Optical voltage imaging and nonviral gene delivery for mechanistic investigation of Parkinson’s disease
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Abstract
Parkinson’s disease (PD) is the most prevalent neurodegenerative movement disorder characterized by the loss of midbrain dopaminergic neurons that project to the striatum. The striatum, a key basal ganglia region associated with movement, has been broadly implicated in PD. However, it remains unclear how striatal neural dynamics regulate voluntary movement and how their disruption contributes to motor pathology in PD. In this dissertation, we performed cell type–specific voltage imaging to investigate circuit-level neural dysfunction in PD and develop a nonviral gene delivery platform that expands the capacity for long-lasting transgene expression in the mammalian brain.In Chapter 2, we selectively expressed the genetically encoded voltage indicator SomArchon in cholinergic interneurons (ChIs) using ChAT-Cre mice to probe the impact of dopamine (DA) loss on ChI cellular dynamics in vivo. Leveraging our lab’s expertise as a leader in voltage imaging, we were able to perform high-resolution recordings of ChI neural activity. We also infused 6-hydroxydopamine (6-OHDA) unilaterally into the mouse striatum to induce DA loss. Although comprising only ~1-2% of striatal neurons, ChIs play a critical role in modulating both the direct and indirect pathways. Using optical voltage imaging, we identified that dopamine depletion exaggerates ChI burst firing, which is crucial for shaping striatal excitability. Additionally, dopamine loss impaired ChI encoding of locomotion, indicating that aberrant patterning of ChI firing contributes to degraded movement signaling in PD. Together, these results provide new mechanistic insight into how striatal interneuron function is altered in dopamine-depleted states and enhance our understanding of striatal circuit-level pathology in PD.
In Chapter 3, we developed a nonviral gene delivery method that targets brain cells, providing a novel tool to modulate the circuits we studied in Chapter 1. We screened lipid nanoparticle (LNP) formulations loaded with modified self-amplifying RNA (saRNA) containing cytidine substitution 5-hydroxymethylcytidine (hm5C), known to reduce innate immune responses. When injected into the striatum of mice, modified saRNA encapsulated in an LNP formulation comprising ALC-0315 (present in Comirnaty®; COVID-19 vaccine by BioNtech/Pfizer) efficiently mediates robust and long-lasting protein expression in brain cells beyond five weeks, with detectable expression in some neurons at three months. hm5C saRNA substantially outperforms N1-methyl-pseudouridine (N1mΨ) mRNA, a modified mRNA used in some COVID-19 vaccines. Intriguingly, in addition to transfecting astrocytes and neurons at the injection site, saRNA-LNPs labels neurons retrogradely. Thus, saRNA-LNPs are a promising nonviral gene transduction method that effectively transduces brain cells with excellent potency and mediates prolonged gene expression.
Together, this dissertation combines mechanistic investigation of striatal circuit dysfunction with the development of a molecular tool towards studying this circuitry. Chapter 2 establishes a powerful optical platform for precision analysis of ChI neural dynamics in motor impairment in PD, while Chapter 3 introduces a gene delivery platform that demonstrates capabilities for long-term gene expression and neuromodulation of striatal circuits in PD. This conceptual framework advances our understanding of PD and provides a foundation for future mechanistic and translational studies.
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2026