Parkinson’s disease (PD) is a multifactorial condition, presenting mitochondrial dysfunction as a hallmark. This mitochondrial impairment incurs bioenergetic and oxidative stress that triggers the demise of dopaminergic neurons. The causes of mitochondrial deficiency are not fully understood, but mitochondrial DNA (mtDNA) disruption likely contributes to this malfunction. Surprisingly, new findings implicate mtDNA as the nexus connecting mitochondrial (dys)function to inflammation in PD. Indeed, under stress conditions, mtDNA can be released from mitochondria into the cytosol or the extracellular milieu, where it activates specific sensors driving the engagement of immunological cascades. Much interest is now being directed at the elucidation of the molecular processes underlying mtDNA extrusion. In this regard, the formation of BAX/BAK macropores, which permeabilize the mitochondrial outer membrane, thus providing an outlet for the extrusion of the inner mitochondrial membrane, has been evidenced. Yet, direct visualization of mtDNA extrusion and the determination of the driving molecular apparatus has not been attained in the context of PD. The Molecular and Functional Neurobiology Group at LCSB (Head: Prof. Anne Grünewald) works on the elucidation of the cause and consequence of mitochondrial dysfunction in both idiopathic and familial forms of PD while putting a significant focus on the role of mtDNA in the etiology of this condition. Hence, we are currently exploring the mechanisms of mtDNA extrusion and its function as a vector for neuroinflammation. Within the framework of this INTERmobility grant, we want to address the following hypothesis: PD-related Parkin dysregulation prompts Bax/Bak macropore formation, promoting increased mtDNA release and subsequent inflammatory responses in patient neurons. Specifically, the project will comprise two research objectives: (i)To determine if Parkin influences the formation of BAX/BAK macropores and the subsequent herniation of mitochondria in PD; (ii)To characterize the composition of the mitochondrial damage-associated molecular patterns (DAMPs) released through inner mitochondrial membrane hernias in PD.To pursue these objectives, we are making use of a unique set of patient-derived neuroepithelial stem cells with Parkin mutations, which we have previously established. To analyze mitochondrial herniation events in a three-dimensional fashion and to study the composition and relative localization of mitochondrial DAMPs, we will apply correlative light and electron microscopy to Parkin-mutant neurons. At LCSB, we have access to STED microscopy, which enables us to intracellularly trace our mitochondrial target molecules (mtDNA, BAX/BAK, cytochrome c etc.) at super-resolution. By teaming up with Prof. Martin Picard (Columbia University, Irving Medical Centre, USA) an expert in quantitative 3D mapping of electron microscopy (EM) images, we will be in the position to integrate our enhanced subcellular localization data with ultrastructure information. We are confident that this combined approach will significantly advance our project, helping us reach our goals.We anticipate that the INTERmobility grant will generate the following outcomes:1)Elucidation of the mechanisms of mtDNA extrusion in PD2)Enhanced collaborative relationship between the two groups and institutes3)Career development by acquiring new innovative knowledge and skills4)Protocols and knowledge that will be transferred to the LCSB 5)High-quality publicationsThe INTERmobility grant will be instrumental to bring together the expertise and resource of both groups to answer a very relevant topic in the study of PD. This project will clarify a link between mitochondrial dysfunction and neuroinflammation helping identify molecular targets and mechanisms for tangible immunomodulatory interventions, which could be applied to slow disease progression.