Cadmium-induced attenuation of coronary blood flow in the perfused rat heart.

The present studies were undertaken to test the hypothesis that cadmium alters myocardial metabolism, producing effects on coronary blood flow. Hearts isolated from adult, male Sprague-Dawley rats were control-equilibrated and then treated with Cd (0.1-100 microM). Myocardial O2 consumption (MVO2), lactate production, contractile activity, and coronary flow were determined at regular intervals. Parallel experiments were also performed in hearts administered 0.5 mM KCN to compare the effects of cadmium on myocardial lactate release with that of a known inhibitor of mitochondrial respiration. Coronary flow decreased significantly in response to Cd. This effect was dose-related (ED50 = 0.4 microM Cd), achieving maximal levels within the initial 5 min of exposure and persisting thereafter. Cd treatment also significantly decreased myocardial contractile activity [ED50 = 2.4 microM Cd (+dP/dt)] and MVO2 in a dose-dependent manner. This latter effect was time-dependent with an ED50 value of 2.1 microM at 5 min, as compared to a value of 0.5 microM Cd at 30 min. Lactate levels measured in the coronary effluent were unaffected by Cd treatment, except at the highest dose. Collectively, the results of this study failed to support a metabolic mechanism as the basis for the observed changes in coronary flow in response to cadmium administration. Instead, these results suggest an alternative hypothesis that Cd disturbs coronary flow via a mechanism involving direct actions on the coronary vasculature.

Differential effects of fatty acids on glycolysis and glycogen metabolism in vascular smooth muscle.

The effects of fatty acids of different chain lengths on aerobic glycolysis, lactic acid production, glycogen metabolism and contractile function of vascular smooth muscle were investigated. Porcine carotid artery segments were treated with 50 microM iodoacetate and perchloric acid tissue extracts were then analyzed by 31P-NMR spectroscopy to observe the accumulation of phosphorylated glycolytic intermediates so that the activity of the Embden-Myerhof pathway could be tracked under various experimental paradigms. Aerobic glycolysis and lactate production in resting arteries were almost completely inhibited with 0.5 mM octanoate, partially inhibited with 0.5 mM acetate and unaffected by 0.5 mM palmitate. Inhibition of glycolysis by octanoate was not attributable to inhibition of glucose uptake or glucose phosphorylation. Basal glycogen synthesis was unchanged with palmitate and acetate, but was inhibited by 52% with octanoate incubation. The characteristic glycogenolysis which occurs upon isometric contraction with 80 mM KCl in the absence of fatty acid in the medium was not demonstrable in the presence of any of the fatty acids tested. Glycogen sparing was also demonstrable in norepinephrine contractions with octanoate and acetate, but not with palmitate. Additionally, norepinephrine-stimulated isometric contraction was associated with enhanced synthesis of glycogen amounting to 6-times the basal rate in medium containing octanoate. Contractile responses to norepinephrine were attenuated by 20% in media containing fatty acids. Thus, fatty acids significantly alter metabolism and contractility of vascular smooth muscle. Fatty acids of different chain lengths affect smooth muscle differentially; the pattern of substrate utilization during contraction depends on the contractile agonist and the fatty acid present in the medium.

Phosphatic metabolites, intracellular pH and free [Mg2+] in single, intact porcine carotid artery segments studied by 31P-NMR.

Superfused porcine carotid artery segments (approximately 7 cm lengths) were analyzed by 31P-NMR spectroscopic methods to characterize the 31P spectrum of arterial smooth muscle and to determine the influence of passive stretch (intraluminal pressurization, 95-100 mmHg) on cellular phosphatic metabolite levels, intracellular pH and free magnesium concentration ([Mg2+free]i). Equilibrated, single, intact arteries were studied under steady-state, constant flow conditions at 37 degrees C. Phosphoethanolamine, phosphocholine, inorganic phosphate (Pi), phosphocreatine (PCr) and nucleoside triphosphates (NTP), primarily ATP, were the principle metabolites detected in the 31P-NMR spectrum of intact arterial smooth muscle. The concentration of these metabolites and intracellular pH, as determined from the referenced chemical shift of Pi, were unaffected by pressurization. The PCr:Pi ratios determined for nonpressurized (flaccid) and pressurized arteries were 1.2 +/- 0.1 and 1.3 +/- 0.3, respectively. Intracellular pH averaged 7.02 +/- 0.02 (mean +/- 1 S.D.) for flaccid arteries vs. 7.03 +/- 0.05 for pressurized arteries. The upfield chemical shift of the beta-ATP peak, which has been described in other types of smooth muscle, was also observed in these experiments. Interestingly, pressurization significantly shifted the resonance position of this peak, which was interpreted to represent a change in [Mg2+free]i. The average [Mg2+free]i of flaccid artery preparations was computed to be 0.54 +/- 0.03 x 10(-3) M, as compared to 0.99 +/- 0.10 x 10(-3) M for pressurized arteries. This change in [Mg2+free]i was evident within the first hour following pressurization and persisted thereafter. These findings suggest that altering the resting length of vascular smooth muscle produces a change in [Mg2+free]i. This shift in free Mg2+ levels may act as a metabolic signal triggering a change in vascular smooth muscle metabolism, an effect which has been reported to occur in smooth muscle in response to stretch.

Effects of altering carbohydrate metabolism on energy status and contractile function of vascular smooth muscle.

Substrate-dependent changes in vascular smooth muscle energy metabolism and contractile function were investigated in isolated porcine carotid arteries. In media containing glucose glycogen catabolism accounted for all the estimated high-energy phosphate turnover that occurred in conjunction with contraction induced by 80 mM KCl. However, in glucose-free media glycogen catabolism accounted for only a portion of the estimated ATP utilization in resting and contracting arteries, even though glycogen stores were not depleted. The glycogenolysis and lactate production that ordinarily accompanies contraction was completely inhibited by 5 mM 2-deoxyglucose (2-DG). However, there was no decrease in the high-energy phosphate levels when compared to control resting arteries similarly treated with 2-DG. The results suggest that an endogenous non-carbohydrate source may be an important substrate for energy metabolism. Treatment of arteries with 50 microM iodoacetate (IA) in media containing glucose resulted in a marked reduction of high energy phosphate levels and an accumulation of phosphorylated glycolytic intermediates, as demonstrated by 31P-NMR spectroscopy. In glucose-free media, 50 microM IA had only a slight effect on high-energy phosphate levels, while glycogenolysis proceeded unhindered. With 1 mM IA in glucose-free media, the oxidative metabolism of glycogen was inhibited as evidenced by the depletion of high-energy phosphates and the appearance of sugar phosphates in the 31P-NMR spectra. Thus, the titration of enzyme systems with IA reveals a structural partitioning of carbohydrate metabolism, as suggested by previous studies.

Species-dependent differences in ATP half-life of glucose deprived crystalline lens.

1. Significant differences were detected by 31P NMR spectroscopic techniques in the intrinsic rate of ATP hydrolysis by crystalline lenses from adult Macaca mulatta, Saimiri sciureus, Felis catus, Sus, Canis familiaris, Cavia rodentia and Oryctalagis cuniculus during exogenous glucose deprivation. 2. These differences were not attributable to differences in endogenous glycogen stores, and appear instead to emanate from comparative species differences in lenticular enzyme activities.

Improved postischemic recovery of cardiac pump function in exercised trained diabetic rats.

This study examined whether exercise training of diabetic rats (streptozocin, 55 mg/kg) would affect the ability of perfused hearts to recover pump function after 75 min of ischemia and 30 min of reperfusion. All hearts were perfused with buffer containing the diabetic plasma concentrations of glucose and free fatty acids. Four groups were studied: sedentary control, trained control, sedentary diabetic, and trained diabetic. Trained control and diabetic rats were exercised on a treadmill at 5% grade, 21 m/min, 90 min/day, 6 days/wk for 8 wk. Sedentary diabetics had significantly lower body weight and elevated plasma glucose, triacylglycerol, and cholesterol relative to both control groups. Hearts from this group exhibited depressed postischemic recovery of pump function during reperfusion. In contrast, trained diabetic rats exhibited significantly lower plasma levels of triacylglycerol and cholesterol relative to sedentary diabetics. The postischemic recovery of cardiac pump function was improved in hearts from trained diabetic rats relative to sedentary diabetics. Exercise training had no effect on control rats. These results suggest that exercise training produces an antihyperlipidemic effect in diabetic rats and improves the tolerance of the diabetic heart to ischemia.

Cardiovascular actions of lead and relationship to hypertension: a review.

Chronic and acute lead poisoning cause overt, clinical symptoms of cardiac and vascular damage with potentially lethal consequences. Morphological, biochemical, and functional derangements of the heart have all been described in patients following exposure to excessive lead levels. Disturbances in cardiac electrical and mechanical activity and postmortem evidence of morphological and biochemical derangements of the myocardium have all been reported following excessive exposure to lead in humans. In addition, signs of vascular degeneration, abnormal vascular smooth muscle function, and altered vessel compliance have been described in humans chronically and acutely exposed to toxic lead levels. Similar cardiovascular complications have been detected following excessive lead exposure in experimental animals. Myocarditis, electrocardiographic disturbances, heightened catecholamine arrhythmogenicity, altered myocardial contractile responsiveness to inotropic stimulation, degenerative structural and biochemical changes affecting the musculature of the heart and vasculature, hypertension, hypercholesterolemia, atherosclerosis, and increased vascular reactivity to alpha-adrenergic agonists have been among the reported cardiovascular disturbances linked to lead poisoning. Less certain are the cardiovascular effects of subclinical lead poisoning. Although controversial, chronic low-level lead exposure has been linked to hypertension and other cardiovascular disturbances in both clinical and experimental studies. In general, it can be concluded that lead over a wide range of exposure intensities can induce significant changes in the function of the cardiovascular system. Evidence points to the involvement of multiple sites of action. Cardiac and vascular sites, as well as sites within the central nervous system, have all been implicated in the sequelae of cardiovascular effects. The exact pathogenic mechanisms that underlie the actions of lead in the cardiovascular system, however, have yet to be elucidated definitively.(ABSTRACT TRUNCATED AT 250 WORDS)

Depressed in vivo myocardial reactivity to dobutamine in streptozotocin diabetic rats: influence of exercise training.

To assess the effects of chronic diabetes on in vivo myocardial reactivity to beta 1 adrenergic receptor stimulation and to evaluate the therapeutic effect of exercise training in preventing the cardiac abnormalities induced by diabetes four groups of rats were studied: sedentary control, trained control, sedentary diabetic, and trained diabetic. Trained rats were adapted to treadmill running before the induction of diabetes with streptozotocin 55 mg.kg-1 iv. The duration, speed, and grade of exercise were then progressively increased during eight weeks of training until the rats could run for 90 min at 18 m/min, 5% grade. A training effect was confirmed by an increase in plantaris muscle cytochrome oxidase activity. In vivo cardiac contractile performance was assessed by intracardiac catheterisation. Heart rate, left intraventricular peak systolic pressure, and positive and negative dP/dt were measured under basal conditions and after the intravenous administration of dobutamine 10(-10) to 5 x 10(-7) mol.kg-1 body weight. Under basal conditions, there were no differences among the four groups in left intraventricular peak systolic pressure, positive dP/dt, and heart rate, but negative dP/dt was lower in both diabetic groups. The response to dobutamine of the sedentary diabetic group, as reflected in the measured cardiodynamic variables, was significantly attenuated compared with that of the sedentary control group. Exercise training tended to improve cardiac function towards the level detected in the sedentary controls; however, the differences between sedentary and trained diabetic groups were not statistically significant. Exercise training also did not significantly alter the response of the control group to dobutamine.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of rat heart mitochondrial respiration by cadmium chloride.

Mitochondria were isolated from hearts obtained from adult male Sprague-Dawley rats by two-part differential centrifugation of heart homogenates. Time-dependent (0-120 sec) and concentration-dependent (0-10 microM CdCl2) effects of cadmium on pyruvate-malate-supported state 3 and state 4 respiration were measured in a constant temperature reaction chamber at 37 degrees C, according to established procedures. The ID50 for cadmium chloride on state 3 respiration was determined to be 4.2 microM. The inhibition produced by cadmium chloride in heart mitochondria was compared, using identical procedures, to the effects induced by two compounds, sodium atractyloside and potassium cyanide, which are known to alter mitochondrial respiration at specific sites. The calculated ID50 values for these agents in heart mitochondria were 1.8 and 16 microM, respectively. The concentration-dependent inhibition of mitochondrial respiration induced by either cadmium chloride or potassium cyanide was maintained in the presence of 50 microM carbonyl cyanide m-chlorophenylhydrazone (CCCP), a known uncoupling agent. In contrast, sodium atractyloside did not block the uncoupling effect of 50 microM CCCP. In addition cadmium chloride was also shown to inhibit CCCP-uncoupled mitochondrial respiration. The cadmium-induced inhibition of mitochondrial respiration was reversed partially by cysteine and completely by 2,3-dimercaptopropanol. The results of the present study indicate that, at all concentrations, cadmium chloride acted solely as an inhibitor of rat heart pyruvate-malate-supported mitochondrial respiration. These findings suggest a possible mechanism for the reported disturbances in myocardial metabolism and function that occur in conjunction with acute and chronic cadmium exposure in humans and experimental animals.

31P nuclear magnetic resonance studies of phosphoglyceride metabolism in developing and degenerating brain: preliminary observations.

31P nuclear magnetic resonance (NMR) studies were conducted on perchloric acid extracts of the brain of one control, two Huntington's disease (HD), one probable Alzheimer's disease (AD), and one AD patient. These studies demonstrated significant elevations (over control) in the levels of phosphomonoesters in all brain areas of the patients with HD and AD even in areas devoid of neuropathological findings. Elevations of phosphodiesters were also observed, but they tended to reflect the degree of neuropathological change. We postulate that the 31P NMR findings represent molecular alterations with corresponding metabolic correlates which either antedate or occur in the absence of changes in cellular morphology or structure. As such the 31P NMR findings may reflect a subcellular "molecular neuropathology."

Myocardial adaptation to endurance exercise training in diabetic rats.

The purpose of this study was to determine whether exercise training would prevent the progressive functional decline in pump function of hearts from diabetic rats. Four groups were studied: sedentary control, trained control, sedentary diabetic, and trained diabetic. Trained rats were adapted to the treadmill prior to induction of diabetes in half of the group streptozotocin injected (50 mg/kg). Thereafter the duration, speed, and grade were then progressively increased until the trained rats could run for 60 min at 27 m/min, 5% grade (wk 8). Cardiac output and work were measured in isolated working hearts perfused at various left atrial filling pressures and with buffer containing the concentrations of glucose and fatty acids found in vivo. Sedentary diabetic rats had lowered body weight, elevated plasma glucose, triacylglycerol, and cholesterol. Exercise training of diabetic rats lowered plasma triacylglycerol levels. Training increased plantaris muscle cytochrome oxidase activity significantly in both the trained control and trained diabetic groups. Cardiac pump function was impaired in hearts from the sedentary diabetic rats perfused with either normal or diabetic substrate conditions, but the impairment was larger under diabetic conditions. Training of diabetic rats prevented this depression. Myocardial carnitine content was decreased in hearts from sedentary diabetic rats. Exercise training increased carnitine content in both control and diabetic rats. This training protocol did not affect cardiac pump function of the trained control group. These results suggest that exercise training may limit the myocardial contractile dysfunction associated with diabetes mellitus.

Effects of vanadate on in vivo myocardial reactivity to norepinephrine in diabetic rats.

Myocardial contractile function is often depressed in patients with diabetes mellitus. Vanadate is an essential trace element that has purportedly an insulin-like action and has been suggested as a therapeutic agent for the treatment of diabetes mellitus. The purpose of the present study was to compare the prophylactic efficacy of oral vanadate therapy (0.8 mg of sodium orthovanadate per milliliter drinking water) to that of insulin treatment (5 units/day s.c.) in terms of its ability to reduce or prevent the progressive cardiodepression that occurs in untreated diabetes mellitus. Diabetes was induced in male rats by i.v. streptozotocin injection (50 mg/kg). Diabetes rats were assigned randomly to one of three regimens for 8 weeks: untreated, insulin-treated or vanadate-treated. Noninjected rats served as controls. In vivo myocardial contractile function was measured under basal conditions and after i.v. norepinephrine infusions in ketamine-xylazine-anesthetized rats using a miniature catheter-tip pressure transducer inserted in the right carotid artery and advanced into the left ventricle. Vanadate and insulin treatment resulted in comparable increases in body weight and reductions in plasma glucose, which were improved relative to untreated diabetics. These findings suggest that vanadium may possess an insulin-like action. Basal in vivo myocardial contractile performance was depressed significantly in untreated diabetic rats as compared to control and insulin-treated diabetic rats. The contractile performance of vanadate-treated diabetic rats was in between untreated diabetic and control groups. In vivo myocardial reactivity to norepinephrine based on assessments of left intraventricular developed pressure, positive and negative dP/dt and delta dP/dt was depressed significantly in untreated diabetic rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Impaired in vivo myocardial reactivity to norepinephrine in diabetic rats.

The effects of long-term diabetes with and without insulin treatment on in vivo myocardial contractile activity were studied under basal conditions and as a function of intravenously infused norepinephrine. Diabetes was induced by iv injection of streptozotocin (50 mg/kg). Insulin-treated diabetic rats received 5 units per day of isophane insulin suspension. The duration of the study was 8 weeks. In vivo myocardial contractility measurements were performed in ketamine-xylazine-anesthetized rats using a miniature catheter-tip pressure transducer advanced through the right carotid artery into the left ventricle. Peak positive dP/dt and intraventricular developed pressure were comparable among the groups when measured under basal conditions; however, the magnitude of the response to variable doses of norepinephrine (6 X 10(-12) to 6 X 10(-8) mole/kg body wt) were significantly diminished in diabetic rats, but the sensitivity was unchanged. Negative dP/dt was decreased under basal conditions and in response to norepinephrine in diabetic rats. Insulin treatment to diabetic rats prevented these changes, but heart rate was elevated. These results demonstrate that the in vivo cardiovascular reactivity of diabetic rats to norepinephrine is significantly attenuated.

31P NMR studies of the intact perfused rat heart: a novel analytical approach for determining functional-metabolic correlates, temporal relationships, and intracellular actions of cardiotoxic chemicals nondestructively in an intact organ model.

Intact hearts isolated from adult male, Sprague-Dawley rats were perfused under standardized conditions in an apparatus designed for use in a high-resolution nuclear magnetic resonance (NMR) spectrometer system. Myocardial phosphate metabolite concentrations (ATP, PCr, Pi, and phosphomonoesters) and intracellular pH were determined sequentially at timed intervals coincident with the functional assessments of the intact heart by phosphorus-31 (31P) NMR spectroscopic methods. Myocardial functional and metabolic parameters were unaffected by sustained control perfusion (2 hr). The negative inotropic actions of cadmium were associated with significant changes in the chemical environment of inorganic phosphate (Pi) within the cells. This initial cellular response to cadmium, which correlated with the onset and magnitude of the contractile disturbances, appeared to represent the formation of an acidic, intracellular Pi pool (pH, 6.0). This pH compartment reached a steady state during the period in which maximal changes in contractile function were manifested, and before cellular ATP and PCr concentrations were altered. These findings are consistent with the interpretation that the functional deficits caused by cadmium originated primarily from changes in the chemical environment experienced by intracellular metabolites, rather than changes in the amounts of cellular high energy substrates. In contrast, the time-dependent negative inotropic effects of arsenate were proportional to the loss of cellular ATP stores. Intracellular pH was not affected in these hearts. A distinctive metabolic finding associated with the cardiotoxicity of arsenate was the time-dependent accumulation of previously undetected phosphate metabolites in the arsenate-treated hearts. Efforts to chemically identify these metabolites proved inconclusive; however, existing evidence suggests the possibility that these phosphorus-containing compounds may be arsenophosphate derivatives of naturally occurring cellular metabolites. The present findings provide experimental evidence demonstrating that toxicologic assessments in an intact organ model are feasible using whole organ 31P NMR spectroscopic methods and that meaningful, new insights regarding the biochemical mechanisms responsible for the cardiotoxic actions of xenobiotic agents can be obtained by this analytical approach.