2013-2014 Post-Doctoral Award
João Paulo Lima Daher, MD, PhD
University of Alabama at Birmingham
Project Title: Defining LRRK2 Action in α-Synuclein Induced Neurodegeneration
(link to)
JP completed medical school in Rio de Janeiro with a focus in experimental pathology, and his
work there led him to pursue biomedical research in the laboratory of Professor Dan Berkowitz
at Johns Hopkins. JP continued onto a Master’s Program jointly in the laboratory of Professor
Daniel Weinreich at the University of Maryland, where he focused on physiology of the
autonomous nervous system. JP next pursued a doctoral degree focused on understanding
molecular mechanisms in Parkinson’s disease in the laboratories of Ted and Valina Dawson.
His research focus was on initial characterizations of transgenic models of LRRK2 and αsynuclein. After successful thesis dissertation defense, JP joined the laboratory of Dr. Andrew
West to further investigate the interaction between LRRK2 and α-synuclein, the two autosomal
dominant causes of Parkinson’s disease.
Objective: I propose to test whether LRRK2 modifies dopaminergic cells neurodegeneration
and neuroinflammatory responses associated with α-synuclein-over-expression in rat brains.
Background: Over-expression of α-synuclein can lead to dopaminergic cells
neurodegeneration and neuroinflammatory responses similar to that associated with
Parkinson’s disease. Having Dr. West’s laboratory recently demonstrated that LRRK2 plays a
critical role in mediating neuroinflammatory responses, I propose to investigate the interaction of
LRRK2 and α-synuclein using a novel research tool, rAAV2-based vector system, to induce
over-expression of α-synuclein into the brain of LRRK2 transgenic rats.
Methods/Design: I propose to investigate the interaction of LRRK2 and α-synuclein using a
novel recombinant adeno-associated virus, serotype 2 (rAAV2)-based vector system to induce
over-expression of α-synuclein into the substantia nigra pars compacta (SNpc) of LRRK2
transgenic rats. I propose to perform unilateral stereotaxic injections of the vector in the SNpc of
3 months-old LRRK2 WT/KO Long Evans rats and LRRK2 WT/G2019S Sprague Dawley rats.
Over-expression of α-synuclein in dopaminergic neurons will be evaluated by doubleimmunofluorescence at 4, 8 and 12 weeks post-injection. Neurodegeneration in these animals
will be evaluated at 4, 8 and 12 weeks post-injection by counting dopaminergic neurons using
unbiased stereology. Neuroinflammatory responses will be evaluated 4, 8 and 12 weeks postinjection by counting microglial cells in close proximity to the SNpc using unbiased stereology.
Moreover, I will perform neuropathological studies to search for α-synuclein aggregates in
midbrain sections. Finally, I will measure dopaminergic fibers density using optical density,
striatal dopaminergic content using HPLC, and locomotor function using a battery of behavioral
tests. The estimated time for the realization of this project is approximately 12 months, and most
of the work will focus on stereotaxic injections, immunohistochemistry, stereology and
neuropathological analysis.
Relevance to Diagnosis/Treatment of Parkinson’s disease: I hope to understand the
potential role of LRRK2 in modifying neuroinflammatory responses and associated
neurodegeneration in α-synuclein-injected rats. This is critically required to improve our
understanding of the molecular mechanisms of PD, particularly in the maintenance/survival of
SNpc dopaminergic neurons. I also hope that this model may reveal whether LRRK2 and αsynuclein function in a common pathogenic pathway and could potentially serve as a relevant
pre-clinical model of PD to test LRRK2 kinase inhibitors in order to halt or slow the underlying
progressive neurodegeneration/neuroinflammation that is characteristic of this neurological
disorder.