Evidence for an intracellular localization of the adenosine A2B receptor in rat cardiomyocytes.
Protection achieved by ischemic preconditioning is dependent on A(2B) adenosine receptors (A(2B)AR) in rabbit and mouse hearts and, predictably, an A(2B)AR agonist protects them. But it is controversial whether cardiomyocytes themselves actually express A(2B)AR. The present study tested whether A(2B)AR could be demonstrated on rat cardiomyocytes. Isolated rat hearts experienced 30 min of ischemia and 120 min of reperfusion. The highly selective, cell-permeant A(2B)AR agonist BAY60-6583 (500 nM) infused at reperfusion reduced infarct size from 40.4 ± 2.0% of the risk zone in control hearts to 19.9 ± 2.8% indicating that A(2B)AR are protective in rat heart as well. Furthermore, BAY60-6583 reduced calcium-induced mitochondrial permeability transition in isolated rat cardiomyocytes. A(2B)AR protein could be demonstrated in isolated cardiomyocytes by western blotting. In addition, message for A(2B)AR was found in individual cardiomyocytes using quantitative RT-PCR. Surprisingly, immunofluorescence microscopy did not show A(2B)AR on the cardiomyocyte's sarcolemma but rather at intracellular sites. Co-staining with MitoTracker Red in isolated cardiomyocytes revealed A(2B)AR are localized to mitochondria. Western blot analysis of a mitochondrial fraction from either rat heart biopsies or isolated cardiomyocytes revealed a strong A(2B)AR band. Thus, the present study demonstrates that activation of A(2B)AR is strongly cardioprotective in rat heart and suppresses transition pores in isolated cardiomyocytes, and A(2B)AR are expressed in individual cardiomyocytes. However, surprisingly, A(2B)AR are present in or near mitochondria rather than on the sarcolemma as are other adenosine receptors. Because A(2B)AR signaling is thought to result in inhibition of mitochondrial transition pores, this convenient location may be important.
Department of Cardiology, Ernst-Moritz-Arndt University, Greifswald, Germany.
Karina
K
Grube
Julia
J
Rüdebusch
Zhelong
Z
Xu
Thomas
T
Böckenholt
Carmen
C
Methner
Tobias
T
Müller
Friederike
F
Cuello
Katrin
K
Zimmermann
Xiulan
X
Yang
Stephan B
SB
Felix
Michael V
MV
Cohen
James M
JM
Downey
Thomas
T
Krieg
D000818:Animals; D015153:Blotting, Western; D005260:Female; D005455:Fluorescent Antibody Technique; D008297:Male; D053078:Membrane Potential, Mitochondrial / Q000502:physiology; D008928:Mitochondria / Q000378:metabolism*; D015428:Myocardial Reperfusion Injury / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D043704:Receptor, Adenosine A2B / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D043704:Receptor, Adenosine A2B
Circ Res. 2009 Jan 2;104(1):15-8
19038864
J Mol Cell Cardiol. 2010 Nov;49(5):886-93
20797398
FEBS Lett. 2006 Oct 16;580(24):5690-6
16996504
J Biochem. 1995 Oct;118(4):683-5
8576078
FEBS Lett. 2007 Dec 22;581(30):5765-8
18037379
Circ Res. 2001 Aug 3;89(3):273-8
11485978
Cell Metab. 2009 Mar;9(3):265-76
19254571
Biochem Pharmacol. 2001 Mar 15;61(6):657-63
11266650
Cardiovasc Res. 2005 Aug 1;67(2):234-44
15919068
J Mol Biol. 2000 Jul 21;300(4):1005-16
10891285
J Neurochem. 2003 Dec;87(6):1427-35
14713298
Am J Physiol. 1995 May;268(5 Pt 2):H1862-8
7771537
Mol Cell Endocrinol. 2009 Mar 5;300(1-2):37-42
19007846
Cardiovasc Res. 2006 Jul 1;71(1):69-78
16631628
J Biol Chem. 2010 Dec 10;285(50):39271-88
20926384
Am J Physiol Heart Circ Physiol. 2006 Jan;290(1):H441-9
16155103
Basic Res Cardiol. 2010 Nov;105(6):771-85
20960209
J Biol Chem. 2004 Jun 11;279(24):25219-25
15070903
Basic Res Cardiol. 2006 Jul;101(4):319-26
16604438
Cardiovasc Res. 2006 May 1;70(2):308-14
16545350
Br J Pharmacol. 2008 May;154(1):25-31
18332860
Biochemistry. 2000 Oct 24;39(42):12828-36
11041847
Br J Pharmacol. 1995 Jun;115(4):648-52
7582485
Biochim Biophys Acta. 2008 Jan;1783(1):1-11
18062929
Circ Res. 1995 Feb;76(2):242-51
7834835
J Cell Biol. 1967 Mar;32(3):719-35
4291912
Proc Natl Acad Sci U S A. 2001 Nov 20;98(24):14126-31
11717466
J Clin Invest. 2010 May;120(5):1441-53
20364086
Am J Physiol Heart Circ Physiol. 2002 Feb;282(2):H437-44
11788390
Basic Res Cardiol. 2010 Mar;105(2):151-4
20066536
Circulation. 2007 Mar 27;115(12):1581-90
17353435
Mol Pharmacol. 1995 Jun;47(6):1126-32
7603451
J Clin Invest. 2006 Jul;116(7):1913-23
16823489
J Mol Cell Cardiol. 2007 Sep;43(3):262-71
17632123
Biochim Biophys Acta. 2009 Jan;1793(1):33-41
18672008
J Mol Cell Cardiol. 2009 Nov;47(5):684-90
19695259
Basic Res Cardiol. 2003 Jul;98(4):235-41
12835952
Am J Physiol Renal Physiol. 2002 Jul;283(1):F41-51
12060585
Circulation. 2008 Nov 4;118(19):1915-9
18981312
Mol Pharmacol. 1999 Oct;56(4):705-13
10496952
J Cell Biochem. 2006 Nov 1;99(4):1165-74
16795051
J Mol Cell Cardiol. 1982 Jul;14(7):397-412
7175947
J Biol Chem. 2001 Aug 10;276(32):30199-207
11333255
Basic Res Cardiol. 2009 Mar;104(2):141-7
19242638
J Cardiovasc Pharmacol. 2005 Dec;46(6):794-802
16306804
Cardiovasc Res. 2004 Feb 15;61(3):448-60
14962476