Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.

The properties of leaky patch currents in whole cell recording of HEK-293T cells were examined as a means to separate these control currents from expressed sodium and calcium leak channel currents from snail NALCN leak channels possessing both sodium (EKEE) and calcium (EEEE) selectivity filters. Le...

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Autores principales:	Adrienne N Boone, Adriano Senatore, Jean Chemin, Arnaud Monteil, J David Spafford
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2014
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/ede0ab7787d044dd8121c6319d466fea
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spelling	oai:doaj.org-article:ede0ab7787d044dd8121c6319d466fea2021-11-18T08:15:04ZGd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.1932-620310.1371/journal.pone.0098808https://doaj.org/article/ede0ab7787d044dd8121c6319d466fea2014-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/24945283/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203The properties of leaky patch currents in whole cell recording of HEK-293T cells were examined as a means to separate these control currents from expressed sodium and calcium leak channel currents from snail NALCN leak channels possessing both sodium (EKEE) and calcium (EEEE) selectivity filters. Leak currents were generated by the weakening of gigaohm patch seals by artificial membrane rupture using the ZAP function on the patch clamp amplifier. Surprisingly, we found that leak currents generated from the weakened membrane/glass seal can be surprisingly stable and exhibit behavior that is consistent with a sodium leak current derived from an expressible channel. Leaky patch currents differing by 10 fold in size were similarly reduced in size when external sodium ions were replaced with the large monovalent ion NMDG+. Leaky patch currents increased when external Ca2+ (1.2 mM) was lowered to 0.1 mM and were inhibited (>40% to >90%) with 10 µM Gd3+, 100 µM La3+, 1 mM Co2+ or 1 mM Cd2+. Leaky patch currents were relatively insensitive (<30%) to 1 mM Ni2+ and exhibited a variable amount of block with 1 mM verapamil and were insensitive to 100 µM mibefradil or 100 µM nifedipine. We hypothesize that the rapid changes in leak current size in response to changing external cations or drugs relates to their influences on the membrane seal adherence and the electro-osmotic flow of mobile cations channeling in crevices of a particular pore size in the interface between the negatively charged patch electrode and the lipid membrane. Observed sodium leak conductance currents in weak patch seals are reproducible between the electrode glass interface with cell membranes, artificial lipid or Sylgard rubber.Adrienne N BooneAdriano SenatoreJean CheminArnaud MonteilJ David SpaffordPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 9, Iss 6, p e98808 (2014)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Adrienne N Boone Adriano Senatore Jean Chemin Arnaud Monteil J David Spafford Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.
description	The properties of leaky patch currents in whole cell recording of HEK-293T cells were examined as a means to separate these control currents from expressed sodium and calcium leak channel currents from snail NALCN leak channels possessing both sodium (EKEE) and calcium (EEEE) selectivity filters. Leak currents were generated by the weakening of gigaohm patch seals by artificial membrane rupture using the ZAP function on the patch clamp amplifier. Surprisingly, we found that leak currents generated from the weakened membrane/glass seal can be surprisingly stable and exhibit behavior that is consistent with a sodium leak current derived from an expressible channel. Leaky patch currents differing by 10 fold in size were similarly reduced in size when external sodium ions were replaced with the large monovalent ion NMDG+. Leaky patch currents increased when external Ca2+ (1.2 mM) was lowered to 0.1 mM and were inhibited (>40% to >90%) with 10 µM Gd3+, 100 µM La3+, 1 mM Co2+ or 1 mM Cd2+. Leaky patch currents were relatively insensitive (<30%) to 1 mM Ni2+ and exhibited a variable amount of block with 1 mM verapamil and were insensitive to 100 µM mibefradil or 100 µM nifedipine. We hypothesize that the rapid changes in leak current size in response to changing external cations or drugs relates to their influences on the membrane seal adherence and the electro-osmotic flow of mobile cations channeling in crevices of a particular pore size in the interface between the negatively charged patch electrode and the lipid membrane. Observed sodium leak conductance currents in weak patch seals are reproducible between the electrode glass interface with cell membranes, artificial lipid or Sylgard rubber.
format	article
author	Adrienne N Boone Adriano Senatore Jean Chemin Arnaud Monteil J David Spafford
author_facet	Adrienne N Boone Adriano Senatore Jean Chemin Arnaud Monteil J David Spafford
author_sort	Adrienne N Boone
title	Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.
title_short	Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.
title_full	Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.
title_fullStr	Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.
title_full_unstemmed	Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.
title_sort	gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.
publisher	Public Library of Science (PLoS)
publishDate	2014
url	https://doaj.org/article/ede0ab7787d044dd8121c6319d466fea
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Gd3+ and calcium sensitive, sodium leak currents are features of weak membrane-glass seals in patch clamp recordings.

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