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The ubiquitin-proteasome system plays an important role in regulating muscle mass. Inducible immunoproteasome subunits LMP-2 and LMP-7 are constitutively expressed in the heart; however, their regulation and functions are poorly understood. We here investigated the hypothesis that immunoproteasomes regulate cardiac muscle mass in diabetic mice. Type 1 diabetes was induced in wildtype mice by streptozotocin. After hyperglycemia developed, insulin and the proteasome inhibitor epoxomicin were used to treat diabetic mice for 6weeks. Isolated mouse hearts were perfused with control or high glucose solution. Catalytic proteasome beta-subunits and proteolytic activities were analyzed in the heart by immunoblotting and fluorogenic peptide degradation assays, respectively. Insulin and epoxomicin blocked loss of heart weight and improved cardiac function in diabetic mice. LMP-7 and its corresponding chymotryptic-like proteasome activity were increased in diabetic hearts and high glucose-treated hearts. Myosin heavy chain protein was decreased in diabetic hearts, which was largely reversed by epoxomicin. High glucose decreased LMP-2 protein levels in perfused hearts. In diabetic hearts, LMP-2 expression was downregulated whereas expression of the phosphatase and tensin homologue deleted on chromosome ten (PTEN) and the muscle atrophy F-box were upregulated. Moreover, mice with muscle-specific knockout of PTEN gene demonstrated increased cardiac muscle mass, while mice with LMP-2 deficiency demonstrated PTEN accumulation, muscle mass loss, and contractile impairment in the heart. Therefore, we concluded that high glucose regulates immunoproteasome subunits and modifies proteasome activities in the heart, and that dysregulated immunoproteasome subunits may mediate loss of cardiac muscle mass in experimental diabetic mice., (Copyright 2010 Elsevier Ltd. All rights reserved.)

Nitric oxide (NO.) generated from nitric oxide synthase (NOS) isoforms bound to cellular membranes may serve to modulate oxidative stresses in cardiac muscle and thereby regulate the function of key membrane-associated enzymes. Ischemia is known to inhibit the function of sarcolemmal enzymes, including the (Na+ + K+)-ATPase, but it is unknown whether concomitant injury to sarcolemma (SL)-associated NOS isoforms may contribute to this process by reducing the availability of locally generated NO. Here we report that nNOS, as well as eNOS (SL NOSs), are tightly associated with cardiac SL membranes in several different species. In isolated perfused rat hearts, global ischemia caused a time-dependent irreversible injury to cardiac SL NOSs and a disruption of SL NO. generation. Pretreatment with low concentrations of the NO. donor 1-hydroxy-2-oxo-3-(N-3-methyl-aminopropyl)-3-methyl-1-triazene (NOC-7) markedly protected both SL NOS and (Na+ + K+)-ATPase functions against ischemia-induced inactivation. Moreover, ischemia impaired SL Na+/K+ binding, and NOC-7 significantly prevented ischemic injury to the ion binding sites on (Na+ + K+)-ATPase. These novel findings indicate that NO. can protect cardiac SL NOSs and (Na+ + K+)-ATPase against ischemia-induced inactivation and suggest that locally generated NO. may serve to regulate SL Na+/K+ ion active transport in the heart.

Nitric oxide (NO*) is produced endogenously from NOS isoforms bound to sarcolemmal (SL) and sarcoplasmic reticulum (SR) membranes. To investigate whether locally generated NO* directly affects the activity of enzymes mediating ion active transport, we studied whether knockout of selected NOS isoforms would affect the functions of cardiac SL (Na+ + K+)-ATPase and SR Ca2+-ATPase. Cardiac SL and SR vesicles containing either SL (Na+ + K+)-ATPase or SR Ca2+-ATPase were isolated from mice lacking either nNOS or eNOS, or both, and tested for enzyme activities. Western blot analysis revealed that absence of single or double NOS isoforms did not interrupt the protein expression of SL (Na+ + K+)-ATPase and SR Ca2+-ATPase in cardiac muscle cells. However, lack of NOS isoforms in cardiac muscle significantly altered both (Na+ + K+)-ATPase activity and SR Ca2+-ATPase function. Our experimental results suggest that disrupted endogenous NO* production may change local redox conditions and lead to an unbalanced free radical homeostasis in cardiac muscle cells which, in turn, may affect key enzyme activities and membrane ion active transport systems in the heart.

This study tested the usefulness of technetium-99m-labeled antigranulocyte monoclonal antibody BW250/183 (AGMAb) for identifying granulocyte accumulation in ischemic/reperfused canine myocardium. In dogs with 90 minutes coronary artery occlusion and 180 minutes reperfusion (n = 8), ischemic/reperfused myocardial samples demonstrated 8.5 +/- 2.4 times more Tc-99m-AGMAb accumulation than nonischemic samples. Dogs given Tc-99m-labeled nonspecific human immunoglobulin instead of Tc-99m-AGMAb (n = 3) had about half as much accumulation (4.5 +/- 1.6, P < .05). Ex vivo myocardial imaging of Tc-99m-AGMAb demonstrated marked uptake in infarcted regions identified by absent triphenyl tetrazolium chloride staining. The amount of uptake was inversely related to the severity of ischemia (determined by radioactive microspheres) and directly correlated with tissue myeloperoxidase activity, a specific marker of granulocyte accumulation. No increase in Tc-99m-AGMAb uptake occurred in dogs with 90 minutes ischemia and no reperfusion (n = 3) or 15 minutes ischemia and 180 minutes reperfusion (n = 2). In conclusion, Tc-99m-AGMAb is taken up in reperfused infarcted myocardium by both nonspecific and specific mechanisms. Because the amount of uptake reflects myocardial granulocyte accumulation, Tc-99m-AGMAb combined with nuclear imaging techniques may be useful for studying inflammatory processes in the heart in experimental animal models and human beings.

To determine whether myocardial necrosis may occur during postischemic reperfusion, electron microscopy was used to identify morphological features of irreversible injury in myocardial samples taken from anesthetized dogs with 90-min ischemia and 0-, 5-, 90-, or 180-min reperfusion. In samples without detectable collateral blood flow, necrosis was almost complete, whether or not the myocardium was reperfused. In samples with collateral flow, necrosis was more frequent after 180-min reperfusion than in the absence of reperfusion, despite similar collateral flows in the two groups. Excess of necrosis after 180-min reperfusion was evident in endocardium (ischemia only: 4 of 13, 180-min reflow: 14 of 20; P = 0. 03) and midwall (ischemia only: 9 of 25, 180-min reflow: 29 of 45; P = 0.02). Multiple logistic regression with variables of collateral flow and transmural position was used to determine risk of irreversible injury in 111 samples from ischemic myocardium without reperfusion (model predictive accuracy = 75%, P < 0.00001) and to predict risk of necrosis in myocardium reperfused for 180 min. Of 65 samples from endocardium and midwall with detectable collateral flow, the model predicted necrosis in 23 samples but necrosis was observed in 43 samples (P < 0.01). Reperfusion duration was a determinant of frequency of irreversible injury. Multiple logistic regression for 186 samples from myocardium reperfused for 5, 90, or 180 min showed that reperfusion duration was an independent predictor of irreversible injury (P = 0.0003) when collateral flow and transmural location were accounted for. These findings are consistent with the occurrence of necrosis during reperfusion in myocardium exposed to substantial, prolonged ischemia but with sufficient residual perfusion to avoid necrosis during the period of flow impairment.

Background: To limit ischemic myocardial injury, it is important to differentiate viable from infarcted myocardium. Three dimensional (3D) tagged MRI has the ability to quantify myocardial 3D deformation and strain (noninvasively and precisely), and can achieve a true comparison of contraction not only from region to region, but also at different levels of function. In this study, we investigated whether regional strain mapping obtained by 3D-tagged MRI can differentiate between viable but stunned myocardium and nonviable myocardium., Methods and Results: We examined 7 dogs 2 days after a 90-minute closed-chest left anterior descending coronary artery occlusion followed by 48 hours of reperfusion. 3D-tagged MR images spanning the entire left ventricle were acquired both at rest and during dobutamine infusion (5 microg. kg-1. min-1 IV). Regional blood flow was measured with radioactive microspheres and used to define risk regions. Infarcted regions were defined as 2,3,5 triphenyltetrazolium chloride negative regions. Strains in infarcted regions were greatly impaired compared with remote regions (P<0.001) and remained unchanged during dobutamine stress. Risk regions showed a dysfunction at rest, with improved function during dobutamine infusion. Receiver operating characteristics analysis showed that radial strain was more accurate for identifying viable regions., Conclusions: When coupled with a stress test, 3D strain mapping by the use of tagged MRI is a sensitive and noninvasive method for characterizing ischemic injury. Regional strain can be used to differentiate between viable but stunned and nonviable myocardium within the postischemic injured myocardium.

NO. is a free radical that modulates heart function and metabolism. We report that a neuronal-type NO synthase (NOS) is located on cardiac sarcoplasmic reticulum (SR) membrane vesicles and that endogenous NO. produced by SR-associated NOS inhibits SR Ca2+ uptake. Ca2+-dependent biochemical conversion of L-arginine to L-citrulline was observed from isolated rabbit cardiac SR vesicles in the presence of NOS substrates and cofactors. Endogenous NO. was generated from the vesicles and detected by electron paramagnetic resonance spin-trapping measurements. Immunoelectron microscopy demonstrated labeling of cardiac SR vesicles by using anti-neuronal NOS (nNOS), but not anti-endothelial NOS (eNOS) or anti-inducible NOS (iNOS) antibodies, whereas skeletal muscle SR vesicles had no nNOS immunoreactivity. The nNOS immunoreactivity also displayed a pattern consistent with SR localization in confocal micrographs of sections of human myocardium. Western blotting demonstrated that cardiac SR NOS is larger than brain NOS (160 vs. 155 kDa). No immunodetection was observed in cardiac SR vesicles from nNOS knockout mice or with an anti-nNOS mu antibody, suggesting the possibility of a new nNOS-type isoform. 45Ca uptake by cardiac SR vesicles, catalyzed by Ca2+-ATPase, was inhibited by NO. produced endogenously from cardiac SR NOS, and 7-nitroindazole, a selective nNOS inhibitor, completely prevented this inhibition. These results suggest that a cardiac muscle nNOS isoform is located on SR of cardiac myocytes, where it may respond to intracellular Ca2+ concentration and modulate SR Ca2+ ion active transport in the heart.

Background: Microvascular obstruction within an area of myocardial infarction indicates worse functional recovery and a higher risk of postinfarction complications. After prolonged coronary occlusion, contrast-enhanced MRI identifies myocardial infarction as a hyperenhanced region containing a hypoenhanced core. Because the time course of microvascular obstruction after infarction/reperfusion is unknown, we examined whether microvascular obstruction reaches its full extent shortly after reperfusion or shows significant progression over the following 2 days., Methods and Results: Seven dogs underwent 90-minute balloon occlusion of the left anterior descending coronary artery (LAD) followed by reflow. Gadolinium-DTPA-enhanced MRI performed at 2, 6, and 48 hours after reperfusion was compared with radioactive microsphere blood flow (MBF) measurements and myocardial staining to define microvascular obstruction (thioflavin S) and infarct size (triphenyltetrazolium chloride, TTC). The MRI hypoenhanced region increased 3-fold during 48 hours after reperfusion (3.2+/-1.8%, 6.7+/-4.4%, and 9.9+/-3.2% of left ventricular mass at 2, 6, and 48 hours, respectively, P<0.03) and correlated well with microvascular obstruction (MBF <50% of remote region, r=0.99 and thioflavin S, r=0.93). MRI hyperenhancement also increased (21.7+/-4.0%, 24.3+/-4.6%, and 28.8+/-5.1% at 2, 6, and 48 hours, P<0.006) and correlated well with infarct size by TTC (r=0.92). The microvascular obstruction/infarct size ratio increased from 13.0+/-4.8% to 22.6+/-8.9% and to 30.4+/-4.2% over 48 hours (P=0.024)., Conclusion: The extent of microvascular obstruction and the infarct size increase significantly over the first 48 hours after myocardial infarction. These results are consistent with progressive microvascular and myocardial injury well beyond coronary occlusion and reflow.

Background: Contrast-enhanced fast magnetic resonance (MR) images of acute, reperfused human infarcts demonstrate regions of hypoenhancement and hyperenhancement. The relations between the spatial extent and time course of these enhancement patterns to myocardial risk, infarct, and no-reflow regions have not been well characterized., Methods and Results: The proximal left anterior descending coronary artery was occluded in 11 closed-chest dogs for 90 minutes followed by 2 days of reperfusion. Regional blood flow was determined by use of radioactive microspheres. The animals were studied at the 2-day time point with contrast-enhanced fast MRI (Signa 1.5 T, General Electric). Thioflavin-S was administered to demarcate no-reflow regions. The hearts were then excised, sectioned into five base-to-apex slices, stained with 2,3,5-triphenyltetrazolium chloride (TTC), and photographed under room light (for TTC) and ultraviolet light (for thioflavin). The spatial extents of thioflavin-negative, TTC-negative, and risk regions were compared planimetrically with MRI hypoenhanced and hyperenhanced regions. The spatial locations of subendocardial hypoenhancement in MR images correlated closely with those of thioflavin-negative regions. Microsphere blood flow in these regions was significantly reduced compared with remote regions (0.37 +/- 0.09 versus 0.88 +/- 0.10 mL/min per gram, respectively, P < .001) and with baseline (0.37 +/- 0.09 versus 0.87 +/- 0.15 mL/min per gram, P < .01). The spatial extent of hyperenhancement was smaller than the risk region (r = .64, slope = 0.48, P < .001) but highly correlated with TTC-negative regions and were, on average, 12% larger (r = .93, slope = 1.12, P = .035)., Conclusions: In contrast-enhanced MR images of 2-day-old reperfused canine infarcts, myocardial regions of hypoenhancement are related to the no-reflow phenomenon. Approximately 90% of the myocardium within hyperenhanced regions is nonviable.

Although ATP derived from glycolysis represents only a small fraction of total myocardial ATP production, metabolic compartmentation may result in preferential use of glycolytic ATP for certain membrane activities, including pumping of Ca2+ from the cytoplasm. We tested this hypothesis by looking for evidence of Ca2+ overload in normoxic perfused rabbit hearts given iodoacetate (IAA, 50 microM) to block glycolysis and isoproterenol (Iso, 0.05 microM) to stimulate Ca2+ entry. The hearts beat isovolumically and were perfused with 16 mM glucose and 5 or 10 mM pyruvate (to preserve oxidative metabolism) in a superconducting magnet for 31P-nuclear magnetic resonance (NMR) measurements of high energy phosphates or 19F-NMR measurements of intracellular free Ca2+ concentration ([Ca2+]i). IAA by itself had no effect on left ventricular (LV) developed pressure, end-diastolic pressure, pressure-rate product, or tissue high-energy phosphates. During exposure to Iso, mean LV end-diastolic pressure increased from 10.7 to 49.3 mmHg in hearts pretreated with IAA (n = 7) but did not change in control hearts (n = 7). During Iso, there were substantial reductions in developed pressure, ATP, and phosphocreatine in IAA-treated hearts but not in control hearts. After exposure to IAA and Iso, a doubling of diastolic [Ca2+]i was observed with 19F-NMR. In IAA-treated hearts, reduction of perfusate Ca2+ concentration from 2.5 to 0.6 mM during Iso exposure (n = 6) prevented the mechanical dysfunction and decrease in high-energy phosphates. These findings suggest that glycolysis is necessary to preserve myocardial Ca2+ homeostasis during beta-adrenergic stimulation.

This study examined the hypothesis that glycolysis is required for functional recovery of the myocardium during reperfusion by facilitating restoration of calcium homeostasis. [Ca2+]i was measured in isolated perfused rabbit hearts by using the Ca2+ indicator 1,2-bis(2-amino-5-fluorophenoxy)ethane-N,N,N',N'-tetraacetic acid (5F-BAPTA) and 19F nuclear magnetic resonance spectroscopy. In nonischemic control hearts, inhibition of glycolysis with iodoacetate did not alter [Ca2+]i. In hearts subjected to 20 minutes of global zero-flow ischemia, [Ca2+]i increased from 260 +/- 80 nM before ischemia to 556 +/- 44 nM after 15 minutes of ischemia (p less than 0.05). After reperfusion with 5 mM pyruvate as a carbon substrate, [Ca2+]i increased further in hearts with intact glycolysis to 851 +/- 134 nM (p less than 0.05 versus ischemia) during the first 10 minutes of reperfusion, before returning to preischemic levels. In contrast, inhibition of glycolysis during the reperfusion period resulted in persistent severe calcium overload ([Ca2+]i, 1,380 +/- 260 nM after 15 minutes of reperfusion, p less than 0.02 versus intact glycolysis group). Furthermore, despite the presence of pyruvate and oxygen, inhibition of glycolysis during early reperfusion resulted in greater impairment of functional recovery (rate/pressure product, 3,722 +/- 738 mm Hg/min) than did reperfusion with pyruvate and intact glycolysis (rate/pressure product, 9,851 +/- 590 mm Hg/min, p less than 0.01). Inhibition of glycolysis during early reperfusion was also associated with a marked increase in left ventricular end-diastolic pressure during reperfusion (41 +/- 5 mm Hg) compared with hearts with intact glycolysis (16 +/- 2 mm Hg, p less than 0.01). The detrimental effects of glycolytic inhibition during early reperfusion were, however, prevented by initial reperfusion with a low calcium solution ([Ca]o, 0.63 mM for 30 minutes, then 2.50 mM for 30 minutes). In these hearts, the rate/pressure product after 60 minutes of reperfusion was 12,492 +/- 1,561 mm Hg/min (p less than 0.01 versus initial reflow with [Ca]o of 2.50 mM). These findings indicate that the functional impairment observed in postischemic myocardium is related to cellular Ca2+ overload. Glycolysis appears to play an important role in restoration of Ca2+ homeostasis and recovery of function of postischemic myocardium.

To determine whether technetium-99m-hexakis-2-methoxyisobutyl isonitrile (SESTAMIBI) remains fixed in the myocardium following its initial uptake or undergoes time-related redistribution, anesthetized dogs underwent occlusion of the anterior descending coronary artery for 6 min, followed by 3-hr reperfusion. Technetium-99m-SESTAMIBI and thallium-201 (201Tl) were injected intravenously after 1 min occlusion and regional myocardial blood flow was measured with radioactive microspheres. Tomographic imaging of Tc-SESTAMIBI revealed a perfusion defect with slight but definite filling in over 2 hr. Quantitative analysis indicated a significant rise in the nadir and decrease in the width of the defect in circumferential profile curves. After 3-hr of reperfusion, Tc-SESTAMIBI activity in the previously ischemic area was always greater than the activity of microspheres injected during coronary occlusion (mean normalized values, 0.32 versus 0.11, p less than 0.0001). Our results indicate that following transient ischemia and reperfusion, Tc-SESTAMIBI clearly undergoes myocardial redistribution, although more slowly and less completely than 201Tl.

The influence of nifedipine on left ventricular ejection fraction, infarct size, and infarct expansion was studied in a prospective, double blind, randomised, placebo controlled trial in 132 patients with low risk acute myocardial infarction of less than 12 hours duration, defined by an initial left ventricular ejection fraction greater than 35% and clinical Killip class of less than or equal to II. Sixty four patients were assigned to nifedipine 120 mg/day and 68 to placebo. Treatment was started on average (SEM) 8.0 (0.2) hours after onset of pain and continued for six weeks. Gated blood pool scans, thallium scans, and cross sectional echocardiograms were performed before treatment and at 10 days. There were no significant differences between the two groups in age, sex, cardiac risk factors, or use of other medications. Mean (SEM) global left ventricular ejection fraction was not different before treatment (nifedipine group 53 (2%), placebo group 55 (2%) and did not differ at 10 days (nifedipine group 54 (2%), placebo group 52 (2%). There were also no differences in regional wall motion or regional ejection fractions. Thallium defects quantified by computer analysis were similar in both groups before treatment (nifedipine 7.8 (0.7), placebo 7.9 (0.7)) and at 10 days (nifedipine 5.3 (0.7) placebo 5.3 (0.7)). In the subgroup of patients with transmural infarction who had good quality echocardiograms and serial studies (n = 30), there was no difference in mean (SEM) baseline infarct segment lengths between the two groups (nifedipine 70 (4) mm, placebo 65 (4) mm); however, the nifedipine group demonstrated no significant change in infarct segment length between the initial and 10 day studies ( + 0.6 (3) mm) while there was a significant increase in the infarct segment length in the placebo group (+ 7.8 (4) mm). The infarct segment length increased by >/= 1 cm in seven (47%) placebo patients but in only one (7%) nifedipine patient. The nifedipine group had a significant initial 10% decrease in mean arterial pressure whereas there was no change in the in the placebo group; this blood pressure difference persisted for 10 days. Thus although the early administration of nifedipine has no detectable effect on clinical outcome and infarction size, it may reduce early infarct expansion via an afterload reduction mechanism in patients with transmural infarction. These initial results must be interpreted with caution and need to be confirmed in a larger trial.

The "no-reflow" phenomenon, the occurrence of areas with very low flow in hearts reperfused after ischemia, is thought to be largely established at the time of reperfusion as a result of microvascular damage induced by ischemia. In the present study we sought to determine whether additional impairment of tissue perfusion might also occur during the course of reperfusion. Open-chest dogs were subjected to 90 minutes of left circumflex coronary artery occlusion and reperfused for 2 minutes (n = 7) or 3.5 hours (n = 8). Myocardial perfusion was visualized in left ventricular slices following in vivo injection of the fluorescent dye thioflavin-S just before killing. The area of impaired perfusion (absent thioflavin) averaged 9.5 +/- 3.0% of the risk region in dogs reperfused for 2 minutes, whereas it was nearly three times as large in dogs reperfused for 3.5 hours (25.9 +/- 8.2% of the risk region, p less than 0.05). Serial measurements of flow by microspheres during reperfusion demonstrated zones within the postischemic myocardium that were hyperemic 2 minutes after reperfusion, with adequate flow still present at 30 minutes, but with a subsequent marked fall in perfusion. After 3.5 hours these areas showed negligible flow (0.13 +/- 0.3 ml/min/g) and no thioflavin uptake. Tissue samples showing postischemic impairment in perfusion has received virtually no collateral flow during ischemia (less than 0.01 ml/min/g), whereas collateral flow was significantly higher in adjacent thioflavin-positive zones (0.04 +/- 0.01 ml/min/g in endocardial samples and 0.07 +/- 0.02 ml/min/g in samples from the midmyocardium, p less than 0.001 vs. thioflavin-negative areas). Areas that showed late impairment of flow invariably demonstrated contraction band necrosis, which contrasted with the pattern of coagulation necrosis observed in areas of "true" (i.e., immediate) no-reflow. Intracapillary erythrocyte stasis and marked intravascular neutrophil accumulation (to levels greater than 20-fold that found after 2 minutes reperfusion) were typically observed in areas of delayed impairment to flow. Obstruction to flow at the capillary level was confirmed in additional dogs in which the heart was injected postmortem with silicone rubber to delineate the microvascular filling pattern. Areas of absent capillary filling were much more extensive after 3.5 hours than after 2 minutes reperfusion. Thus, this study shows that the occurrence of areas of markedly impaired perfusion in postischemic myocardium is related only in part to an inability to reperfuse certain areas on reflow. A more important factor is represented by a delayed, progressive fall in flow to areas that initially received adequate reperfusion.(ABSTRACT TRUNCATED AT 400 WORDS)

The cumulative effects of repeated, brief episodes of regional ischemia on myocardial function and dimension were examined in 14 open-chest dogs. The left anterior descending coronary artery was occluded for 5 minutes, followed by 10 minutes of reflow, repeated 16 times, and then 1 hour recovery. Systolic function decreased progressively in segments made repetitively ischemic and remained depressed even after 1 hour of recovery. Average systolic shortening decreased 20% from baseline after recovery from the first occlusion, 82% after the 8th, 91% after the 16th, and 104% after the 1 hour recovery (p less than 0.015, analysis of variance). End-diastolic segment length progressively increased in regions made repetitively ischemic, lengthening 4% after the first occlusion, 10% after the third occlusion, 19% after the sixteenth occlusion, and 16% after 1 hour of recovery (p less than 0.02). Nonischemic end-diastolic segment length also showed a smaller but parallel increase, while non-ischemic systolic function showed compensatory improvement. After the dogs were killed, myocardial staining with triphenyl tetrazolium chloride revealed no necrosis. Electron microscopy, performed in 5 dogs, showed scattered mitochondrial swelling in both postischemic and nonischemic regions, but no evidence of irreversible injury. The ratio of myocardial blood flow in the region made repetitively ischemic to nonischemic flow, as measured with microspheres, was 1.00 +/- 0.02 before the occlusions and 0.90 +/- 0.03 just before death (difference not significant). Thus, in the dog progressively abnormal regional systolic function and regional and global diastolic dilatation can be produced by repetitive, brief, coronary occlusions, which are not associated with histochemical or ultrastructural evidence of myocardial necrosis.

Although thallium-201 is known to redistribute slowly into regions of ischemic myocardium after restoration of blood flow, it is not clear to what extent normalization of flow is an essential requirement for the redistribution process. In a search for a flow-independent component of thallium redistribution, 12 dogs with stenosis of the circumflex coronary artery underwent atrial pacing for either 20 minutes (group I, 6 dogs) or 2 hours (group II, 6 dogs). Radioactive thallium and radioactive microspheres, 7 to 10 mu, were injected after 10 minutes of atrial pacing in both groups. Pacing resulted in a 40 percent reduction in subendocardial blood flow to the circumflex-perfused myocardium in both groups I and II. This relative reduction in flow was maintained at a stable level over the 2 hour pacing period in group II. Thallium activity in the relatively ischemic zone was significantly greater in dogs with 2 hours of pacing (group II) than in those with 10 minutes of pacing (group I). Redistribution of thallium occurred despite the continued presence of reduced flow in circumflex-perfused endocardial tissue. These data suggest that a significant component of thallium redistribution may be flow-independent.

Although electrical ablation of ventricular tachycardia via percutaneous catheters has been recently accomplished in human beings, little is known of its pathologic or arrhythmogenic effects. We studied 21 open-chest anesthetized dogs in which an endocardial electrode catheter was percutaneously introduced into the left ventricle. Direct current (DC) shock was delivered by a standard defibrillator through the distal electrode to a back paddle. Cross-sectional two-dimensional echocardiographic studies were performed in the plane of the catheter (confirmed by epicardial metal beads), and blood flow was determined by the microsphere technique before DC shock and when the animals were killed 2 to 8 days later. Of 11 dogs receiving a total of 100 to 400 J, only three survived 48 hr compared with nine of 10 receiving 50 J and all three control dogs. Holter monitoring demonstrated sustained ventricular tachycardia (VT) (greater than or equal to 30 sec) in all 11 dogs monitored (six received greater than or equal to 100 J), beginning within 5 hr of the DC shock; three control dogs had no VT. Two dogs that died suddenly while being monitored showed ventricular fibrillation. Histologic examination revealed hemorrhagic contraction band necrosis in the shock zone, a type of injury similar to that observed in reperfusion necrosis. Necrosis of the left ventricle was 0.5% to 5%. There was no significant difference in necrosis between dogs receiving 100 J or more and those receiving 50 J (2.5 vs 1.7 g; p greater than .10). Percent systolic thickening determined in eight equally divided regions around the left ventricle showed no difference between the shock zone, perishock zone, or remote normal zone in dogs receiving 50 J.(ABSTRACT TRUNCATED AT 250 WORDS)

This study examined the possibility that the postischemic mechanical depression observed in the "stunned" myocardium is a result of an alteration in the control of intracellular calcium. Regional myocardial stunning was produced in five open-chest dogs by eight to twelve 5-minute occlusions of the left anterior descending coronary artery, alternated with 10-minute reflow periods and followed by a final 60-minute period of reperfusion. Systolic segment shortening in the postischemic zone, measured by sonomicrometry, fell from 14.9% at baseline to -1.1% at the end of reperfusion. Sarcoplasmic reticulum isolated from stunned myocardium demonstrated a 17% reduction in oxalate-supported 45Ca2+ transport compared with sarcoplasmic reticulum from normal myocardium (0.93 vs. 1.12 mumol Ca2+/mg protein/min, p less than 0.005). There was also a 20% decrease in the maximal activation by Ca2+ of the sarcoplasmic reticulum Ca2+, Mg2+-ATPase (2.46 vs. 1.96 mumol Pi/mg protein/min, p less than 0.005), and a downward shift in the Ca2+-activation curve of the Ca2+, Mg2+-ATPase. These results indicate that myocardial stunning is associated with damage to the calcium-transport system of the sarcoplasmic reticulum. Altered intracellular control may contribute to the inability of the stunned heart to maintain normal mechanical function.

Eight open chest dogs underwent 25 min of coronary occlusion to determine whether brief myocardial ischemia disrupts the normal myocardial inotropic response to sympathetic nervous stimulation. If so, this could represent a mechanism contributing to postischemic myocardial dysfunction. Myocardial segment shortening was measured using ultrasonic dimension crystals before and after coronary artery occlusion and reperfusion. Left ansa subclavia stimulation and systemic norepinephrine (NE) infusion were used to test the myocardial inotropic response to neural stimulation and direct exposure to the sympathetic mediator, respectively. Before coronary artery occlusion, base-line preischemic segment shortening (12.5 +/- 1.6%) (SEM) increased during both sympathetic stimulation (20.2 +/- 1.4%) and NE infusion (19.7 +/- 1.1%). The control segment responded similarly. After ischemia and reperfusion there was no significant change in heart rate, aortic or left ventricular pressures, nor changes in control segment shortening. In contrast, shortening in the postischemic segment was markedly reduced compared to baseline (4.1 +/- 2.4%), and no longer responded to sympathetic stimulation (2.4 +/- 2.8%), while responsiveness to systemic NE was maintained (12.9 +/- 2.0%), P less than 0.001, which suggested injury to the sympathetic-neural axis during the period of ischemia. This reduced response to neural stimulation was persistent for up to 2 h after reperfusion. Left atrial or intracoronary infusion of bretylium tosylate, which releases norepinephrine from nerve terminals, resulted in an immediate inotropic response in the postischemic segment, which indicated that total depletion of NE from nerve terminals during the ischemic period had not occurred. Disruption of sympathetic neural responsiveness is likely a component of the mechanism of postischemic myocardial dysfunction whenever there is appreciable sympathetic drive to the heart.

To study the regional function of nonischemic myocardium after the onset of regional ischemia, graded circumflex coronary arterial stenosis was induced in 18 open-chest anesthetized dogs. Two-dimensional echocardiographic views were obtained at each degree of occlusion in a cross-sectional plane marked by two to three metal beads sewn to the left ventricular epicardium. Percent systolic thickening was measured at 16 equally spaced points around the left ventricle and correlated with microsphere-determined regional myocardial blood flow. Baseline thickening averaged 44.9 +/- 6.4%. During transmural ischemia percent systolic thickening decreased to -16.1 +/- 4.0% in the ischemic region and also decreased in adjacent nonischemic regions (to 2.4 +/- 2.4% in segments closest to the ischemic region [adjacent 1] and to 15.5 +/- 3.9 in segments further away [adjacent 2]), but was unchanged in segments directly opposite the ischemic region (remote region). During subendocardial ischemia, percent systolic thickening fell only in the ischemic and adjacent 1 regions (1.4 +/- 5.2% and 24.9 +/- 5.0%, respectively). Dipyridamole, 0.21 to 0.42 mg/min iv, given to seven dogs during transmural ischemia, caused a three- to fivefold increase in flow to the nonischemic and no change in flow to the ischemic region; function was not altered in any region. Propranolol, 0.1 mg/kg iv, was given to five dogs during transmural ischemia to depress contractility in the remote region. Percent systolic thickening fell in the remote (from 50.0 +/- 7.7% to 34.6 +/- 5.6%), but increased in adjacent 1 (from -0.25 +/- 3.7% to 15.2 +/- 3.9%) and in adjacent 2 (from 17.4 +/- 2.8% to 33.4 +/- 3.9%) regions, and remained unchanged in the ischemic region. We conclude the following: During transmural ischemia percent systolic thickening is markedly impaired in nonischemic myocardium immediately adjacent to the ischemic region, and is impaired to a lesser degree in regions located relatively far from the ischemic border. Dysfunction therefore overestimates the extent of regional ischemia after total coronary occlusion. During subendocardial ischemia function ceases in the ischemic region and functional impairment of nonischemic myocardium is restricted to immediately adjacent regions. Dysfunction of adjacent regions is not caused by "relative ischemia" related to increased local oxygen demands or to a steal phenomenon. Mechanical tethering of nonischemic myocardium adjacent to ischemic regions, secondary to changes in left ventricular shape during contraction, may contribute to the impairment of systolic thickening in adjacent regions during transmural ischemia.

The effect of a single dose (10 mg/kg) of intravenous thiopental (TP), during acute myocardial infarction, on infarct size was studied in conscious dogs randomized 10 minutes after left circumflex coronary artery occlusion to either the TP group (n = 10) or a control group given 0.9% saline solution (n = 10). During the first hour following therapy, myocardial blood flow (microspheres), arterial pressure, left atrial pressure, and arterial blood gases were similar in the two groups, but the heart rate (140 +/- 3 vs 110 +/- 3 bpm; p less than 0.001) and rate-pressure product (15,090 vs 12,210 bpm X mm Hg; p less than 0.025) were greater in the TP group. Infarct size (planimetry) and occluded bed size (postmortem coronary arteriography) measured 2 days later revealed that: the slope of the relation between infarct and occluded bed mass, as a percentage of the left ventricle (% LV) was greater with TP than with saline solution (1.10 vs 0.61; p less than 0.001); excluding hearts (four TP and three saline solution) with small occluded beds (less than 22% LV), infarcts were also larger with TP (n = 6) than with saline solution (n = 7), both as a percentage of the left ventricle (26.4 vs 12.2%; p less than 0.02) or occluded bed (61.5 vs 28.9%; p less than 0.005); and transmural and endocardial extents of the infarcts on topographic maps were greater with TP than with saline solution. In 12 other conscious dogs, increasing the heart rate between 10 and 70 minutes after left circumflex coronary artery occlusion to the average rate of the TP group (140 bpm) by atrial pacing resulted in infarcts larger than those in control dogs but similar to those in the TP group. Thus, TP therapy after left circumflex occlusion increased infarct size in dogs. This effect appeared to be due mainly to the increased heart rate, probably via increased myocardial oxygen demands.

Although oxygen consumption closely parallels mechanical work in the normal heart, previous studies have found that stunned myocardium may have normal or even increased oxygen consumption despite depressed function. In this study we used microspectrophotometry to measure the oxygen saturations within arteries and veins of less than 100 micron diameter in quick-frozen biopsy samples from normal and regionally stunned myocardium of 10 open-chest anesthetized dogs. Regional myocardial blood flow, measured by radioactive microspheres, was similar in stunned and normal regions, as was mean arteriolar oxygen saturation. However, mean venous oxygen saturation was lower in the stunned region (epicardium 38.0% vs 43.8%, p less than .02; endocardium 36.2% vs 39.5%, p = .12), indicating increased oxygen extraction and consumption, despite a marked reduction in mean systolic segmental shortening from 14.4% to 0.5%. In addition, there was greater vein-to-vein heterogeneity of oxygen saturation in the stunned region, with an excess of veins having low saturations (statistically significant in epicardium, nonsignificant trend in endocardium). Microvascular injections studies with Microfil or drafting ink revealed filling of over 95% of arterioles and 85% of capillaries in the stunned region, similar to the findings in the normal region. Our results are consistent with an inefficient transfer of energy into myocyte contraction or an increased use of energy for noncontractile activities in stunned myocardium. In addition, the finding of increased heterogeneity of oxygen extraction suggests that the injury to stunned myocardium may not be uniform to all contractile elements, but instead may be focally and irregularly distributed.