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Voltage-dependent gating and block by internal spermine of the murine inwardly rectifying K+ channel, Kir2.1.
J Physiol. 2003 Apr 15; 548(Pt 2):361-71.JP

Abstract

The mechanism of inward rectification was investigated by recording single-channel currents through an inwardly rectifying K+ channel (Kir2.1). cDNA encoding a wild-type (WT) channel, a mutant replacing Asp 172 with Asn (D172N), and a tandem tetramer WT-(D172N)2-WT, was transfected into COS-1 cells using the liposome method, and after 48-72 h single-channel currents were recorded in the inside-out configuration at 150 mM internal and external K+. Steady-state open probability of outward currents decreased with larger depolarizations. The activation curve was fitted with a single Boltzmann equation. The voltages of half-activation in the absence of spermine were +35.9 mV (WT), +55.0 mV (WT-(D172N)2-WT) and +76.7 mV (D172N). Open-time and zero-current-time histograms were constructed. The open-time histogram was fitted with a single exponential function. Two exponential functions were necessary to fit the closed-time histogram. In each channel, internal spermine at a concentration of 1-100 nM reduced the open time of the outward currents in a concentration-dependent manner and produced one blocked state without affecting the inward currents, suggesting that spermine acts as an open channel blocker. The normalized steady-state open probability-spermine concentration curve was fitted by saturation kinetics with a Hill coefficient of 1. On the assumption of the linear sequential state model, the unblock and blocking rates were estimated in each channel. Unblock rates depended on the number of D172N mutant subunits, but blocking rates did not. The results suggest that closing gates work independently of the spermine block and D172 is involved in both intrinsic gating and the spermine block.

Authors+Show Affiliations

Department of Physiology, Kansai Medical University, Moriguchi, Osaka, Japan. matsudah@takii.kmu.ac.jpNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

12640008

Citation

Matsuda, Hiroko, et al. "Voltage-dependent Gating and Block By Internal Spermine of the Murine Inwardly Rectifying K+ Channel, Kir2.1." The Journal of Physiology, vol. 548, no. Pt 2, 2003, pp. 361-71.
Matsuda H, Oishi K, Omori K. Voltage-dependent gating and block by internal spermine of the murine inwardly rectifying K+ channel, Kir2.1. J Physiol. 2003;548(Pt 2):361-71.
Matsuda, H., Oishi, K., & Omori, K. (2003). Voltage-dependent gating and block by internal spermine of the murine inwardly rectifying K+ channel, Kir2.1. The Journal of Physiology, 548(Pt 2), 361-71.
Matsuda H, Oishi K, Omori K. Voltage-dependent Gating and Block By Internal Spermine of the Murine Inwardly Rectifying K+ Channel, Kir2.1. J Physiol. 2003 Apr 15;548(Pt 2):361-71. PubMed PMID: 12640008.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Voltage-dependent gating and block by internal spermine of the murine inwardly rectifying K+ channel, Kir2.1. AU - Matsuda,Hiroko, AU - Oishi,Keiko, AU - Omori,Koichiro, Y1 - 2003/03/14/ PY - 2003/3/18/pubmed PY - 2003/12/5/medline PY - 2003/3/18/entrez SP - 361 EP - 71 JF - The Journal of physiology JO - J Physiol VL - 548 IS - Pt 2 N2 - The mechanism of inward rectification was investigated by recording single-channel currents through an inwardly rectifying K+ channel (Kir2.1). cDNA encoding a wild-type (WT) channel, a mutant replacing Asp 172 with Asn (D172N), and a tandem tetramer WT-(D172N)2-WT, was transfected into COS-1 cells using the liposome method, and after 48-72 h single-channel currents were recorded in the inside-out configuration at 150 mM internal and external K+. Steady-state open probability of outward currents decreased with larger depolarizations. The activation curve was fitted with a single Boltzmann equation. The voltages of half-activation in the absence of spermine were +35.9 mV (WT), +55.0 mV (WT-(D172N)2-WT) and +76.7 mV (D172N). Open-time and zero-current-time histograms were constructed. The open-time histogram was fitted with a single exponential function. Two exponential functions were necessary to fit the closed-time histogram. In each channel, internal spermine at a concentration of 1-100 nM reduced the open time of the outward currents in a concentration-dependent manner and produced one blocked state without affecting the inward currents, suggesting that spermine acts as an open channel blocker. The normalized steady-state open probability-spermine concentration curve was fitted by saturation kinetics with a Hill coefficient of 1. On the assumption of the linear sequential state model, the unblock and blocking rates were estimated in each channel. Unblock rates depended on the number of D172N mutant subunits, but blocking rates did not. The results suggest that closing gates work independently of the spermine block and D172 is involved in both intrinsic gating and the spermine block. SN - 0022-3751 UR - https://www.unboundmedicine.com/medline/citation/12640008/Voltage_dependent_gating_and_block_by_internal_spermine_of_the_murine_inwardly_rectifying_K+_channel_Kir2_1_ L2 - https://doi.org/10.1113/jphysiol.2003.038844 DB - PRIME DP - Unbound Medicine ER -