Western blot – alpha-synuclein
By onThis protocol describes how to detect alpha-synuclein in protein derived from STC-1 cells by western blot using DAB/peroxidase or ECL to visualize the bands.
Mouse Brain Tissue Collection and Analysis
By onThis protocol describes the dissection and collection of coronal sections of the striatum and midbrain from a mouse brain. The tissue can be used in a number of applications and here we describe two: the measurement catecholamine levels using high-performance liquid chromatography (aided by a neurochemistry core) and the distribution of the vesicular monoamine transporter (VMAT2) using a radioligand binding assay.
Challenging Beam Test
By onThe challenging beam test is used to measure motor performance, coordination, and balance.
The emerging role of LRRK2 in tauopathies
By onParkinson’s disease (PD) is conventionally described as an α-synuclein aggregation disorder, defined by Lewy bodies and neurites, and mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common autosomal dominant cause of PD. However, LRRK2 mutations may be associated with diverse pathologies in patients with Parkinson’s syndrome including tau pathology resembling progressive supranuclear palsy (PSP). The recent discovery that variation at the LRRK2 locus is associated with the progression of PSP highlights the potential importance of LRRK2 in tauopathies. Here, the authors review the emerging evidence and discuss the potential impact of LRRK2 dysfunction on tau aggregation, lysosomal function, and endocytosis and exocytosis.
H9 ES AAVS1-NGN2 FAM134C-/-
By onES cells manipulated with CRISPR/Cas9 lack FAM134C receptor for ER-phagy. NEUROG2 construct introduced via AAVS1 locus in human embryonic stem cells, creating iNeurons. Derived from blastocyst stage, female, Homo sapiens.
Astrocyte extraction from brain organoids V.2
By onProtocol for astrocyte extraction from brain organoids.
Therapeutic deep brain stimulation disrupts movement-related subthalamic nucleus activity in Parkinsonian mice
By onSubthalamic nucleus deep brain stimulation (STN DBS) relieves many motor symptoms of Parkinson's Disease (PD), but its underlying therapeutic mechanisms remain unclear. Since its advent, three major theories have been proposed: (1) DBS inhibits the STN and basal ganglia output; (2) DBS antidromically activates motor cortex; and (3) DBS disrupts firing dynamics within the STN. Previously, stimulation-related electrical artifacts limited mechanistic investigations using electrophysiology. We used electrical artifact-free GCaMP fiber photometry to investigate activity in basal ganglia nuclei during STN DBS in parkinsonian mice. To test whether the observed changes in activity were sufficient to relieve motor symptoms, we then combined electrophysiological recording with targeted optical DBS protocols. Our findings suggest that STN DBS exerts its therapeutic effect through the disruption of movement-related STN activity, rather than inhibition or antidromic activation. These results provide insight into optimizing PD treatments and establish an approach for investigating DBS in other neuropsychiatric conditions.
Tet-off FLTau (HEK293T)
By onAssociated with the following preprint: Saha et al., 2022 (published on biorxiv on Feb 19th, 2022) https://www.biorxiv.org/content/10.1101/2022.02.18.481043v1.full
A unifying model for the role of the ATG8 system in autophagy
By onThe core ATG8 system is one of the most-studied autophagy components. Here, authors reconcile prior observations of the core ATG8 system into a unifying model.
Cell culture, transfection and imaging
By onThis protocol details the general preparation of cells for imaging and also for imaging experiments involving cellular hypotonic shock and cytosolic Ca2+ changes as they were performed in https://doi.org/10.1083/jcb.202010004.
Halo-LC3B processing assay to assess autophagy
By onThis protocol details Halo-LC3B processing assay to assess autophagy.
Insect Cell Protocol for LRRK1 and LRRK2 Expression
By onProtocol for expressing LRRK1 and LRRK2 in insect cells.
Structural basis for ATG9A recruitment to the ULK1 complex in mitophagy initiation
By onHere, the authors examine the structural interaction between ATG9A and components of the ULK1 complex to better understand the process of the PINK1- and Parkin- dependent mitophagy pathway implicated in Parkinson's disease.
Immunocytochemical analysis
By onAssociated with preprint: https://www.biorxiv.org/content/10.1101/2022.11.02.514817v1
Bone Marrow Derived Macrophage (BMDM) differentiation and maintenance
By onThis protocol describes the isolation of bone marrow derived macrophages.
Proteomics analysis of autophagy cargos reveals distinct adaptations in PINK1 and LRRK2 models of Parkinson disease
By onAutophagy is crucial for neuronal health. Studies on PD mouse models show adaptive pathways are activated to maintain brain balance, despite reduced autophagic flux in PINK1-/- mice and impaired autophagosomal function in LRRK2G2019S mice.
Wire Hang Test
By onThis behavioral test is designed to assess early motor deficits associated with muscle and grip strength.
