ALTERATIONS TO HIPPOCAMPAL NEUROPLASTICITY ACROSS THE LIFESPAN IN A RODENT ...

324 Pages·2016·5.36 MB·English

by  Karen Boschen

ALTERATIONS TO HIPPOCAMPAL NEUROPLASTICITY ACROSS THE
LIFESPAN IN A RODENT MODEL OF FASD
by
Karen E. Boschen

A dissertation submitted to the Faculty of the University of Delaware in partial
fulfillment of the requirements for the degree of Doctor of Philosophy in Psychology

Spring 2016

© 2016 Karen E. Boschen
All Rights Reserved
ProQuest Number: 10156560
All rights reserved
INFORMATION TO ALL USERS
The quality of this reproduction is dependent upon the quality of the copy submitted.
In the unlikely event that the author did not send a complete manuscript
and there are missing pages, these will be noted. Also, if material had to be removed,
a note will indicate the deletion.
ProQuest 10156560
Published by ProQuest LLC (2016). Copyright of the Dissertation is held by the Author.

All rights reserved.
This work is protected against unauthorized copying under Title 17, United States Code
Microform Edition © ProQuest LLC.
ProQuest LLC.
789 East Eisenhower Parkway
P.O. Box 1346
Ann Arbor, MI 48106 - 1346
ALTERATIONS TO HIPPOCAMPAL NEUROPLASTICITY ACROSS THE
LIFESPAN IN A RODENT MODEL OF FASD

by
Karen E. Boschen

Approved: __________________________________________________________
Robert Simons, Ph.D.
Chair of the Department of Psychology

Approved: __________________________________________________________
George H. Watson, Ph.D.
Dean of the College of Arts & Sciences

Approved: __________________________________________________________
Ann L. Ardis, Ph.D.
Senior Vice Provost for Graduate and Professional Education
I certify that I have read this dissertation and that in my opinion it meets
the academic and professional standard required by the University as a
dissertation for the degree of Doctor of Philosophy.

Signed: __________________________________________________________
Anna Y. Klintsova, Ph.D.
Professor in charge of dissertation

I certify that I have read this dissertation and that in my opinion it meets
the academic and professional standard required by the University as a
dissertation for the degree of Doctor of Philosophy.

Signed: __________________________________________________________
Tania L. Roth, Ph.D.
Member of dissertation committee

I certify that I have read this dissertation and that in my opinion it meets
the academic and professional standard required by the University as a
dissertation for the degree of Doctor of Philosophy.

Signed: __________________________________________________________
Jeffrey B. Rosen, Ph.D.
Member of dissertation committee

I certify that I have read this dissertation and that in my opinion it meets
the academic and professional standard required by the University as a
dissertation for the degree of Doctor of Philosophy.

Signed: __________________________________________________________
Jaclyn M. Schwarz, Ph.D.
Member of dissertation committee
I certify that I have read this dissertation and that in my opinion it meets
the academic and professional standard required by the University as a
dissertation for the degree of Doctor of Philosophy.

Signed: __________________________________________________________
Alexandre E. Medina, Ph.D.
Member of dissertation committee
ACKNOWLEDGEMENTS
First, I would like to thank my advisor and mentor, Dr. Anna Klintsova. Thank
you for your support, for challenging me to always be a better scientist, and,
ultimately, for your trust in me. You have always listened to my opinion and ideas as
an equal, which allowed me to gain confidence in my own judgment and abilities –
thank you. I would also like to thank each member of my committee. Thank you to Dr.
Tania Roth for collaborating with me, allowing me to freely use your lab and
equipment, and supporting me during my dissertation; I always appreciate your advice
and words of optimism. Thank you to Dr. Jaclyn Schwarz for her guidance as we
began exploring the neuroimmune system in our lab and the use of IL-1β primers.
Thank you to Dr. Jeffrey Rosen for his support during my time at UD and,
specifically, his advice pertaining to Specific Aim 3. Thank you to Dr. Alexandre
Medina for your encouragement during this process and professional advice.
Thank you to my labmates over the years, the ones who were on the “front
lines” with me. Specifically, thank you to Dr. Gillian Hamilton for teaching me
everything I know and being an example of a great friend and lab partner. Thank you
to Kerry Criss for being my lab treasure and an amazing friend. Thank you to Zachary
Gursky, I know I am leaving the lab in capable hands with you. Thank you to Dr.
Vitaly Palamarchouk for establishing the ELISA technique in our lab and his cheerful
spirit. A huge thank you to all of my undergraduate research assistants over the years:
Michael Ruggiero (who counted microglia in Aim 1), Sarah McKeown (who
v
performed DCX immunohistochemistry in Aim 3), Shaqran Shareeq (who ran agarose
gels and spectrophotometry), Zubin Hussain (who assisted with the Ki-67
immunohistochemistry), Samuel Modlin, Mia Castiglione, James Delorme, Alejandro
Morales, Shannon Houlihan, Brie Gerry, Liz Hetterly, Julia Johansson, Jackie King,
Natalie Ginn, Emma Spillman, and Fiona Flowerhill. Each one of you is an invaluable
asset to the lab and helped make lab feel like home.
Thank you to my parents, Paula Boschen and Dr. John and Rosa Boschen, and
my brothers and sisters, Andrew, Sally, Kelly, Kaitlin, and Gordon. I feel eternally
grateful to have such a wonderful, supportive family. Thank you to the friends I have
made here at Delaware, your patient ears and laughter during times of stress and
uncertainty meant a lot. Thank you to Kate Tubbs Blank and Mary Goldsberry Troyer
who remind me that true friends are always just a phone call away. Thanks to my cat
Nixie for the companionship during long days of writing. And finally, thank you to my
partner Tom Gamage for the love, friendship, and your belief in me – I am truly
grateful for your presence in my life.

vi
TABLE OF CONTENTS
LIST OF TABLES ....................................................................................................... xii
LIST OF FIGURES ..................................................................................................... xiii
LIST OF ABBREVIATIONS ...................................................................................... xx
ABSTRACT .............................................................................................................. xxiii

Chapter

1 INTRODUCTION .............................................................................................. 1

1.1 Brief Introduction to Neuroplasticity in the Hippocampus ...................... 2
1.2 Brief Introduction Fetal Alcohol Spectrum Disorders and Hippocampal
Dysfunction............................................................................................... 3
1.3 Brief Introduction to Behavioral Interventions and Neuroplasticity in the
Hippocampus ............................................................................................ 5
1.4 Dissertation Questions .............................................................................. 6
1.5 Dissertation Summary .............................................................................. 9

2 FETAL ALCOHOL SPECTRUM DISORDERS (FASD) AND THE
HIPPOCAMPUS .............................................................................................. 11

2.1 Fetal Alcohol Spectrum Disorders (FASD) ............................................ 12
2.2 Modeling Developmental Alcohol Effects in Experimental Animals .... 15
2.3 Overview of Hippocampal Anatomy and Development ........................ 20
2.4 Developmental Alcohol Exposure and Hippocampal Dysfunction ........ 23
2.5 Developmental Alcohol Exposure Alters Learning, Anxiety- and
Depression-like Behaviors ...................................................................... 31
2.6 Conclusions ............................................................................................ 34

3 NEUROTROPHINS, NEUROIMMUNE FUNCTION, AND
ENVIRONMENTAL MANIPULATIONS: IMPACT ON HIPPOCAMPAL
DEVELOPMENT AND NEUROPLASTICITY ............................................. 36

3.1 Role of Neurotrophic Factors in Brain Development and
Neuroplasticity ........................................................................................ 37

3.1.1 Expression of Neurotrophic Factors During Development ............. 37
3.1.2 Neurotrophins Influence Hippocampal Structure and Function ...... 39
3.1.3 Neurotrophins Enhance Learning and Reduce Anxiety- and
Depressive-like Behaviors ............................................................... 45

vii
3.2 Role of the Neuroimmune System on Brain Development and
Neuroplasticity ........................................................................................ 47

3.2.1 Microglial Colonization, Morphology, and Cytokine Expression in
the Developing CNS ........................................................................ 48
3.2.2 Neuroimmune Activation during Development has Long-term
Consequences on Brain Function .................................................... 52
3.2.3 Neurotrophins and the Neuroimmune Response ............................. 54

3.3 Exercise and Environmental Complexity Impact Hippocampal Structure
and Function ........................................................................................... 56

3.3.1 Impact of Exercise ........................................................................... 56
3.3.2 Impact of Environmental Complexity (EC) .................................... 59

3.4 Conclusions ............................................................................................ 62

4 FOUNDATIONAL STUDIES.......................................................................... 64

4.1 Neonatal Alcohol Exposure Negatively Impacts Hippocampal Adult
Neurogenesis and Dendritic Morphology ............................................... 64
4.2 Neurotrophins as Targets of Alcohol Exposure...................................... 70
4.3 Neurotrophins as Neuroprotective Molecules in the Developing Brain . 73
4.4 Neonatal Alcohol Exposure Induces the Neuroimmune Response ........ 76
4.5 Behavioral Interventions Reverse Hippocampal Dysfunction in Rodents
Models of FASDs ................................................................................... 77
4.6 Bdnf Gene Methylation as a Novel Approach to Assess Long-Lasting
Changes to BDNF in the Hippocampus. ................................................ 81
4.7 Conclusions ............................................................................................ 83

5 GENERAL PROCEDURES ............................................................................. 85

5.1 Subjects ................................................................................................... 85
5.2 Alcohol Exposure ................................................................................... 87
5.3 Blood Alcohol Concentration (BAC) Analysis ...................................... 88
5.4 Adolescent and Adult Housing Conditions ............................................ 88

5.4.1 Wheel Running (WR) ...................................................................... 89
5.4.2 Environmental Complexity (EC) ..................................................... 90

5.5 Immunohistochemistry ........................................................................... 91
5.6 Analysis of Immunolabeled Hippocampal Tissue .................................. 92

viii
5.6.1 Cell Count Estimates Using Unbiased Stereology .......................... 92
5.6.2 Microglial Cell Territory Analysis .................................................. 93

5.7 Gene Expression Analysis ...................................................................... 97
5.8 Protein Expression Analysis ................................................................... 97
5.9 DNA Methylation Analysis .................................................................... 99
5.10 Statistical Analyses ............................................................................... 100

6 SPECIFIC AIM 1: IMPACT OF NEONATAL ALCOHOL EXPOSURE ON
BDNF EXPRESSION AND THE NEUROIMMUNE RESPONSE IN THE
DEVELOPING RAT HIPPOCAMPUS ......................................................... 102

6.1 Introduction........................................................................................... 102
6.2 Experiment 1: Impact of Neonatal Alcohol Exposure on BDNF and
TrkB Protein Levels, Bdnf Exon-Specific Gene Expression, and Bdnf
DNA Methylation in the Developing Rat Hippocampus ...................... 103
6.2.1 Materials and Methods .................................................................. 106
6.2.2 Results ........................................................................................... 110
6.2.3 Discussion...................................................................................... 118

6.3 Experiment 2: Impact of Neonatal Alcohol Exposure on Number and
Activation State of Microglia in the Developing Rat Hippocampus .... 124

6.3.1 Materials and Methods .................................................................. 127
6.3.2 Results ........................................................................................... 134
6.3.3 Discussion...................................................................................... 141

6.4 General Discussion ............................................................................... 149

6.4.1 Neuroprotection in the Developing Brain ..................................... 149
6.4.2 Role of Intubation Stress in the Current Findings ......................... 151

6.5 Conclusion ............................................................................................ 154

7 SPECIFIC AIM 2: INFLUENCE OF AEROBIC EXERCISE ALONE OR
FOLLOWED BY ENVIRONMENTAL COMPLEXITY ON HIPPOCAMPAL
BDNF EXON-SPECIFIC GENE EXPRESSION, PROTEIN, AND DNA
METHYLATION IN ADULT RATS NEONATALLY EXPOSED TO
ALCOHOL ..................................................................................................... 159

7.1 Introduction........................................................................................... 159
7.2 Materials and Methods ......................................................................... 162

ix