Calcium influx through presynaptic calcium channels triggers transmitter release, but many of the details that underlie calcium-triggered secretion are not well understood. In an attempt to increase our understanding of this process, synaptic transmission at the frog neuromuscular junction has been investigated using physiological experiments and computational modeling. Pharmacological manipulations ((R)roscovitine and DAP) were used as tools to modulate presynaptic calcium influx and study effects on transmitter release. I showed that (R)-roscovitine predominately slowed deactivation kinetics of calcium current (by 427%), and as a result, increased the integral of calcium channel current evoked by a physiological action potential waveform (by 44%). (R)-roscovitine also increased the quantal content of acetylcholine released from the motor nerve terminals (by 149%) without changing paired-pulse facilitation under low calcium conditions. In contrast, exposure to 3,4-diaminopyridine (which affects transmitter release evoked by partially blocking potassium channels, altering the amplitude of the presynaptic action potential, and indirectly increasing calcium entry) increased paired-pulse facilitation (by 23%). In normal calcium conditions, both pharmacological treatments showed relatively similar effects on paired-pulse facilitation. I used a computational model, constrained by previous reports in the literature and my physiological measurements, to simulate my experimental data. This model faithfully reproduced calcium current with a single action potential, the average number of released synaptic vesicles, and the effects of (R)-roscovitine and DAP on calcium influx and vesicle release. Using this model, I made several predictions about the mechanisms underlying transmitter release. First, calcium ions originating from one or two voltage-gated calcium channels most often contributed to cause the fusion of each vesicle. Second, the calcium channel closest to a vesicle that fuses, provides 77% of calcium ions. My simulation of paired-pulse facilitation using the present model needed more adjustments, and in the process of adjusting the model parameters, various hypotheses that might explain observed short-term synaptic plasticity, including the effects of changes in buffer conditions, the effects of uneven calcium channel distribution, reducing terminal volume by adding vesicles to a storage pool, changes in the second action potential waveform, and possible persistent changes in vesicle release machinery were explored.
| Identifer | oai:union.ndltd.org:PITT/oai:PITTETD:etd-08092007-204525 |
| Date | 20 September 2007 |
| Creators | Cho, Soyoun |
| Contributors | Nathaniel N. Urban, German Barrionuevo, Guo-Qiang Bi, Karl Kandler, Joel R. Stiles, Harold L. Atwood, Stephen D. Meriney |
| Publisher | University of Pittsburgh |
| Source Sets | University of Pittsburgh |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.pitt.edu/ETD/available/etd-08092007-204525/ |
| Rights | restricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to University of Pittsburgh or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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