Effect of fetal anaemia on myocardial ischaemia-reperfusion injury and coronary vasoreactivity in adult sheep.

AIMS: We investigated whether chronic fetal anaemia affects myocardial infarct in adulthood and elicits functional modifications in adult coronary vasoreactivity. METHODS: Seven-month-old sheep that were made anaemic in utero and transfused to normal haematocrit before birth were studied. Infarct size was determined by tetrazolium after 1-h ischaemia (occlusion of the mid of left anterior descending artery) and 2-h reperfusion. The dose-response to vasoconstrictors and vasodilators was assessed in small resistance coronary arteries. RESULTS: There were no significant differences between the animals previously subjected to in utero anaemia and the control animals regarding the percentage infarct size and the area-at-risk to the left ventricle. The ventricular function (dP/dt) was preserved. The percentage infarct size of the area-at-risk (70.7 +/- 3.5%) was larger than that in the controls (49.8 +/- 4.5%) (P = 0.006). The vascular responses were not altered. Endothelium-dependent relaxation to bradykinin (96.0 +/- 2.6% vs. 98.8 +/- 1.0%) was not affected by PGI(2) inhibitor (94.6 +/- 2.6% vs. 98.5 +/- 1.0%) but significantly reduced by the inhibition of nitric oxide (NO) in both anaemic (P < 0.05) and control (P < 0.001) groups with a significant right shift of EC(50) (P < 0.01). The non-NO-non-PGI(2)-mediated relaxation was slightly potentiated in anaemic animals. CONCLUSIONS: Exposing fetal sheep to in utero anaemia in late gestation for 3 weeks may increase the susceptibility of adult hearts to ischaemia-reperfusion injury without major alterations in coronary vasomotor responsiveness. The impact of in utero anaemia at earlier period of pregnancy and on the earlier or later life of the adult is yet to be further investigated.

Myocardial infarction stimulates galanin expression in cardiac sympathetic neurons.

Cardiac ischemia-reperfusion alters sympathetic neurotransmission in the heart, but little is known about its effect on neuropeptide expression in sympathetic neurons. Ischemia followed by reperfusion induces the production of inflammatory cytokines in the heart, including interleukin-6 and cardiotrophin-1. These cytokines and related molecules inhibit the expression of neuropeptide Y (NPY), and stimulate the expression of vasoactive intestinal peptide (VIP), substance P (SubP), and galanin (GAL) in cultured sympathetic neurons. Therefore, we quantified NPY, VIP, SubP, and GAL mRNA in neurons of the stellate ganglia 1 week after ischemia-reperfusion to determine if neuropeptide expression was altered in cardiac sympathetic neurons. NPY, VIP, and SubP mRNAs were unchanged compared to unoperated control animals, but GAL mRNA was increased significantly. The increased GAL mRNA was not accompanied by elevated GAL peptide content in the stellate ganglia. Galanin content was increased significantly in the heart, however, indicating that elevated GAL mRNA led to increased peptide production. GAL content was increased in the left ventricle below the coronary artery ligation, but was not increased significantly in the atria or the base of the heart above the ligation. The buildup of GAL specifically in the damaged left ventricle is consistent with previous reports that GAL is transported to regenerating nerve endings after axon damage.

Manipulations in glycogen metabolism and the failure to influence infarct size in the ischaemic rabbit heart.

BACKGROUND AND OBJECTIVE: Myocardial ischaemic preconditioning is characterized by a reduction in the rate of glycolysis. Brief myocardial ischaemia also reduces the glycogen content of the heart. The first objective was to determine whether augmenting glucose oxidation by activation of the pyruvate dehydrogenase complex would prevent the infarct limitation of ischaemic preconditioning. The second part of the study evaluates whether glycogen depletion before ischaemia mimics the infarct-limiting effect of ischaemic preconditioning. METHODS: Dichloroacetate (300 + 150 mg kg(-1)), an activator of the pyruvate dehydrogenase complex, was administered intravenously in the anaesthetized open-chest rabbit. All animals underwent 45 min of regional ischaemia and 3 h of reperfusion. Ischaemic preconditiong was elicited by 5 min of coronary occlusion. Control rabbits, those with ischaemic preconditioning with no dichloroacetate, received a saline vehicle. An isolated perfused rabbit heart model was employed to test the second hypothesis. Hearts were depleted of glycogen by perfusing them with a substrate-free buffer. Infarction was assessed by triphenyl tetrazolium chloride and area at risk determined with fluorescent particles. RESULTS: (a) Pyruvate dehydrogenase complex activation experiments. Treatment with dichloroacetate alone did not alter infarct size (58 +/- 7% control vs. 60 +/- 5% dichloroacetate). Addition of dichloroacetate did not attenuate the infarct-limiting effect of ischaemic preconditioning as evidenced by a similar reduction in infarct size in the ischaemic preconditioning group (22 +/- 5%) and in the ischaemic preconditioning + dichloroacetate group (27 +/- 7%). (b) Glycogen depletion experiments. Compared with control hearts with a normal glycogen content (4.84 +/- 0.15 mg g(-1) wet weight), glycogen depleted and ischaemic preconditioning hearts had reduced glycogen content before ischaemia (2.15 +/- 0.26, 1.62 +/- 0.17 mg g(-1) wet weight, respectively; P < 0.01). Glycogen depletion did not reduce infarct size: 25.0 +/- 4.5% cf. 27.9 +/- 3.4% in the control group. However, ischaemic preconditioning resulted in a significant reduction of infarct size (11.5 +/- 2.3% vs. 27.9 +/- 3.4% control; P < 0.01). CONCLUSIONS: Augmentation of oxidative glycolysis by dichloroacetate in in situ rabbit hearts does not alter the effect of ischaemic preconditioning, and glycogen depletion in the isolated rabbit heart does not influence infarct size after subsequent coronary occlusion.

Propidium iodide compares favorably with histology and triphenyl tetrazolium chloride in the assessment of experimentally-induced infarct size.

Quantitative assessment of infarct size after histology processing or by tetrazolium staining requires long reperfusion times to facilitate the delineation of the injured tissue. We evaluated the nucleic acid stain propidium iodide as an alternate technique for assessment of infarct size because it does not require extended reperfusion and is accomplished with more simple tissue processing. Eight mice underwent 45 min of coronary artery occlusion and 24 h of reperfusion, after which propidium iodide was administered and allowed to circulate for 15 min. Hearts were excised, sliced into transverse slices approximately 620 micro m thick, and photographed for determination of infarction with propidium iodide. Slices were processed by standard histology techniques, and infarction was evaluated using classical criteria of necrosis. In four other mice, infarct size was assessed by both triphenyl tetrazolium chloride and propidium iodide staining. A total of 46 slices were analysed and infarction was described as a percentage of the cross-sectional area of the slice. The correlation (r=0.94), orthogonal regression line (y=0.9x+1.8), and the percentage of the cross-section infarcted (histology 17.5+/-3.0%v propidium iodide 18.5+/-3.0%), demonstrates that infarct size assessment after propidium iodide staining or histology processing yields the same results. Similarly, propidium iodide measurement of infarct size was comparable to that obtained with TCC staining (22.5+/-4.1%v 25.4+/-5.4%, respectively, r=0.84). We conclude that propidium iodide staining yields accurate infarct size assessment and is a simpler alternative to tetrazolium staining or histologic processing.

Met5-enkephalin protects isolated adult rabbit cardiomyocytes via delta-opioid receptors.

In rats and rabbits, endogenous opioid peptides participate in ischemic preconditioning. However, it is not known which endogenous opioid(s) can trigger cardioprotection. We examined preconditioning-induced and opioid-induced limitation of cell death in isolated, calcium-tolerant, adult rabbit cardiomyocytes. Cells were subjected to simulated ischemia by pelleting and normothermic hypoxic incubation. Preconditioning was elicited with 15 min of simulated ischemia followed by 15 min of resuspension and reoxygenation. All cells underwent 180 min of simulated ischemia. Cell death was assessed by trypan blue permeability. Morphine protected cells, as did preconditioning; naloxone blocked the preconditioning-induced protection. Exogenous Met5-enkephalin (ME) induced protection, but exogenous beta-endorphin did not. ME-induced protection was blocked by the delta-selective antagonist naltrindole. Additionally, two other proenkephalin products, Leu5-enkephalin and Met5-enkephalin-Arg-Phe, provided protection equipotent to ME. These data suggest that one or more proenkephalin products interact with delta-opioid receptors to endogenously trigger opioid-mediated protection.

Ischemic preconditioning limits infarct size following regional ischemia-reperfusion in in situ mouse hearts.

OBJECTIVE: Ischemic preconditioning has been demonstrated in a wide variety of animals, from dogs to rats. Experimentally-induced murine myocardial ischemia-reperfusion has been described, but ischemic preconditioning has not been reported in mouse hearts. To test the hypothesis that mouse hearts exhibit preconditioning-induced protection, experiments were conducted in anesthetized open chest mice subjected to regional myocardial ischemia-reperfusion. METHODS: Following barbiturate anesthesia the FVB and C57BL/6J mice underwent a tracheostomy and were mechanically ventilated. The heart was exposed via a left thoracotomy performed with the aid of a dissecting microscope. A 7-0 silk suture on a curved taper needle was passed under the proximal left anterior descending coronary artery to form a snare. Mice were then randomly assigned to receive either no preconditioning or preconditioning. All mice were subjected to 60 min regional myocardial ischemia followed by 2.5 h of reperfusion. Ischemic preconditioning (IP) was induced with two (FVB mice) or three (C57BL/6J mice) cycles of 5 min coronary occlusion and 5 min reperfusion. Control animals did not receive preconditioning ischemia. Area-at-risk was assessed with fluorescent particles. Infarct size was assessed with triphenyl tetrazolium chloride, and is expressed below as a percentage of the area-at-risk. RESULTS: In FVB mice preconditioning reduced infarct size 49%, from 36.7 +/- 4.5% to 18.9 +/- 2.8% (P < 0.05). In C57BL/6J mice preconditioning reduced infarct size by 66%, from 56.4 +/- 8.3% to 18.9 +/- 4.2% (P < 0.05). CONCLUSION: From these data we conclude that the infarct limiting effect of ischemic preconditioning is demonstrable in murine hearts.

Naloxone blockade of myocardial ischemic preconditioning does not require central nervous system participation.

OBJECTIVE: The hypothesis that naloxone blockade of ischemic preconditioning (IP)-induced infarct limitation does not require central nervous system participation was evaluated using quaternary naloxone in anesthetized rabbits (Study I) and naloxone hydrochloride in isolated rabbit hearts (Study II). METHODS: In Study I, rabbits underwent 30 min coronary artery occlusion and 180 min reperfusion. IP was elicited with a 5 min coronary artery occlusion beginning 15 min before the 30 min occlusion. Intravenous naloxone methiodide, 12.9 mg/kg, was bolused 10 or 1 min before IP. In Study II, rabbit hearts underwent 45 min coronary artery occlusion and 120 min reperfusion. IP was elicited with 2 cycles of 5 min coronary artery occlusion plus 5 min reperfusion, beginning 20 min before the 45 min occlusion. Naloxone hydrochloride, 1 mumol/L or 100 mumol/L, was added to the buffer perfusate for 25 min preceding the long coronary artery occlusion. In both studies, infarct size was assessed with tetrazolium, normalized to risk volume, and analyzed using ANOVA. RESULTS: In both studies, IP reduced infarct size compared to control (6.3 +/- 2.3 vs. 29.5 +/- 4.4, P = 0.007, Study I; 11.8 +/- 4.7 vs. 47.7 +/- 6.7, P = 0.03, Study II). In Study I, IP was not blocked when naloxone methiodide was given 10 min before IP (13.8 +/- 4.8 vs. 42.3 +/- 5.4, P = 0.004) but was blocked when given 1 min before IP (25.3 +/- 7.2 vs. 28.4 +/- 5.0, P = ns). In Study II, infarct size was intermediate in the 1 mumol/L naloxone hydrochloride + IP group (19.0 +/- 6.5 vs. 48.9 +/- 8.4, P = ns) but IP was blocked by 100 mumol/L naloxone hydrochloride (62.6 +/- 4.5 vs. 56.2 +/- 6.7, P = ns). CONCLUSION: Naloxone blockade of IP-induced infarct limitation involves a cardiac mechanism.

Hypothermia increases the threshold for ischemic preconditioning.

OBJECTIVES: Both hypothermia and ischemic preconditioning are known to provide tolerance to myocardial ischemia and reperfusion. The aim of this study was to determine whether hypothermia during the ischemic preconditioning period attenuates the protective effect of ischemic preconditioning. METHODS: Experiments were performed in buffer-perfused isolated rabbit hearts. All hearts underwent 45 minutes of regional ischemia, followed by 2 hours of reperfusion. Ischemic preconditioning was elicited by either one or four periods of 5 minutes of regional ischemia. Hypothermia (25 degrees C) was induced beginning either 20 or 50 minutes before the 45-minute period of regional ischemia; normothermia (38 degrees C) was restored 10 minutes before the 45-minute period of regional ischemia. Except for the hypothermic periods noted, hearts were maintained at 38 degrees C. RESULTS: Normothermic ischemic preconditioning with either one or four cycles of 5 minutes of coronary occlusion resulted in a profound reduction of infarct size (58% reduction with one cycle, p < 0.05; 95% reduction with four cycles, p < 0.01). Hypothermic ischemic preconditioning with one cycle of 5-minute coronary occlusion resulted in no reduction of infarct size but hypothermic ischemic preconditioning with four cycles of 5-minute coronary occlusions resulted in a 94% reduction of infarct size (p < 0.01). Myocardial glycogen and lactate levels were maintained near control levels during hypothermic ischemia. CONCLUSIONS: From these data we conclude that hypothermia during the preconditioning period increases the threshold for eliciting the infarct limitation of ischemic preconditioning.

Altered heart rate baroreflex during pregnancy: role of sympathetic and parasympathetic nervous systems.

Two studies were performed to determine whether the attenuation of baroreflex control of heart rate during late pregnancy in conscious rabbits is due to changes in parasympathetic (Para) or sympathetic (Sym) control of the heart. In the first, baroreflex relationships between arterial pressure and heart rate were generated before and after treatment with propranolol (Pro) to block Sym or with methscopolamine (Meth) to block Para. Each rabbit was studied in both the pregnant and nonpregnant state. Pregnancy decreased maximum baroreflex gain from 14.9 +/- 4.0 to 4.8 +/- 0.9 beats.min-1.mmHg-1 (P < 0.01) and decreased heart rate range from 177 +/- 6 to 143 +/- 10 beats/min (P < 0.01), primarily by increasing minimum heart rate (114 +/- 6 to 134 +/- 8 beats/min; P < 0.01). The difference between pregnant and nonpregnant rabbits in baroreflex gain was not altered by Meth but was abolished by Pro, suggesting that it is due to decreased Sym control of the heart. The elevated minimum heart rate of pregnancy persisted after Pro, but was abolished by Meth, suggesting that it is mediated by decreased Para control of the heart. In the second study, isolated buffer-perfused hearts from pregnant and nonpregnant rabbits were treated with increasing doses of isoproterenol (0.3-300 mM) or acetylcholine (0.3-10,000 microM), and the heart rate responses were determined. Hearts from pregnant rabbits were more sensitive to isoproterenol (P < 0.05), but less responsive to acetylcholine (P < 0.05). In conclusion, pregnancy-induced decreases in cardiac reflex gain and range appear to be mediated by alterations in Sym and Para, respectively. The change in Sym occurs proximal to the heart, whereas the decreased contribution of Para may be due, at least in part, to decreased sensitivity of the heart to acetylcholine.

High spinal anesthesia does not alter experimental myocardial infarction size or ischemic preconditioning.

OBJECTIVE: The role of the central nervous system in the development of myocardial infarction and ventricular fibrillation in virgin and ischemically preconditioned myocardium was investigated. DESIGN: Infarct size and ventricular arrhythmias were assessed after regional ischemia-reperfusion. Animals were randomly assigned to four groups: (1) preconditioned, central nervous system intact; (2) nonpreconditioned, nervous system intact; (3) preconditioned, nervous system blocked; and (4) nonpreconditioned, nervous system blocked. Differences in hemodynamics and infarct size were assessed with analysis of variance, and differences in ventricular fibrillation were assessed with the Kruskal-Wallis test. SETTING: Experiments were performed in the Anesthesiology Research Laboratory at a medical center. PARTICIPANTS: Anesthetized open-chest New Zealand white rabbits were used for these studies. INTERVENTIONS: Rabbits underwent 30 minutes of coronary artery occlusion and 3 hours of reperfusion. The central nervous system was blocked with total spinal anesthesia. Ischemic preconditioning was elicited with 5 minutes of coronary artery occlusion and 10 minutes of reperfusion. Infarction was assessed with tetrazolium and expressed as a percentage of the risk zone (mean +/- SEM). MEASUREMENTS AND MAIN RESULTS: Preconditioning resulted in infarct size limitation compared with the control (8% +/- 4% v 43% +/- 5%; p < 0.001) and delayed the onset of fibrillation (15.5 minutes v 11 minutes; p = 0.001). Spinal blockade neither altered nonpreconditioned infarct size nor attenuated preconditioning (32% +/- 7% v 8% +/- 3%; p = 0.04), but it was associated with ventricular fibrillation in 24/25 rabbits as compared with 6/14 rabbits without blockade. In blocked animals, preconditioning resulted in a decreased duration of fibrillation (2.5 minutes v 12.5 minutes; p = 0.0004). However, spinal blockade eliminated the preconditioning-induced delay in fibrillation (10 minutes v 12 minutes; p = NS). CONCLUSIONS: It is concluded that (1) activation of efferent sympathetic nerves is not necessary for ischemic preconditioning; (2) preconditioning delays the onset of ventricular arrhythmias; and (3) spinal blockade exacerbates ischemia-induced ventricular arrhythmias.

Naloxone blockade of myocardial ischemic preconditioning is stereoselective.

This study tested the hypothesis that endogenous opioids are involved in the infarct limitation of myocardial ischemic preconditioning (IP). Blockade of IP-induced infarct limitation by (-)naloxone hydrochloride (-NAL) or its receptor-inactive stereoisomer (+)naloxone (+NAL) was evaluated. Fifty-two pentobarbitone-anesthetized, open-chest rabbits underwent 30 min coronary artery occlusion and 180 min reperfusion. Treatment groups were: control (n = 9), i.p. (n = 8), -NAL (n = 9) and -NAL/i.p. (n = 12), or +NAL (n = 6) and +NAL/i.p. (n = 8). i.p. was elicited with 5 min regional ischemia, beginning 15 min before the 30 min coronary occlusion. -NAL or +NAL, 3 mg/kg i.v. bolus, was given 25 min before the 30 min coronary occlusion. Infarct size was assessed with tetrazolium and expressed as a percentage of area-at-risk. There were no significant intergroup differences of area-at-risk. IP resulted in marked infarct limitation compared to control (control, 32.9 +/- 7.6% v i.p., 5.8 +/- 4.5%; P = 0.04). Neither -NAL nor +NAL alone altered infarct size compared to control, but -NAL did block the infarct limitation of i.p. (-NAL, 31.4 +/- 6.7% v -NAL/i.p., 24.3 +/- 6.2%) whereas +NAL did not (+NAL, 40.5 +/- 5.0% v +NAL/i.p., 13.7 +/- 3.6%; P = 0.02). In conclusion, naloxone blockade of i.p.-induced cardioprotection is stereospecific and therefore likely to be opioid receptor-mediated.

Adrenergic activation confers cardioprotection mediated by adenosine, but is not required for ischemic preconditioning.

BACKGROUND: The aim of this study was to determine whether (1) adrenergic activation is cardioprotective, (2) adrenergic cardioprotection occurs via adenosine receptor activation, and (3) ischemic preconditioning requires alpha-adrenergic activation. METHODS: Anesthetised open chest rabbits underwent 30 min coronary occlusion and 3 h reperfusion. Ischemic preconditioning was elicited with 5 min coronary occlusion and 10 min reperfusion. Activation of adrenergic receptors with endogenous norepinephrine was achieved with tyramine (0.28 mg/kg/min intravenously for 5 min). Adenosine receptors were blocked with 8-p-sulfophenyl theophylline (10 mg/kg intravenously), alpha 1-adrenergic receptors were selectively blocked with prazosin (0.1 mg/kg intravenously), and alpha-adrenergic receptors were blocked with phentolamine (4 mg/kg intravenously). RESULTS: Ischemic preconditioning reduced risk-adjusted infarct volume by 79% (P < 0.0005). This protection was attenuated by adenosine receptor blockade. Tyramine infusion resulted in a 1305% change from baseline plasma norepinephrine concentration (P < or = 0.01), and reduced infarct volume by 55% (P = 0.01). Adenosine receptor blockade abolished this protection. Blockade of alpha 1-adrenergic receptors with prazosin failed to abolish ischemic preconditioning (79 versus 89% reduction in infarct volume, without and with prazosin, respectively). Similarly, non-selective blockade of alpha-adrenergic receptors also failed to abolish ischemic preconditioning (79 versus 57% reduction without and with phentolamine, respectively). CONCLUSIONS: We conclude that the cardioprotection of ischemic preconditioning and alpha-adrenergic activation both involve adenosine, but ischemic preconditioning does not require alpha-adrenergic activation.

Dichloroacetate reduces plasma lactate levels but does not reduce infarct size in rabbit myocardium.

Dichloroacetate (DCA), an activator of pyruvate dehydrogenase (PDHC), enhances postischemic mechanical recovery of isolated hearts. It is not known whether this is secondary to reduced infarction or preservation of contractile function in viable cardiomyocytes. This study investigated the effect of DCA on myocardial infarct size. Anesthetized open chest rabbits underwent regional coronary occlusion and reperfusion. DCA (300 mg/kg plus 150 mg/kg 1 h later) was administered intravenously either before occlusion (DCA-O; n = 8) or at reperfusion (DCA-R; n = 7). Control rabbits (n = 8) received saline vehicle. Myocardial PDHC activity was measured after administration of 300 mg/kg i.v. DCA in 10 separate rabbits. DCA reduced plasma lactate levels and increased PDHC activity by 76%, from 2.79 +/- .30 mumol/min.g-1 to 4.92 +/- .44 mumol/min.g-1 (p < .005). However, infarct size in DCA-treated animals was not significantly different from Control (60 +/- 5% DCA-O, 57 +/- 6% DCA-R, 58 +/- 7% Control). We conclude that stimulation of pyruvate dehydrogenase does not limit infarct size.

Fluorescent vs. radioactive microsphere measurement of regional myocardial blood flow.

OBJECTIVES: This study compared simultaneous regional myocardial blood flow (RMBF) measurements using fluorescent microspheres (FM) and radiolabeled microspheres (RM). The utility of an internal standard during processing was also examined. METHODS: Paired FM and RM were injected into the left atrium of 9 anaesthetised rabbits. RMBF was altered by use of either regional ischaemia or (-)-N6-(2-phenylisopropyl)-adenosine. Radioactivity of blood reference and tissue samples was quantitated using standard methods. Samples were then digested with potassium hydroxide and microspheres recovered by vacuum filtration, with an additional label of FM as the internal standard. FM labels were extracted using Carbitol acetate and quantitated using fluorescence spectroscopy. Agreement between the fluorescent and radioactive methods was assessed using both orthogonal regression and difference-against-mean analyses. RESULTS: Using recovery-uncorrected data, the slope of the orthogonal regression of RM and FM-determined RMBF was not statistically different from 1, but the intercept was statistically different from 0 [-0.03(0.01), P = 0.005] and the mean RMBF by each method differed from one another [1.24(0.08) vs. 1.17(0.08) ml.min-1.g-1, P = 0.0002]. The mean +/- 2 s.d. of the differences of RMBF (RM minus FM) was +0.07 +/- 0.30 ml.min-1.g-1. Although recovery of FM from tissue averaged 97.6(1.2)%, use of the internal standard to correct for losses substantially improved the agreement between RM and FM-determined RMBF: the orthogonal regression slope was not statistically different from 1, the intercept was not statistically different from 0, and the means of the flows were not different. The mean +/- 2 s.d. of the differences of RMBF was -0.01 +/- 0.22 ml.min-1.g-1. The internal standard also improved RMBF estimates from samples with simulated large spillage during processing. CONCLUSION: Fluorescent microspheres are an equivalent alternative to radiolabeled microspheres for the estimation of RMBF. Although the overall recovery of microspheres using this technique was high, use of an internal standard is recommended for correction of random losses.

Anaesthetics alter the magnitude of infarct limitation by ischaemic preconditioning.

OBJECTIVE: The aim was to determine whether three commonly used animal anaesthetics alter the magnitude of infarct limitation achieved with ischaemic preconditioning. METHODS: Eighty four anaesthetised non-preconditioned and preconditioned open chest rabbits underwent a 30 min coronary occlusion followed by 3 h reperfusion. Ischaemic preconditioning was achieved with 5 min coronary occlusion beginning 15 min before the 30 min coronary occlusion. The anaesthetics studied were: pentobarbitone (30 mg.kg-1 intravenously +30-50 mg.kg-1.h-1 intravenously), isoflurane (1.5-2.5% end expiratory), and ketamine/xylazine (cocktail of 67 mg ketamine and 6.7 mg xylazine.ml-1, 1 ml.kg-1 intramuscularly +0.3-1.3 ml.kg-1.h-1 intramuscularly). Area at risk was delineated with ZnCdS particles and infarction assessed with tetrazolium. RESULTS: There were no significant differences in area at risk, heart rate, arterial pressure, and temperature between non-preconditioned and preconditioned hearts. Although infarct size was not significantly different among non-preconditioned hearts for each anaesthetic regimen (p = NS), the magnitude of infarct limitation with preconditioning varied with the anaesthetic employed (decrease in infarct size from control values of 81%, 44%, and 33% for pentobarbitone, isoflurane and ketamine/xylazine, respectively, p = 0.0145 for comparison of the three magnitudes, two factor ANOVA). CONCLUSION: Anaesthetic regimens affect the degree of infarct size limitation seen with ischaemic preconditioning.

"Normothermic range" temperature affects myocardial infarct size.

OBJECTIVES: This study tested the hypothesis that small changes in temperature above the hypothermic range may alter myocardial infarct size after acute coronary occlusion-reperfusion. A secondary hypothesis, that a correlation between temperature and infarct size may be independent of an associated change of heart rate, was also evaluated. METHODS: Eighteen pentobarbitone-anaesthetised, open chest rabbits underwent 30 min coronary artery occlusion and 3 h reperfusion at blood temperatures ranging from 35-42 degrees C, achieved and maintained using surface methods (not paced, group NP). In a second group of 11 animals, heart rate was held constant across the same range of temperatures (paced, group P), before and throughout coronary artery occlusion-reperfusion. Infarct sizes were assessed by the tetrazolium method. RESULTS: Target temperature was effectively controlled over the duration of the experimental protocol to within +/- 0.25 degrees C. Area at risk did not vary with temperature. Infarct size, normalised to area at risk, was correlated with temperature in both groups (infarct size = 7.9 x temp-250.0, r = 0.75, p = 0.0003, group NP; infarct size = 11.7 x temp-404.5, r = 0.88, p = 0.0004, group P). There was no significant difference between the slopes of these two lines (p = 0.18), indicating that the positive correlation between infarct size and temperature is not related to changes of heart rate. CONCLUSION: Temperatures in the range of 35-42 degrees C affect myocardial infarct size significantly, independent of heart rate.

Cardioprotection provided by adenosine receptor activation is abolished by blockade of the KATP channel.

Adenosine agonists and openers of the ATP-sensitive potassium (KATP) channel have been reported to limit infarct size (IS). We tested the hypothesis that these phenomena are interdependent. Anesthetized swine underwent 60 min of coronary artery occlusion and 90 min of reperfusion. Preconditioning was elicited by two cycles comprising 10 min of occlusion and 10 min of reperfusion (n = 7 swine). An intracoronary infusion of adenosine (Ado; n = 10) or (-)-N6-(2-phenylisopropyl)-adenosine (R-PIA; n = 7) replaced preconditioning ischemia. KATP channels were blocked with sodium 5-hydroxydecanoate (5-HD) in the absence (n = 6) or presence (n = 8) of R-PIA. Control pigs (n = 7) received saline vehicle. IS was assessed with tetrazolium and normalized as percentage of area at risk. Preconditioning resulted in a reduced IS compared with Control (3.9 +/- 1.8 vs. 43.5 +/- 6.9%, respectively; P < 0.0005). Ado and R-PIA also reduced IS [21.1 +/- 6.8 (P < 0.01) and 11.2 +/- 7.4% (P < 0.005), respectively]. 5-HD alone did not alter IS, but it abolished R-PIA-induced cardioprotection (IS 5-HD + R-PIA = 48.6 +/- 13.2%). Thus Ado A1-receptor agonists mimicked the cardioprotection of ischemic preconditioning. The Ado-induced limitation of IS was abolished by blockade of the KATP channel. We conclude that both Ado A1 receptors and KATP channels may be involved in ischemic preconditioning.

Whole body heat stress fails to limit infarct size in the reperfused rabbit heart.

OBJECTIVE: It has recently been shown that induction of heat stress proteins by whole body heat stress confers myocardial protection in the isolated in vitro rat and rabbit heart. This study extends the above studies by examining the effects of stress protein synthesis on the limitation of infarct size in the in vivo rabbit heart model. METHODS: 30 male New Zealand white rabbits were used. Six rabbits were used for measurement of heat stress protein; 10 were used for infarct size determination in a heat stress group (HS); 14 were used for infarct size determination in a control group. There were 10 exclusions. Under anaesthesia, body temperature was raised to 42 degrees C for 15 min in the HS group. Following 24 hours of recovery rabbits were reanaesthetised and the hearts subjected to a 45 min period of regional ischaemia followed by 3 h reperfusion. The risk zone was defined with fluorescent particles and the infarct area determined by tetrazolium staining. Western blotting showed an increase in the 72 KD heat stress protein in hearts in the HS group. RESULTS: Infarct size as a percent of risk area was 61.4 (SEM 6.4)% (n = 14) in control hearts and 71.8(7.3)% (n = 10) in the HS hearts. These results were not statistically significant. CONCLUSIONS: No protective effect of heat stress could be seen when infarct size was used as the end point. Either the protection seen in earlier studies using the Krebs perfused isolated heart model does not accurately reflect protection against myocardial infarction, or heat stress itself may induce injurious factors in the blood which will negate any direct protective effect to the myocardium in this model.