Effects of H_1tn2O_1tn2 at rat myenteric neurones in culture [Elektronische Ressource] / by Pouokam Kamgne Ervice Vidal

Pouokam Kamgne, Ervice Vidal

Informations

• Publié par: justus-liebig-universitat_giessen
• Publié le: 01 janvier 2009
• Nombre de lectures: 25
• Langue: English
• Poids de l'ouvrage: 2 Mo

Extrait

Effects of H O at rat myenteric neurones in 2 2
culture


Inaugural Dissertation
submitted to the
Faculty of Veterinary Medicine
in partial fulfilment of the requirements
for the PhD Degree
of the Faculties of Veterinary Medicine and Medicine
of the Justus Liebig University Giessen
Germany

by

Pouokam Kamgne Ervice Vidal

from

Douala, Cameroon


Giessen 2009



From the Institute For Veterinary Physiology
of the Justus Liebig University Giessen






First Supervisor and Committee Member : Prof. Dr. Martin Diener
Second Supervisor and Committee Member : Prof. Dr. Winfried Neuhuber
Chairman of the oral panel : Prof. Dr. Norbert Weissmann
Examiner : Prof. Dr. Stefan Arnhold





stDate of the Doctoral Defense : March 1 , 2010. Pouokam K.E.V. Dedication i


Dedication

This thesis is dedicated to my parents Kamgne Jacques and Metchuiam Elise, my
brothers and sisters Kamche Pelagie, Kamsu Sylvain, Feutseu Yves, Kouam Alliance,
and Tameu Jacques, my nephew Mouam Gilles and the entire Taguieguem family.

Pouokam K.E.V. Table of contents ii
Table of contents

Dedication.................................................................................................................................i
Table of contents....................................................................................................................ii
List of Figures........................................................................................................................v
List of Tables.......................................................................................................................iiiv
List of Abbreviations............................................................................................................ix

1 Introduction................................................................................................................1
1.1 Short description of the enteric nervous system................................................1
1.2 Types and functions of enteric neurones.............................................................4
1.3 Synaptic behavior of myenter.........................................................10
1.3.1 Fast excitatory postsynaptic potentials............................................................10
1.3.2 Slow excitatory postsynaptic potentials..........................................................10
1.3.3 Slow inhibitory postsynaptic potentials............................................................11
1.3.4 Presynaptic inhibition........................................................................12
1.4 Ionic channels and excitability in neurones......................................................12
1.4.1 Voltage-gated sodium channels........................................................................12
1.4.2 Potassium channel..............................................................................................16
1.4.3 Calcium channels...............................................................................................16
1.4.3.1 The voltage-gated calcium channels...............................................................17
2+1.4.3.2 Ca release channels: ryanodine receptors and IP receptors.....................17 3
1.4.3.3 Mitochondrial and nuclear ion channels.........................................................18
2+ 1.4.3.4 Regulation of Ca fluxes..................................................................................18
1.5 Enteric nervous system and gastrointestinal motility.....................................21
1.6 Aim of the study................................................................................................22 Pouokam K.E.V. Table of contents iii
2 Materials and Methods………………….………………....………...………..24
2.1 Materials………………………….................………………………………..24
2.1.1 Chemicals...............................................................................................................24
2.1.2 Antibiotics............................................................................................26
2.1.3 Enzymes.................................................................................................................27
2.1.4 Biological material..............................................................................................27
2.1.5 Buffers and solutions..........................................................................................27
2.1.5.1 Myenteric plexus culturing solutions................................................................27
2.1.5.2 Solutions for patch-clamping and imaging......................................................28
2.2 Methods.....................................................................................................................29
2.2.1 Isolation and culture of the myenteric neurones............................................29
2.2.2 Patch-clamp experiments..................................................................................32
2.2.2.1 Technique......................................................................................................….32
2.2.2.2 Isolation of sodium currents and effects of H O ..........................................35 2 2
2.2.3 Calcium imaging........................................................................................…...35
2.2.4 Statistics................................................................................................................38
3 Results.......................................................................................................39
2+3.1 Effects of H O on the membrane potential and the cytosolic Ca 2 2
Concentration..........................................................................................................39
+3.2 Inhibition of the Na currents...........................................................49
3.3 Alteration of the excitability................................................................................57
+3.4 Mediation of the H O effects on Na currents.................................................59 2 2
4 Discussion.................................................................................................................65
4.1 Mechanism of the hyperpolarization evoked by H O .................................65 2 2
2+4.1.1 Ca fluxes affected by H O ..........................................................................65 2 2
2+4.1.2 The role of Ca in the hyperpolarization....................................68
4.2 Alteration of the excitability................................................................................69 Pouokam K.E.V. Table of contents iv
4.3 Mediation of H O effects, implication of thiol groups and 2 2
role of the kinases/phosphatases system............................................................71
4.4 Functional significance of the H O effects and 2 2
pathophysiological consequences........................................................................76
4.5 Conclusion..............................................................................................80
5 Summary.................................................................................................83
6 Zusammenfassung................................................................................85
7 References...............................................................................................87
8 Declaration...........................................................................................103
9 Acknowledgements...............................................................................................104
Publications..........................................................................................106




Pouokam K.E.V. List of Figures v
List of Figures

Figure 1.1A: Schematic drawing of the enteric nervous system showing the
muscle layers and the plexuses…………………………………………3

Figure 1.1B: Drawing depicting the communication between the ENS and
the parasympathetic and the sympathetic systems……………………..3

Figure 1.2: Electrophysiological properties and classification of enteric
neurones…………………………………………………………....…..6

Figure 1.3: Pharmacological properties and classification of myenteric
neurones…………………………………………………………….…7

Figure 1. 4 Molecular structure of voltage-gated sodium channels……….……..15

Figure 1.5A: Overview of regulation of calcium fluxes in cells…………………...20

Figure 1.5B: Regulation of calcium fluxes in neurones…………………………....20

Figure 2.1: Isolation of myenteric plexus………………………………………...31

Figure 2.2: Schematic representation of the patch-clamp
technique …………………………………………………………....34

Figure 2.3: Fast sodium inward current in neurones…………………………......34