The morphology and electrophysiology of leech neurones in vitro: A study using microfabricated devices

Wilson, Richard James Alfred (1992) The morphology and electrophysiology of leech neurones in vitro: A study using microfabricated devices. PhD thesis, University of Glasgow.

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1. This thesis considers the possibility of investigating neuronal information processing by making mulitisite recordings either from individual isolated neurones or from small neuronal networks of controlled design. 2. Isolated leech neurones were used (a) to study the effects of isolation on electrophysiological properties (Chapter 2); (b) to provide the first demonstration of the use of planar extracellular electrode arrays to measure electrophysiological properties of individual isolated neurones (Chapter 3); and (c) to investigate the use of topographical features for controlling the outgrowth of neurones in culture (Chapters 4 & 5). 3.The study into the effects of isolation confirmed and extended previous reports which showed that (a) the action potentials recorded from the cell body of isolated Retzius neurones are generally similar to action potentials recorded in vivo and (b) isolation causes an increase in input resistance. 4. The results of an experiment designed to show a correlation between input resistance and the length of processes, suggest that the removal of processes during extraction is not the main cause of the high input resistance of isolated cells. One possibility is that the input resistance is a direct result of a change in membrane properties. 5. This conclusion is supported by the demonstration in isolated neurones of a slow inward transient (known as anomolous rectification) that occurs shortly after the onset of large hyperpolarising current injections. This transient was not observed in dissected ganglia and has not been previously reported in leech neurones. 6. The electrophysiology of isolated nevirones was also explored using extracellular electrode arrays. Specifically the electrode arrays were used to show that: (a) the conduction velocity of action potentials in P cells is faster than that of action potentials in Retzius cells (which correlates with the difference in rise time); (b) the action potentials of isolated neurones propagate from the tip of the extracted process towards the cell body (indicating that the concentration of Na+ channels is greater at the tip of the extracted process than in other regions); and (c) the action potential in the extracted process may have a faster rise time than that in the cell body, suggesting a difference in the average concentration of Na+ channels. 7. The use of extracellular electrode arrays in the above experiments demonstrates: (a) the first multisite extracellular recordings of isolated neurones; (b) the first extracellular recordings made of the electrical activity that results from extracellular stimulation of the same cell; and (c) the feasibility of using these devices to investigate information processing in single cells. 8. In order to control the morphology of single neurones, and the connectivity of groups of neurones, the influence of the substratum in determining the morphology of cultured neurones was investigated. The principal result demonstrates for the first time that topography can influence the outgrowth morphology of large identified invertebrate neurones. The dimensions of the topographical features were similar to those required to align the neurites of much smaller vertebrate neurones. 9. The results also show that: (a) on planar Con A-coated substrata the outgrowth of Retzius neurones tends to be dominated by large lamellae, whereas on leech extracellular matrix (ECM)-coated substrata cells produced an elaborate network of neurites (confirming previous reports), (b) interference reflection microscopy (IRM) revealled that the neurites made a series of very close but intermittent contacts with the substratum, whereas the lamella was characterised by a large region of uniform close contact, and (c) whereas neurites were strongly influenced by topographical features the lamellar-type outgrowth was only partially aligned. 10. Based on these results a new hypothesis (the Topographical Modulating Hypothesis) is presented. The hypothesis proposes that the influence of topographical discontinuities in determining the morphology of neurones, or the orientation of migrating cells, is modulated by the molecular nature of the substrate.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Keywords: Physiology, morphology.
Colleges/Schools: College of Medical Veterinary and Life Sciences
Supervisor's Name: Blackshaw, Dr. Susanna and Dow, Dr. Julian
Date of Award: 1992
Depositing User: Enlighten Team
Unique ID: glathesis:1992-72974
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 11 Jun 2019 11:06
Last Modified: 20 Jul 2021 11:54

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