The Role of β-Catenin in Long-Term Memory Formation Open Access

Maguschak, Kimberly Ann (2010)

Permanent URL: https://etd.library.emory.edu/concern/etds/ks65hc57g?locale=en
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Abstract

Abstract
The Role of β-Catenin in Long-Term Memory Formation
By Kimberly A. Maguschak
The ability of connections between neurons to change as a function of learning is commonly referred to as synaptic plasticity. Such changes in synaptic morphology and organization are thought to underpin long term memory formation. Among the most studied structural changes has been the elaboration of new synaptic architecture following a learning event. This process has been postulated by many investigators to be a physiologically-relevant means of synaptic potentiation. The processes governing dendritic morphogenesis are many and varied, but much work has focused on the role of β-catenin in allowing the remodeling of synapses in an activity-dependent way. β-Catenin is expressed broadly in the adult mammalian brain and plays a role in both cadherin-mediated cell-cell adhesion and Wnt signaling. Although β-catenin has been implicated in neuronal synapse regulation and plasticity, all of this work has been done in vitro, and not in intact animals. Here we have examined the role of β-catenin in the adult mouse brain and its role in amygdala- and hippocampal-dependent learning and memory. We found that β-catenin is highly expressed in these brain regions and is dynamically regulated at the post-translational level with fear learning. Such alterations correlated with a change in the association of β-catenin with cadherin. Genetically, the role of β-catenin was confirmed with site-specific deletions of loxP-flanked Ctnnb1 (encoding β-catenin) in the amygdala or dorsal hippocampus. In both cases, the manipulation affected the consolidation, but not acquisition, of memory. Notably, Ctnnb1 deletion did not affect a number of other behaviors, including locomotor or anxiety-related behavior. Furthermore, we found that Wnt signaling may play a role in mediating β-catenin dependent memory formation. Interfering with Wnt signaling prevented consolidation and altered the transient destabilization and re-stabilization of β-catenin-cadherin interactions during memory formation. Therefore, our findings suggest a general role for β-catenin in the synaptic remodeling and stabilization underlying long-term memory in adults.

Table of Contents


TABLE OF CONTENTS

CHAPTER 1...1

GENERAL INTRODUCTION...1

INTRODUCTION...2

Molecular mechanisms of synaptic plasticity...2
Morphological changes associated with synaptic plasticity...3
Cell adhesion molecules and synaptic plasticity...4

β-CATENIN STRUCTURE AND FUNCTION...5

Structure of β-catenin...5
Functional roles of β-catenin...6
Posttranslational modifications regulate β-catenin function...9
Role of β-catenin in presynaptic structure and function...11
Role of β-catenin in postsynaptic structure and function...15
Activity-dependent regulation of β-catenin...17
β-catenin and memory related pathology...19
β-catenin and Learning and Memory in the Adult Brain...21

CHAPTER 2...22
Β-CATENIN IS REQUIRED FOR MEMORY CONSOLIDATION...22

ABSTRACT...23
INTRODUCTION...23
RESULTS...25

β-catenin mRNA expression in the adult mouse brain...25
β-catenin mRNA increases in the amygdala following fear conditioning...27
β-catenin is post-translationally regulated following fear conditioning...29
Increasing β-catenin functional stability results in an enhancement in learning...35
Ctnnb1 deletion in the BLA does not alter baseline measures...41

β-catenin is required for fear-memory consolidation...47
β-catenin is not required for expression of fear memory...51

DISCUSSION...54
METHODS...59

Animals...59
Immunoblotting...59
Immunoprecipitation...61
In situ hybridization...61
Lentiviral constructs and virus production...62
Surgery and injection of virus...62
Behavior...63

Elevated plus maze...63
Open field...63
Fear Conditioning Apparatus...63
Fear conditioning...64
Freezing apparatus...65

Data analysis...66

CHAPTER 3...67
WNT SIGNALING IN AMYGDALA-DEPENDENT LEARNING AND MEMORY...67

ABSTRACT...68
INTRODUCTION...68
RESULTS...71

Decreasing Wnt mediated signaling impairs memory formation...71
Dkk-1 prevents learning-dependent re-stabilization between β-catenin and cadherin...76
Genes in Wnt-mediated signaling are altered during memory formation...78
Wnt1 gene expression changes with learning...82
Increasing Wnt1 impairs fear memory formation...86

Wnt1 prevents learning-dependent disassociation between β-catenin and cadherin...90

DISCUSSION...93
METHODS...97

Animals...97
Stereotaxic surgery and infusion of peptides...97
Immunoprecipitation and Immunoblotting...98
Immunohistochemistry...99
Behavioral studies...99

Open Field Behavior...99
Fear Conditioning Apparatus...100
Fear Conditioning...100

RNA extraction and cDNA synthesis...101
Real-time PCR...101
In situ hybridization...102
Data analysis...103

CHAPTER 4...104
Β-CATENIN IS REQUIRED FOR HIPPOCAMPAL-DEPENDENT LEARNING IN ADULT MICE...104

ABSTRACT...105
INTRODUCTION...105
RESULTS...107
β-catenin deletion in the hippocampus does not alter baseline activity or anxiety measures...107

β-catenin in the hippocampus is required for context-dependent, but not cue-dependent fear memory consolidation...111
β-Catenin deletion impairs object recognition memory...113
β-Catenin deletion impairs consolidation, but not acquisition, of spatial memory...115

DISCUSSION...117
METHODS...120

Animals...120
In situ hybridization...121
Lentiviral vectors and virus infection...121
Behavioral Studies...122

Open field...122
Elevated plus maze...122
Context-dependent fear conditioning...122
Cue-dependent fear conditioning...123
Novel object recognition...123
Morris Water Maze...125

Data analysis...126

CHAPTER 5...127
CONCLUDING REMARKS...127

SUMMARY OF FINDINGS...128

Dynamic Regulation of Cadherin/β-catenin Interaction...135
Subcellular Distribution of β-Catenin...138
The Stabilization and De-Stabilization of Memory...139
Implications for Alzheimer's Disease...141
Role of β-catenin in development to disease...143

REFERENCES...144

TABLE OF FIGURES

FIGURE 2.3 - β-CATENIN EXPRESSION IN MICE...30
FIGURE 2.3- PHOSPHORYLATION STATE OF GSK-3β IS ALTERED AFTER FEAR CONDITIONING...31
FIGURE 2.4 - PHOSPHORYLATION STATE OF β-CATENIN IS ALTERED AFTER FEAR CONDITIONING...32
FIGURE 2.5 - COIMMUNOPRECIPITATION OF CADHERIN AND β-CATENIN...34
FIGURE 2.6 - LICL DECREASES GSK-3β-MEDIATED β-CATENIN PHOSPHORYLATION IN THE AMYGDALA...37
FIGURE 2.7 - LICL TREATMENT ENHANCES LEARNING...38
FIGURE 2.8 - LICL TREATMENT ENHANCES LEARNING...40
FIGURE 2.9 - REGION-SPECIFIC DELETION OF β-CATENIN IN THE ADULT BRAIN...42
FIGURE 2.10 - CRESYL VIOLET (NISSL) STAINING OF INFECTED BASOLATERAL AMYGDALA...44
FIGURE 2.11 - AMYGDALA-SPECIFIC β-CATENIN DELETIONS DO NOT AFFECT BASELINE ANXIETY OR ACTIVITY MEASURES...46
FIGURE 2.12 - AMYGDALA-SPECIFIC CTNNB1 DELETION DOES NOT PREVENT THE EXPRESSION, OF CONDITIONED FEAR...48
FIGURE 2.13 - AMYGDALA-SPECIFIC CTNNB1 DELETION PREVENTS THE CONSOLIDATION OF CONDITIONED FEAR...50
FIGURE 2.14 - AMYGDALA-SPECIFIC CTNNB1 DELETION DOES NOT AFFECT THE EXPRESSION OF CONDITIONED FEAR...53
FIGURE 3.1 - EXPRESSION OF DKK-1 IN THE AMYGDALA...72
FIGURE 3.2 - THE EFFECT OF DKK-1 IN THE AMYGDALA ON BASELINE EMOTIONAL BEHAVIOR...73
FIGURE 3.3 - THE EFFECT OF DKK-1 IN THE AMYGDALA ON FEAR LEARNING AND MEMORY...74
FIGURE 3.4 - THE EFFECT OF DKK-1 IN THE AMYGDALA ON FEAR LEARNING AND MEMORY WHEN GIVEN PRIOR TO ACQUISITION...75
FIGURE 3.5 - THE EFFECT OF DKK-1 IN THE AMYGDALA ON FEAR LEARNING AND MEMORY WHEN GIVEN IMMEDIATELY POST-ACQUISITION...76
FIGURE 3.6 - THE EFFECT OF FEAR CONDITIONING AND DKK-1 ADMINISTRATION ON THE INTERACTION BETWEEN β-CATENIN AND CADHERIN...78
FIGURE 3.7 - TEMPORAL CHANGES IN WNT GENE EXPRESSION AFTER FEAR CONDITIONING...81
FIGURE 3.8 - WNT1 MRNA EXPRESSION IN THE AMYGDALA...83
FIGURE 3.9 - WNT1 MRNA EXPRESSION IN THE AMYGDALA...85
FIGURE 3.10 - WNT1 EXPRESSION IN THE AMYGDALA...87
FIGURE 3.11 - THE EFFECT OF WNT1 IN THE AMYGDALA ON BASELINE ANXIETY...88
FIGURE 3.12 - THE EFFECT OF WNT1 IN THE AMYGDALA ON FEAR LEARNING AND MEMORY...89
FIGURE 3.13 - THE EFFECT OF WNT1 IN THE AMYGDALA ON FEAR LEARNING AND MEMORY...90
FIGURE 3.14 - THE EFFECT OF FEAR CONDITIONING AND WNT1 ADMINISTRATION ON THE INTERACTION BETWEEN β-CATENIN AND CADHERIN...92
FIGURE 4.1 - HIPPOCAMPAL-SPECIFIC β-CATENIN DELETIONS DO NOT AFFECT BASELINE ACTIVITY OR ANXIETY MEASURES AS MEASURED BY OPEN FIELD...109
FIGURE 4.2 - HIPPOCAMPAL-SPECIFIC β-CATENIN DELETIONS DO NOT AFFECT BASELINE ACTIVITY AS MEASURED BY ELEVATED PLUS MAZE...110
FIGURE 4.3 - HIPPOCAMPAL-SPECIFIC β-CATENIN DELETION PREVENTS THE CONSOLIDATION, BUT NOT ACQUISITION OF CONTEXT-DEPENDENT FEAR MEMORIES...112
FIGURE 4.4 - HIPPOCAMPAL-SPECIFIC β-CATENIN DELETION DOES NOT AFFECT THE ACQUISITION OF CONSOLIDATION OF CUE-DEPENDENT FEAR CONDITIONING...113
FIGURE 4.5 - HIPPOCAMPAL-SPECIFIC β-CATENIN DELETION IMPAIRS OBJECT MEMORY...114
FIGURE 4.6 - HIPPOCAMPAL-SPECIFIC β-CATENIN DELETION DOES NOT AFFECT THE ACQUISITION OF SPATIAL MEMORY...115
FIGURE 4.7 - HIPPOCAMPAL-SPECIFIC β-CATENIN DELETION DOES IMPAIR CONSOLIDATED SPATIAL MEMORIES...117
FIGURE 5.1 - SCHEMATIC REPRESENTATION OF THE ROLE OF β-CATENIN IN PRODUCING THE LABILE AND STABLE PHASES OF MEMORY FORMATION...130
FIGURE 5.2 - SCHEMATIC REPRESENTATION OF THE EFFECT OF A WNT ANTAGONIST AND WNT AGONIST ON THE LABILE AND STABLE PHASES OF MEMORY FORMATION...133
FIGURE 5.3 - SCHEMATIC REPRESENTATION SHOWING A POSSIBLE INTERACTION BETWEEN BDNF SIGNALING AND NMDAR ACTIVATION...137


TABLE OF TABLES
TABLE 1.1 EFFECTS OF LOSS OF FUNCTION OR OVEREXPRESSION OF β-CATENIN ON PRESYNAPTIC AND POSTSYNAPTIC FUNCTION...12
TABLE 3.1 - TEMPORAL CHANGES, EXPRESSED AS FOLD REGULATION, IN THE EXPRESSION OF 84 GENES RELATED TO WNT-MEDIATED SIGNALING FOLLOWING FEAR CONDITIONING...80

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