Reduced vesicular storage of catecholamines in the pathogenesis of Parkinson's Disease Open Access

Taylor, Tonya (2010)

Permanent URL: https://etd.library.emory.edu/concern/etds/gq67jr59h?locale=en
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Abstract

Parkinson's disease (PD) is a neurodegenerative disorder which affects millions of people
worldwide. Although the cause of PD is unknown, it is thought to be due to a combination of
genetic and environmental factors; many therapies aim to restore dopamine (DA) within the
brain. Animal models, both chemical and transgenic, have been used to uncover potential
mechanisms of PD, but none have successfully recapitulated the symptoms of PD on an
appropriate timescale. Genetic perturbation of the vesicular monoamine transporter (VMAT2)
yielded the VMAT2-deficient mice, which display many of the motor and non-motor symptoms
associated with PD. With a 95% reduction in VMAT2 expression, VMAT2-deficient animals
have decreased novelty-induced locomotor activity and a shortened stride length at older ages.
VMAT2-deficient animals also displayed progressive deficits in olfactory discrimination without
changes in other sensory systems, which appear to be possibly correlated with previously seen
age-dependent reduction in dopamine transporter expression. Moreover, VMAT2-deficient mice
have a shorter latency to behavioral signs of sleep, delayed gastric emptying, anxiety-like
behaviors at younger ages, and a progressive depressive-like phenotype. Taken together, these
data suggest that reduced storage of monoamines may contribute to the development of many
features of Parkinson's disease. Progressive neurodegeneration in the substantia nigra (SNpc),
locus coeruleus (LC), and dorsal raphe (DR) has been observed, accompanied by α-synuclein
accumulation. Moreover, primary cultures from the SNpc, LC, and DR of postnatal mice
exposed to physiological concentrations of monoamines were found to undergo increased
oxidative stress via formation of H2O2 and other oxidative species; whereas, cultures from
wildtype mice were unaffected. These studies demonstrate that reduced vesicular storage may
play a role in the pathogenesis of parkinsonian symptoms and neurodegeneration. Perturbing the
catecholaminergic cytosolic environment may ultimately lead to the development of several
behavioral phenotypes and neuronal death. Using the VMAT2-deficient mice as a new model of
PD, could potentially lead to new therapeutic strategies beyond DA replacement therapy.

Table of Contents

CONTENTS

CHAPTER 1: INTRODUCTION AND BACKGROUND

Parkinson's Disease……………………………………………………………………...2

Dopamine and PD……………………………………………………………..…………3

Non-motor symptoms of PD…………………………………………………………..…5

Norepinephrine in PD………………………………………………………………..…..6

Serotonergic involvement in PD…………………………………………………..…….8

Hypotheses of PD……………………………………………………………………….10

Vesicular Monoamine Transporter……………………………………………………13

Introduction to Specific Aims………………………………………………...………..15

CHAPTER 2: BEHAVIORAL PHENOTYPING OF MOUSE MODELS OF PD

Abstract…………………………………………………………………………………18

Introduction……………………………………………………………………………..19

General Health………………………………………………………………………….20

Motor phenotype of PD……………………………………………………………..….22

Olfactory deficits in PD……………………………………………………………..….23

Sleep abnormalities in PD…………………………………………………………...…25

Gastrointestinal dysfunction in PD……………………………………………………27

Anxiety and depression…………………………………………………………...……30

Cognitive deficits in PD……………………………………………………………..….34

Conclusion………………………………………………………………………………35

CHAPTER 3: NON-MOTOR SYMPTOMS OF PD REVEALED IN MICE WITH REDUCED VESICULAR MONOAMINE STORAGE

Abstract………………………………………………………………………………….40

Introduction………………………………………………………..…………..………..41

Materials and Methods…………………………………………………………………42

Results…………………………………………………………………………………...48

Discussion……………………………………………………………………………….53

Acknowledgments………………………………………………………………………58

CHAPTER 4: PROGRESSIVE NORADRENERGIC DEGENERATION PRECEDES NIGRAL CELL LOSS IN A MOUSE MODEL OF PD

Abstract………………………………………………………………………………….76

Introduction……………………………………………………………………………..77

Materials and Methods……………………………………………………………..…..78

Results………………………………………………………………………………...…82

Discussion…………………………………………………………………………….....87

Acknowledgments……………………………………………………………………....90

CHAPTER 5: SEROTONERGIC DYSFUNCTION IN A MOUSE MODEL OF PARKINSON'S DISEASE

Abstract……………………………………………………………………..………….106

Introduction…………………………………………………………………..………..107

Materials and Methods………………………………………………………..………108

Results………………………………………………………………………………….111

Discussion……………………………………………………………………...………113

Acknowledgments……………………………………………………………………..116

CHAPTER 6: SUMMARY AND CONCLUSIONS

Introduction……………………………………………………………………..……..125

Genetic Manipulation of VMAT2…………………………………………………….127

The VMAT2 Hypomorph Mouse……………………………………………….……129

Perfected Perturbation: VMAT2-deficient mice………………………………...…..132

Conclusions………………………………………………………………………...…..136

Future Directions…………………………………………………………………...…137

Final Thoughts….……………………………………………………………………..139

APPENDIX A: THE EFFECT OF INCREASED CYTOSOLIC CATECHOLAMINES ON CELL TOXICITY

Summary……………………………………………………………………………….145

REFERENCES…………………………………………………………………………..……..153



LIST OF FIGURES

CHAPTER 3: NON-MOTOR SYMPTOMS OF PD REVEALED IN MICE WITH REDUCED VESICULAR MONOAMINE STORAGE

Figure 3-1. VMAT2-deficient mice display widespread reductions in monoamines at 12-15 months of age……………………………………………………..…………59

Figure 3-2. VMAT2-deficient animals display progressive deficits in olfactory discrimination…………………………………………………………..……...61

Figure 3-3. VMAT2-deficient mice display age-dependent deficits in non-social olfactory acuity……………………………………………………………………………63

Figure 3-4. VMAT2-deficient mice do not display deficits in general sensory behavioral tests……………………………………………………………………….……..65

Figure 3-5. VMAT2-deficient animals display normal circadian activity but a premature shortened latency to behavioral signs of sleep…………………………..……67

Figure 3-6. VMAT2-deficient animals have delayed gastric emptying……………...…..69

Figure 3-7. VMAT2-deficient animals display an anxiety-like and a progressive depressive-like phenotype…………………………………………….……….71

Figure 3-8. VMAT2-deficient mice have decreased retinal DA but normal vision…..…73

CHAPTER 4: PROGRESSIVE NORADRENERGIC DEGENERATION PRECEDES NIGRAL CELL LOSS IN A MOUSE MODEL OF PD

Figure 4-1. Immunohistochemical analysis of tyrosine hydroxylase in substantia nigra of aged VMAT2-deficient mice……………………………………..……………91

Figure 4-2. Immunohistochemical analysis of tyrosine hydroxylase in locus coeruleus of aged VMAT2-deficient mice……………………………………..……………93

Figure 4-3. VMAT2-deficient mice show evidence of catecholaminergic degeneration at 24 months of age……………………………………………………..…………95

Figure 4-4 . TH and hematoxylin cell counts in the SNpc and LC of 18, 24, and 30 month wildtype andVMAT2-deficient mice………………………………………….97

Figure 4-5 . Evidence of dopaminergic-induced oxidative stress in VMAT2-deficient SNpc primary neurons…………………………………………………..…….97

Figure 4-6 . Evidence of noradrenergic-induced oxidative stress in VMAT2-deficient LC primary neurons…………………………………………………………...…101

Figure 4-7. Immunohistochemical analysis of α -synuclein in substantia nigra and locus coeruleus of aged VMAT2-deficient mice…………………………..……….103

CHAPTER 5: SEROTONERGIC DYSFUNCTION IN A MOUSE MODEL OF PARKINSON'S DISEASE

Figure 5-1. Evidence of serotonergic-induced oxidative stress in VMAT2-deficient DR primary neurons………………………………………………...……………116

Figure 5-2. VMAT2-deficient mice show evidence of monoaminergic degeneration at 24 months of age…………………………………………………….……………118

Figure 5-3. VMAT2-deficient mice show evidence of monoaminergic degeneration at 24 months of age…………………………………………………….……………120

Figure 5-4. VMAT2-deficient animals display a progressive depressive-like phenotype that is sensitive to fluoxetine…………………………………………………122

CHAPTER 6: SUMMARY AND CONCLUSIONS

Figure 6-1. VMAT2-deficient animals display impaired stride length at older ages….141

Figure 6-2. Timeline of parkinsonian features observed in the VMAT2-deficient mice……………………………………………………………………………143

APPENDIX A: THE EFFECT OF INCREASED CYTOSOLIC CATECHOLAMINES ON CELL TOXICITY

Figure A-1. Manipulation of DAT and VMAT2 do not enhance intracellular oxidative damage after exposure to exogenous catecholamines in vitro………..…….147

Figure A-2. Toxicity of catecholamines and their precursors to SN4741 cells…………149

Figure A-3. SN4741 cells exposed to various concentrations of catecholamines do not exhibit increased oxidative stress as measured by DCF……………………151

LIST OF TABLES

CHAPTER 2: BEHAVIORAL PHENOTYPING OF MOUSE MODELS OF PD

Table 2-1: L-DOPA responsive motor phenotypes in parkinsonian mouse model…….36

Table 2-2: Behavioral phenotypes in mouse models of Parkinson's disease…………..37

Table 2-3: Summary of parkinsonian behavioral analyses……………………...………38

CHAPTER 6: SUMMARY AND CONCLUSIONS

Table 6-1: Summary of L-DOPA responsive parkinsonian symptoms………..………140

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