autophagy is inhibited by activated calcineurin.
In Aim 2 we will determine whether the protective effect of CsA/FK506 in HD is through autophagy by looking at whether the protective effect is abolished with inhibition of autophagy and whether the rate of autophagy in a neuron is predictive of neuronal death by apoptosis.
Our research seeks to understand what controls the natural process of autophagy in neurons. Autophagy can remove damaged cellular components and ameliorate aspects of the Huntington's disease, a disease where a misfolded protein can lead to neuron death. Understanding and subsequent exploiting these natural mechanisms will allow for the design of novel treatment strategies for this currently untreatable disease.