Gutekunst, C.-A. N. | Search bioRxiv

bioRxiv2022

Chronic activation of dopaminergic neurons via bioluminescence-optogenetics provides neuroprotection in a rodent model of Parkinson's disease

Previous studies in human patients and rodent models of Parkinsons disease (PD) have established neuroprotection of dopaminergic (DA) neurons in substantia nigra pars compacta (SNC) by physical exercise, but the precise origin of this neuroprotective effect has yet to be elucidated. In this study, we tested a hypothesis that enhanced activity of DA neurons in SNC results in neuroprotection using the unilateral 6-hydroxydopamine (6-OHDA) injection model in mice. To increase activity of DA neurons chronically and specifically, we injected an adeno-associated viral vector carrying a step-function luminopsin (SFL) - a fusion protein of light-emitting Gaussia luciferase and light-sensing step-function channelrhodopsin 2 - into SNC ipsilateral to 6-OHDA using the pan-neuronal human synapsin I promoter or the Cre-lox system in transgenic mice expressing the recombinase under control of the tyrosine hydroxylase (TH) promoter. Upon application of SFL substrate, coelenterazine (CTZ), the luciferase moiety of luminopsin emits bioluminescence which in turn activate the opsin moiety. Daily injection of CTZ for 4 weeks ameliorated a stereotypical behavior, namely ipsiversive rotations, induced by unilateral 6-OHDA. In addition, postmortem immunohistochemistry against TH revealed less severe neurodegeneration of DA neurons compared to vehicle-injected control animals. Furthermore, when mice were pretreated with ANA-12, a selective antagonist for tropomyosin receptor kinase B (TrkB), the behavioral improvement and neuroprotective effect were diminished. These results suggest that increased neuronal activity of DA neurons provides neuroprotection against 6-OHDA injury and alleviates its symptoms through the brain-derived neurotrophic factor-TrkB pathway.

neuroscience↗

bioRxiv2021

Kinetic Monitoring Of Neuronal Stress Response To Proteostasis Dysfunction

Proteostasis dysfunction and activation of the unfolded protein response (UPR) are characteristic of all major neurodegenerative diseases. Nevertheless, although the UPR and proteostasis dysfunction has been studied in great detail in model organisms like yeast and mammalian cell lines, it has not yet been examined in neurons. In this study, we applied a viral vector-mediated expression of a reporter protein based on a UPR transcription factor, ATF4, and time-lapse fluorescent microscopy to elucidate how mouse primary neurons respond to pharmacological and genetic perturbations to neuronal proteostasis. In in vitro models of endoplasmic reticulum (ER) stress and proteasome inhibition, we used the ATF4 reporter to reveal the time course of the neuronal stress response relative to neurite degeneration and asynchronous cell death. We showed how potential neurodegenerative disease co-factors, ER stress and mutant -synuclein overexpression, impacted neuronal stress response and overall cellular health. This work therefore introduces a viral vector-based reporter that yields a quantifiable readout suitable for non-cell destructive kinetic monitoring of proteostasis dysfunction in neurons by harnessing ATF4 signaling as part of the UPR activation.

cell biology↗