INTRODUCTION: To ensure adequate energy supply in the brain, iron levels are precisely regulated, as it is vital for the mitochondrial electron transport chain (ETC). In neurodegenerative diseases like Parkinson's, iron dysregulation can trigger ferroptosis, leading to neuronal loss and fueling disease progression. Using zebrafish (Danio rerio), an excellent in vivo model due to their optical transparency, genetic tractability, and conserved neurochemical architecture, we aim to establish strategies for inducing this pathological iron accumulation.

AIM(S): We aim to develop approaches to induce pathological iron accumulation and determine if ferroptotic stress directly mimics Parkinsonian-like dopaminergic neuronal death. To model Parkinson’s and ferroptosis-like conditions using MPTP, RSL3, and transgenic PINK1 lines, while also developing a CRISPR-Cas9-based LRRK2 knock-in line with Ms. Sofia Baranykova.

METHOD(S): Our approach involves generating models of Parkinson's Disease and ferroptosis using PINK1 mutant transgenic lines, alongside creating and characterizing chemically induced models. We expose the 5dpf wild-type larvae to varying concentrations of MPTP, and the PINK1 models to mimic PD-like environment while RSL3/Erastin are aimed at disrupting iron homeostasis. To investigate the molecular mechanisms of ferroptosis, we will examine the expression of key markers like GPX4, HMOX1, HMGB1, and MCU. Furthermore, in our various models, we utilize Perl's Prussian Blue staining to identify iron buildup that correlates with ferroptotic progression.

RESULTS: Initial lab findings show that PINK1 mutants exhibit DA neuronal loss at 3 dpf, which is mitigated by inactivation of the mitochondrial calcium uniporter (MCU).

CONCLUSIONS: These coordinated strategies will lay the groundwork for establishing a zebrafish-based platform for studying neurodegeneration and ferroptosis ultimately facilitating future pharmacological screening. As the project is in its early stages, definitive conclusions are yet to be drawn.

FINANCIAL SUPPORT: This study is supported by the National Science Centre (NCN), OPUS 25, project no. 2023/49/B/NZ4/02744 to JK.