Charge immobilization caused by modification of internal cysteines in squid Na channels

Kamran Khodakhah, Alexey Melishchuk, Clay M. Armstrong

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

We studied the effects of modification of native cysteines present in squid giant axon Na channels with methanethiosulfonates. We find that intracellular, but not extracellular, perfusion of axons with positively charged [(2-trimethylammonium)-ethyl]methanethiosulfonate (MTSET), or 3(triethylammonium)propyl]methanethiosulfonate (MTS-PTrEA) irreversibly reduces sodium ionic (I(Na)) and gating (I(g)) currents. The rate of modification of Na channels was dependent on the concentration of the modifying agent and the transmembrane voltage. Hyperpolarized membrane potentials (e.g., -110 mV) protected the channels from modification by MTS- PTrEA. In addition to reducing the amplitudes of I(Na) and I(g), MTS-PTrEA also altered their kinetics such that the remaining/(Na) did not appear to inactivate, whereas I(g) was made sharper and declined to baseline more quickly. The shape and amplitude of I(g) after modification of channels with MTS-PTrEA appeared to be 'charge-immobilized,' as if the modified channels were inactivated. MTS-PTrEA did not affect/(Na) or I(g) when inactivation was removed by internal perfusion of the axon with pronase. In addition, we find that the steady-state inactivation curve of modified Na channels is made much shallower and is markedly shifted to hyperpolarized potentials. The rates of activation, deactivation, or open-state inactivation were not altered in MTS- PTrEA-modified channels. The uncharged sulfhydryl reagent methymethanethiosulfonate (MMTS) did not affect either I(Na) or I(g), but prevented the irreversible effects of MTS-PTrEA or MTSET on Na channels. It is proposed that the positively charged methanethiosulfonates MTS-PTrEA and MTSET modify a native internal cysteine(s) in squid Na channels, and by doing so promote inactivation from closed states, resulting in charge immobilization and reduction of I(Na).

Original languageEnglish (US)
Pages (from-to)2821-2829
Number of pages9
JournalBiophysical Journal
Volume75
Issue number6
StatePublished - Dec 1998
Externally publishedYes

Fingerprint

Decapodiformes
Immobilization
Cysteine
Axons
Perfusion
Sulfhydryl Reagents
Pronase
Membrane Potentials
Sodium

ASJC Scopus subject areas

  • Biophysics

Cite this

Charge immobilization caused by modification of internal cysteines in squid Na channels. / Khodakhah, Kamran; Melishchuk, Alexey; Armstrong, Clay M.

In: Biophysical Journal, Vol. 75, No. 6, 12.1998, p. 2821-2829.

Research output: Contribution to journalArticle

Khodakhah, Kamran ; Melishchuk, Alexey ; Armstrong, Clay M. / Charge immobilization caused by modification of internal cysteines in squid Na channels. In: Biophysical Journal. 1998 ; Vol. 75, No. 6. pp. 2821-2829.
@article{68e1e8be3c2c4d94ad2f278a3064c132,
title = "Charge immobilization caused by modification of internal cysteines in squid Na channels",
abstract = "We studied the effects of modification of native cysteines present in squid giant axon Na channels with methanethiosulfonates. We find that intracellular, but not extracellular, perfusion of axons with positively charged [(2-trimethylammonium)-ethyl]methanethiosulfonate (MTSET), or 3(triethylammonium)propyl]methanethiosulfonate (MTS-PTrEA) irreversibly reduces sodium ionic (I(Na)) and gating (I(g)) currents. The rate of modification of Na channels was dependent on the concentration of the modifying agent and the transmembrane voltage. Hyperpolarized membrane potentials (e.g., -110 mV) protected the channels from modification by MTS- PTrEA. In addition to reducing the amplitudes of I(Na) and I(g), MTS-PTrEA also altered their kinetics such that the remaining/(Na) did not appear to inactivate, whereas I(g) was made sharper and declined to baseline more quickly. The shape and amplitude of I(g) after modification of channels with MTS-PTrEA appeared to be 'charge-immobilized,' as if the modified channels were inactivated. MTS-PTrEA did not affect/(Na) or I(g) when inactivation was removed by internal perfusion of the axon with pronase. In addition, we find that the steady-state inactivation curve of modified Na channels is made much shallower and is markedly shifted to hyperpolarized potentials. The rates of activation, deactivation, or open-state inactivation were not altered in MTS- PTrEA-modified channels. The uncharged sulfhydryl reagent methymethanethiosulfonate (MMTS) did not affect either I(Na) or I(g), but prevented the irreversible effects of MTS-PTrEA or MTSET on Na channels. It is proposed that the positively charged methanethiosulfonates MTS-PTrEA and MTSET modify a native internal cysteine(s) in squid Na channels, and by doing so promote inactivation from closed states, resulting in charge immobilization and reduction of I(Na).",
author = "Kamran Khodakhah and Alexey Melishchuk and Armstrong, {Clay M.}",
year = "1998",
month = "12",
language = "English (US)",
volume = "75",
pages = "2821--2829",
journal = "Biophysical Journal",
issn = "0006-3495",
publisher = "Biophysical Society",
number = "6",

}

TY - JOUR

T1 - Charge immobilization caused by modification of internal cysteines in squid Na channels

AU - Khodakhah, Kamran

AU - Melishchuk, Alexey

AU - Armstrong, Clay M.

PY - 1998/12

Y1 - 1998/12

N2 - We studied the effects of modification of native cysteines present in squid giant axon Na channels with methanethiosulfonates. We find that intracellular, but not extracellular, perfusion of axons with positively charged [(2-trimethylammonium)-ethyl]methanethiosulfonate (MTSET), or 3(triethylammonium)propyl]methanethiosulfonate (MTS-PTrEA) irreversibly reduces sodium ionic (I(Na)) and gating (I(g)) currents. The rate of modification of Na channels was dependent on the concentration of the modifying agent and the transmembrane voltage. Hyperpolarized membrane potentials (e.g., -110 mV) protected the channels from modification by MTS- PTrEA. In addition to reducing the amplitudes of I(Na) and I(g), MTS-PTrEA also altered their kinetics such that the remaining/(Na) did not appear to inactivate, whereas I(g) was made sharper and declined to baseline more quickly. The shape and amplitude of I(g) after modification of channels with MTS-PTrEA appeared to be 'charge-immobilized,' as if the modified channels were inactivated. MTS-PTrEA did not affect/(Na) or I(g) when inactivation was removed by internal perfusion of the axon with pronase. In addition, we find that the steady-state inactivation curve of modified Na channels is made much shallower and is markedly shifted to hyperpolarized potentials. The rates of activation, deactivation, or open-state inactivation were not altered in MTS- PTrEA-modified channels. The uncharged sulfhydryl reagent methymethanethiosulfonate (MMTS) did not affect either I(Na) or I(g), but prevented the irreversible effects of MTS-PTrEA or MTSET on Na channels. It is proposed that the positively charged methanethiosulfonates MTS-PTrEA and MTSET modify a native internal cysteine(s) in squid Na channels, and by doing so promote inactivation from closed states, resulting in charge immobilization and reduction of I(Na).

AB - We studied the effects of modification of native cysteines present in squid giant axon Na channels with methanethiosulfonates. We find that intracellular, but not extracellular, perfusion of axons with positively charged [(2-trimethylammonium)-ethyl]methanethiosulfonate (MTSET), or 3(triethylammonium)propyl]methanethiosulfonate (MTS-PTrEA) irreversibly reduces sodium ionic (I(Na)) and gating (I(g)) currents. The rate of modification of Na channels was dependent on the concentration of the modifying agent and the transmembrane voltage. Hyperpolarized membrane potentials (e.g., -110 mV) protected the channels from modification by MTS- PTrEA. In addition to reducing the amplitudes of I(Na) and I(g), MTS-PTrEA also altered their kinetics such that the remaining/(Na) did not appear to inactivate, whereas I(g) was made sharper and declined to baseline more quickly. The shape and amplitude of I(g) after modification of channels with MTS-PTrEA appeared to be 'charge-immobilized,' as if the modified channels were inactivated. MTS-PTrEA did not affect/(Na) or I(g) when inactivation was removed by internal perfusion of the axon with pronase. In addition, we find that the steady-state inactivation curve of modified Na channels is made much shallower and is markedly shifted to hyperpolarized potentials. The rates of activation, deactivation, or open-state inactivation were not altered in MTS- PTrEA-modified channels. The uncharged sulfhydryl reagent methymethanethiosulfonate (MMTS) did not affect either I(Na) or I(g), but prevented the irreversible effects of MTS-PTrEA or MTSET on Na channels. It is proposed that the positively charged methanethiosulfonates MTS-PTrEA and MTSET modify a native internal cysteine(s) in squid Na channels, and by doing so promote inactivation from closed states, resulting in charge immobilization and reduction of I(Na).

UR - http://www.scopus.com/inward/record.url?scp=0031736454&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0031736454&partnerID=8YFLogxK

M3 - Article

C2 - 9826604

AN - SCOPUS:0031736454

VL - 75

SP - 2821

EP - 2829

JO - Biophysical Journal

JF - Biophysical Journal

SN - 0006-3495

IS - 6

ER -