We investigated the effect of A(1) adenosine receptor overexpression, which has been reported to increase myocardial tolerance to ischemia-reperfusion injury, on sarcoplasmic reticulum (SR) Ca(2+) ...handling.
Transgenic mouse hearts (approximately 300-fold A(1) adenosine receptor overexpression) and wild-type mouse hearts were perfused in the Langendorff mode and subjected either to 80 min of aerobic perfusion or to 30 min of aerobic perfusion, 20 min of global ischemia and 30 min of reperfusion. The hearts were then homogenized and used to assay SR oxalate-supported 45Ca(2+) uptake and 3H-ryanodine binding.
Transgenic hearts showed increased resistance to ischemia-reperfusion, as shown by lower diastolic tension (1.5 +/- 0.2 vs. 2.6 +/- 0.1 g, P<0.05) and higher recovery of developed tension (45 +/- 3 vs. 30 +/- 4% of the baseline, P<0.05) following ischemia-reperfusion. Under baseline conditions, oxalate-supported 45Ca(2+) uptake was lower in transgenic hearts, owing to reduced V(max) (10.6 +/- 2.0 vs. 17.8 +/- 2.7 nmol/min per mg of protein, P<0.05), and the difference was preserved after ischemia-reperfusion (10.0 +/- 1.0 vs. 15.7 +/- 2.5 nmol/min per mg of protein, P<0.05). No significant difference in 3H-ryanodine binding was observed.
A(1) adenosine receptor overexpression is associated with a decreased rate of active Ca(2+) transport into the SR. We hypothesize that changes in SR function may cause a depletion of the SR Ca(2+) pool, which might protect from ischemic injury by delaying the development of cytosolic Ca(2+) overload during ischemia.
The aim of the present study was to assess the effects of A(1)-adenosine receptor (A1-AR) stimulation in ventricle of A(1)-adenosine receptor overexpressing mice (transgenic mice, TG).
Effects of the ...A(1)-adenosine receptor agonist R-PIA ((-)-N(6)-phenylisopropyladenosine) on phosphorylation of phospholamban (PLB), Ca(2+) transients, Ca(2+) currents and cell shortening were studied in isolated ventricular cardiomyocytes.
R-PIA alone did not affect contractility in isolated electrically stimulated cardiomyocytes from wild-type mice (WT) or TG. However, after pre-stimulation of beta-adrenoceptors by isoproterenol, R-PIA reduced contractility in cardiomyocytes from WT but increased contractility in TG. Under the same conditions, R-PIA reduced isoproterenol-stimulated currents through L-type Ca(2+) channels, Ca(2+) transients and phosphorylation of PLB in cardiomyocytes from WT. In contrast, R-PIA diminished phospholamban phosphorylation induced by isoproterenol but augmented isoproterenol-elevated currents through L-type Ca(2+) channels, and isoproterenol-heightened Ca(2+) transients in cardiomyocytes from TG.
We suggest that A(1)-adenosine receptor overexpression reverses the interaction of beta-adrenergic and A(1)-adenosine receptor stimulation, at least in part. Hence, the receptor/effector coupling is dependent on receptor density in this model.
To characterize effects of A(3) adenosine receptor (A(3)AR) activation and gene knock-out on responses to ischemia-reperfusion in mouse heart.
Perfused hearts from wild-type and A(3)AR gene knock-out ...(A(3)AR KO) mice were subjected to 20 min ischemia and 30 min reperfusion. Functional responses were assessed and changes in energy metabolism and cytosolic pH monitored via 31P-NMR spectroscopy.
Selective A(3)AR agonism with 100 nM 2-chloro-N(6)-(3-iodobenzyl)-adenosine-5'-N-methyluronamide (chloro-IB-MECA) enhanced post-ischemic contractile recovery without altering contracture development in wild-type hearts, an effect unrelated to non-selective activation of A(1) or A(2) adenosine receptors. Chloro-IB-MECA also improved recovery in hearts overexpressing A(1)ARs. Paradoxically, post-ischemic recovery was enhanced by A(3)AR KO. Developed pressure, +dP/dt, and -dP/dt all recovered to higher levels in A(3)AR KO (70-80% of pre-ischemia) vs. wild-type hearts (45-50% of pre-ischemia) (P<0.05). Enhanced recovery was unrelated to recoveries of ATP, phosphocreatine (PCr), inorganic phosphate (P(i)), energy state (ATP/ADP x P(i), DeltaG(ATP)) or cytosolic pH.
Selective A(3)AR activation is cardioprotective in wild-type hearts and hearts overexpressing A(1)ARs, yet A(3)AR gene deletion generates an ischemia-tolerant phenotype without altering energy metabolism or pH. This may be due to compensatory changes or undefined genotypic differences in A(3)AR KO vs. wild-type hearts.
Adenosine participates in the coupling of cerebral blood flow to oxygen consumption in the brain during such stimuli as hypoxia, ischemia, and seizures. It has been suggested that it also ...participates in the regulation of cerebral blood flow during somatosensory stimulation, a condition during which cerebral blood flow and oxygen consumption appear to be uncoupled. Interstitial adenosine was estimated by the microdialysis technique and cerebral blood flow was measured by hydrogen clearance in the hindlimb sensory-motor cortex during sciatic nerve stimulation. Cerebral blood flow increased from 102 to 188 ml min-1 100 g-1 (p less than 0.001) in the cortex contralateral to the stimulated leg without an associated increase in interstitial adenosine (baseline 0.624 microM, stimulation 0.583 microM). Infusion of the adenosine antagonist 8-sulfophenyltheophylline failed to block an increase in cerebral blood flow during central sciatic nerve stimulation, but decreased basal cerebral blood flow (69 ml min-1 100 g-1). These results suggest that adenosine does not mediate changes in cerebral blood flow during somatosensory stimulation, but may participate in the regulation of cerebral blood flow in the basal state.
1 Department of Pediatrics and the Cardiovascular Research
Center, University of Virginia Health Sciences Center, Charlottesville,
Virginia 22908, and 2 The Rotary Center for Cardiovascular
...Research, Griffith University Gold Coast Campus, Southport, Q 4217 Australia
The role of A 1
adenosine receptors (A 1 AR) in ischemic preconditioning was
investigated in isolated crystalloid-perfused wild-type and transgenic
mouse hearts with increased A 1 AR. The effect of preconditioning on postischemic myocardial function, lactate
dehydrogenase (LDH) release, and infarct size was examined. Functional
recovery was greater in transgenic versus wild-type hearts (44.8 ± 3.4% baseline vs. 25.6 ± 1.7%). Preconditioning improved
functional recovery in wild-type hearts from 25.6 ± 1.7% to
37.4 ± 2.2% but did not change recovery in transgenic hearts
(44.8 ± 3.4% vs. 44.5 ± 3.9%). In isovolumically
contracting hearts, pretreatment with selective A 1 receptor
antagonist 1,3-dipropyl-8-cyclopentylxanthine attenuated the improved
functional recovery in both wild-type preconditioned (74.2 ± 7.3% baseline rate of pressure development over time untreated vs.
29.7 ± 7.3% treated) and transgenic hearts (84.1 ± 12.8%
untreated vs. 42.1 ± 6.8% treated). Preconditioning wild-type
hearts reduced LDH release (from 7,012 ± 1,451 to 1,691 ± 1,256 U · l 1 · g 1 · min 1 )
and infarct size (from 62.6 ± 5.1% to 32.3 ± 11.5%).
Preconditioning did not affect LDH release or infarct size in hearts
overexpressing A 1 AR. Compared with wild-type hearts,
A 1 AR overexpression markedly reduced LDH release (from
7,012 ± 1,451 to 917 ± 1,123 U · l 1 · g 1 · min 1 )
and infarct size (from 62.6 ± 5.1% to 6.5 ± 2.1%). These
data demonstrate that murine preconditioning involves endogenous
activation of A 1 AR. The beneficial effects of
preconditioning and A 1 AR overexpression are not additive.
Taken with the observation that A 1 AR blockade equally
eliminates the functional protection resulting from both preconditioning and transgenic A 1 AR overexpression, we
conclude that the two interventions affect cardioprotection via common mechanisms or pathways.
mouse; heart; lactose dehydrogenase; infarct size
Activation of A
1
adenosine receptors (A
1
ARs) may be a crucial step in protection against myocardial ischemia-reperfusion (I/R) injury; however, the use of pharmacological A
1
AR antagonists to ...inhibit myocardial protection has yielded inconclusive results. In the current study, we have used mice with genetically modified A
1
AR expression to define the role of A
1
AR in intrinsic protection and ischemic preconditioning (IPC) against I/R injury. Normal wild-type (WT) mice, knockout mice with deleted (A
1
KO
−/−
) or single-copy (A
1
KO
+/−
) A
1
AR, and transgenic mice (A
1
TG) with increased cardiac A
1
AR expression underwent 45 min of left anterior descending coronary artery occlusion, followed by 60 min of reperfusion. Subsets of each group were preconditioned with short durations of ischemia (3 cycles of 5 min of occlusion and 5 min of reperfusion) before index ischemia. Infarct size (IF) in WT, A
1
KO
+/−
, and A
1
KO
−/−
mice was (in % of risk region) 58 ± 3, 60 ± 4, and 61 ± 2, respectively, and was less in A
1
TG mice (39 ± 4, P < 0.05). A strong correlation was observed between A
1
AR expression level and response to IPC. IF was significantly reduced by IPC in WT mice (35 ± 3, P < 0.05 vs. WT), A
1
KO
+/−
+ IPC (48 ± 4, P < 0.05 vs. A
1
KO
+/−
), and A
1
TG + IPC mice (24 ± 2, P < 0.05 vs. A
1
TG). However, IPC did not decrease IF in A
1
KO
−/−
+ IPC mice (63 ± 2). In addition, A
1
KO
−/−
hearts subjected to global I/R injury demonstrated diminished recovery of developed pressure and diastolic function compared with WT controls. These findings demonstrate that A
1
ARs are critical for protection from myocardial I/R injury and that cardioprotection with IPC is relative to the level of A
1
AR gene expression.
Adenosine, acting via A1 receptors, modulates heart rate and contractility, and provides myocardial protection during times of stress. A transgenic model of cardiac A1 overexpression was produced and ...it demonstrated cardiac protection from ischemia. Since A1 receptor stimulation can inhibit contractility under some conditions, the present study was undertaken to determine the effects of transgenic A1 overexpression on intrinsic contractility and the response to catecholamine stimulation. Isolated working mouse hearts were subjected to volume- and pressure-loading protocols to assess intrinsic contractility, and isoproterenol infusions to assess catecholamine response. Basal heart rates were lower in transgenic (Trans) hearts than controls (Ctrl), but with pacing baseline cardiac function and contractility (as measured by +dP/dt) were similar. Volume and pressure loading of Ctrl and Trans hearts were also similar along the entire range tested. No differences were seen in the sensitivity to isoproterenol infusion, but at maximal doses there was a decrease in maximum +dP/dt in Trans hearts compared to Ctrl (maximum +dP/dt 152 +/- 6% baseline for Ctrl, 131 +/- 2% baseline for Trans, P < 0.05). In summary, overexpression of A1 receptors does not produce untoward effects on ventricular function or sensitivity to catecholamine stimulation, but does dampen the contractile response at high doses of catecholamines. These data suggest that even with 1000-fold overexpression of A1 adenosine receptors, adenosine plays little or no role in regulating intrinsic myocardial contractility in the sympathectomized isolated working heart, only modulating contractility as the heart becomes stressed during exposure to higher catecholamine levels.
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Available for:
IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Experimental Physiology Reichelt, Melissa E; Willems, Laura; Peart, Jason N ...
Experimental physiology,
01/2007, Volume:
92, Issue:
1
Journal Article
Peer reviewed
While inhibition of ischaemic contracture was one of the first documented cardioprotective actions of exogenously applied adenosine, it is not known whether this is a normal function of endogenous ...adenosine generated during ischaemic stress. Additionally, the relevance of delayed contracture to postischaemic outcome is unclear. We tested the ability of endogenous versus exogenous adenosine to modify contracture (and postischaemic outcomes) in C57/Bl6 mouse hearts. During ischaemia, untreated hearts developed peak contracture (PC) of 85 ± 5 mmHg at 8.9 ± 0.8 min, with time to reach 20 mmHg (time to onset of contracture; TOC) of 4.4 ± 0.3 min. Adenosine (50 μm) delayed TOC to 6.7 ± 0.6 min, as did pretreatment with 10 μm 2-chloroadenosine (7.2 ± 0.5 min) or 50 nm of A1 adenosine receptor (AR) agonist N6-cyclohexyladenosine (CHA) (6.7 ± 0.3 min), but not A2AAR or A3AR agonists (20 nm 2-4-(2-carboxyethyl) phenethylamino-5' N-methylcarboxamidoadenosine (CGS21680) or 150 nm 2-chloro-N6-(3-iodobenzyl)-adenosine-5'-N-methyluronamide (Cl-IB-MECA), respectively). Adenosinergic contracture inhibition was eliminated by A1AR gene knockout (KO), mimicked by A1AR overexpression, and was associated with preservation of myocardial ATP. This adenosine-mediated inhibition of contracture was, however, only evident after prolonged (10 or 15 min) and not brief (3 min) pretreatment. Ischaemic contracture was also insensitive to endogenously generated adenosine, since A1AR KO, and non-selective and A1AR-selective antagonists (50 μm 8-sulphophenyltheophylline and 150 nm 8-cyclopentyl-1, 3-dipropylxanthine (DPCPX), respectively), all failed to alter intrinsic contracture development. Finally, delayed contracture with A1AR agonism/overexpression or ischaemic 2,3-butanedione monoxime (BDM; 5 μm to target Ca2+ cross-bridge formation) was linked to enhanced postischaemic outcomes. In summary, adenosinergic inhibition of contracture is solely A1AR mediated; the response is 'supraphysiological', evident only with significant periods of pre-ischaemic AR agonism (or increased A1AR density); and ischaemic contracture appears insensitive to locally generated adenosine, potentially owing to the rapidity of contracture development versus the finite time necessary for expression of AR-mediated cardioprotection.
Full text
Available for:
BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
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