UUnniivveerrssiittyy ooff PPeennnnssyyllvvaanniiaa SScchhoollaarrllyyCCoommmmoonnss Publicly Accessible Penn Dissertations Summer 2011 RRoollee OOff HHiippppooccaammppaall NNeeuurrooggeenneessiiss IInn TThhee EEttiioollooggyy AAnndd TTrreeaattmmeenntt OOff MMoooodd && AAnnxxiieettyy DDiissoorrddeerrss Jennifer L. Onksen [email protected] Follow this and additional works at: https://repository.upenn.edu/edissertations Part of the Behavioral Neurobiology Commons, Behavior and Behavior Mechanisms Commons, Mental Disorders Commons, and the Pharmacology Commons RReeccoommmmeennddeedd CCiittaattiioonn Onksen, Jennifer L., "Role Of Hippocampal Neurogenesis In The Etiology And Treatment Of Mood & Anxiety Disorders" (2011). Publicly Accessible Penn Dissertations. 387. https://repository.upenn.edu/edissertations/387 This paper is posted at ScholarlyCommons. https://repository.upenn.edu/edissertations/387 For more information, please contact [email protected]. RRoollee OOff HHiippppooccaammppaall NNeeuurrooggeenneessiiss IInn TThhee EEttiioollooggyy AAnndd TTrreeaattmmeenntt OOff MMoooodd && AAnnxxiieettyy DDiissoorrddeerrss AAbbssttrraacctt Mood disorders are associated with deficits in hippocampal size and function. Neurogenesis in the dentate gyrus of the hippocampus has been implicated in the behavioral efficacy of some antidepressants. However, the precise function of newborn hippocampal neurons in the etiology and treatment of mood disorders remains unclear. Anxiety, which is highly comorbid with mood disorders, also appears to be influenced by neurogenesis. The intricacies of this influence are poorly understood. Here, the role of neurogenesis in the pathophysiology and treatment of mood and anxiety disorders is investigated in a transgenic mouse exhibiting partial suppression of hippocampal neurogenesis. This reduction is induced by hippocampal microinjection of a Cre-expressing adeno-associated virus to delete ATR, a cell cycle checkpoint kinase, from the hippocampus of adult mice. Subsequent to hippocampal ATR deletion, behavior is examined in mouse models of anxiety, depressive-like behavior, antidepressant efficacy, and susceptibility to stress. In addition, the necessity of heightened neurogenesis in exercise- induced anxiety is examined. ATR deletion resulted in an anhedonic phenotype in a sucrose drinking task but did not increase the likelihood of developing behavioral despair in the learned helplessness paradigm. Furthermore, mice lacking hippocampal ATR exhibited reduced anxiety in a number of behavioral tests. In the novelty-induced hypophagia task, the ability of chronic antidepressants to alleviate hyponeophagia was absent in ATR-deleted mice, and this effect was linked to an attenuation of the ability of chronic antidepressants to stimulate dendritic spine density. Finally, ATR deletion attenuated the neurogenic effect of voluntary wheel running, and concurrently blocked some of the anxiogenic effects of wheel running. In conclusion, neurogenesis appears to be an essential regulator of behaviors indicative of mood and anxiety disorders. These observations contribute to the body of work investigating the etiology and treatment of mood disorders for the purpose of guiding pharmaceutical drug discovery towards the eventual identification of antidepressant compounds with greater efficacy and fewer side effects. DDeeggrreeee TTyyppee Dissertation DDeeggrreeee NNaammee Doctor of Philosophy (PhD) GGrraadduuaattee GGrroouupp Pharmacology FFiirrsstt AAddvviissoorr Dr. Julie A. Blendy KKeeyywwoorrddss neurogenesis, antidepressants, depression, anxiety, stress, exercise SSuubbjjeecctt CCaatteeggoorriieess Behavioral Neurobiology | Behavior and Behavior Mechanisms | Mental Disorders | Pharmacology This dissertation is available at ScholarlyCommons: https://repository.upenn.edu/edissertations/387 ROLE OF HIPPOCAMPAL NEUROGENESIS IN THE ETIOLOGY AND TREATMENT OF MOOD & ANXIETY DISORDERS Jennifer L Onksen A DISSERTATION in Pharmacology Presented to the Faculties of the University of Pennsylvania in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy 2011 Supervisor of Dissertation Signature ________________________________ Julie A Blendy, Ph.D Professor of Pharmacology Graduate Group Chairperson Signature_________________________________ Vladimir Muzykantov, MD, Ph.D Professor of Pharmacology Dissertation Committee Steven Thomas, MD, Ph.D, Associate Professor of Pharmacology Eric Brown, Ph.D, Assistant Professor of Cancer Biology Irwin Lucki, Ph.D, Professor of Pharmacology Brenda Porter, MD, Ph.D, Assistant Professor of Pediatrics and Neurology ABSTRACT ROLE OF HIPPOCAMPAL NEUROGENESIS IN THE ETIOLOGY AND TREATMENT OF MOOD & ANXIETY DISORDERS Jennifer L Onksen Advisor: Julie A Blendy, Ph.D Mood disorders are associated with deficits in hippocampal size and function. Neurogenesis in the dentate gyrus of the hippocampus has been implicated in the behavioral efficacy of some antidepressants. However, the precise function of newborn hippocampal neurons in the etiology and treatment of mood disorders remains unclear. Anxiety, which is highly comorbid with mood disorders, also appears to be influenced by neurogenesis. The intricacies of this influence are poorly understood. Here, the role of neurogenesis in the pathophysiology and treatment of mood and anxiety disorders is investigated in a transgenic mouse exhibiting partial suppression of hippocampal neurogenesis. This reduction is induced by hippocampal microinjection of a Cre- expressing adeno-associated virus to delete ATR, a cell cycle checkpoint kinase, from the hippocampus of adult mice. Subsequent to hippocampal ATR deletion, behavior is examined in mouse models of anxiety, depressive-like behavior, antidepressant efficacy, and susceptibility to stress. In addition, the necessity of heightened neurogenesis in exercise-induced anxiety is examined. ATR deletion resulted in an anhedonic phenotype in a sucrose drinking task but did not increase the likelihood of developing behavioral despair in the learned helplessness paradigm. Furthermore, mice lacking hippocampal ATR exhibited reduced anxiety in a number of behavioral tests. In the novelty-induced hypophagia task, the ability of chronic antidepressants to alleviate hyponeophagia was absent in ATR-deleted mice, and this effect was linked to an attenuation of the ability of ii chronic antidepressants to stimulate dendritic spine density. Finally, ATR deletion attenuated the neurogenic effect of voluntary wheel running, and concurrently blocked some of the anxiogenic effects of wheel running. In conclusion, neurogenesis appears to be an essential regulator of behaviors indicative of mood and anxiety disorders. These observations contribute to the body of work investigating the etiology and treatment of mood disorders for the purpose of guiding pharmaceutical drug discovery towards the eventual identification of antidepressant compounds with greater efficacy and fewer side effects. iii Table of Contents I. Introduction ..................................................................................................1 A. Mood and anxiety disorders.................................................................................. 1 i. Mood disorders ii. Anxiety disorders B. Rodent models..................................................................................................... 3 i. Modeling depression ii. Modeling anxiety C. Hippocampal neurogenesis ................................................................................... 8 i. Characterization of hippocampal neurogenesis ii. Neurogenesis in the etiology of mood and anxiety disorders iii. Neurogenesis in the treatment of mood and anxiety disorders D. Manipulating neurogenesis in rodents...................................................................12 i. Cytotoxic chemicals ii. X-ray irradiation iii. Genetic manipulation E. Crelox-conditional ATR knockout..........................................................................15 i. ATR, a cell cycle checkpoint kinase ii. ATR deletion in the adult mouse iii. Targeting ATR in the hippocampus F. References..........................................................................................................17 II. Selective deletion of ATR reduces neurogenesis and alters responses in rodent models of behavioral affect.................................................................29 A. Abstract..............................................................................................................30 B. Introduction........................................................................................................31 C. Materials and Methods.........................................................................................33 D. Results...............................................................................................................42 E. Figures...............................................................................................................47 F. Discussion...........................................................................................................58 G. References.........................................................................................................64 iv III. Low neurogenesis does not increase susceptibility to stress-induced or corticosterone-induced depressive behaviors in mice....................................96 A. Abstract..............................................................................................................70 B. Introduction........................................................................................................71 C. Materials and Methods.........................................................................................73 D. Results...............................................................................................................80 E. Figures...............................................................................................................83 F. Discussion...........................................................................................................87 G. References.........................................................................................................91 IV. Mice deficient in hippocampal neurogenesis do not develop exercise- induced anxiety...............................................................................................96 A. Abstract..............................................................................................................97 B. Introduction........................................................................................................98 C. Materials and Methods.........................................................................................99 D. Results.............................................................................................................105 E. Figures.............................................................................................................110 F. Discussion.........................................................................................................117 G. References.......................................................................................................122 V. Conclusions and future directions ............................................................126 A. Discussion........................................................................................................126 B. References .......................................................................................................132 v List of Illustrations Chapter II Figure 1. Hippocampal microinjection of AAV.Cre deletes ATR....................................47 Figure 2. Hippocampal ATR deletion inhibits hippocampal neurogenesis......................48 Figure 3. Hippocampal ATR deletion is associated with reduced anxiety in the elevated zero maze (EZM).........................................................................50 Figure 4. ATR deletion is associated with reduced anxiety..........................................51 Figure 5. In the NIH paradigm, ATR-deleted mice exhibit altered antidepressant response............................................................................52 Figure 6. Hippocampal CA1 dendritic spine density following DMI treatment................54 Figure s1. Hippocampal ATR deletion did not affect general locomotor activity or HPA axis activation following an acute stress.........................................55 Figure s2. Fear conditioning is not altered by hippocampal ATR deletion.....................56 Figure s3. ATR deletion does not induce impairments in spatial learning and memory in the Morris water maze......................................................57 Chapter III Figure 1. Sucrose preference and fluid intake............................................................83 Figure 2. Learned helplessness.................................................................................84 Figure 3. Behavioral and endocrine consequences of chronic corticosterone................85 Chapter IV Figure 1. Experimental design................................................................................110 Figure 2. ATRΔHipp mice exhibit deficits in running-induced neurogenesis....................111 Figure 3. Running-induced anxiety in the NIH test is attenuated in ATRΔHipp mice.......112 Figure 4. Running-induced anxiety in an open field is attenuated in ATRΔHipp mice......113 Figure 5. Running-induced anxiety in the light-dark box is attenuated in ATRΔHipp mice..........................................................................................115 vi CHAPTER I. INTRODUCTION A. Mood and anxiety disorders Mood and anxiety disorders are highly prevalent mental illnesses afflicting a significant proportion of adults in the US each year. Among affected individuals, these disorders are highly variable in their etiology and treatment outcome. A better understanding of the pathophysiology of mood and anxiety disorders will not only improve disease prognosis but also ease the large financial burden these illnesses impart due to high healthcare and economic costs (Greenberg et al., 2003). Mood disorders Approximately 10% of adults in the US suffer from a mood disorder each year (Kessler et al., 2005). The majority suffer from dysthymic or major depressive disorder, both of which are characterized by persistent, depressive-like symptoms and differ in regards to severity. The DSM-IV further subdivides depressive disorders into numerous clinical subtypes, supporting the hypothesis that depression may represent a diverse set of pathophysiologies presenting with similar, overlapping symptoms (Akiskal, 1989; Fagiolini and Kupfer, 2003). Depression is defined as the presence of at least one two- week episode of dysphoria and/or anhedonia plus a multitude of additional symptoms including altered sleep/appetite, fatigue, agitation, inability to concentrate, and recurrent thoughts of death or suicide (American Psychiatric Association. and American Psychiatric Association. Task Force on DSM-IV., 2000). Depression is thought to be brought on by an interaction between stressful life experiences and genetic susceptibility factors (Goldberg, 2006), including alterations in the serotonin transporter gene (Caspi et al., 1 2003). Treatment for depression includes cognitive behavioral therapy, pharmacotherapy, and electroconvulsive therapy for severe, treatment resistant depression. Current pharmacotherapies act to increase levels of monoamine transmitters in the brain, either through blocking their reuptake at the synapse or through inhibiting their degradation. Because these drugs require 4-6 weeks to relieve symptoms of depression, they are thought to induce downstream changes in gene transcription and neuronal plasticity following chronic administration (Hyman and Nestler, 1996). Current pharmacotherapies often produce unwanted side effects and less than half of patients show full remission of their disease, even following trials of multiple drugs (Berton and Nestler, 2006). Anxiety Disorders Approximately 18% of adults in the US experience an anxiety disorder each year (Kessler et al., 2005), including general anxiety, panic disorder, social anxiety, post- traumatic stress disorder, and obsessive-compulsive disorder. Anxiety disorders are marked by pervasive worry, tension and avoidance (Allgulander, 2006) and as such, components of fear circuitry in the brain, including the amygdala, nucleus accumbens, hippocampus and insular cortex are implicated in animal models (Kent and Rauch, 2003; Davis, 2006; Maren, 2008). Treatment for anxiety includes benzodiazepines, anticonvulsants, and the serotonin and norepinephrine reuptake inhibitors commonly used in the treatment of depression (Baldwin et al., 2011). Depression and anxiety disorders are highly comorbid; up to 60% of depressed individuals also suffer from anxiety symptoms (Kessler et al., 2003). This comorbidity, in addition the ability of 2
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