Dietary carbohydrate influences tissue fatty acid and lipid composition in the copper-deficient rat

Fructose and copper have been shown independently to influence long chain fatty acid metabolism. Since fructose feeding exacerbates copper deficiency, their possible interaction with respect to tissue long chain fatty acid and lipid composition was studied. Weanling male Sprague-Dawley rats were given diets containing 0.6 or 6 mg/kg copper. The carbohydrate source (627 g/kg) was either fructose or corn starch. After 3 wk, fatty acid profiles and total lipids in heart and liver were analyzed. Copper-deficient rats fed fructose had more severe signs of copper deficiency than those fed starch, according to heart/body wt ratio, hematocrit, and liver copper content. The fatty acid composition of heart and liver triacylglycerol was significantly different between groups, but the changes did not correlate with the severity of copper deficiency. In heart, phosphatidylinositol and phosphatidylserine, arachidonic acid and docosapentaenoic acid (n-6) were increased 193 and 217%, respectively, p<0.05) in rats given the copper-deficient diet containing fructose. Changes in the long chain fatty acids in heart phospholipids may be related to the higher mortality commonly observed in rats fed a copper-deficient diet containing fructose.     

Effects of fatty acids on phosphatidylcholine biosynthesis in isolated hamster heart

The effects of stearic, oleic, and arachidonic acids on phosphatidylcholine biosynthesis in the hamster heart were investigated. When hamster hearts were perfused with labelled choline in the presence of fatty acids, biosynthesis of phosphatidylcholine was stimulated only by stearic acid. Stearic acid was found to accumulate in unesterified (free) form in the hamster heart after perfusion. The stimulation by stearic acid was mediated in vivo by an enhancement of CTP:phosphocholine cytidylyltransferase activity in the microsomal fraction of the hamster heart and the enzyme activity in the cytosolic fraction was not affected. In contrast with the observations in rat hepatocytes, cytidylyltransferase from the hamster heart was not stimulated directly by stearic acid. The selective activation of the microsomal enzyme when the heart was perfused with stearic acid suggests that activation of the enzyme was mediated via the modification of the membrane by stearic acid.     

Protection of the ischemic diabetic heart by L-propionylcarnitine therapy

Diabetics suffer from an increased incidence of myocardial infarction and are less likely to survive an ischemic insult. Since L-propionylcarnitine (LPC) has been shown to protect against ischemic/reperfusion injury, we hypothesized that LPC may be of even greater benefit to the diabetic heart. Diabetes was induced by i.v. streptozotocin, 60 mg/kg; duration: 12 wks. The chronic effect of LPC was determined by daily i.p. injections (100 mg/kg) for 8 wks. The acute effects of LPC were determined by adding it to the perfusion medium (5 mM) of control and diabetic hearts. Initial cardiac contractile performance of isolated perfused working hearts was assessed by varying left atrial filling pressure. Hearts were then subjected to 90 min of low flow global ischemia followed by 30 min reperfusion. Chronic LPC treatment had no effect on initial cardiac performance in either control or diabetic hearts. Acute addition of LPC to the perfusion medium enhanced pump performance of control hearts, but had no effect in diabetic hearts. Both acute and chronic LPC significantly improved the ability of control and diabetic hearts to recover cardiac contractile performance after ischemia and reperfusion, however, chronic treatment was more effective in diabetic hearts.     

Accumulation of specific ceramides in ischemic/reperfused rat heart; effect of ischemic preconditioning.

Ceramide signalling has been implicated in the mechanism of myocardial ischemia/reperfusion injury (IR). This study tested the hypothesis that ceramides containing a specific amino-linked acyl residue mediate the injury, and that ischemic preconditioning (IPC) affords myocardial protection because it prevents increased ceramide accumulation in IR myocardium. Perfused rat hearts were subjected either to the sham perfusion or to 30 min global ischemia, 30 min ischemia/30 min reperfusion (IR) or were preconditioned prior to the standard IR. The ventricles were harvested for biochemical assay that involved transmethylation of ceramide amino-linked acyl residues, and gas liquid chromatography measurement of acyl methyl esters. Fourteen ceramides containing myrystic, palmitic, palmitoleic, stearic, oleic, linoleic, linolenic, arachidic, arachidonic, eicosapentaenoic, behenic, docosapentaenoic, docosahexaenoic or nervonic acid were identified in the myocardium of rats. The total basal ceramide concentration in the myocardium was 135 nmol/g tissue, and it was increased by 14.1% and 48.4% in the ischemia and IR group, respectively. However, in fact, IR increased the accumulation of only 7 out of 14 ceramides identified in the heart (i.e., those containing palmitic, stearic, oleic, linoleic, and arachidonic acid), and the relative magnitude of these increases varied between the particular ceramides and was independent from their basal tissue concentration. IPC improved postischemic hemodynamic recovery and partially prevented the reperfusion-induced increases in these 7 ceramides, while the other ceramides were unaffected by IPC. These results support the role of the specific ceramide signalling in the mechanism of myocardial IR injury. We speculate that by preventing tissue accumulation of certain ceramides, IPC attenuates this signalling, that adds to the mechanism of myocardial protection afforded by IPC.     

Cell cycle-dependent changes in arachidonic acid and glycerol metabolism in Swiss 3T3 cells stimulated by platelet-derived growth factor

Quiescent Swiss 3T3 cells stimulated to divide by human platelet-derived growth factor (PDGF) were used to investigate cell cycle-dependent changes in arachidonic acid, stearic acid, and glycerol metabolism. PDGF at 12 ng/ml stimulated incorporation of labeled arachidonic and stearic acid into phosphatidic acid and phosphatidylinositol within 60 min. With similar kinetics PDGF stimulated glycerol incorporation into phosphatidic acid and phosphatidylinositol indicating early growth factor-dependent stimulation of de novo phosphatidylinositol synthesis. This early effect of PDGF was specific for the phosphatidylinositol synthesis pathway since no comparable changes were noted in other glycerolipids. After a lag of 4-6 h, PDGF strongly stimulated arachidonic acid incorporation into triacylglycerol: at 6 h, arachidonate radioactivity in triacylglycerol exceeded that in phosphatidylcholine, phosphatidylethanolamine, and phosphatidylinositol. This effect of PDGF was not associated with de novo triacylglycerol synthesis since no increase in the rate of glycerol incorporation into this lipid was noted. Finally, PDGF stimulated incorporation of glycerol into all major phospholipids and triacylglycerol during S-phase. These results disclose three novel effects of PDGF on glycerolipid metabolism in Swiss 3T3 cells: 1) early selective activation of the phosphatidylinositol synthesis pathway; 2) delayed strong stimulation of arachidonic acid incorporation into triacylglycerol; and 3) late induction of de novo phosphatidylcholine, phosphatidylethanolamine, and triacylglycerol synthesis. These PDGF effects are likely to play important roles in phosphatidylinositol metabolism, membrane biosynthesis, and fatty acid turnover in rapidly growing cells.     

Influence of low extracellular Ca2+ ions on the cardiac function changes induced by verapamil in the perfused rabbit heart

We tested the hypothesis that the myocardial effects of verapamil (VER) could be enhanced by decreasing the extracellular Ca2+ concentration ([Ca2+]o) in the isolated rabbit heart at 37 degrees C. After perfusion with standard Krebs - bicarbonate solution containing 1.27 mM Ca2+, for a 30-min period of stabilization and 15 min of control, groups of hearts were perfused for an additional 60 min with solutions containing one of the following: 1.27 mM Ca2+ (control group), 0.23 mM Ca2+ (low [Ca2+]o group), 1.27 mM Ca2+ plus 10(-7) M VER (VER group), or 0.23 mM Ca2+ plus 10(-7) M VER (combination, CBN group). These concentrations of [Ca2+]o and VER produce submaximal responses in our preparation. We found that the heart rate - LV pressure product (RPP) in the CBN group fell rapidly to 0 in the first 2-3 min of perfusion, this response being significantly lower than in the other two groups for the first 15 min. Electromechanical dissociation (EMD) appeared in one of six hearts at 60 min and in four of six hearts at 30 min in the low [Ca2+]o and VER groups, respectively, whereas it occurred in the CBN group in all hearts at 3 min. Depolarization rate (DR) fell by 10% in the low [Ca2+]o and VER groups versus a reduction of 45% in the CBN group (P less than 0.05) during the last 45 min of perfusion. The PR interval increased by 300% in the CBN group, a much greater and significant change (P less than 0.05) than in the hearts exposed to VER or low [Ca2+]o.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of sucrose diet on insulin secretion in vivo and in vitro and on triglyceride storage and mobilisation of the heart of rats

Basal heart triacylglycerol (TG) (mumole triacylglycerol/g of dry weight) (- before "in vitro" Langendorff perfusion -) was significantly higher in animals rendered chronically hypertriglyceridaemic (H) by a 63% sucrose-rich diet than in controls (C, standard diet); 28 +/- 2.6 means + SEM vs. 19.3 +/- 1.2; respectively (p less than 0.01). After 40' perfusion with Krebs-Henseleit buffer + 5.5 mM glucose, 2.5 mM Ca++, TG content fell to 14.2 +/- 0.6 in C and 14.9 +/- 1.9 in H (n.S.). Administration of 1 n mol x min-1 of glucagon (Gn) from min 20 to 40 reduced TG to 9.0 +/- 0.5 in C (p less than 0.05). In contrast no effect of Gn was observed in H (TG at min 40: 16.7 +/- 2.5). Glycogen (Gly) content (mumol/g of dry weight) after Gn perfusion fell from 30 +/- 1.9 to 17 +/- 2.1 (p less than 0.01) in C, while again no effect was recorded in H. "In vivo" plasma glucose fractional coefficient disappearance rate was lower (p less than 0.001) in H: 1.01 x 10(-2) +/- 0.09 x 10(-2) vs 2.61 x 10(-2) +/- 0.14 x 10(-2) in C, in spite of H showing hyperinsulin secretion. Hyperinsulinism was further documented by "in vitro" Iri release studies from incubated pancreas pieces. In the absence of glucose (G) from the incubation medium H produced 541 +/- 19.8 mU/mg weight Tissue/20', while C produced 91.2 +/- 12.7 (p less than 0.001). With 100 mg% G, H released 1058 +/- 259 and C 377 +/- 82.5 (p less than 0.001). It is suggested that hyperinsulin secretion plus insulin resistance may account for the above findings.     

Properties of phosphatidate phosphohydrolase and diacylglycerol acyltransferase activities in the isolated rat heart. Effect of glucagon, ischaemia and diabetes.

Myocardial triacylglycerol hydrolysis is subject to product inhibition. After hydrolysis of endogenous triacylglycerols, the main proportion of the liberated fatty acids is re-esterified to triacylglycerol, indicating the importance of fatty acid re-esterification in the regulation of myocardial triacylglycerol homoeostasis. Therefore, we characterized phosphatidate phosphohydrolase (PAP) and diacylglycerol acyltransferase (DGAT) activities, enzymes catalysing the final steps in the re-esterification of fatty acids to triacylglycerols in the isolated rat heart. The PAP activity was mainly recovered in the microsomal and soluble cell fractions, with an apparent Km of 0.14 mM for both the microsomal and the soluble enzyme. PAP was stimulated by Mg2+ and oleic acid. Oleic acid, like a high concentration of KCl, stimulated the translocation of PAP activity from the soluble to the particulate (microsomal) fraction. Myocardial DGAT had an apparent Km of 3.8 microM and was predominantly recovered in the particulate (microsomal) fraction. Both enzyme activities were significantly increased after acute streptozotocin-induced diabetes, PAP from 15.6 +/- 1.1 to 28.1 +/- 3.6 m-units/g wet wt. (P less than 0.01) and DGAT from 2.23 +/- 0.11 to 3.01 +/- 0.11 m-units/g wet wt. (P less than 0.01). In contrast with diabetes, low-flow ischaemia during 30 min did not affect PAP and DGAT activity in rat hearts. Perfusion with glucagon (0.1 microM) during 30 min did not affect total PAP activity, but changed the subcellular distribution. More PAP activity was recovered in the particulate fraction. DGAT activity was lowered by glucagon treatment from 0.37 +/- 0.03 to 0.23 +/- 0.02 m-unit/mg of microsomal protein (P less than 0.05). The role of PAP and DGAT activity and PAP distribution in the myocardial glucose/fatty acid cycle is discussed.     

Dietary coenzyme Q(10) supplement renders swine hearts resistant to ischemia-reperfusion injury

To examine whether nutritional supplementation of coenzyme Q(10) (CoQ(10)) can reduce myocardial ischemia-reperfusion injury, a group of swine was fed a regular diet supplemented with CoQ(10) (5 mg x kg(-1) x day(-1)) for 30 days. Another group of pigs that were fed a regular diet supplemented with placebo served as a control. After 30 days, isolated in situ pig hearts were prepared and hearts were perfused with a cardiopulmonary pump system. Each heart was subjected to 15 min of regional ischemia by snaring of the left anterior descending coronary artery, followed by 60 min of hypothermic cardioplegic global ischemia and 120 min of reperfusion. After the experiments were completed, myocardial infarct size was measured by triphenyltrazolium chloride staining methods. Postischemic left ventricular contractile function was better recovered in the CoQ(10) group than in the control group of pigs. CoQ(10)-fed pigs revealed less myocardial infarction and less creatine kinase release from the coronary effluent compared with control pigs. The experimental group also demonstrated a smaller amount of malonaldehyde in the coronary effluent and a higher content of the endogenous antioxidants ascorbate and thiol. Significant induction of the expression of ubiquitin mRNA was also found in the hearts of the CoQ(10)-fed group. The results of this study demonstrate that nutritional supplementation of CoQ(10) renders the hearts resistant to ischemia-reperfusion injury, probably by reducing the oxidative stress.     

Studies on the involvement of lipolytic enzymes in endogenous lipolysis of the isolated rat heart

Rat hearts were depleted in vivo from both the heparin-releasable lipoprotein lipase and heparin-resistant tissue neutral triacylglycerol lipase activity by treatment of the animals with cycloheximide (2 mg/kg body weight), intraperitoneally injected 2.5 and 5 h prior to perfusion. The tissue acid lipase, mono- and diacylglycerol lipase activities were not affected by cycloheximide-induced inhibition of protein synthesis. Myocardial basal and glucagon-stimulated lipolysis, determined by the rate of glycerol production and release from the isolated hearts, was not significantly different in control and cycloheximide-treated rats. Tissue triacylglycerols were recovered with the highest relative specific distribution in the lysosomal fraction isolated from heart homogenates. Upon prolongation of the perfusion-duration the relative specific distribution of triacylglycerols in the lysosomal fraction decreased. In addition, the specific lysosomal triacylglycerol content (micrograms/mg protein) dropped significantly, indicating an important role of lysosomes in myocardial triacylglycerol turnover. Our data strongly suggest that the heparin-resistant neutral triacylglycerol lipase activity may not be the only determinant of endogenous lipolysis in the isolated rat heart and indicate that lipolysis may additionally be mediated by the lysosomal, acid lipase in concert with the microsomal mono-and diacylglycerol lipase.     

Mechanical function and fatty acid oxidation in the neonatal pig heart with ischemia and reperfusion

We investigated mechanical function and exogenous fatty acid oxidation in neonatal pig hearts subjected to ischemia, followed by reperfusion. Isolated, isovolumically-beating hearts, from pigs 12 h to 2 days of age, were perfused with an erythrocyte-enriched (hematocrit approximately 15%) solution (37 degrees C). All hearts were studied for 30 min. with a perfusion pressure of 60 mmHg (pre-ischemia). One group of hearts (low-flow ischemia, N = 12) was then perfused for 30 min. with a perfusion pressure of approximately 12 mmHg. In the other group (no-flow ischemic arrest, N = 9), the perfusion pressure was zero for 30 min. Following ischemia in both groups, the perfusion pressure was restored to 60 mmHg for 40 min. (reperfusion). Pre-ischemia parameters for all hearts averaged: left ventricular peak systolic pressure, 99.0 +/- 2.0 mmHg; end diastolic pressure, 1.9 +/- 0.2 mmHg; coronary flow, 3.4 +/- 0.1 ml/min per g; myocardial oxygen consumption, 56.6 +/- 1.6 microliter/min per g and fatty acid oxidation, 33.4 +/- 1.4 nmol/min per g. During low-flow ischemia, hearts released lactate, and the corresponding parameters decreased to: 30.7 +/- 0.9 mmHg; 1.2 +/- 0.3 mmHg; 0.8 +/- 0.1 ml/min per g; 26.6 +/- 2.3 microliters/min per g and 12.9 +/- 1.1 nmol/min per g, respectively. Early in reperfusion in both groups, all parameters, except for fatty acid oxidation, exceeded pre-ischemia values, before recovering to near pre-ischemia values. Late in reperfusion, however, rates of fatty acid oxidation exceeded pre-ischemia rates by approximately 60%. Thus, the neonatal pig heart demonstrated similar recovery following 30 min of low-flow ischemia or no-flow ischemic arrest.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mice deficient in mitochondrial glycerol-3-phosphate acyltransferase-1 have diminished myocardial triacylglycerol accumulation during lipogenic diet and altered phospholipid fatty acid composition

Glycerol-3-phosphate acyltransferase-1 (GPAT1), which is located on the outer mitochondrial membrane comprises up to 30% of total GPAT activity in the heart. It is one of at least four mammalian GPAT isoforms known to catalyze the initial, committed, and rate-limiting step of glycerolipid synthesis. Because excess triacylglycerol (TAG) accumulates in cardiomyocytes in obesity and type 2 diabetes, we determined whether lack of GPAT1 would alter the synthesis of heart TAG and phospholipids after a 2-week high-sucrose diet or a 3-month high-fat diet. Even in the absence of hypertriglyceridemia, TAG increased 2-fold with both diets in hearts from wildtype mice. In contrast, hearts from Gpat1(-/-) mice contained 20-80% less TAG than the wildtype controls. In addition, hearts from Gpat1(-/-) mice fed the high-sucrose diet incorporate 60% less [(14)C]palmitate into heart TAG as compared to wildtype mice. Because GPAT1 prefers 16:0-CoA to other long-chain acyl-CoA substrates, we determined the fatty acid composition of heart phospholipids. Compared to wildtype littermate controls, hearts from Gpat1(-/-)(-/-) mice contained a lower amount of 16:0 in phosphatidylcholine, phosphatidylethanolamine, and phosphatidylserine/phosphatidylinositol and significantly more C20:4n6. Phosphatidylcholine and phosphatidylethanolamine from Gpat1(-/-)(-/-) hearts also contained higher amounts of 18:0 and 18:1. Although at least three other GPAT isoforms are expressed in the heart, our data suggest that GPAT1 contributes significantly to cardiomyocyte TAG synthesis during lipogenic or high-fat diets and influences the incorporation of 20:4n6 into heart phospholipids.     

The ischemic rat heart releases S100B

S100B is an astrocytic protein assessed in cerebrospinal fluid and serum as a biochemical marker of cerebral injuries. However, increasing evidences suggest the influence of extra cerebral sources on its serum levels. Since it was reported that the injured myocardium expresses S100B, we investigated whether the isolated heart releases this protein. The rat hearts were excised and perfused by the Langendorff technique of isolated heart perfusion. After stabilization, 10 hearts (ischemic group) were submitted to 20 minutes of ischemia and 30 minutes of reperfusion, and 5 hearts (control group) were submitted to 50 minutes of perfusion. The perfusion fluid was collected at pre-ischemia, and 0, 5, 10, 15 and 30 min after ischemia (or equivalent in controls) for S100B and cardiac troponin T (a heart injury marker) assays. In the ischemic group, S100B and troponin T levels increased significantly at time 0 min: S100B values [mug/L, median (IQ25/IQ75)] increased from < or = 0.02 (< or = 0.02/0.03) to 0.38 (0.22/0.84), while troponin T values [mug/L, median (IQ25/IQ75)] increased from 0.31 (0.15/0.45) to 2.84 (2.00/3.63). Our results point to the ischemic heart as an extra cerebral source of S100B.     

Heat stress pretreatment mitigates postischemic arachidonic acid accumulation in rat heart

Heat stress pretreatment of the heart is known to protect this organ against an ischemic/reperfusion insult 24 h later. Degradation of membrane phospholipids resulting in tissue accumulation of polyunsaturated fatty acids, such as arachidonic acid, is thought to play an important role in the multifactorial process of ischemia/reperfusion-induced damage.The present study was conducted to test the hypothesis that heat stress mitigates the postischemic accumulation of arachidonic acid in myocardial tissue, as a sign of enhanced membrane phospholipid degradation. The experiments were performed on hearts isolated from rats either 24 h after total body heat treatment (42°C for 15 min) or 24 h after sham treatment (control). Hearts were made ischemic for 45 min and reperfused for another 45 min.Heat pretreatment resulted in a significant improvement of postischemic hemodynamic performance of the isolated rat hearts. The release of creatine kinase was reduced from 30 ± 14 (control group) to 17 ± 5 units/g wet wt per 45 min (heat-pretreated group) (p < 0.05). Moreover, the tissue content of the inducible heat stress protein HSP70 was found to be increased 3-fold 24 h after heat treatment. Preischemic tissue levels of arachidonic acid did not differ between heat-pretreated and control hearts. The postischemic ventricular content of arachidonic acid was found to be significantly reduced in heat-pretreated hearts compared to sham-treated controls (6.6 ± 3.3. vs. 17.8 ± 12.0 nmol/g wet wt). The findings suggest that mitigation of membrane phospholipid degradation is a potential mechanism of heat stress-mediated protection against the deleterious effects of ischemia and reperfusion on cardiac cells.     

Roles of ferritin and iron in ischemic preconditioning of the heart

Iron and copper play major roles in biological systems, catalyzing free radical production and consequently causing damage. The relatively high levels of these metals, which are mobilized into the coronary flow following prolonged ischemia, have been incriminated as key players in reperfusion injury to the heart. In the present communication we investigated other roles of iron - providing protection to the ischemic heart via preconditioning (PC). PC was accomplished by subjecting isolated rat hearts to three episodes of 2 min ischemia separated by 3 min of reperfusion. Prolonged ischemia followed the PC phase. PC hearts (group I) were compared to hearts subjected to normal perfusion (group II, no ischemia) and to ischemia without PC (group III). Group I showed a marked improvement in the recovery of hemodynamic function vs. group III. Biochemical parameters further substantiated the PC protection provided to group I against prolonged ischemia. Correspondingly, group I presented markedly lower re-distribution and mobilization of iron and copper into the coronary flow, following prolonged ischemia, as evinced from the decrease in total levels, and in the 'free' fraction of iron and copper. During the PC phase no loss of cardiac function was observed. A small wave of re-distribution and mobilization of iron (typically less than 4-8% of the value of 35 min ischemia) was recorded. The cellular content of ferritin (Ft) measured in the heart was significantly higher in group I than in group III (0.90 and 0.54 microg/mg, respectively). Also, iron-saturation of Ft was significantly lower for PC hearts, compared to both groups II and III (0.22 vs. 0.32 and 0.31 microg/mg, for 35 min ischemia, respectively). These findings are in accord with the proposal that intracellular re-distribution and mobilization of small levels of iron, during PC, cause rapid accumulation of ferritin - the major iron-storage protein. It is proposed that iron play a dual role: (i) It serves as a signaling pathway for the accumulation of Ft following the PC phase. This iron is not involved in cardiac injury, but rather prepares the heart against future high levels of 'free' iron, thus reducing the degree of myocardial damage after prolonged ischemia. (ii) High levels of iron (and copper) are mobilized following prolonged ischemia and cause tissue damage.     

Reversibility of acute alcohol cardiac depression: 31P NMR in hamsters

Isolated hamster hearts were perfused with 2% ethanol for 30 min and then reequilibrated with control medium. One group of hamsters was pretreated with verapamil. Another group received diltiazem. Myocardial verapamil levels were 9.5 +/- 0.7 mg/g dry wt; diltiazem levels were 22 +/- 7 mg/g dry wt. Energy metabolites were assessed by using 31P NMR standardized with high-pressure liquid chromatography of freeze-clamped tissue. Intracellular calcium was measured by atomic absorption spectrophotometry, marking the extracellular space with K(CoEDTA). After 30 min of perfusion, untreated hamster hearts showed a 74% decrease in developed pressure, a marked increase in end-diastolic pressure, a decrease of ATP from 9.8 to 8.8 mmol, and an increase of Pi from 6.7 to 9.8 mmol, but no change of phosphocreatine (PCr) or intracellular pH (pHi). Verapamil pretreatment partially prevented cardiac depression during alcohol perfusion. Whereas diltiazem had no protective effect. After reequilibration, developed pressure and oxygen consumption significantly exceeded control values. ATP decreased to 8 mmol; pHi, PCr, and Pi showed no significant change. Verapamil-pretreated hearts showed better performance than untreated hearts without change in PCr and Pi, whereas ATP dropped slightly to 8.7 mmol. Thus, functional cardiac depression resulting from acute alcohol exposure is reversible. Increased intracellular calcium levels during alcohol exposure normalized after the removal of alcohol. There was no major change in high-energy phosphates during alcohol exposure or after the removal of alcohol. Verapamil protects the heart from functional depression during alcohol exposure without affecting energy resources.     

Lesions to the Corticostriatal Pathways Ameliorate Hypoglycemia-Induced Arachidonic Acid Release

The concentrations of free fatty acids (FFAs) in the neostriatum of control rats and rats subjected to unilateral cortical ablation were measured during and following severe insulin-induced hypoglycemia. The total FFA concentration in the caudate nucleus contralateral to the lesion increased to approximately 1.5 and 3 times the control level after 5 and 30 min of isoelectricity, respectively, and was similar to the control value following 1 h of recovery. After 5 min of isoelectricity, the total FFA pool was significantly smaller in the decorticated striatum. No difference between hemispheres was noted after 30 min of isoelectricity. After 5 min of isoelectricity the levels of stearic and arachidonic acid were selectively increased whereas palmitic acid and oleic acid remained at control levels. In the decorticated striatum of lesioned animals the arachidonic acid concentration was significantly lower, whereas the level of stearic acid was not significantly different from the control value. After 30 min of isoelectricity the levels of all four FFA species were increased. Apart from a significantly lower level of oleic acid on the decorticated side, there were no interhemispheric differences in the FFA levels. Since the early interhemispheric differences in the FFA levels. Since the early interhemispheric hemispheric differences in the levels of arachidonic and stearic acids coincide with a selective decrease in the levels of glutamate and a decreased energy utilization on the decorticated side, the results suggest that glutamate release during hypoglycemia induces an early receptor-mediated degradation of phospholipids, presumably via the phosphatidylinositol cycle.     

Roles of ferritin and iron in ischemic preconditioning of the heart

Iron and copper play major roles in biological systems, catalyzing free radical production and consequently causing damage. The relatively high levels of these metals, which are mobilized into the coronary flow following prolonged ischemia, have been incriminated as key players in reperfusion injury to the heart. In the present communication we investigated other roles of iron – providing protection to the ischemic heart via preconditioning (PC).PC was accomplished by subjecting isolated rat hearts to three episodes of 2 min ischemia separated by 3 min of reperfusion. Prolonged ischemia followed the PC phase. PC hearts (group I) were compared to hearts subjected to normal perfusion (group II, no ischemia) and to ischemia without PC (group III). Group I showed a marked improvement in the recovery of hemodynamic function vs. group III. Biochemical parameters further substantiated the PC protection provided to group I against prolonged ischemia. Correspondingly, group I presented markedly lower re-distribution and mobilization of iron and copper into the coronary flow, following prolonged ischemia, as evinced from the decrease in total levels, and in the 'free' fraction of iron and copper.During the PC phase no loss of cardiac function was observed. A small wave of re-distribution and mobilization of iron (typically less than 4–8% of the value of 35 min ischemia) was recorded. The cellular content of ferritin (Ft) measured in the heart was significantly higher in group I than in group III (0.90 and 0.54 g/mg, respectively). Also, iron-saturation of Ft was significantly lower for PC hearts, compared to both groups II and III (0.22 vs. 0.32 and 0.31 g/mg, for 35 min ischemia, respectively). These findings are in accord with the proposal that intracellular re-distribution and mobilization of small levels of iron, during PC, cause rapid accumulation of ferritin – the major iron-storage protein.It is proposed that iron play a dual role: (i) It serves as a signaling pathway for the accumulation of Ft following the PC phase. This iron is not involved in cardiac injury, but rather prepares the heart against future high levels of 'free' iron, thus reducing the degree of myocardial damage after prolonged ischemia. (ii) High levels of iron (and copper) are mobilized following prolonged ischemia and cause tissue damage.     

β(2)-adrenoreceptor mediates the cardioprotection of ischemic preconditioning on myocardial contraction in rats subjected to ischemia/reperfusion injury

The aim of the present study is to investigate the role of beta(2)-adrenoreceptor (beta(2)-AR) in ischemic preconditioning (IP) in isolated rat heart model of ischemia/reperfusion (I/R). Sprague-Dawley rat hearts were quickly removed, mounted on Langendorff apparatus, and perfused with Krebs-Henseleit (KH) solution. After the initial stabilization period, the rats were randomly divided into 6 groups including control group (perfused for an additional 20 min), IP group (4 cycles of 5 min of ischemia followed by 5 min of reflow), isoproterenol (ISO) group (10 nmol/L ISO perfusion for 5 min followed by 5 min washout), IP + ICI118551 group (55 nmol/L ICI118551 perfusion for 5 min before and throughout IP), ISO + ICI118551 group (55 nmol/L ICI118551 perfusion for 5 min before and throughout ISO treatment), ICI118551 group (55 nmol/L ICI118551 perfusion for 20 min). After these treatments, all hearts were followed by 30 min of no-flow ischemia and 30 min of reperfusion. A computer-based electrophysiological recorder system was used to measure changes of the maximal rate of pressure increase in systole phase (+dp/dt(max)), maximal rate of pressure decrease in diastole phase (-dp/dt(max)), and difference of left ventricular pressure (DeltaLVP). Then cardiomyocytes from these hearts were isolated by 5 min of Ca(2+)-free buffer perfusion and 25 min of collagenase perfusion. The ventricles were chopped and filtered. The myocytes were resuspended in KB buffer. The contraction and the viability of cardiomyocytes were measured. Lactate dehydrogenase (LDH) concentration in coronary effluent was assayed with assay kit. The results showed that both IP and ISO significantly increased the values of +/-dp/dt(max), DeltaLVP, the contraction and viability of cardiomyocytes, shortened the time-to-peak contraction (TTP), and decreased the release of LDH in coronary effluent. ICI118551, a selective beta(2)-AR antagonist, blocked these effects. Either the time-to-50% relaxation (R(50)) or the time-to-100% relaxation (R(100)) had no significant differences between groups. Our results indicate that the cardioprotection of IP was mediated by beta(2)-AR in isolated rat hearts subjected to I/R injury.     

Effects of insulin on the metabolism of the isolated working rat heart perfused with undiluted rat blood

Working rat hearts were perfused with either buffer or with defibrinated, undiluted rat blood dialyzed to remove vasoconstrictor factors. With precautions taken for sterility in the preparation of the perfusate and the apparatus, hearts were obtained which were stable as judged by stroke rate and cardiac output. In these hearts, cardiac output and coronary flow averaged 46.0 and 1.7 ml/g heart per min, respectively. Perfusion with erythrocyte-free buffer depressed cardiac output by 30%, while coronary flow averaged 8.8 ml/g of heart per min. The mean stroke rate of blood-perfused hearts was 300 beats/min but only 240 beats/min during buffer perfusion. In blood-perfused hearts, insulin did not alter stroke rate but significantly lowered coronary flow. The hormone caused a transient increase in cardiac output in hearts perfused with buffer. Insulin did not alter glucose uptake in buffer-perfused hearts but increased lactate release in perfusions with blood. Both serum fatty acids and triacylglycerol fatty acids were significant metabolic fuels in hearts perfused with undiluted blood. The preparation described would appear to be potentially useful for the study of myocardial metabolism in vitro.