Potassium channels for high-frequency action potential generation in GABAergic interneurons of rat hippocampus [Elektronische Ressource] / vorgelegt von Cheng-Chang Lien

Lienchen

Sujets

• Biologie

Informations

• Publié par: albert-ludwigs-universitat_freiburg
• Publié le: 01 janvier 2003
• Nombre de lectures: 15
• Langue: English
• Poids de l'ouvrage: 19 Mo

Extrait

Aus dem Physiologischen Institut der Albert-Ludwigs-Universität
Freiburg im Breisgau



Title Page



Potassium Channels for High-Frequency Action
Potential Generation in GABAergic Interneurons of Rat
Hippocampus









INAUGURAL-DISSERTATION
zur
Erlangung des Medizinischen Doktorgrades

der Medizinischen Fakultät
der Albert-Ludwigs-Universität
Freiburg im Breisgau





Vorgelegt 2003
von
Cheng-Chang Lien
geboren in Taipei
































Dekan Prof. Dr. Martin Schumacher
1. Gutachter Prof. Dr. Peter Jonas
2. Gutachter Prof. Dr. Klaus Starke
Jahr der Promotion 2003




* Gedruckt mit Unterstützung des Deutschen Adademischen Austauschdienstes iii

























Dedicated to my family






iv

Citations to Own Previously Published Work

Parts of this thesis have been previously published:

Journal articles:

• LIEN, C.-C., MARTINA, M., SCHULTZ, J., EHMKE, H. & JONAS, P.
(2002). Gating, modulation, and subunit composition of voltage-
+gated K channels in dendritic inhibitory interneurones of rat
hippocampus. Journal of Physiology 538, 405-419.

• LIEN, C.-C. & JONAS, P. (2003). Kv3 potassium conductance is
necessary and kinetically optimized for high-frequency action
potential generation in hippocampal interneurons. Journal of
Neuroscience 23, 2058-2068.

Abstracts:

• LIEN, C.-C., MARTINA, M. & JONAS, P. (2001). Functional
+identification and modulation of K channels in oriens-alveus
interneurons of rat hippocampus. Pflügers Archiv 441, R181.

• LIEN, C.-C. & JONAS, P. (2002). Dynamic clamp analysis of the
+functional impact of Kv3-like K channels on the action potential
phenotype of hippocampal interneurons. Pflügers Archiv 443,
S279.

+• LIEN, C.-C. & JONAS, P. (2002). How do Kv3-like K channels in
hippocampal interneurons facilitate fast spiking? Program No.
546.12.2002 Abstract Viewer and Itinerary Planner. Washington,
DC: Society for Neuroscience, 2002. Online.







Table of Contents v
Table of Contents
TITLE PAGE I
CITATIONS TO OWN PREVIOUSLY PUBLISHED WORK IV
TABLE OF CONTENTS V
LIST OF FIGURES VIII
ZUSAMMENFASSUNG IX
ABSTRACT X
1 INTRODUCTION 1
1.1 THE HIPPOCAMPUS (AMMON’S HORN) 1
1.2 NEURONAL ORGANIZATION OF THE RAT HIPPOCAMPUS 2
1.3 KV CHANNELS IN GABAERGIC INHIBITORY INTERNEURONS 6
1.4 IONIC CHANNELS 7
1.5 FUNCTIONAL AND MOLECULAR DIVERSITY OF KV CHANNELS 8
+1.5.1 CLASSIFICATION AND NOMENCLATURE OF K CHANNELS 8
1.5.2 STRUCTURE OF KV CHANNELS 10
1.5.3 FUNCTIONAL PROPERTIES OF KV3 SUBFAMILY 12
1.5.4 GENE EXPRESSION AND CHANNELS 13
1.6 THE AIMS OF THIS WORK 14
2 MATERIALS AND METHODS 15
2.1 PREPARATION OF ACUTE BRAIN SLICES
2.2 IDENTIFICATION OF NEURONS WITH IR-DIC VIDEO MICROSCOPY 16
2.3 PATCH-CLAMP TECHNIQUE IN ACUTE BRAIN SLICES 16
2.3.1 PATCH-CLAMP SETUP 16
2.3.2 NUCLEATED PATCH CONFIGURATION 17
2.4 SINGLE-CELL RT-PCR ANALYSIS OF KV SUBUNIT EXPRESSION 19
2.4.1 PREPARATION FOR WHOLE-CELL RECORDING AND HARVESTING 19
2.4.2 ISOLATION OF RNA FROM SINGLE CELLS
2.4.3 EXPELLING AND RT REACTION OF SINGLE-CELL RNA 20
2.4.4 PCR AMPLIFICATION OF CDNAS 22
2.5 THE IMPLEMENTATION OF THE DYNAMIC-CLAMP TECHNIQUE 24
2.5.1 DESCRIPTION OF KV3 CHANNEL GATING WITH A HH-TYPE MODEL 24
2.5.2 FAST DYNAMIC-CLAMP SYSTEM 25
2.5.3 ADDITION AND SUBTRACTION OF KV3 CONDUCTANCE 28 Table of Contents vi

2.6 DATA ANALYSIS 29
2.6.1 ANALYSIS OF NUCLEATED PATCH DATA 29
2.6.1.1 Activation, inactivation curves, and recovery from inactivation 29
2.6.1.2 Activation and deactivation time constants 30
2.6.1.3 Dose-response curves 30
2.6.2 ANALYSIS OF DYNAMIC-CLAMP DATA 31
2.6.3 CONVENTIONS
2.7 SOLUTIONS AND CHEMICALS 32
2.8 BIOYTIN STAINING AND MORPHOLOGICAL ANALYSIS 34
3 RESULTS 35
3.1 PART I: GATING, MODULATION, AND SUBUNIT COMPOSITION OF KV CHANNELS IN
DENDRITIC INHIBITORY INTERNEURONS OF RAT HIPPOCAMPUS 35
3.1.1 AP PATTERNS AND MORPHOLOGICAL PROPERTIES OF OA INTERNEURONS 35
3.1.2 PHARMACOLOGICAL DISSECTION OF KINETICALLY DISTINCT KV CURRENT COMPONENTS 39
+3.1.3 GATING PROPERTIES OF THE THREE TYPES OF K CHANNELS IN OA INTERNEURONS 43
+3.1.4 SELECTIVE MODULATION OF FAST DELAYED RECTIFIER K CHANNELS 48
+3.1.5 SINGLE-CELL RT-PCR ANALYSIS OF K CHANNEL SUBUNIT TRANSCRIPTS 50
3.1.6 TEMPORAL RELATIONSHIP BETWEEN KV3-LIKE CURRENTS AND AP TRAJECTORIES 52
3.1.7 KV3-LIKE CURRENTS FACILITATE FS AND PREVENT SPIKE BROADENING 54
3.2 PART II: KV3 POTASSIUM CONDUCTANCE IS NECESSARY AND KINETICALLY OPTIMIZED
FOR HIGH-FREQUENCY AP GENERATION IN HIPPOCAMPAL INTERNEURONS 56
3.2.1 IMPLEMENTATION OF KV3 CHANNEL GATING MODEL IN A DYNAMIC-CLAMP SYSTEM 56
3.2.2 ADDITION OF KV3 CONDUCTANCE AFTER PHARMACOLOGICAL BLOCK RESCUES THE FS
PHENOTYPE 59
3.2.3 SUBTRACTION OF KV3 CONDUCTANCE MIMICS THE EFFECTS OF BLOCKERS 61
3.2.4 KV3 IS SUFFICIENT FOR FS, INDEPENDENTLY OF CELLULAR BACKGROUND 63
3.2.5 INTERMEDIATE DEACTIVATION RATE IS OPTIMAL FOR THE FS PHENOTYPE 65
3.2.6 IMPACT OF ACTIVATION THRESHOLD ON AP PHENOTYPES 67
3.2.7 IMPACT OF FAST INACTIVATION OF KV3 ON AP DURATIONS AND AP PHENOTYPES 69
4 DISCUSSION 71
4.1 A KV3.1/3.2-LIKE CHANNEL IS THE MAJOR KV CHANNEL IN OA INTERNEURONS 71
4.2 KV3-LIKE CURRENTS ACTIVATED PREFERENTIALLY DURING APS 72
+4.3 K CHANNEL EXPRESSION DETERMINES THE AP PATTERNS OF NEURONS 73
4.4 KV3 CHANNELS ARE NECESSARY, AND PERHAPS SUFFICIENT, FOR THE FS PHENOTYPE 74
4.5 IMPACT OF INDIVIDUAL KV3 GATING PROPERTIES ON AP PATTERNS 75
4.6 NEUROMODULATORY CONTROL OF KV3 CHANNELS 76
+4.7 A POSSIBLE ROLE OF INTERNEURON K CHANNELS IN DENDRITIC INHIBITION 79
4.8 FUNCTIONAL SIGNIFICANCE FOR SYNAPTIC INHIBITION AND NETWORK ACTIVITY 80
4.9 PATHOPHYSIOLOGICAL RELEVANCE 81
4.9.1 OSCILLATIONS AND EPILEPTIC SEIZURE 81
4.9.2 PARKINSON’S DISEASE 82
5 REFERENCES 84 Table of Contents vii

ACKNOWLEDGEMENTS 100
CURRICULUM VITAE 101
APPENDIX 102

























List of Figures viii
List of Figures
List of Figures
FIG. 1 ANATOMY OF THE RAT HIPPOCAMPAL FORMATION......................................................3
FIG. 2 SYNAPTIC ORGANIZATION AND WIRING DIAGRAM OF THE EXCITATORY PATHWAYS
OF THE HIPPOCAMPUS.......................................................................................................4
FIG. 3 DOMAIN-SPECIFIC INNERVATIONS OF INTERNEURONS IN THE HIPPOCAMPAL CA1-
CA3 REGION........................................................................................................................5
+FIG. 4 CLASSIFICATION AND NOMENCLATURE OF K CHANNELS ...........................................9
FIG. 5 KV CHANNEL STRUCTURE .............................................................................................11
FIG. 6 THE PATCH-CLAMP SETUP FOR ELECTROPHYSIOLOGICAL RECORDINGS ................18
FIG. 7 FLOW DIAGRAM OF THE STEPS OF THE SINGLE-CELL RT-PCR ANALYSIS.................21
FIG. 8 SCHEMATIC ILLUSTRATION OF THE DYNAMIC-CLAMP SYSTEM..................................27
FIG. 9 RECORDING FROM OA INTERNEURONS IN HIPPOCAMPAL SLICES.............................35
FIG. 10 CORRELATION BETWEEN FUNCTIONAL CHARACTERISTICS AND MORPHOLOGY OF
OA INTERNEURONS ..........................................................................................................37
FIG. 11 IDENTIFICATION OF OA INTERNEURONS BY INTRACELLULAR FILLING WITH
BIOCYTIN...........................................................................................................................38
+FIG. 12 PHARMACOLOGICAL PROPERTIES OF K CHANNELS IN NUCLEATED PATCHES
FROM OA INTERNEURONS ...............................................................................................40
FIG. 13 OCCLUSION BETWEEN THE EFFECTS OF LOW CONCENTRATIONS OF 4-AP AND TEA
...........................................................................................................................................41
FIG. 14 CURRENT SUBTRACTION ANALYSIS SUGGESTS THE PRESENCE OF THREE
+KINETICALLY DISTINCT K CURRENT COMPONENTS IN NUCLEATED PATCHES...........42
+FIG. 15 GATING PROPERTIES OF THE FAST