Abnormal dopamine signaling in a mouse model of dystonia Público

Abstract

Dystonia is a neurological movement disorder characterized by involuntary twisting movements and unnatural postures. Current treatment options for dystonia are palliative or inadequate for many patients. The limited effectiveness of current treatment options highlights our incomplete understanding of underlying pathomechanisms. Studies of acquired, inherited, and idiopathic dystonias have led to the identification of some common etiologies, specifically, abnormal dopamine signaling in the basal ganglia. Historically, dystonia has been viewed as a disorder of the basal ganglia and studies also directly support a role for postsynaptic dopamine dysfunction in the striatum, the primary input nucleus of the basal ganglia, in many forms of dystonia. Dopamine exerts opposite physiological effects on neural activity via activation of dopamine D1 (D1Rs) or D2 receptors (D2Rs), which are expressed on the primary output neurons of the striatum, so-called medium spiny neurons. However, the precise nature of abnormal postsynaptic dopamine-mediated intracellular signaling pathways in dystonia is unknown. Until recently, it was not possible to study abnormal postsynaptic dopamine-mediated signaling pathways in dystonia because suitable animal models that exhibit dystonia caused by dysfunction in dopamine neurotransmission were not available. However, studies in a recently developed mouse model of dystonia (DRD mice) have identified specific abnormalities in dopamine-mediated signaling implicated in dystonia. I utilized the DRD mouse model to determine the precise nature of striatal dysfunction that may distinguish dystonia from other abnormal motor phenotypes. My findings showed that dysfunction of both D1Rs and D2Rs is necessary for the expression of dystonia. I determined that DRD mice exhibit blunted striatal D2R-mediated intracellular signaling and supersensitive striatal D1R-mediated intracellular signaling. My results suggest that abnormal striatal D1R-mediated signaling may be necessary, but not sufficient, for the expression of dystonia. Determining the relationship between D1R- and D2R-mediated effects of abnormal striatal intracellular signaling may ultimately reveal the precise nature of dysfunction in dopamine neurotransmission that gives rise to dystonia. 

Table of Contents

Chapter 1: Introduction

1.1. Historical perspective 1

1.2. Epidemiology 2

1.3. Phenomenology 2

1.3.1. Clinical characteristics 3

1.4. Treatments 4

1.4.1. Botulinum toxin 4

1.4.2. Small molecule medications 5

1.4.3. Surgical procedures 7

1.5. Etiology 8

1.5.1. Brain areas involved in dystonia 9

1.5.2. Evidence supporting basal ganglia dysfunction in dystonia 10

1.5.3. Evidence supporting dopamine signaling dysfunction in dystonia 11

1.6. Anatomy of the basal ganglia 15

1.7. Dopamine signaling in the striatum 17

1.7.1. Second messenger-dependent protein phosphorylation signaling 20

1.7.2. Transcriptional regulation 22

1.8. Abnormal postsynaptic dopamine-mediated signaling pathways in dystonia 23

1.9. Overall objective 28

Chapter 2: A novel method using QuPath to automate counts of immunoreactive cells and define striatal regions

2.1. Abstract 30

2.2. Introduction 31

2.3. Methods 32

2.4. Results 34

2.4.1. Confirmation of 6-OHDA-lesion 34

2.4.2. Immunoreactive cell count automation 35

2.4.3. Defining striatal regions 39

2.5. Discussion 39

Chapter 3: Abnormal dopamine receptor-mediated intracellular signaling in a mouse model of DOPA-responsive dystonia

3.1. Abstract 41

3.2. Introduction 42

3.3. Methods 43

3.4. Results 50

3.4.1. Behavioral analysis of DRD mice expressing fluorescent reporter proteins 50

3.4.2. Distribution of D1R-tdTom and D2R-EGFP striatal neurons 51

3.4.3. D1R agonist-mediated signaling 53

3.4.4. D2R agonist-mediated signaling 59

3.4.5. D2R antagonist-mediated signaling 63

3.5. Discussion 69

3.5.1. Distribution of D1R- and D2R-MSNs 70

3.5.2. Regional differences in dopamine signaling in the dorsal striatum 70

3.5.3. D1R-mediated signaling 71

3.5.4. D2R-mediated signaling 73

3.5.5. Implications of blunted D2R inhibition in DRD mice 74

3.5.6. Conclusions 75

Chapter 4: Determining the therapeutic mechanism of L-DOPA in a mouse model of DOPA-responsive dystonia

4.1. Abstract 76

4.2. Introduction 77

4.3. Methods 77

4.4. Results 79

4.4.1 L-DOPA-induced p-PKA-sub immunoreactivity 79

4.4.2 L-DOPA-induced p-ERK immunoreactivity 81

4.4.3. L-DOPA-induced c-Fos immunoreactivity 83

4.4.3. Correlation of L-DOPA-induced p-PKA-sub, p-ERK, and c-Fos immunoreactivity 85

4.5. Discussion 87

4.5.1. L-DOPA-induced postsynaptic intracellular signaling cascades 87

4.5.2. Regional differences in L-DOPA-induced ERK phosphorylation 89

4.5.3. Conclusions 90

Chapter 5: Determining the role of D1Rs and D2Rs in the therapeutic mechanism of L-DOPA in a mouse model of DOPA-responsive dystonia

5.1. Abstract 92

5.2. Introduction 93

5.3. Methods 93

5.4. Results 96

5.4.1. Role of D1Rs in L-DOPA-induced abnormal striatal signaling in DRD mice 96

5.4.2. Role of D2Rs in L-DOPA-induced abnormal striatal signaling in DRD mice 98

5.4.3 Effects of dopamine receptor antagonist treatment on baseline behaviors 100

5.4.4. Role of D1Rs in L-DOPA-induced behavioral effects 102

5.4.5. Role of D2Rs in L-DOPA-induced behavioral effects 103

5.4.6. Role of D1R plus D2R-activation in L-DOPA-induced behavioral effects 105

5.5. Discussion 106

5.5.1. Role of D1Rs and D2Rs in the therapeutic effect of L-DOPA 107

5.5.2. Significance of D1R-antagonism in dystonia 108

5.5.3. Significance of D2R-antagonism in dystonia 108

5.5.4. Conclusions 109

Chapter 6: General discussion

6.1. Blunted D2R-mediated signaling in DRD mouse striatum 111

6.2. Supersensitive D1R-mediated signaling in DRD mouse striatum 114

6.3. Significance of L-DOPA-induced abnormal signaling in dystonia 115

6.4. Mechanism of L-DOPA-induced abnormal signaling in dystonia 119

6.5. Regional differences in postsynaptic intracellular signaling cascades in dystonia 121

6.6. Conclusions and future directions 123

Appendix: Assessment of the striatal proteome in a mouse model of DOPA-responsive dystonia

A.1. Abstract 124

A.2. Introduction 125

A.3. Methods 125

A.4. Results 130

A.5. Discussion 140

References

References 145

About this Dissertation

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School	Laney Graduate School
Department	Biological and Biomedical Sciences
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Biology, Neuroscience
Palavra-chave	Dopamine Dystonia
Committee Chair / Thesis Advisor	Hess, Ellen, Emory University
Committee Members	Pardue, Machelle, Georgia Tech Hepler, John, Emory University Weinshenker, David, Emory University Jinnah, H.A., Emory University Smith, Yoland, Emory University

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