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Conference Paper: Patch-Clamp Study of Single Ryanodine Receptor Channels in the Outer Nuclear Membrane

TitlePatch-Clamp Study of Single Ryanodine Receptor Channels in the Outer Nuclear Membrane
Authors
Issue Date2012
Citation
The 2012 Hong Kong-Taiwan Physiology Symposium and Joint Scientific Meeting of Hong Kong Society of Neurosciences (HKSN) & The Biophysical Society of Hong Kong (BPHK), The Chinese University of Hong Kong, Hong Kong, China, 14-15 June 2012, p. 52, abstract no. P15 How to Cite?
AbstractModulation of cytoplasmic free calcium (Ca2+) concentration is a universal signaling pathway that regulates numerous cellular processes. Ubiquitous intracellular Ca2+ release channels – inositol 1,4,5-trisphosphate receptor (InsP3R) and ryanodine receptor (RyR) channels – localized in the sarco/endoplasmic reticulum (ER) play a central role in this pathway in all animal cells. Electrophysiological study of the single-channel conductance and gating properties of these Ca2+ release channels with conventional patch-clamp approach has been hindered by their intracellular localization. To overcome this limitation, patch-clamp electrophysiology has been applied on isolated nuclei where these Ca2+ release channels are found abundantly in the outer nuclear envelope. We have successfully uterlized this nuclear membrane electrophysiology to study the gating properties of single InsP3R channels in several cellular systems. Whereas, all the current single channel data, including channel conductance, permeation properties, and ligand regulation, of the RyR channels were done exclusively by reconstituting the channels into artificial planar lipid bilayers. To gain insights into the single channel properties of the RyR in its native membrane milieu, we applied nuclear membrane electrophysiological study on isolated nuclei from stable-inducible mouse RyR2 HEK-293 cells. Using potassium as charge carrier, caffeine activated single channel current with conductance of 750 pS in isolated nuclei. This caffeine activated current showed a linear current/voltage relationship under symmetrical ionic conditions and was sensitive to non-specific RyR inhibitor, ruthenium red. Furthermore, the single RyR channels recorded from the outer nuclear membrane exhibited bi-phasic Ca2+ regulation. In conclusion, we demonstrated, for the first time, that single RyR channels recordings from isolated nuclei and our results suggested that the nuclear membrane electrophysiology could be a sensitive and robust technique to study the gating properties of intracellular channels, including the InsP3R and RyR.
DescriptionPoster presentation
Persistent Identifierhttp://hdl.handle.net/10722/153111

 

DC FieldValueLanguage
dc.contributor.authorCheung, KHen_US
dc.contributor.authorTong, CKBen_US
dc.contributor.authorFoskett, JKen_US
dc.date.accessioned2012-07-16T09:57:06Z-
dc.date.available2012-07-16T09:57:06Z-
dc.date.issued2012en_US
dc.identifier.citationThe 2012 Hong Kong-Taiwan Physiology Symposium and Joint Scientific Meeting of Hong Kong Society of Neurosciences (HKSN) & The Biophysical Society of Hong Kong (BPHK), The Chinese University of Hong Kong, Hong Kong, China, 14-15 June 2012, p. 52, abstract no. P15en_US
dc.identifier.urihttp://hdl.handle.net/10722/153111-
dc.descriptionPoster presentation-
dc.description.abstractModulation of cytoplasmic free calcium (Ca2+) concentration is a universal signaling pathway that regulates numerous cellular processes. Ubiquitous intracellular Ca2+ release channels – inositol 1,4,5-trisphosphate receptor (InsP3R) and ryanodine receptor (RyR) channels – localized in the sarco/endoplasmic reticulum (ER) play a central role in this pathway in all animal cells. Electrophysiological study of the single-channel conductance and gating properties of these Ca2+ release channels with conventional patch-clamp approach has been hindered by their intracellular localization. To overcome this limitation, patch-clamp electrophysiology has been applied on isolated nuclei where these Ca2+ release channels are found abundantly in the outer nuclear envelope. We have successfully uterlized this nuclear membrane electrophysiology to study the gating properties of single InsP3R channels in several cellular systems. Whereas, all the current single channel data, including channel conductance, permeation properties, and ligand regulation, of the RyR channels were done exclusively by reconstituting the channels into artificial planar lipid bilayers. To gain insights into the single channel properties of the RyR in its native membrane milieu, we applied nuclear membrane electrophysiological study on isolated nuclei from stable-inducible mouse RyR2 HEK-293 cells. Using potassium as charge carrier, caffeine activated single channel current with conductance of 750 pS in isolated nuclei. This caffeine activated current showed a linear current/voltage relationship under symmetrical ionic conditions and was sensitive to non-specific RyR inhibitor, ruthenium red. Furthermore, the single RyR channels recorded from the outer nuclear membrane exhibited bi-phasic Ca2+ regulation. In conclusion, we demonstrated, for the first time, that single RyR channels recordings from isolated nuclei and our results suggested that the nuclear membrane electrophysiology could be a sensitive and robust technique to study the gating properties of intracellular channels, including the InsP3R and RyR.-
dc.languageengen_US
dc.relation.ispartofHong Kong-Taiwan Physiology Symposium and HKSN / BOHK Joint Scientific Meetingen_US
dc.rightsCreative Commons: Attribution 3.0 Hong Kong License-
dc.titlePatch-Clamp Study of Single Ryanodine Receptor Channels in the Outer Nuclear Membraneen_US
dc.typeConference_Paperen_US
dc.identifier.emailCheung, KH: ckingho@hku.hken_US
dc.identifier.authorityCheung, KH=rp01463en_US
dc.description.naturepublished_or_final_version-
dc.identifier.hkuros200823en_US
dc.identifier.spage52, abstract no. P15-
dc.identifier.epage52, abstract no. P15-

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