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Achieving Maximal Speed of Solution Exchange for Patch Clamp Experiments

BACKGROUND: Resolving the kinetics of agonist binding events separately from the subsequent channel gating processes requires the ability of applying and removing the agonist before channel gating occurs. No reported system has yet achieved pulses shorter than 100 µs, necessary to study nicotinic AC...

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Autores principales: Auzmendi, Jerónimo, Fernández Do Porto, Darío, Pallavicini, Carla, Moffatt, Luciano
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3411769/
https://www.ncbi.nlm.nih.gov/pubmed/22879927
http://dx.doi.org/10.1371/journal.pone.0042275
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author Auzmendi, Jerónimo
Fernández Do Porto, Darío
Pallavicini, Carla
Moffatt, Luciano
author_facet Auzmendi, Jerónimo
Fernández Do Porto, Darío
Pallavicini, Carla
Moffatt, Luciano
author_sort Auzmendi, Jerónimo
collection PubMed
description BACKGROUND: Resolving the kinetics of agonist binding events separately from the subsequent channel gating processes requires the ability of applying and removing the agonist before channel gating occurs. No reported system has yet achieved pulses shorter than 100 µs, necessary to study nicotinic ACh receptor or AMPA receptor activation. METHODOLOGY/PRINCIPAL FINDINGS: Solution exchange systems deliver short agonist pulses by moving a sharp interface between a control and an experimental solution across a channel preparation. We achieved shorter pulses by means of an exchange system that combines a faster flow velocity, narrower partition between the two streams, and increased velocity and bandwidth of the movement of the interface. The measured response of the entire system was fed back to optimize the voltage signal applied to the piezoelectric actuator overcoming the spurious oscillations arising from the mechanical resonances when a high bandwidth driving function was applied. Optimization was accomplished by analyzing the transfer function of the solution exchange system. When driven by optimized command pulses the enhanced system provided pulses lasting 26 ± 1 µs and exchanging 93 ± 1% of the solution, as measured in the open tip of a patch pipette. CONCLUSIONS/SIGNIFICANCE: Pulses of this duration open the experimental study of the molecular events that occur between the agonist binding and the opening of the channel.
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spelling pubmed-34117692012-08-09 Achieving Maximal Speed of Solution Exchange for Patch Clamp Experiments Auzmendi, Jerónimo Fernández Do Porto, Darío Pallavicini, Carla Moffatt, Luciano PLoS One Research Article BACKGROUND: Resolving the kinetics of agonist binding events separately from the subsequent channel gating processes requires the ability of applying and removing the agonist before channel gating occurs. No reported system has yet achieved pulses shorter than 100 µs, necessary to study nicotinic ACh receptor or AMPA receptor activation. METHODOLOGY/PRINCIPAL FINDINGS: Solution exchange systems deliver short agonist pulses by moving a sharp interface between a control and an experimental solution across a channel preparation. We achieved shorter pulses by means of an exchange system that combines a faster flow velocity, narrower partition between the two streams, and increased velocity and bandwidth of the movement of the interface. The measured response of the entire system was fed back to optimize the voltage signal applied to the piezoelectric actuator overcoming the spurious oscillations arising from the mechanical resonances when a high bandwidth driving function was applied. Optimization was accomplished by analyzing the transfer function of the solution exchange system. When driven by optimized command pulses the enhanced system provided pulses lasting 26 ± 1 µs and exchanging 93 ± 1% of the solution, as measured in the open tip of a patch pipette. CONCLUSIONS/SIGNIFICANCE: Pulses of this duration open the experimental study of the molecular events that occur between the agonist binding and the opening of the channel. Public Library of Science 2012-08-03 /pmc/articles/PMC3411769/ /pubmed/22879927 http://dx.doi.org/10.1371/journal.pone.0042275 Text en © 2012 Auzmendi et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Auzmendi, Jerónimo
Fernández Do Porto, Darío
Pallavicini, Carla
Moffatt, Luciano
Achieving Maximal Speed of Solution Exchange for Patch Clamp Experiments
title Achieving Maximal Speed of Solution Exchange for Patch Clamp Experiments
title_full Achieving Maximal Speed of Solution Exchange for Patch Clamp Experiments
title_fullStr Achieving Maximal Speed of Solution Exchange for Patch Clamp Experiments
title_full_unstemmed Achieving Maximal Speed of Solution Exchange for Patch Clamp Experiments
title_short Achieving Maximal Speed of Solution Exchange for Patch Clamp Experiments
title_sort achieving maximal speed of solution exchange for patch clamp experiments
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3411769/
https://www.ncbi.nlm.nih.gov/pubmed/22879927
http://dx.doi.org/10.1371/journal.pone.0042275
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