Functional mapping of the entorhinal cortex reveals a hub for synaptic excitability in the young 3xTg mouse model for Alzheimer's disease A THESIS SUBMITTED TO THE UNIVERSITY OF MANCHESTER FOR THE DEGREE OF DOCTOR OF PHILOSOPHY IN THE FACULTY OF BIOLOGY, MEDICINE AND HEALTH 2019 Francesca Mandino SCHOOL OF BIOLOGICAL SCIENCES Neuroscience & Experimental Psychology Table of Contents List of Figures ......................................................................................................................... 5 List of Tables .......................................................................................................................... 7 Alternative Format Structure ............................................................................................... 8 Abstract ................................................................................................................................. 10 Declaration ............................................................................................................................ 11 Copyright statement ............................................................................................................. 11 Acknowledgements ............................................................................................................... 12 List of main abbreviations ................................................................................................... 13 ~Chapter 1 ~ ......................................................................................................................... 15 General Introduction ........................................................................................................... 15 1.1 Alzheimer's disease: a general overview .................................................................. 15 1.1.1 Classification ............................................................................................................ 16 1.2 Anatomy of the medial temporal lobe ...................................................................... 18 1.2.1 Entorhinal Cortex: function and relevance in AD ............................................... 20 1.2.2 Lateral entorhinal cortex: cytoarchitecture and projections .............................. 21 1.3 Alzheimer’s disease .................................................................................................... 23 1.3.1 Types of AD: Early- and Late-Onset ..................................................................... 23 1.3.2 Genetic risk factors ................................................................................................. 24 1.3.3 Molecular hallmarks ............................................................................................... 25 1.3.4 Unclear aetiology of AD .......................................................................................... 27 1.4 Animal models in preclinical research ..................................................................... 29 1.4.1 Limitations of animal models ................................................................................. 31 1.4.2 3xTgAD mouse model ............................................................................................. 33 1.5 Structure of the present work ................................................................................... 35 1.6 Neuroimaging ............................................................................................................. 35 1.6.1 Positron Emission Tomography ............................................................................. 36 1.6.2 Structural Magnetic Resonance Imaging .............................................................. 37 1.6.3 Functional magnetic resonance imaging ............................................................... 38 1.7 Functional Connectivity in Alzheimer’s disease ..................................................... 40 1.7.1 Evidence from human fMRI .................................................................................. 40 1.7.2 Evidence from rodent fMRI ................................................................................... 42 1.8 Optogenetics................................................................................................................ 43 1.8.1 Optogenetics in AD ................................................................................................. 46 1.9 Electrophysiology ....................................................................................................... 47 1 1.9.1 Fundamentals of electrophysiology ....................................................................... 47 1.9.2 Memory formation .................................................................................................. 49 1.9.3 Neuronal Changes in AD Models ........................................................................... 50 1.10 Anaesthesia ............................................................................................................... 53 1.11 The present work ...................................................................................................... 55 ~ Chapter 2 ~ ........................................................................................................................ 57 General Methods .................................................................................................................. 57 2.1 Animals ....................................................................................................................... 57 2.2 fMRI ............................................................................................................................ 58 2.2.1 fMRI signal .............................................................................................................. 59 2.2.2 BOLD signal-to-noise .............................................................................................. 59 2.2.3 Animal preparation for imaging ............................................................................ 60 2.2.4 RsfMRI and ofMRI: data acquisition ................................................................... 61 2.2.5 RsfMRI and ofMRI: data pre-processing ............................................................. 62 2.2.6 RsfMRI: data analysis ............................................................................................ 63 2.2.7 OfMRI: optogenetic surgery .................................................................................. 66 2.2.8 OfMRI: stimulation protocols ................................................................................ 67 2.2.9 OfMRI: data analysis .............................................................................................. 69 2.2.10 Statistics and data availability ............................................................................. 69 2.2.11 Ex vivo processing ................................................................................................. 70 2.3 In vivo electrophysiology ............................................................................................ 74 2.3.1 Anaesthesia and surgery ......................................................................................... 75 2.3.2 Electrode placement ................................................................................................ 76 2.3.3 Stimulation protocols .............................................................................................. 79 2.3.4 Data analysis ............................................................................................................ 80 2.3.5 Perfusion .................................................................................................................. 83 2.3.6 Histology................................................................................................................... 83 ~ Chapter 3 ~ ........................................................................................................................ 84 Paper 1 .................................................................................................................................. 84 Rationale behind Paper 1 ................................................................................................ 84 3.1 Abstract ....................................................................................................................... 86 3.2 Introduction ................................................................................................................ 87 3.3 Method ........................................................................................................................ 89 3.3.1 Animals..................................................................................................................... 89 3.3.2 Optogenetic surgery ................................................................................................ 89 2 3.3.3 Animal preparation for imaging ............................................................................ 90 3.3.4 Data acquisition and stimulation protocols .......................................................... 91 3.3.5 fMRI analysis ........................................................................................................... 92 3.3.6 Ex vivo processing ................................................................................................... 93 3.3.7 Statistics and data availability ..............................................................................