Disruption of sulfonylurea receptor-2 protects against ischemia and reperfusion injury via...

Disruption of sulfonylurea receptor-2 protects against ischemia and

reperfusion injury via modulation of mitochondrial bioenergetic phenotype

Pravdić D.1,3, Aggarwal N.2, Mcnally M. E.4, Bošnjak Ž.3, Nian-Qing Shi2, Makielski J.2

Mostar School of Medicine, Mostar, BIH 1

University of Wisconsin, Madison, Madison, WI, USA 2

Medical College of Wisconsin, Milwaukee, WI, USA 3

University of Chicago, Chicago, IL, USA 4

• First discovered in the heart by Noma (Nature 1983; 305: 147-8).

• “An outward current of unknown nature increases significantly when cardiac cells are treated with cyanide or subjected to hypoxia, and decreases on intracellular injection of ATP.”

• ATP-regulated K+ channels

KATP Channels

• Following studies identified KATP channels in

other tissues:

• Pancreatic β-cells

• Vascular smooth muscle

• Skeletal muscle

• Brain

• Kidney

• etc.

KATP Channels

Cardiac sarcKATP Channels

• Under normal metabolic conditions closed.

• During metabolic stress they open - cellular metabolic sensors.

SUR2A Kir6.2

COO-

NH2

ATP ADPH+

-+

Cardiac KATP Channels

Mitochondrial KATP channel

K+

K+

K+

K+

Sarcolemmal KATP

channel

K+ K+K+ K+

K+

K+

K+

The Mechanism of Protection

Ca2+

Extracellular

K+

SarcKATP Channel

MitoKATP channels and cellular protection

• MitoKATP channel activator

diazoxide reduced the

severity of ischemia/

reperfusion damage in rat

hearts (Garlid et al., Circ Res

1997;81:1072-82)Fig. 4

Vehicle Diazo Diazo+5-HD

• Diazoxide protected rabbit

cardiomyocytes from ischemia

in a 5-HD dependent manner (Liu et al., Circulation 1998;97:2463-9)

• Suppression of mitochondrial Ca2+ overload

Proposed mechanisms of protection

ΔΨm -200 mV ΔΨm depolarized

Ca2+ Ca2+

Mitochondrial K+ channels openCa2+ Ca2+

K+

• Mitochondrial swelling and improved oxidative phosphorylation

ATPMitochondrial K+ channels open

H+ A-

Pi-

K+

K+

OH-

• Freshly isolated cardiac myocytes

– Effect on mitochondrial membrane potential (ΔΨm)

• Isolated cardiac mitochondria

– Mitochondrial bioenergetics

– Ca2+ loading

– Resistance to stress

Experimental Design

Animals

• A transgenic mouse was created previously (SUR2 mutant) where the gene encoding SUR2 was disrupted by removing exons 14 to 18 encoding the first nucleotide binding domain.

• These SUR2 mutant mice lack pinacidil, diazoxide and glybenclamide sensitive sarcKATP currents in the cardiac, smooth and skeletal muscle and they are hypertensive, arrhythmic, and exhibit coronary vasospasm and sudden cardiac death

• SUR2 mutant mice have increased protection against both acute adrenergic stress and ischemia compared to control

Stoller D, Kakkar R, Smelley M, Chalupsky K, Earley JU, Shi NQ, Makielski JC, McNally EM.

Mice lacking sulfonylurea receptor 2 (SUR2) ATP-sensitive potassium channels

are resistant to acute cardiovascular stress. J Mol Cell Cardiol. 2007;43:445-454.

Mitochondrial membrane potential

Mitochondrial swelling

Resistance to Ca2+ loading

Effect of hypoxia-reoxygenation on mitochondrial respiration

Myocite Resistance to metabolic inhibition

Conclusions

• SUR2 mutant mitochondria had more depolarized ΔΨm compared to wild type

• Tolerance to Ca2+ loading was increased in SUR2 mutant mitochondria

• Mitochondria swelling, an indicator of K+ influx, was greater in SUR2 mutants

• SUR2 mutant mitochondria recovered better from hypoxia-reoxygenation injury than Wild type

Acknowledgements

Zeljko J. Bosnjak, PhD

Nitin Aggarwal, PhD

Martin Bienengraeber, PhD

Jonatan Makielski, MD

Elizabeth McNally, MD

Chiaki Kwok