Fatty acid effects on calcium influx and efflux in sarcoplasmic reticulum vesicles from rabbit skeletal muscle.

Low concentrations of fatty acids inhibited initial Ca uptake by sarcoplasmic reticulum vesicles, the extent of inhibition varying with chain length and unsaturation in a series of C14-C20 fatty acids. Oleic acid was a more potent inhibitor of initial Ca uptake than stearic acid at 25 degrees C, whereas at 5 degrees C there was less difference between the inhibitory effects of low concentrations of these fatty acids. When the fatty acids were added later, during the phase of spontaneous Ca release that follow Ca uptake in reactions carried out at 25 degrees C 1-4 microM oleic and stearic acids caused Ca content to increase. This effect was due to marked inhibition of Ca efflux and slight stimulation of Ca influx. At concentrations of greater than 4 microM, both fatty acids inhibited the Ca influx that occurs during spontaneous Ca release; in the case of oleic acid, this inhibition resembled that of initial Ca uptake at 5 degrees C. The different effects of fatty acids at various times during Ca uptake reactions may be explained in part if alterations in the physical state of the membranes occur during the transition from the phase of initial Ca uptake to that of spontaneous Ca release.

Oxidative stress during reperfusion of human hearts: potential sources of oxygen free radicals.

OBJECTIVE: The aim was to examine the role of neutrophil activation in the genesis of oxidative stress during the early phases of reperfusion after ischaemia in patients subjected to aortocoronary bypass grafting. METHODS: Ten selected patients were studied. All had normal ejection fraction and normal left ventricular end diastolic pressures before operation. Each patient required at least three grafts, so that the duration of aortic crossclamping exceeded 30 min, the minimum ischaemic period required to detect oxidative stress upon reperfusion. Oxidative stress was assessed by measuring the formation and release of oxidised glutathione (GSSG) in the coronary sinus 1 min before and 3 min after the start of the cardiopulmonary bypass, and then 1, 5, 10, and 20 min after removal of the aortic clamp, and again 5 and 10 min after the end of the cardiopulmonary bypass. The arterial-coronary sinus difference for neutrophils, elastase-alpha 1 protease complex (elastase), and creatine phosphokinase was also monitored at the same intervals. RESULTS: Before clamping GSSG was undetectable in arterial and coronary sinus blood. There was no significant arterial-coronary sinus difference for neutrophils or elastase [53(SEM 66) cell.ml-1 and 1.10(2.49) micrograms.litre-1, respectively[. Five minutes after re-establishment of coronary blood flow, there was both a release of GSSG into the coronary sinus [arterial-coronary sinus difference: 11(2.6) nmol.dl-1] and an accumulation of neutrophils in the heart [arterial-coronary sinus difference: 262(33), P < 0.01 cell.ml-1], whereas no elastase release from the heart was measured [arterial-coronary sinus difference 7.6(4.46) microgram.litre-1, NS]. The arterial levels of elastase increased progressively during the operation from 48(5) microgram.litre-1 (preclamping) to 405(62) microgram.litre-1, P < 0.01 (end of the cardiopulmonary bypass). CONCLUSIONS: These data indicate that, in man, neutrophils do accumulate in the myocardium during early reperfusion. However, they are not activated when oxidative stress occurs. It is unlikely that the neutrophil localisation in the heart has pathological significance in the production of oxygen free radicals during early reperfusion. Free radical accumulation in the coronary vessels may contribute to disorders of coronary flow associated with reperfusion.

Cellular actions and pharmacology of the calcium channel blocking drugs.

The calcium channel blockers represent a group of diverse chemical structures that block calcium-selective channels in the plasma membranes of a variety of excitable cells. As the calcium fluxes carried by these channels allow the calcium ion (Ca2+) to gain access to the cell interior, where calcium serves as an activator messenger, calcium channel blockers generally act to inhibit cell function. By reducing the depolarizing currents caused by the entry of positively charged Ca2+ into the negatively charged interior of resting cells, the calcium channel blockers also inhibit excitatory processes that depend on calcium entry across the plasma membrane. These principles account for most of the effects of calcium channel blockers on the cardiovascular system. The calcium channel blockers inhibit contractile function in the heart and vascular smooth muscle and, because the initial depolarizing currents in the sinoatrial and atrioventricular nodes are carried by calcium channels, slow the heart rate and prolong atrioventricular conduction. The negative inotropic and vasodilatory effects of the calcium channel blockers, both of which can reduce systemic blood pressure, offer a theoretic basis for their potential use in the treatment of hypertension. The tissue specificity exhibited by some of the calcium channel blockers may enhance their therapeutic value in selected hypertensive patients. Of the three calcium channel blockers now available for use in the United States (diltiazem, nifedipine, and verapamil), diltiazem and verapamil are approximately equipotent in inhibiting calcium channel function in the heart and vascular smooth muscle, whereas nifedipine is more potent in smooth muscle. This tissue specificity can be used to advantage in the management of hypertension. These pharmacologic principles underlie the growing appreciation of the potential value of the calcium channel blockers in the treatment of hypertension.

Ventricular ectopic activity: prevalence and risk.

Ambulatory electrocardiographic monitoring and the frequent use of stress electrocardiography have been important tools in characterizing the prevalence and prognostic importance of ventricular ectopic activity in both healthy persons and patients with organic heart disease. These studies have demonstrated that ventricular ectopy is not uncommon in persons with no evidence of heart disease. However, it is rarely of high density or repetitive, and even when frequent or repetitive, or both, carries little, if any, risk of sudden death in patients without syncope. However, in patients with organic heart disease and in certain clinical settings, frequent and repetitive ventricular ectopy identifies a population at high risk for arrhythmia-induced syncope or sudden death. These rhythm disturbances have particular prognostic importance in ischemic heart disease with depressed left ventricular function and hypertrophic cardiomyopathy. Patients with presyncope or syncope and structural heart disease who demonstrate frequent and repetitive ventricular ectopy are also a high-risk group. Therefore, individual risk stratification is important in deciding whether and how to treat patients with ventricular ectopy.

Combined assessment of myocardial perfusion and left ventricular function with exercise technetium-99m sestamibi gated single-photon emission computed tomography can differentiate between ischemic and nonischemic dilated cardiomyopathy.

The purpose of this study was to determine whether exercise technetium-99m sestamibi gated single-photon emission computed tomography (SPECT) accurately distinguishes between patients with ischemic cardiomyopathy and patients with nonischemic left ventricular systolic dysfunction. Noninvasive tests have previously failed to accurately separate patients with ischemic cardiomyopathy from those with nonischemic cardiomyopathy. Technetium-99m gated SPECT imaging offers advantages that have the potential to overcome the limitations of previous studies. Thirty-seven adults with a left ventricular ejection fraction < or = 35%, including 24 patients with nonischemic cardiomyopathy and 13 patients with ischemic cardiomyopathy, were prospectively evaluated using symptom-limited metabolic exercise treadmill testing with technetium-99m sestamibi gated SPECT imaging. Interpretation of myocardial perfusion and regional wall motion was performed, using a 17-segment model. Summed stress, rest, and reversibility perfusion defect scores were determined, and the variance of segmental wall motion scores was computed. Summed stress, rest, and reversibility perfusion defect scores were significantly lower in nonischemic cardiomyopathy patients, compared with those with ischemic cardiomyopathy (summed stress defect score: 6.9 +/- 3.8 vs 32.9 +/- 7.7, respectively, p <0.001). Variability in segmental wall motion was also significantly lower in patients with nonischemic cardiomyopathy compared with those with ischemic cardiomyopathy (variance: 0.3 +/- 0.3 vs 1.2 +/- 0.8, respectively, p <0.001). Thus, assessment of myocardial perfusion and regional ventricular function with exercise technetium-99m sestamibi gated SPECT imaging can reliably distinguish between patients with ischemic cardiomyopathy and patients with nonischemic dilated cardiomyopathy.

Verapamil interaction with the muscarinic receptor: stereoselectivity at two sites.

Verapamil, in addition to blocking calcium channels, exhibits such "non-specific" effects on myocardium as inhibition of sodium and potassium conductances and modifications of muscarinic receptor-ligand interactions. To characterize further the effects of verapamil on the cardiac muscarinic receptor, we examined the abilities of the enantiomers of verapamil to modify the binding of the muscarinic antagonist [3H]quinuclidinyl benzilate ([3H]QNB) to purified canine sarcolemmal vesicles. Membranes were incubated with [3H]QNB and various concentrations of racemic, (+)-, or (-)- verapamil (25 or 37 degrees, pH 7.4), and reactions were terminated by rapid filtration. (-)-Verapamil (Ki of 5.3 +/- 0.2 microM) was twice as potent an inhibitor of equilibrium binding as (+)-verapamil (Ki of 11.4 +/- 0.6 microM), and this effect resulted from the ability of each enantiomer to slow [3H]QNB-receptor association. This degree of stereoselectivity, albeit at nanomolar concentrations, was similar to that observed for each enantiomer to compete for the specific phenylalkylamine site in this preparation. Verapamil also inhibited [3H]QNB-receptor dissociation, but this effect required high concentrations and demonstrated stereoselectivity opposite to that observed for association. These findings support the view that verapamil interacts with two distinct sites, possibly within membrane lipid, each with a different affinity and preference for (+)- and (-)-verapamil, to modify the muscarinic receptor.

Effect of phospholipid hydrolysis by phospholipase A2 on the kinetics of antagonist binding to cardiac muscarinic receptors.

Activation of phospholipases during prolonged myocardial ischemia could contribute to the functional derangement of myocardial cells by altering the phospholipid environment of a number of membrane bound proteins including receptors. The present study examined the kinetics of muscarinic receptor antagonist [3H]quinuclidinyl benzilate binding ([3H]QNB) to muscarinic receptors of highly purified sarcolemmal membranes under control conditions and after treatment with phospholipase A2 (PLA2; EC 3.1.1.4). Initial binding rates of QNB exhibited saturation kinetics, when plotted against the ligand concentration in control and PLA2 treated sarcolemmal membranes. This kinetic behaviour of QNB-binding is consistent with at least a two step binding mechanism. According to this two step binding hypothesis an unstable intermediate receptor-QNB complex (R*QNB) forms rapidly, and this form undergoes a slow conversion to the high affinity ligand-receptor complex R-QNB. The Michaelis constant Km of R-QNB formation was 1.8 nM, whereas the dissociation constant Kd obtained from equilibrium measurements was 0.062 nM. After 5 min exposure of sarcolemmal membranes to PLA2QNB binding capacity (Bmax) was reduced by 62%, and the affinity of the remaining receptor sites was decreased by 47% (Kd = 0.116 nM). This PLA2-induced increase of Kd was accompanied by a corresponding increase of Km, whereas the rate constants k2 and k-2 of the hypothetical slow conversion step (second reaction step) remained unchanged. These results suggest that binding of QNB to cardiac muscarinic receptors induces a transition in the receptor-ligand configuration, which is necessary for the formation of the final high affinity R-QNB complex. PLA2-induced changes of the lipid environment result in the inability of a part of the receptor population to undergo this transition, thereby inhibiting high affinity QNB-binding.

Utility of transesophageal echocardiography for the characterization of cardiovascular anomalies associated with Turner's syndrome.

This case illustrates the complementary value of transesophageal echocardiography to routine transthoracic echocardiography in an asymptomatic adult patient with Turner's syndrome. The combined findings of bicuspid aortic valve, severe aortic dilation, coarctation of the aorta, and type A aortic dissection were clearly delineated by transesophageal echocardiography.

Differential diagnosis of intracavitary tumors obstructing the right ventricular outflow tract.

Three cases of right ventricular outflow tract obstruction caused by 3 distinct tumors-myxoma, sarcoma, and presumed metastatic tumor-diagnosed by transthoracic and transesophageal echocardiography are presented. The differences among these 3 types of tumors with similar clinical and echocardiographic findings are highlighted, and a review of the pertinent literature is discussed. By applying the approximate frequencies of cardiac tumors categorized by type and site, statistically, an intracavitary right ventricular outflow tract tumor is 70 to 140 times more likely to be malignant than benign; furthermore, if it is a primary cardiac tumor, it is approximately 2 times more likely to be a sarcoma than a myxoma.

Aliphatic chain saturation and polar region influence amphiphile effects on sarcoplasmic reticulum calcium sequestration.

Calcium sequestration by skeletal muscle sarcoplasmic reticulum vesicles was enhanced by the addition of C18 saturated fatty acid to the reaction, while the cis-monounsaturated fatty acid and corresponding amine were potent inhibitors. The effects of these amphiphiles were found to depend on the degree of aliphatic chain saturation, the nature of the polar group, and the ratio of amphiphile:membrane phospholipid present in the reaction.

The possible role of endogenous amphiphiles in the membrane abnormalities of ischemic and reperfused myocardium.

Calcium entry into cardiac cells is believed to be controlled by transmembrane-voltage dependent, protein regulated "channels." The sarcoplasmic reticulum participates in the regulation of cytosolic calcium by ATP dependent Ca2+ sequestration during diastole, and by action potential stimulated calcium release. Massive calcium overloading occurs during reperfusion following myocardial ischemia. Calcium overloading activates phospholipases, which may activate another mechanism involved in lethal cellular injury, that is, the accumulation of long chain fatty acids and their derivatives. These compounds are soluble amphiphiles, and once liberated, they may insert into biological membranes and change membrane composition, physiology, and response to ions and drugs. Sarcoplasmic reticulum vesicles were used as an in vitro model to study the effects of palmitic acid, oleic acid, and palmitylcarnitine on the ability of this membrane system to sequester calcium within the vesicles. In the absence of phosphate, palmitic acid enhanced the ability of the vesicles to sequester calcium. Oleic acid and palmitylcarnitine inhibited calcium sequestration. In the presence of phosphate palmitic acid also inhibited the sequestration of calcium by sarcoplasmic reticulum, although not as severely as oleic acid and palmitylcarnitine. These results suggest that the disturbances in cellular calcium homeostasis following ischemia may be due, in part, to the incorporation of accumulated long chain fatty acids into membranes.

Hemodynamic complications of acute myocardial infarction.

These complications can be broken down into three major categories: the direct effects of ischemia, the effects of myocardial rupture or similar structural loss secondary to ischemic insult, and the effects of inhibitory autonomic reflexes triggered by infarction. It is critical to correlate the hemodynamic problem with its etiology in order to choose the appropriate treatment.

Effects of propranolol and timolol on calcium uptake by sarcoplasmic reticulum vesicles.

The effects of the beta-adrenergic receptor blocking agents propranolol and timolol on the initial calcium uptake velocity of sarcoplasmic reticulum vesicles of rabbit skeletal muscle were studied. Racemic d- and l-propranolol had similar inhibitory effects on initial calcium uptake velocity, which was inhibited 50% by 5--7 X 10(-4) M racemic propranolol. Timolol was a much less potent inhibitor of initial calcim uptake velocity; 50% inhibition occurred at approximately 10(-2) M timolol. Both drugs inhibited maximal calcium uptake velocity; however, KCa (the Ca2+ concentration at which calcium uptake was half-maximal) was modified differently. Propranolol increased KCa, whereas timolol caused the KCa to decrease. Addition of either drug to an ongoing calcium uptake reaction at the time that calcium content became maximal caused renewed calcium uptake. The relative potencies of propranolol and timolol as negative inotropic agents are similar to their potencies as inhibitors of sarcoplasmic reticulum calcium uptake, but dissimilar to their beta-adrenergic receptor blocking potencies. Timolol, which has been reported to have less negative inotropic effect than propranolol, is approximately 5 time more potent than propranolol as a beta-adrenergic receptor blocking agent but 15 times less potent as an inhibitor of sarcoplasmic reticulum calcium uptake. Inhibition of sarcoplasmic reticulum calcium uptake may thus characterize negative inotropic potencies of new beta-adrenergic receptor blocking agents.

Lipids and membrane function: implications in arrhythmias.

Increasing attention is being devoted to the relevance of lipid-membrane interactions to arrhythmias and their therapy. Particular emphasis is placed on alterations of transmembrane ion transport that can result from closure of protein channels, perhaps mediated by chemically induced changes in the phospholipid membrane bilayer. Such alterations may underlie antiarrhythmic drug actions.

Mechanisms of fatty acid effects on sarcoplasmic reticulum. I. Calcium-fatty acid interaction.

To elucidate the mechanism for the previously reported increase in calcium sequestration by sarcoplasmic reticulum vesicles in the presence of palmitic acid, the ability of this fatty acid to bind calcium was investigated by dual wavelength spectrophotometry using the calcium indicator arsenazo III,, and by Millipore filtration using 45CaCl2. In the presence of 120 mM KCl, 1 mM MgCl2, 15 microM CaCl2, and 40 mM histidine (pH 6.8, 25 degrees C), calcium binding to 10-80 microM palmitic acid occurred slowly (approximately 3-7 min halftime) and was accompanied by an increase in turbidity (measured by difference spectrophotometry) and a decrease in exchangeable calcium. The stoichiometry of calcium binding to palmitic acid under this condition was 0.15 mol of calcium/mol of palmitate, but increased to approximately 0.4 mol of calcium/mol of palmitate in the presence of 0.03-3 mM calcium. The rate of calcium binding to palmitic acid also increased with calcium concentration such that in the presence of 1-3 mM calcium, palmitic acid bound approximately 0.4 mol of calcium/mol of palmitate within 15 s. Thus, palmitic acid may represent a calcium-precipitating anion in regions of the cell, such as the lumen of the sarcoplasmic reticulum, where calcium concentration is high. Analyses of sarcoplasmic reticulum calcium sequestration in the presence of various palmitic acid concentrations support this hypothesis and suggest that 10% of the palmitic acid is accessible to luminal calcium.

Effects of palmitic acid and palmityl carnitine on calcium sequestration by rabbit skeletal sarcoplasmic reticulum vesicles.

A number of long-chain fatty acids and fatty acid derivatives accumulate in the cytosol of ischemic myocardium. Although the functional significance of this accumulation in vivo remains unclear, these amphiphilic compounds may alter the functional properties of a variety of biological membranes in vitro. In this study, we investigated the effect of palmitic acid and palmityl carnitine on calcium sequestration by sarcoplasmic reticulum vesicles in the absence of calcium-precipitating anions. Palmitic acid, at micromolar concentrations, enhanced calcium sequestration in a concentration-dependent manner when present from the onset of the reaction or when added to calcium-filled vesicles. Under identical conditions, similar concentrations of palmityl carnitine inhibited calcium sequestration when present at the onset of the reaction and caused a rapid release of accumulated calcium when added to calcium-filled vesicles. Low concentrations of palmitic acid decreased the sensitivity of the sarcoplasmic reticulum to the inhibitory effects of palmityl carnitine. These results suggest that calcium pump function in the sarcoplasmic reticulum can be altered by the presence of amphiphilic compounds and that this alteration is dependent on both the structure and number of amphiphiles present.