Imaging the Development of Ventral Midbrain Nuclei

Rajeshwar Awatramani, Ph.D.

Northwestern University, Chicago, IL
Website

Grant Program:

David Mahoney Neuroimaging Program

Funded in:

June 2006, for 2 years

Funding Amount:

$100,000

Lay Summary

Neuronal Development Clues May Help Advance Stem Cell Treatment for Parkinson’s Disease

Northwestern University investigators will use molecular imaging in mice to learn how the brain cells that are destroyed in patients with Parkinson’s disease are initially developed. This insight may lead to an improved ability to develop stem cell therapy for patients with this degenerative disease.

Cells that produce the neurotransmitter dopamine in one region of the midbrain are selectively destroyed in patients who develop Parkinson’s disease. The anatomic, molecular, and functional characteristics of dopamine-producing cells in these regions are distinct. The researchers will determine whether they can develop molecular imaging techniques that can show, in mice, where embryonic progenitor cells migrate to within the midbrain to become dopamine-producing neurons. Specifically, the investigators will see if they can fluorescently tag the progenitor cells and use two-photon imaging to visualize the cells’ migratory paths. If so, the investigators then would be able to test their hypothesis that subsets of progenitor cells receive distinct genetic commands that instruct them to migrate to specific regions of the midbrain.

If that is the case, the research could lead to methods to identify the genes in the subgroup of cells affected by Parkinson’s disease. Identification of genes, in turn, could lead to development of stem cells designed to replace the dopamine-producing neurons that are destroyed in Parkinson’s disease. The imaging method developed in this project also could be used to determine whether those stem cells reach their intended destination.

Significance: This molecular imaging feasibility study is a first step in efforts to develop stem therapy for Parkinson’s disease. The technique also could be used, once stem cells are designed, to determine whether the stem cells migrate to the affected brain area in patients.

Abstract

Imaging the Development of Ventral Midbrain Nuclei

Parkinson's disease, a severely debilitating adult-onset neurodegenerative condition, is caused by a substantial depletion of a specific subset of midbrain dopaminergic neurons located in the substantia nigra pars compacta. Central to understanding the pathophysiology of Parkinson's disease and the formulation of effective stem cell based therapeutics, is a clear grasp of the regulatory molecules and mechanisms governing the specification and development of distinct midbrain dopaminergic neuron types. Towards this end, my goal is to define how embryonic midbrain neuroepithelial progenitor cells are programmed by combinations of gene products to give rise to distinct dopaminergic neuron types.

In Specific Aim 1 I will develop broadly applicable "second generation" intersectional genetic tools that enable the simultaneous fluorescent visualization of subsets of progenitor cells and their descendents in live tissue. In Specific Aim 2, I will test the hypothesis that the anatomically, molecularly and functionally distinct dopaminergic nuclei have distinct embryonic origins. Using the tools developed in Aim 1, I will dissect the ventral midbrain progenitor pool, by genetically tagging subsets of progenitors with a fluorescent protein (FP) and tracking tagged cells through development. Together, these studies will likely illuminate critical pathways in dopaminergic neuron development and will define the developmental basis for heterogeneity in midbrain dopaminergic populations.

Investigator Biographies

Rajeshwar Awatramani, Ph.D.

Assistant Professor of Neurology, Northwestern University

Hypothesis

Hypothesis:
Several anatomically and functionally distinct dopaminergic neuron types are present in the adult mammalian brain. We hypothesize that embryonic midbrain neuroepithelial progenitors are parceled into distinct domains that express unique combinations of regulatory molecules, which, at least in part, results in the specification of uniquedopaminergic types.

Goals:
We will determine how midbrain progenitors are parceled into molecularly distinct groups that give rise to disparate dopaminergic neuron types. Previous studies have defined molecular subdivisions within the midbrain tegmental neuroepithelium. Using genes known to subdivide the midbrain ventricular zone, to drive the expression of Cre and Flpe recombinase, we will use intersectional fate mapping approaches to simultaneously track the fate of distinct progenitor populations. We expect to shed light on the origin of several important midbrain populations, including the dopaminergic neurons of the substantia nigra. Together, these studies will address the hypothesis that dopaminergic diversity is a result of early embryonic specification events, and will thus have implications for programmed stem cell based therapies for Parkinson’s disease.

Methods:
We will utilize genetic lineage tracing methodologies to track the origins of dopamine neurons. The recent development of recombinase-mediated reporter gene activation has facilitated lineage analyses in mice. To improve the resolution of these approaches, we have developed a dual-recombinase mediated intersectional fate mapping strategy, in which a reporter is permanently activated only in progenitors located at the intersection of two gene expression domains. By developing new intersectional fluorescent reporters, additional capabilities such as live imaging, FACS sorting, and electrophysiological characterization of labeled cells will be possible.

Selected Publications

Kittappa R., Chang W.W., Awatramani R.B., and McKay R.D. The foxa2 gene controls the birth and spontaneous degeneration of dopamine neurons in old age. PLoS Biol. 2007 Dec;5(12):e325 .

Farago, A., Awatramani, R., and Dymecki, S. Assembly of the brainstem cochlear nuclear complex is revealed by intersectional and subtractive genetic fate maps. Neuron. 2006 Apr 20;50(2):205-18 .

Awatramani R., Soriano P., Rodriguez C., Mai J.J., and Dymecki S. Cryptic boundaries in roof plate and choroid plexus identified by intersectional gene activation. Nat Genet. 2003 Sep;35(1):70-5.