Striatal melanocortin-4 receptor control of action flexibility Open Access

Heaton, Elizabeth (Fall 2023)

Permanent URL: https://etd.library.emory.edu/concern/etds/bc386k73k?locale=en
Published

Abstract

More than half of all individuals in treatment for substance use disorder (SUD) will relapse. Inflexibility in selecting between familiar, habitual behaviors that have been rewarded in the past (drug seeking) and novel strategies that might be more advantageous (rehabilitation) may be a factor that preserves SUD. The dorsomedial striatum (DMS) is a brain region that receives and integrates glutamatergic input from cortical and subcortical regions required for goal-directed action selection. However, the factors in the DMS responsible for coordinating this incoming information remain incompletely understood. This dissertation begins by describing the extra-hypothalamic functions of the melanocortin-4 receptor (MC4R), a receptor that is well-positioned in the DMS to control flexible, goal-directed action. Next, I report that MC4R in the DMS appears to propel familiar reward-seeking behavior (habit), even when it is not fruitful, and moderating MC4R presence improves the capacity for goal-directed behavior. I then demonstrate that this process requires inputs from the orbitofrontal cortex, a brain region canonically associated with response strategy switching. Then, I further investigate how striatal melanocortin systems propel familiar behaviors, particularly via interaction with the central nucleus of the amygdala (CeA). I demonstrate that MC4R-expressing cells in the DMS are 1) predominantly expressed on dopamine D1-type receptor-expressing medium spiny neurons and 2) are necessary and sufficient for controlling the capacity of mice to arbitrate between actions and habits. I next use site-selective gene silencing and pharmacological techniques to reveal that MC4R presence suppresses goal seeking. I also find that MC4R-expressing neurons are functionally integrated into an amygdalo-striatal circuit that suppresses action flexibility in favor of routinized behaviors. Additionally, I use publicly available spatial transcriptomics datasets to reveal differences in the gene transcript correlates of Mc4r across the striatum, with considerable co-variation in dorsal structures. Guided by these results, I lastly discovered that MC4R function in the dorsolateral striatum complements that in the DMS, here suppressing habitual behavior. Together, these findings provide insight into the molecular and circuit-level mechanisms by which MC4R in the DMS propels habitual behavior. This dissertation thus illuminates mechanistic factors that support the development of automatized routines when flexible decision making is no longer adaptive, which may provide insight into therapeutic targets for neuropsychiatric disorders in which decision making is impaired. 

Table of Contents

CHAPTER 1: Extra-hypothalamic functions of the melanocortin-4 receptor...... 1

1.1      Context and author's contribution                                                                2

1.2      Abstract                                                                                                        2

1.3      Introduction                                                                                                 2

1.3.1      Overview of the central melanocortin system                                    3

1.4      Functions of MC4R outside of the hypothalamus                                       5

1.4.1      Cortex                                                                                                  5

1.4.2      Hippocampus                                                                                      7

1.4.3      Amygdala                                                                                          11

1.4.4      Striatum                                                                                             12

1.5      Discussion                                                                                                  14

1.6      Framework and overview of the dissertation                                            15

CHAPTER 2: Selective breeding reveals control of reward-related action selection by the melanocortin-4 receptor................................................................. 21

2.1      Context, author's contribution, and acknowledgement of reproduction    22

2.2      Abstract                                                                                                      22

2.3      Introduction                                                                                               23

2.4      Results                                                                                                       25

2.4.1      Individual differences in reward-related response strategies in mice 25

2.4.2      Individual differences in instrumental response strategies are associated with striatal protein composition                                     27

2.4.3      MC4R control of action strategies                                                    28

2.4.4      MC4R control of action strategies via the OFC                                31

2.5      Discussion                                                                                                  33

2.6      Methods                                                                                                     38

2.6.1      Subjects                                                                                             38

2.6.2      Ages of mice at testing                                                                      39

2.6.3      Test of action strategies                                                                    39

2.6.4      Breeding strategy                                                                              40

2.6.5      Reinforcer devaluation                                                                      40

2.6.6      Intracranial surgery and viral vectors                                               42

2.6.7      CNO administration and timing in DREADDs experiments            43

2.6.8      Assessments of food intake                                                               43

2.6.9      Histology                                                                                           43

2.6.10   Immunoblotting                                                                                 44

2.6.11   Dendritic spine imaging and reconstruction                                     45

2.6.12   Statistics and reproducibility                                                             46

2.7      Funding                                                                                                      48

2.8      Acknowledgements                                                                                   48

CHAPTER 3: Striatal melanocortin systems propel familiar actions via interaction with the central nucleus of the amygdala............................................. 57

3.1      Context and author's contribution                                                              58

3.2      Abstract                                                                                                      58

3.3      Introduction                                                                                               59

3.4      Results                                                                                                       60

3.4.1      MC4R-expressing cells in the DMS bidirectionally control action flexibility                                                                                           60

3.4.2      Reducing Mc4r in the DMS prompts flexible behavior                    62

3.4.3      MC4R in amygdalo-DMS circuits controls action selection            64

3.4.4      Spatial transcriptomics reveals a diversity of Mc4r expression co-variates across the striatum                                                               66

3.4.5      MC4R acts as a molecular brake on DLS function in prompting habit-like behavior                                                                            68

3.5      Discussion                                                                                                  69

3.5.1      Striatal MC4R controls action flexibility                                          70

3.5.2      MC4R and amygdalo-striatal interactions suppress action flexibility 72

3.5.3      Conclusions                                                                                       74

3.6      Methods                                                                                                     75

3.6.1      Animals                                                                                             75

3.6.2      RNAScope                                                                                        76

3.6.3      Surgery and viral vectors                                                                  76

3.6.4      Instrumental response training                                                          79

3.6.5      Test for response flexibility                                                              79

3.6.6      CNO administration                                                                          80

3.6.7      Cocaine administration                                                                     81

3.6.8      Setmelanotide administration                                                           81

3.6.9      Locomotion and ad libitum feeding                                                  81

3.6.10   Histology                                                                                           82

3.6.11   Immunofluorescence imaging                                                           82

3.6.12   Synaptoneurosome preparation                                                         82

3.6.13   Western blotting                                                                                83

3.6.14   Trans-synaptic retrograde tracing                                                     84

3.6.15   Statistical analysis                                                                             84

3.7      Funding                                                                                                      85

3.8      Acknowledgements                                                                                   86

CHAPTER 4: Concluding remarks....................................................................... 113

4.1      Abstract                                                                                                    114

4.2      Etiological motivations of decision-making behavior                             114

4.3      Physiological motivation: Hunger                                                           115

4.3.1      The cycle of hunger                                                                        115

4.3.2      Hunger and behavioral flexibility                                                   117

4.4      Security motivation: Stress                                                                      118

4.4.1      The hypothalamic-pituitary-adrenal axis                                        119

4.4.2      Stress and behavioral flexibility                                                      119

4.5      Social motivation: Pair bonding                                                              121

4.5.1      The neurobiology of pair bonding                                                  121

4.5.2      Pair bonding and behavioral flexibility                                          122

4.6      Conclusions                                                                                             123

APPENDIX A: Publications to which the author has contributed..................... 126

REFERENCES......................................................................................................... 127

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