Investigation of the subunit composition and the pharmacology of the mitochondrial ATP-dependent K+ channel in the brain

Zsombor Lacza, James A. Snipes, Béla Kis, Csaba Szabo, Gary Grover, David W. Busija

Research output: Contribution to journalArticle

74 Citations (Scopus)

Abstract

Selective activation of mitoKATP channels can protect the brain or cultured neurons against a variety of anoxic or metabolic challenges. However, little is known about the subunit composition or functional regulation of the channel itself. In the present study, we sought to characterize the mitoKATP channel in the mouse brain using overlapping approaches. First, we determined that mitochondria contain the pore-forming Kir6.1 and Kir6.2 subunits with Western blotting, immunogold electron microscopy and the identification of mitochondrial transport sequences. In contrast, we found no evidence for the presence of either known sulfonylurea receptors (SUR1 or SUR2) in the mitochondria. However, the ATP-dependent K (KATP) channel inhibitor glibenclamide specifically binds to mitochondria in both neurons and astrocytes, and small molecular weight SUR2-like proteins were concentrated in mitochondria. In addition to mice, similar results were found in rats and pigs. Second, live respiring mitochondria were stained with the membrane potential sensitive dye MitoFluorRed and visualized by confocal microscopy. We investigated the effects of pharmacological closing and opening of the channel with glibenclamide and the specific mitoKATP openers diazoxide and BMS-191095. Closing of the channel inhibited the energization of the mitochondria, which was reversed by the application of the mitoKATP openers. We also found that blocking mitochondrial peroxynitrite formation with FP15 has a similar effect to blocking the mitoKATP channels. We conclude that brain mitochondria contain functional KATP channels. The pore-forming subunit of the channel can be either Kir6.1 or Kir6.2, and the SUR subunit may be a SUR2 splice variant or a similar protein.

Original languageEnglish (US)
Pages (from-to)27-36
Number of pages10
JournalBrain Research
Volume994
Issue number1
DOIs
StatePublished - Dec 19 2003
Externally publishedYes

Fingerprint

Mitochondria
Pharmacology
Brain
KATP Channels
Glyburide
Sulfonylurea Receptors
Diazoxide
Neurons
Peroxynitrous Acid
mitochondrial K(ATP) channel
Confocal Microscopy
Astrocytes
Membrane Potentials
Electron Microscopy
Proteins
Coloring Agents
Swine
Adenosine Triphosphate
Molecular Weight
Western Blotting

Keywords

  • ATP-dependent potassium channel
  • Ischemic preconditioning
  • Mitochondrial nitric oxide synthase
  • mtNOS
  • Sulfonylurea receptor

ASJC Scopus subject areas

  • Neuroscience(all)

Cite this

Investigation of the subunit composition and the pharmacology of the mitochondrial ATP-dependent K+ channel in the brain. / Lacza, Zsombor; Snipes, James A.; Kis, Béla; Szabo, Csaba; Grover, Gary; Busija, David W.

In: Brain Research, Vol. 994, No. 1, 19.12.2003, p. 27-36.

Research output: Contribution to journalArticle

Lacza, Zsombor ; Snipes, James A. ; Kis, Béla ; Szabo, Csaba ; Grover, Gary ; Busija, David W. / Investigation of the subunit composition and the pharmacology of the mitochondrial ATP-dependent K+ channel in the brain. In: Brain Research. 2003 ; Vol. 994, No. 1. pp. 27-36.
@article{b5534fcf3c75414da0860e15a0137f01,
title = "Investigation of the subunit composition and the pharmacology of the mitochondrial ATP-dependent K+ channel in the brain",
abstract = "Selective activation of mitoKATP channels can protect the brain or cultured neurons against a variety of anoxic or metabolic challenges. However, little is known about the subunit composition or functional regulation of the channel itself. In the present study, we sought to characterize the mitoKATP channel in the mouse brain using overlapping approaches. First, we determined that mitochondria contain the pore-forming Kir6.1 and Kir6.2 subunits with Western blotting, immunogold electron microscopy and the identification of mitochondrial transport sequences. In contrast, we found no evidence for the presence of either known sulfonylurea receptors (SUR1 or SUR2) in the mitochondria. However, the ATP-dependent K (KATP) channel inhibitor glibenclamide specifically binds to mitochondria in both neurons and astrocytes, and small molecular weight SUR2-like proteins were concentrated in mitochondria. In addition to mice, similar results were found in rats and pigs. Second, live respiring mitochondria were stained with the membrane potential sensitive dye MitoFluorRed and visualized by confocal microscopy. We investigated the effects of pharmacological closing and opening of the channel with glibenclamide and the specific mitoKATP openers diazoxide and BMS-191095. Closing of the channel inhibited the energization of the mitochondria, which was reversed by the application of the mitoKATP openers. We also found that blocking mitochondrial peroxynitrite formation with FP15 has a similar effect to blocking the mitoKATP channels. We conclude that brain mitochondria contain functional KATP channels. The pore-forming subunit of the channel can be either Kir6.1 or Kir6.2, and the SUR subunit may be a SUR2 splice variant or a similar protein.",
keywords = "ATP-dependent potassium channel, Ischemic preconditioning, Mitochondrial nitric oxide synthase, mtNOS, Sulfonylurea receptor",
author = "Zsombor Lacza and Snipes, {James A.} and B{\'e}la Kis and Csaba Szabo and Gary Grover and Busija, {David W.}",
year = "2003",
month = "12",
day = "19",
doi = "10.1016/j.brainres.2003.09.046",
language = "English (US)",
volume = "994",
pages = "27--36",
journal = "Brain Research",
issn = "0006-8993",
publisher = "Elsevier",
number = "1",

}

TY - JOUR

T1 - Investigation of the subunit composition and the pharmacology of the mitochondrial ATP-dependent K+ channel in the brain

AU - Lacza, Zsombor

AU - Snipes, James A.

AU - Kis, Béla

AU - Szabo, Csaba

AU - Grover, Gary

AU - Busija, David W.

PY - 2003/12/19

Y1 - 2003/12/19

N2 - Selective activation of mitoKATP channels can protect the brain or cultured neurons against a variety of anoxic or metabolic challenges. However, little is known about the subunit composition or functional regulation of the channel itself. In the present study, we sought to characterize the mitoKATP channel in the mouse brain using overlapping approaches. First, we determined that mitochondria contain the pore-forming Kir6.1 and Kir6.2 subunits with Western blotting, immunogold electron microscopy and the identification of mitochondrial transport sequences. In contrast, we found no evidence for the presence of either known sulfonylurea receptors (SUR1 or SUR2) in the mitochondria. However, the ATP-dependent K (KATP) channel inhibitor glibenclamide specifically binds to mitochondria in both neurons and astrocytes, and small molecular weight SUR2-like proteins were concentrated in mitochondria. In addition to mice, similar results were found in rats and pigs. Second, live respiring mitochondria were stained with the membrane potential sensitive dye MitoFluorRed and visualized by confocal microscopy. We investigated the effects of pharmacological closing and opening of the channel with glibenclamide and the specific mitoKATP openers diazoxide and BMS-191095. Closing of the channel inhibited the energization of the mitochondria, which was reversed by the application of the mitoKATP openers. We also found that blocking mitochondrial peroxynitrite formation with FP15 has a similar effect to blocking the mitoKATP channels. We conclude that brain mitochondria contain functional KATP channels. The pore-forming subunit of the channel can be either Kir6.1 or Kir6.2, and the SUR subunit may be a SUR2 splice variant or a similar protein.

AB - Selective activation of mitoKATP channels can protect the brain or cultured neurons against a variety of anoxic or metabolic challenges. However, little is known about the subunit composition or functional regulation of the channel itself. In the present study, we sought to characterize the mitoKATP channel in the mouse brain using overlapping approaches. First, we determined that mitochondria contain the pore-forming Kir6.1 and Kir6.2 subunits with Western blotting, immunogold electron microscopy and the identification of mitochondrial transport sequences. In contrast, we found no evidence for the presence of either known sulfonylurea receptors (SUR1 or SUR2) in the mitochondria. However, the ATP-dependent K (KATP) channel inhibitor glibenclamide specifically binds to mitochondria in both neurons and astrocytes, and small molecular weight SUR2-like proteins were concentrated in mitochondria. In addition to mice, similar results were found in rats and pigs. Second, live respiring mitochondria were stained with the membrane potential sensitive dye MitoFluorRed and visualized by confocal microscopy. We investigated the effects of pharmacological closing and opening of the channel with glibenclamide and the specific mitoKATP openers diazoxide and BMS-191095. Closing of the channel inhibited the energization of the mitochondria, which was reversed by the application of the mitoKATP openers. We also found that blocking mitochondrial peroxynitrite formation with FP15 has a similar effect to blocking the mitoKATP channels. We conclude that brain mitochondria contain functional KATP channels. The pore-forming subunit of the channel can be either Kir6.1 or Kir6.2, and the SUR subunit may be a SUR2 splice variant or a similar protein.

KW - ATP-dependent potassium channel

KW - Ischemic preconditioning

KW - Mitochondrial nitric oxide synthase

KW - mtNOS

KW - Sulfonylurea receptor

UR - http://www.scopus.com/inward/record.url?scp=0344012088&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0344012088&partnerID=8YFLogxK

U2 - 10.1016/j.brainres.2003.09.046

DO - 10.1016/j.brainres.2003.09.046

M3 - Article

C2 - 14642445

AN - SCOPUS:0344012088

VL - 994

SP - 27

EP - 36

JO - Brain Research

JF - Brain Research

SN - 0006-8993

IS - 1

ER -