MOLECULAR MECHANISMS UNDERLYING SYNAPTIC DYSFUNCTION IN AGING ALZHEIMER'S ...

197 Pages·2011·1.8 MB·English

by  George Crandell

MOLECULAR MECHANISMS UNDERLYING SYNAPTIC DYSFUNCTION IN AGING
ALZHEIMER’S DISEASE AND DIABETES

by

Thiruchelvam Kariharan

A dissertation submitted to the Graduate Faculty of
Auburn University
in partial fulfillment of the
requirements for the Degree of
Doctor of Philosophy

Auburn, Alabama
December 12, 2011

Key words: Cognitive dysfunction, Synaptic plasticity,
Aging, Alzheimer’s disease, Diabetes, PPARγ

Copyright 2011 by Thiruchelvam Kariharan

Approved by

Vishnu Suppiramaniam, Co-chair, Associate Professor of Pharmacal sciences
Rajesh Amin, Co-chair, Assistant Professor of Pharmacal sciences
Murali Dhanasekaran, Associate Professor of Pharmacal sciences
Randall Clark, Professor of Pharmacal sciences
Jay Ramapuram, Associate Professor of Pharmacal sciences
Kevin Huggins, Associate Professor of Nutrition and food science
ABSTRACT

Alzheimer’s disease (AD) and diabetes (DB) contribute to the development of early brain
aging and cognitive dysfunction. Mechanistic basis of cognitive dysfunction in AD and DB is
not well elucidated. Resistance to age related cognitive decline has been observed in growth
hormone deficient, long-lived Ames dwarf mice. These mice show reduced circulating levels of
growth hormone (GH) and insulin growth factor-1 (IGF-1); however, they have increased levels
of these hormones in the hippocampus, a part of the brain where memory is encoded and
consolidated. The hippocampal glutamatergic and cholinergic systems play a vital role in
synaptic plasticity mechanisms of learning and memory. Therefore, we investigated the
expression of glutamtergic synaptic markers and cholinergic and inflammatory enzymatic
activity in the hippocampus of the Ames dwarf mice. We found an increase in postsynaptic
glutamate receptor expression as well as increased cholineacetyl transferase and lactate
dehydrogenase activity. We have also found elevated acetylcholine and enhanced postsynaptic
glutamatergic activity in the hippocampus of these dwarf mice, which results in the development
of improved synaptic plasticity and cognitive function. AD is associated with the early aging of
the brain and results in the development of cognitive impairment. Amyloid beta (Aβ) is one of
the major hallmarks of AD; the molecular mechanism by which Aβ alters glutamatergic system
and impairs cognitive function in the early stages of AD is not well understood. To better
understand the effects of Aβ upon the development of AD, we intracranially infused Aβ (1-42)
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into wild type mice and found that diminished postsynaptic glutamate receptors that are crucial
for learning and memory. Such finding implicates that Aβ, directly disrupts cognitive function
and synaptic plasticity via attenuating postsynaptic glutamatergic transmission prior to
development of neurodegeneration. Recent studies suggested direct link between AD and
diabetes type 2. To better understand the mechanism of how insulin resistance impairs cognitive
function, we utilized a transgenic type 2 diabetic mouse model (Leptin receptor knock out db/db
mice). The nuclear receptor PPARγ plays a key role in metabolic regulation by modulating
whole body glucose homeostasis and insulin sensitivity. Recently, PPARγ has also been
implicated to have anti-inflammatory and neuroprotective effects in mouse models of AD.
Consequently, we investigated the effects of PPARγ activation on diabetes-induced deficits in
spatial and recognition memory in db/db mice. We found that pharmacological activation of
PPARγ ameliorates diabetes-induced impairment of spatial and recognition memory and also
synaptic transmission and LTP. These data can be explained by our findings that PPARγ
activation enhanced the postsynaptic glutamate receptor expression in the db/db mouse model. In
addition, we also found an increase in the insulin substrate-2 protein (IRS-2) and its effector
proteins, which are involved in the insulin signaling cascade. Moreover, we found that synaptic
plasticity master regulatory transcription factor CREB binding protein (CBP) also increased by
PPARγ activation. These data offers a plausible hypothesis that PPARγ enhances synaptic
plasticity and cognitive function by modulating insulin signaling pathway and synaptic plasticity
associated transcription factors leading to improved glutamatergic synaptic transmission. Our
findings suggest that molecular targets such as PPARγ may offer potential therapeutic targets for
DB, AD and aging induced cognitive impairment.

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ACKNOWLEDGEMENTS

I would like to thank my mentors Dr.Raj Amin and Dr. Vishnu Suppiramaniam for their
continuous support and guidance in my professional development throughout my doctoral
program. Dr. Amin enlightened me towards the health disparity of diabetes, which grips our
nation. He further showed me how novel molecular techniques involving PPAR research helps to
ameliorate severe complications associated with diabetes-induced neurological disorders. I
would like to thank Dr.Vishnu for introducing me into the neuroscience research and teach me
neurophysiology and synaptic plasticity mechanisms and enlightened me with
electrophysiological techniques and applying them to better understand the mechanisms of
synaptic dysfunction and cognitive deficits in neurogenerative diseases. I like to thank Dr.Murali
Dhanasekaran for his help and support on behavioral studies in his lab and Dr. Kevin Huggins
for his help and guidance for molecular techniques I learnt in his lab. I appreciate all the helps
and guidance from my graduate committee members Dr. Randall Clark and Dr.Jay Ramapuram.
I would like to express my thanks to Dr. Robert Judd for serving in my graduate committee as an
external reader for my dissertation. I like to thank my senior colleagues Dr. Parameshwaran, Dr.
Uthayathas, Dr. Shonesy, Dr. Karuppagounder for their support and guidance in various
experimental techniques. Especially I would like to thank Dr. Parameshwaran, Dr. Shonesy and
Ms. Abdul Rahman for their help in electrophysiological experiments. I would also like to
express my sincerest appreciation for their guidance and support and encouragement to my lab
colleagues Dr.Nilmini Vishwaprakash and Gayani. I would like to express my thanks to Inseya
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for proof reading my entire manuscript. Lastly, I would like to thank for all the support and help
provided by our department staff and graduate colleagues and friends. This thanks would be
especially extended to Dean Evans, Dr. William Ravis and Dr. Charlene McQueen for their
support, guidance and mentoring throughout the doctoral program. Finally, and most
importantly, I would like to thank my father; Thiruchelvam Vairamuthu and my mother;
Thavamani Devy Thiruchelvam, whom I cannot offer enough thanks for their patience, love and
support.

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TABLE OF CONTENTS

Abstract ......................................................................................................................................... ii
Acknowledgments........................................................................................................................ iv
List of Tables ................................................................................................................................ x
List of Figures .............................................................................................................................. xi
1. Introduction .............................................................................................................................. 1
2. Review of Literature ................................................................................................................. 4
Aging and molecular mechanisms of synaptic dysfunction ............................................. 4
Prevalence of aging .................................................................................................... 4
Aging theories and physiological changes ................................................................. 4
Aging and cognitive dysfunction ............................................................................... 5
Synaptic deficit in aging brain ................................................................................... 6
Gene expression changes in aging brain .................................................................... 8
Signaling cascades of brain aging ............................................................................ 10
Animal models of aging ........................................................................................... 12
References ................................................................................................................ 12
Long-lived Ames dwarf mice .................................................................................. 16
References ................................................................................................................ 20
Synaptic Plasticity and Glutamatergic system .............................................................. 24
Hippocampus and synaptic plasticity....................................................................... 24
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Glutamate Receptors ................................................................................................ 27
AMPAR ................................................................................................................... 28
NMDAR .................................................................................................................. 30
References ............................................................................................................... 32
Alzheimer’s disease and synaptic dysfunction ............................................................ 36
Alzheimer’s disease (AD) ....................................................................................... 37
Gene mutations in AD ............................................................................................ 37
AD and Amyloid beta pathology ............................................................................ 37
Amyloid beta and synaptic dysfunction .................................................................. 38
Animal models of AD ............................................................................................. 40
Exogenous Aβ induced rodent model of AD .......................................................... 40
Aβ and Glutamatergic dysfunction ......................................................................... 41
References ............................................................................................................... 43
Diabetes and synaptic dysfunction............................................................................... 47
Diabetes Mellitus .................................................................................................... 47
Pathophysiology of diabetes ................................................................................... 48
Type 2 diabetes and cognitive impairment ............................................................. 48
Mechanism of synaptic deficit in type 2 diabetes ................................................... 49
Insulin signaling and synaptic plasticity ................................................................. 50
Animal models of diabetes induced cognitive dysfunction .................................... 53
Diabetes and Glutamatergic dysfunction ................................................................ 54
References ............................................................................................................... 56
PPARγ and synaptic plasticity .................................................................................... 62
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PPARs ..................................................................................................................... 62
PPARγ ..................................................................................................................... 63
Molecular mechanisms of PPARγ activity ............................................................. 64
PPARs expression in the brain ................................................................................ 65
Effect of PPARγ in the brain .................................................................................. 66
Thiazolidinediones (TZD) ...................................................................................... 67
References ............................................................................................................... 68
3. Molecular mechanisms of memory enhancement in long-lived Ames dwarf mice ................ 73
Abstract ................................................................................................................... 73
Introduction ............................................................................................................. 74
Materials and Methods ............................................................................................ 80
Results ..................................................................................................................... 83
Figures and Figure Legends .................................................................................... 86
Discussion ............................................................................................................... 91
References ............................................................................................................... 97
4. Amyloid beta induced cognitive dysfunction ....................................................................... 102
Abstract ................................................................................................................. 102
Introduction ........................................................................................................... 103
Materials and Methods .......................................................................................... 107
Results ................................................................................................................... 113
Figures and Figure Legends .................................................................................. 116
Discussion ............................................................................................................. 126
References ............................................................................................................. 131
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5. PPARγ ameliorates diabetes induced cognitive dysfunction ................................................ 135
Abstract ................................................................................................................. 135
Introduction ........................................................................................................... 138
Materials and Methods .......................................................................................... 141
Results ................................................................................................................... 154
Figures and Figure Legends .................................................................................. 161
Discussion ............................................................................................................. 172
References ............................................................................................................. 180
6. Summary and Conclusions ................................................................................................... 184

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LIST OF TABLES

Table 4.1 Change of inflammatory and cholinergic enzymatic activity in icv-Aβ infused
Rodent hippocampus ................................................................................................. 126

Table 5.1 Primer sequences of synaptic plasticity genes .......................................................... 150

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