Multivariate assessment of a preclinical mouse model of Parkinson s disease: Connecting the dots to the human disease

1. Multivariate assessment of a preclinical mouse model of Parkinson�s disease: �Connecting the dots� to the human disease state Diana L. Price, Ph.D. National Center for Microscopy and Imaging Research University of California, San Diego Most of you heard my presentation at the Mouse BIRN AHM this past spring, so I will not well on the background. We�ve made some progress since that time primarily in the area of MRI studies with Dr. Johnson�s group.Most of you heard my presentation at the Mouse BIRN AHM this past spring, so I will not well on the background. We�ve made some progress since that time primarily in the area of MRI studies with Dr. Johnson�s group.

2. Deliverables mGluR5 manuscript: 2006 D1/D2 immuno: confirm. w/ larger N MRI analyses: 2006 MRI analyses: T1 & T2: baseline measures

3. The alpha-synuclein project: A multi-site, multi-disciplinary collaboration NCMIR � UCSD (M.H. Ellisman) Advanced light and electron microscopy Diana Price Van Phung Maryann Martone Heather Nguyen Andrea Thor Experimental Neuropathology Laboratory -UCSD (E. Masliah) Animal models & Parkinson�s strategy Edward Rockenstein Mike Mante This project is a BIRN testbed project which involves many sites with multi-disciplinary contributions towards characterizing mouse models of PD Our group at NCMIR is using multiscale imaging methods to characterize transgenic mouse models of PD produced by Dr. Eliezer Masliah�s group here at UCSD. We�ve also worked with Dr. Masliah�s group to acquire behavioral and molecular biology data on these same mouse models. Dr. Al Johnson�s group at Duke University�s Center for In Vivo Microscopy is producing high resolution microscopic MRI data. The multimodal data we are collecting will allow us to evaluate therapeutics. Some of these approaches will involve pharmacological treatments (such as the orally active Apomorphine targeting on-off syndrome in medicated PD patients developed by John Neumeyer at Harvard University). Others may include immunization strategies (as demonstrated by Dr. Masliah�s group earlier this year) and targeted delivery (such as the nanoparticle delivery mechanism developed by Julia George at the University of Illinois. This project is a BIRN testbed project which involves many sites with multi-disciplinary contributions towards characterizing mouse models of PD Our group at NCMIR is using multiscale imaging methods to characterize transgenic mouse models of PD produced by Dr. Eliezer Masliah�s group here at UCSD. We�ve also worked with Dr. Masliah�s group to acquire behavioral and molecular biology data on these same mouse models. Dr. Al Johnson�s group at Duke University�s Center for In Vivo Microscopy is producing high resolution microscopic MRI data. The multimodal data we are collecting will allow us to evaluate therapeutics. Some of these approaches will involve pharmacological treatments (such as the orally active Apomorphine targeting on-off syndrome in medicated PD patients developed by John Neumeyer at Harvard University). Others may include immunization strategies (as demonstrated by Dr. Masliah�s group earlier this year) and targeted delivery (such as the nanoparticle delivery mechanism developed by Julia George at the University of Illinois.

4. Parkinson�s Disease & Alpha-synuclein Neuronal cell death Lewy Bodies Alpha-synuclein protein (a-SYN) Mouse model overexpressing a-SYN The pathological features of Parkinson�s disease include nigrostriatal neuronal cell death, and protein occlusions called Lewy bodies. A major component of these Lewy bodies is alpha-synuclein protein To recreate these features in an animal model of PD, Dr. Masliah and colleagues produced a transgenic mouse that overexpresses h-alpha-synuclein. The pathological features of Parkinson�s disease include nigrostriatal neuronal cell death, and protein occlusions called Lewy bodies. A major component of these Lewy bodies is alpha-synuclein protein To recreate these features in an animal model of PD, Dr. Masliah and colleagues produced a transgenic mouse that overexpresses h-alpha-synuclein.

5. Challenges in PD research Disease heterogeneity Complex etiology (genetics, environment) Model systems Connecting the dots PD research has substantial challenges associated with making connections between the in vitro-preclinical models-and human disease state. First, Parkinson�s disease is actually a spectrum of disorders (Disease heterogeneity) Second, the different disease states result from a complex etiology (genetics, environment, or interaction) Third, there are no perfect model systems. Instead, there are models which recapitulate certain features of the disease state. Here you see a disease map of certain pathological features. Some of these features are found in both the human and animal model (solid line � point) and others are not (dashed line) As a collaborative group, we are using multi-disciplinary methods to connect the dots PD research has substantial challenges associated with making connections between the in vitro-preclinical models-and human disease state. First, Parkinson�s disease is actually a spectrum of disorders (Disease heterogeneity) Second, the different disease states result from a complex etiology (genetics, environment, or interaction) Third, there are no perfect model systems. Instead, there are models which recapitulate certain features of the disease state. Here you see a disease map of certain pathological features. Some of these features are found in both the human and animal model (solid line � point) and others are not (dashed line) As a collaborative group, we are using multi-disciplinary methods to connect the dots

6. How well does the a-syn mouse recreate features of PD? Widespread a-syn aggregates in CNS Neurochemistry mGluR5 receptor expression Dopamine D1/D2 receptors MRI studies: volume & composition Cognitive and motor deficits As with any model, we ask how well does the a-syn mouse recreate features of PD? These animals have several key PD-like pathological features such as: Widespread alpha-synuclein immunopositive aggregates Altered mGluR5 and dopamine receptor expressions These animals also have a wide range of PD-like cognitive and motor neurological deficits.As with any model, we ask how well does the a-syn mouse recreate features of PD? These animals have several key PD-like pathological features such as: Widespread alpha-synuclein immunopositive aggregates Altered mGluR5 and dopamine receptor expressions These animals also have a wide range of PD-like cognitive and motor neurological deficits.

7. Multimodal studies of PD animal models The characterization of the a-syn tg mouse model of PD includes multi-modal data including: Behavioral data Assessment of cognitive and motor function Pathological correlates with PD-like symptoms (severity, motor vs. cognitive associated neural circuitry) Evaluation of putative PD-therapeutics Multi-scale imaging compare regional changes in signal intensity with other data, such as large scale brain maps of alpha-synuclein immunoreactivity determine of how pathology visible at the light microscopic level is manifested in images obtained by MRI methods Chemistry and genetics Protein techniques, QTL analyses (i.e. looking at differences in gene expression levels), and ligand binding studies can be used to guide multiscale studiesThe characterization of the a-syn tg mouse model of PD includes multi-modal data including: Behavioral data Assessment of cognitive and motor function Pathological correlates with PD-like symptoms (severity, motor vs. cognitive associated neural circuitry) Evaluation of putative PD-therapeutics Multi-scale imaging compare regional changes in signal intensity with other data, such as large scale brain maps of alpha-synuclein immunoreactivity determine of how pathology visible at the light microscopic level is manifested in images obtained by MRI methods Chemistry and genetics Protein techniques, QTL analyses (i.e. looking at differences in gene expression levels), and ligand binding studies can be used to guide multiscale studies

8. Multiscale imaging studies at NCMIR Electron microscopy Multiphoton and confocal microscopy At NCMIR we are conducting multi-scale imaging studies using electron microscopy and light microscopies such as (multiphoton and confocal microscopy) to study the ultrastructural features and widespread distributions of PD-related CNS markers in the alpha-synuclein tg mouse. We are correlating our findings with the other types of data that I described in the previous slide to give us a more complete picture of the animal model. These data give us a a complete set of �baseline� pathological and behavioral features that will allow us to evaluate therapeutics. At NCMIR we are conducting multi-scale imaging studies using electron microscopy and light microscopies such as (multiphoton and confocal microscopy) to study the ultrastructural features and widespread distributions of PD-related CNS markers in the alpha-synuclein tg mouse. We are correlating our findings with the other types of data that I described in the previous slide to give us a more complete picture of the animal model. These data give us a a complete set of �baseline� pathological and behavioral features that will allow us to evaluate therapeutics.

9. New MRI imaging techniques for improved resolution and contrast Clinical imaging limitations Evaluation of preclinical models High resolution T1 weighted imaging High contrast T2 weighted imaging Correlation histological data 12 T1 weighted datasets (6 female & 6 males) and correlated T2 weighted datasets Facilitation of processing procedures Pre-processing underway Volumetric analyses & regional intensities Goal: Extension to non-invasive clinical imaging Towards this limitation, Dr. Al Johnson of Duke University�s Center for In Vivo Microscopy is developing new MRI imaging techniques for improved resolution and contrast. Clinical imaging limitations include magnet size and contrast agents. We are using Dr. Johnson�s techniques to evaluate whether the a-syn mice have changes in regional volumes or tissue composition. We�ve just completed the acquisition of a high-resolution set of images. Preliminary assessments demonstrate signal intensity differences in tg mice which correlate to areas with increased a-syn immunolabeling. If confirmed by complete analyses, we plan to assess whether passive immunization results in a �normalization� of transgenic images. The techniques used to generate these images should provide information that can be applied to clinical imaging of PD.Towards this limitation, Dr. Al Johnson of Duke University�s Center for In Vivo Microscopy is developing new MRI imaging techniques for improved resolution and contrast. Clinical imaging limitations include magnet size and contrast agents. We are using Dr. Johnson�s techniques to evaluate whether the a-syn mice have changes in regional volumes or tissue composition. We�ve just completed the acquisition of a high-resolution set of images. Preliminary assessments demonstrate signal intensity differences in tg mice which correlate to areas with increased a-syn immunolabeling. If confirmed by complete analyses, we plan to assess whether passive immunization results in a �normalization� of transgenic images. The techniques used to generate these images should provide information that can be applied to clinical imaging of PD.

10. Non-invasive MRI imaging of a-syn tg mice Here are examples of recent MRI data Examples of T1 weighted images of the striatal region demonstrate an apparent difference in tissue composition (larger �patches�). These same brains were also imaged using another pulse sequence These �T2� weighted images reveal further evidence of differences in tissue composition as we see here by an overlay of corresponding pseudo colored T2 images. Here are examples of recent MRI data Examples of T1 weighted images of the striatal region demonstrate an apparent difference in tissue composition (larger �patches�). These same brains were also imaged using another pulse sequence These �T2� weighted images reveal further evidence of differences in tissue composition as we see here by an overlay of corresponding pseudo colored T2 images.

11. Non-invasive MRI imaging of a-syn tg mice Here are examples of recent MRI data Examples of T1 weighted images of the striatal region demonstrate an apparent difference in tissue composition (larger �patches�). These same brains were also imaged using another pulse sequence These �T2� weighted images reveal further evidence of differences in tissue composition as we see here by an overlay of corresponding pseudo colored T2 images. Here are examples of recent MRI data Examples of T1 weighted images of the striatal region demonstrate an apparent difference in tissue composition (larger �patches�). These same brains were also imaged using another pulse sequence These �T2� weighted images reveal further evidence of differences in tissue composition as we see here by an overlay of corresponding pseudo colored T2 images.

12. Opportunities for evaluating potential therapies Pharmacological Dopaminergic: Dopamine D1/D2 receptor agonists Glutamatergic: mGluR5 receptor antagonists Immunization Passive immunization: clearing of protein aggregates Multivariate assessment allows us to evaluate these therapeutics on pathological and behavioral levels for correlation with clinical observations Several opportunities exist for us to evaluate these potential therapeutics including pharmacological (eg. dopaminergic and glutamateric ligands) and immunization strategies. Our multi-modal strategy will allow us to completely assess the efficacy of these therapeutics on several levels relevant to the clinical disease state. One challenge of clinical PD research includes the limits of in vivo assessments of neuropathological features.Several opportunities exist for us to evaluate these potential therapeutics including pharmacological (eg. dopaminergic and glutamateric ligands) and immunization strategies. Our multi-modal strategy will allow us to completely assess the efficacy of these therapeutics on several levels relevant to the clinical disease state. One challenge of clinical PD research includes the limits of in vivo assessments of neuropathological features.

13. Publications D.L. Price, S.K. Chow, N.A.B. MacLean, H. Hakozaki, S. Peltier, M.E. Martone and M.H. Ellisman (In Press) High-Resolution Large-Scale Mosaic Imaging Using Multiphoton Microscopy to Characterize Transgenic Mouse Models of Human Neurological Disorders, Neuroinformatics, Special Issue: Integration and modeling of imaging in the phenotype-genotype problem D.L. Price, E. Rockenstein, N.A.B. MacLean, M. Mante, V. Phung, D. Askay, E. Masliah, and M.H. Ellisman (In preparation) Increased mGluR5 immunoreactivity and behavioral deficits in transgenic mice overexpressing alpha-synuclein