PERSISTENT FIRING AND DEPOLARIZATION BLOCK IN RAT CA1 PYRAMIDAL NEURONS by Beate Knauer A thesis submitted in partial fulfilment of the requirements for the degree of Philosophiae Doctoris (PhD) in Neuroscience from the International Graduate School of Neuroscience Ruhr University Bochum March 31st 2015 This research was conducted at the Neural Dynamics Lab within the Faculty of Psychology in association with the Mercator Research Group 1 at the Ruhr University under the supervision of Jun.-Prof. Dr. Motoharu Yoshida Printed with the permission of the International Graduate School of Neuroscience, Ruhr University Bochum "Die allererste und größte Aufgabe im Leben besteht darin, das Gleichgewicht zu halten." Wiesmann (2012) [1] Statement I certify herewith that the dissertation included here was completed and written independently by me and without outside assistance. References to the work and theories of others have been cited and acknowledged completely and correctly. The “Guidelines for Good Scientific Practice” according to § 9, Sec. 3 of the PhD regulations of the International Graduate School of Neuroscience were adhered to. This work has never been submitted in this, or a similar form, at this or any other domestic or foreign institution of higher learning as a dissertation. The abovementioned statement was made as a solemn declaration. I conscientiously believe and state it to be true and declare that it is of the same legal significance and value as if it were made under oath. Beate Knauer / Signature Bochum, 14.07.2015 PhD Commission Chair: Prof. Dr. Stefan Herlitze 1st Internal Examiner: Jun.-Prof. Dr. Motoharu Yoshida 2nd Internal Examiner: Prof. Dr. Denise Manahan-Vaughan External Examiner: Prof. Dr. Maxim Volgushev Non-Specialist: Prof. Dr. med. Dr. rer. nat. habil. Konstanze F. Winklhofer Date of Final Examination: June 30, 2015 PhD Grade Assigned: magna cum laude CONTENTS Contents I List of figures ................................................................................................................... IV II List of tables .................................................................................................................... VI III List of abbreviations ................................................................................................. VII IV List of compounds ...................................................................................................... IIX V Abstract .............................................................................................................................. 1 1 Introduction ....................................................................................................................... 3 1.1 The hippocampus ........................................................................................................................ 4 1.1.1 Short-term maintenance of information in the hippocampus ................................................. 6 1.1.2 The role of the hippocampus in seizures and epilepsy ........................................................... 7 1.2 The hippocampal cholinergic system ......................................................................................... 9 1.2.1 The role of the cholinergic system in short-term maintenance of information .................... 10 1.2.2 The role of the cholinergic system in seizures and epilepsy ................................................ 13 1.3 Electrophysiological properties of CA1 pyramidal neurons related to cholinergic signaling .................................................................................................................................................... 14 1.3.1 The role of muscarinic-dependent intracellular processes in short-term maintenance of information .......................................................................................................................... 19 1.3.2 The role of muscarinic-dependent intracellular processes in seizure-related epileptiform activity ................................................................................................................................. 22 1.4. Summary and hypotheses ........................................................................................................ 24 2 Method ............................................................................................................................... 25 2.1 Animals....................................................................................................................................... 26 2.2 Chemicals and solutions............................................................................................................ 27 2.3 Preparation of acute brain slices .............................................................................................. 28 2.4 Recording procedures ............................................................................................................... 29 2.5 Selection of neurons for data analysis ..................................................................................... 32 2.5.1 Only pyramidal neurons were analyzed ............................................................................... 32 2.5.2 Silent RMP was required for the inclusion of cells .............................................................. 32 2.6 Identification of the cell's anatomical location ....................................................................... 34 I CONTENTS 2.6.1 Dorso-ventral axis ................................................................................................................ 34 2.6.2 Proximo-distal axis ............................................................................................................... 36 2.7 Raw data analysis ...................................................................................................................... 37 2.7.1 Passive properties ................................................................................................................. 38 2.7.2 Hyperpolarization activated current ..................................................................................... 39 2.7.3 Action potential properties ................................................................................................... 40 2.7.4 Post-burst properties ............................................................................................................. 43 2.7.5 Excitability ........................................................................................................................... 44 2.7.6 Persistent activity parameters ............................................................................................... 47 2.8 Statistical comparison of group data ....................................................................................... 50 2.8.1 Comparison with previously reported data .......................................................................... 51 2.8.2 Comparison of categorical data ............................................................................................ 52 3 Results ................................................................................................................................ 54 3.1 The effect of carbachol on intrinsic electrophysiological properties and its modulation by intercellular variability ............................................................................................................. 55 3.1.1 The effect of carbachol on intrinsic electrophysio-logical properties .................................. 55 3.1.2 Modulation of intrinsic properties by intercellular variability ............................................. 67 3.1.3 Summary of the cholinergic effects on CA1 pyramidal neurons ......................................... 79 3.2. Persistent firing ........................................................................................................................ 81 3.2.1 In vitro CA1 pyramidal neurons engaged in persistent firing .............................................. 81 3.2.2 Persistent firing was intrinsic to individual neurons ............................................................ 85 3.2.3 Intrinsic persistent firing may be a physiologically plausible phenomenon ........................ 91 3.2.4 Mechanism underlying intrinsic persistent firing in CA1 pyramidal neurons ..................... 95 3.2.5 Sensitivity of persistent firing to the carbachol concentration ........................................... 104 3.2.6 Sensitivity of persistent firing to inter-cellular variability within CA1 ............................. 105 3.2.7 Summary of persistent firing in CA1 pyramidal neurons .................................................. 106 3.3 Depolarization block ............................................................................................................... 108 3.3.1 The mechanism underlying the switch between persistent firing and depolarization block ........................................................................................................................................... 108 3.3.2 Sensitivity of depolarization block to inter-cellular variability .......................................... 122 3.3.3 Aspects of inter-cellular variability modulating the occurrence of persistent firing and depolarization block .......................................................................................................... 124 3.3.4 Summary