A new mouse model of L-DOPA-responsive dystonia Open Access

Rose, Samuel (2015)

Permanent URL: https://etd.library.emory.edu/concern/etds/js956g52f?locale=en


Dystonia is a neurological movement disorder characterized by sustained or intermittent muscle contractions causing abnormal, often repetitive, movements or postures. Abnormal dopamine neurotransmission is associated with many different dystonic disorders. For instance, mutations in genes critical for the synthesis of dopamine, including GTP cyclohydrolase 1 and tyrosine hydroxylase cause l-DOPA-responsive dystonia. Despite evidence that implicates abnormal dopamine neurotransmission in dystonia, the precise nature of the dopaminergic defects that result in dystonia is not known. To better understand these defects, we generated a knockin mouse model of l-DOPA-responsive dystonia that recapitulates the human p.381Q>K TH mutation (c.1141C>A). Mice homozygous for this mutation (DRD mice) had reduced TH activity throughout the brain and striatal dopamine concentration that were ~1% of normal. Although the gross anatomy of the nigrostriatal dopaminergic neurons was normal in DRD mice, the microstructural target of corticostriatal synapses was affected; corticostriatal input in DRD mice showed a shift away from synapses on dendritic spines towards dendrites themselves. DRD mice displayed the core behavioral features of the human disorder, including dystonia that worsened throughout the course of the active phase, and improvement in the dystonia in response to both l-DOPA and trihexyphenidyl. Administration of D1- or D2-type dopamine receptor agonists reduced the dystonic movements while administration of D1- or D2-type dopamine receptor antagonists worsened the dystonia, suggesting that both receptors mediate the dystonic movements. Further, D1-dopamine receptors were supersensitive; adenylate cyclase activity, locomotor activity and stereotypy were exaggerated in DRD mice in response to the D1-dopamine receptor agonist SKF 81297. D2-dopamine receptors responses were blunted or altered in DRD mice with an increase in adenylate cyclase activity and blunted behavioral responses after challenge with the D2-dopamine receptor agonist quinpirole. Together, the findings here implicate developmental dopamine loss within a specific range in the development of dystonia. Further, they implicate maladaptive changes to dopamine receptor responses as important factors for this disorder.

Table of Contents

Table of Contents

Chapter 1: Introduction

Dystonia____________________________________________________________ 1

Neuroanatomical substrates of dystonia___________________________________ 4

Basal ganglia function and dysfunction in dystonia___________________________ 5

Cerebellar dysfunction in dystonia________________________________________ 10

Dysfunctional DA neurotransmission as a common molecular pathway in dystonia__11

Tyrosine hydroxylase__________________________________________________15

DA receptors________________________________________________________ 16

DA and the development of the basal ganglia_______________________________19

Evidence for DA dysfunction in dystonia___________________________________20

DRD as a disorder prototypical of dystonia arising from DA dysfunction___________23

DRD: Clinical features__________________________________________________24

Mutations in tetrahydrobiopterin synthesizing enzymes________________________25

Mutations in TH_______________________________________________________25

Animal models closely resembling DRD____________________________________27

Neonatal 6-OHDA-treated rats____________________________________________27

MPTP-treated primates__________________________________________________29

Mice modeling impaired BH4 synthesis_____________________________________29

Mutant TH lines________________________________________________________31

Summary and guiding questions of thesis work_______________________________31

Chapter 2: In vivo molecular, neurochemical, and anatomical effects of the p.382Q>K mutation in TH





PCR conformation of DRD knockin allele______________________________42

Generating homozygous DRD mice__________________________________46

DRD mice exhibit reduced TH activity and brain catecholamines___________50

Normal gross anatomy of midbrain and striatum in DRD mice______________54

Microstructural changes to corticostriatal and thalamostriatal connectivity____56


Chapter 3: Behavioral phenotype of the DRD mice: special emphasis on diurnal fluctuations and drug responses.





Dystonic movements in DRD mice___________________________________70

Neurochemistry correlates with diurnal fluctuations______________________75

Behavioral responses to indirect catecholamine agonists_________________79


Chapter 4: DA receptor-mediated responses in DRD mice





DA receptors mediate dystonic movements___________________________100

Abnormal DA receptor responses in DRD mice________________________102

Ex vivo DA receptor expression and second messenger responses________107


Chapter 5: General Discussion

DRD mice: contribution to the field________________________________________115

Dystonia vs. parkinsonism______________________________________________117

DA receptor mechanisms specific to dystonia_______________________________120

Future directions______________________________________________________121


List of Figures

1. Structures of the basal ganglia and related circuitry in mouse___________________6

2. Schematic of presynaptic DA terminal____________________________________14

3. Conformation of knockin and resolution of Neo_____________________________44

4. RNA expression of DRD allele__________________________________________45

5. Pre and postnatal death of DRD mice on a C57BL6/J background______________47

6. Viability of DRD mice on F2D2B6 background______________________________49

7. In vivo TH activity in DRD mice_________________________________________52

Table 1. Regional monoamine concentration_________________________________53

8. Gross anatomy of nigrostriatal pathway in DRD mice________________________55

9. Glutamatergic cortico- and thalamo-striatal immunostraining___________________58

10. Glutamatergic cortico- and thalamo-striatal terminals in DRD mice_____________59

11. Dystonic movements in DRD mice______________________________________72

12. Time of day-dependent differences in motor behavior_______________________73

13. Diurnal fluctuations in DA metabolism___________________________________77

14. Amphetamine response in DRD mice____________________________________80

15. l-DOPA response in DRD mice________________________________________82

Table 2. Effect of l-DOPA and l-DOPS on monoamines________________________83

16. Regional specificity of l-DOPA effects___________________________________85

17. l-DOPS response in DRD mice________________________________________85

18. THP response in DRD mice___________________________________________86

19. Abnormal movement response to DA receptor-specific agonists and antagonists_101

20. Behavioral sensitivity to D1DAR agonism in DRD mice_____________________104

21. Behavioral responses to D2DAR agonism in DRD mice____________________105

22. Cataleptic response to DA receptor antagonists in DRD mice________________106

Table 3. qRT-PCR for DA receptors_______________________________________108

Table 4. DA receptor binding____________________________________________108

23. Ex vivo striatal adenylate cyclase activity________________________________109

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