Biomedical subjects

N S Dhalla

Publications and source records attributed to N S Dhalla.

At least 217 records · Page 12Linked to original sources

PubMed1985

Evidence for three catalytic sites in heart sarcolemmal phospholipid N-methylation.

Conversion of phosphatidylethanolamine to phosphatidylcholine through sequential transmethylation reactions was studied in rat cardiac sarcolemmal membrane prepared by the hypotonic shock-LiBr treatment method. Three catalytic sites were identified on the basis of their requirement for divalent cations and kinetic parameters. Site I (Km = 0.1 +/- 0.01 microM) required Mg2+ as a cofactor and had a very high affinity for [3H]-S-adenosyl-L-methionine (AdoMet). Sites II (Km = 2.9 +/- 0.3 microM) and III (Km = 112 +/- 6 microM) did not require any divalent cation and showed comparatively lower affinities for AdoMet. Developmental-related differences were found to exist among these three catalytic sites when phospholipid methylation was carried out in 1- to 12-week-old rats. The results suggest that three distinct methyltransferase sites are involved in cardiac sarcolemmal phospholipid methylation.

Animals↗

PubMed1984

Alterations in adenylate cyclase activity due to streptozotocin-induced diabetic cardiomyopathy.

The effect of chronic experimental diabetes on the adenylate cyclase system (AC) in the rat heart was investigated. Rats were made diabetic by an intravenous injection of streptozotocin (65 mg/kg), hearts were removed 8 weeks later and washed cell particles were isolated. AC activity was measured in the absence and presence of different concentrations of forskolin, NaF, GTP analogue [Gpp(NH)p] or epinephrine. A significant depression in the epinephrine stimulated AC activity was observed in diabetic hearts. Basal AC activity and stimulation of AC with forskolin, NaF and Gpp(NH)p were not significantly different between control and diabetic preparations. These results indicate no apparent alterations in the regulatory or catalytic properties of AC in hearts from chronic diabetic rats. The observed depression in epinephrine stimulated AC activity may account for the depressed inotropic action of catecholamines in the diabetic cardiomyopathy.

Adenylyl Cyclases↗

PubMed1984

Characterization of heart sarcolemmal phospholipid methylation.

The transmethylation of phosphatidylethanolamine (PE) to phosphatidylcholine (PC) was studied in rat heart sarcolemmal membrane. Kinetically, three apparent Km values for S-adenosyl-L-methionine (AdoMet) were obtained when the total [3H]methyl groups incorporation into the phospholipids was examined in the presence of 0.01-250 microM AdoMet. A first methyltransferase active site having a very low Km (0.1 microM) for AdoMet showed a partial requirement for Mg2+ and an optimum pH of 8.0 with a major formation of phosphatidyl-N-monomethylethanolamine (PMME). Both Ca2+ and K+ were inhibitory to this site. A second active site with a Km of 3.6 microM showed an optimum pH of 7.0 with predominant formation of phosphatidyl-N,N-dimethylethanolamine (PDME) and no Mg2+ requirement; in addition, transmethylation activity was also observed over a broad alkaline pH range (9-11) with an optimum at pH 10.5. This site was insensitive to Ca2+ but was stimulated by Na+, while K+ had an inhibitory effect. A third active site with a Km of 119 microM showed an optimum pH of 10.5 with major formation of PC and no Mg2+ requirement. This site was also insensitive to Ca2+ but markedly inhibited by both K+ and Na+. Under optimal conditions, the activities of all three methyltransferase sites were linear for at least 30 min of incubation and the sensitivity to the inhibitory effect of S-adenosyl-L-homocysteine was different for each site. Addition of exogenous PMME and PDME as substrates enhanced the synthesis of the corresponding methylated products by 3-5-fold and 3-8-fold, respectively. In contrast, exogenous PE failed to increase methyltransferase activity. These results provide evidence for the existence of three distinct methyltransferase active sites in rat heart sarcolemma.

Animals↗

PubMed1984

Defect in the adrenergic receptor-adenylate cyclase system during development of catecholamine-induced cardiomyopathy.

We have investigated alterations in adrenergic receptors and adenylate cyclase activity in cardiac membranes from rats injected with 40 mg/kg intraperitoneal isoproterenol. Reduction in the number of beta-adrenergic and alpha-adrenergic receptors, as assessed by changes in specific binding of 3H-dihydroalprenolol (DHA) and 3H-dihydroergocryptine (DHE), was observed only at 9 and 24 hours after isoproterenol injection, respectively. On the other hand, epinephrine-stimulated, NaF-stimulated, and Gpp (NH)p-stimulated adenylate cyclase activity was decreased as early as 3 hours after isoproterenol treatment without changes in the basal adenylate cyclase activity. These results demonstrate a defect in the adrenergic receptor-adenylate cyclase system during the development of catecholamine-induced cardiomyopathy and may partly explain the attenuated inotropic adrenergic response of the heart under stressful situations.

Adenylyl Cyclases↗

PubMed1984

Involvement of calcium in coronary vasoconstriction due to prolonged hypoxia.

In this study we employed a hypoxic rat heart model for investigating the mechanisms of coronary spasm. Perfusion of the isolated heart with hypoxic medium resulted in initial coronary dilatation followed by sustained constriction. Pretreatment of rats with 5 mg/kg reserpine did not alter the rate of the magnitude of constriction during 60 minutes of hypoxic perfusion. The hypoxia-induced vasoconstriction was not affected by the inclusion of either 1 or 10 micrograms/ml phentolamine in the perfusion buffer, but phenoxybenzamine (0.1 and 1 microgram/ml) significantly attenuated the degree of constriction. Since phenoxybenzamine and phentolamine were equally effective in preventing the phenylephrine-induced coronary constriction, it is unlikely that the effect of phenoxybenzamine was due to an alpha-receptor blocking property but instead may be accounted for by its calcium channel blocking action. Two calcium channel blockers, verapamil and D-600 (0.1 and 1 microgram/ml), were also effective. The rate of rise in coronary pressure was substantially reduced by decreasing the concentration of calcium and was increased by elevating the concentration of calcium in the perfusion medium, but the magnitude of hypoxia-induced constriction was not affected. These results are consistent with the suggestion that the coronary constriction seen during hypoxia does not involve adrenergic mechanisms but is dependent upon the availability of calcium.

Adrenergic alpha-Antagonists↗

PubMed1984

Sarcolemmal alterations during the development of genetically determined cardiomyopathy.

The purpose of this study was to identify alterations in specific enzyme and Ca2+ binding activities in cardiac sarcolemmal fractions from UM-X7.1 myopathic Syrian hamsters during the development of cardiomyopathy. Experimental and healthy control animals were examined from 25 to 200 days of age. Sarcolemmal Na+, K+-ATPase activity was depressed in the myopathic hamsters throughout the time course of this study. Sarcolemmal ATP-independent Ca2+ binding was found to be depressed in experimental animals as early as 55 days of age. Ca2+ -stimulated, Mg2+ -dependent ATPase activity was depressed in the experimental animals by 90 days of age and this decrease in enzyme activity was accompanied by a decrease in ATP-dependent Ca2+ binding capacity of the sarcolemmal membranes. Mg2+ -ATPase and Ca2+ -ATPase activities were only affected in the latter stages of the disease (155 to 200 days old). NaF, epinephrine and Gpp(NH)p stimulation of the sarcolemmal adenylate cyclase activity was also observed to be attenuated during the latter stages of the disease. These defects in adenylate cyclase system of the sarcolemmal fraction appeared specific since basal adenylate cyclase activity was not altered at any age studied. The results demonstrate that the earliest lesions in sarcolemmal activity in myopathic hamster heart occur in Na+, K+-ATPase and ATP-independent Ca2+ binding capacity. These defects correspond temporally to the initial stages of cardiac necrotic development in this strain of myopathic hamster.

Adenosine Triphosphatases↗

PubMed1984

Sarcoplasmic reticular Ca2+-pump adaptation in cardiac hypertrophy due to pressure overload in pigs.

Ca2+-transport activities of the sarcoplasmic reticulum isolated from the non-failing hypertrophied left ventricle due to supravalvular banding of aorta for 4 and 8 weeks in pigs were compared with those from the sham operated controls. The rate of Ca2+-uptake, but not the capacity, in 4 weeks hypertrophied heart preparations was increased. On the other hand, both the rate and capacity of sarcoplasmic reticulum to accumulate Ca2+ were decreased at 8 weeks of heart hypertrophy. The Ca2+-binding and Ca2+-stimulated Mg2+ dependent ATPase activities of the reticular fractions from 4 and 8 weeks hypertrophied hearts were not significantly different from the control values. No changes in the sarcoplasmic reticular phospholipid contents were evident except that sphingomyelin was significantly decreased in 4 weeks and diphosphatidylglycerol was increased in both 4 and 8 weeks hypertrophied preparations. These results suggest that adaptation of the sarcoplasmic reticulum function in physiologic and pathologic hypertrophied hearts may occur by changing the efficiency of Ca2+-pump system during the development of cardiac hypertrophy.

Animals↗

PubMed1984

Progress and problems in understanding the involvement of calcium in heart function.

In this article we have briefly reviewed the role of Ca2+ in the excitation contraction coupling in the myocardium and have indicated that cardiac contraction and relaxation are initiated upon raising and lowering the intracellular concentration of free Ca2+, respectively. Different mechanisms for the entry of Ca2+ through sarcolemma as well as release of Ca2+ from sarcoplasmic reticulum and possibly mitochondria have been outlined for initiating cardiac contraction. Relaxation of the cardiac muscle appears to be intimately dependent upon efflux of Ca2+ through sarcolemma as well as sequestration of Ca2+ by the intracellular storage sites, particularly sarcoplasmic reticulum and possibly mitochondria. The actions of some pharmacological and pathophysiological interventions have been explained on the basis of changes in subcellular Ca2+ movements in myocardium. Quinidine, which produced an initial positive inotropic action on rat heart was also found to increase sarcolemmal Ca2+-ATPase activity without any changes in the Na+-K+ ATPase. Other antiarrhythmic agents, procainamide and lidocaine, also increased sarcolemmal Ca2+-ATPase activity without affecting the Na+-K+ ATPase. On the other hand, both Ca2+-ATPase and Na+-K+ ATPase activities were increased in heart sarcolemma obtained from cardiomyopathic hamsters. In this model the increased Ca2+-ATPase activity may promote the occurrence of intracellular Ca2+ overload in the cardiac cell whereas the increased Na+-K+ ATPase activity may increase Ca2+ efflux through Na+-Ca2+ exchange systems as an adaptive mechanism. It has been suggested that some caution should be exercised while interpreting the data from in vitro experiments in terms of functional changes in the myocardium. Furthermore, it has been proposed that the pathophysiology and pharmacology of Ca2+ movements at different membrane sites be understood fully in normal and diseased myocardium in order to improve the therapy of heart disease.

Adenosine Triphosphate↗

PubMed1984

Biochemical alterations in heart after exhaustive swimming in rats.

This study investigated alterations in glycogen, catecholamines, and the function of various subcellular membranes of the heart after exhaustive swimming in rats. The rats were exhausted by daily exercise over 1, 3, or 7 consecutive days. Glycogen content of the heart and three selected skeletal muscles was depleted after a single bout of exhaustive exercise. Repeated bouts of exhaustive swimming elicited a depletion of glycogen in only the plantaris and gastrocnemius skeletal muscles. Plasma norepinephrine and epinephrine levels were highly elevated, and cardiac concentrations of these hormones were significantly depleted immediately after all exercise sessions. Cardiac sarcoplasmic reticulum (SR) Ca2+ transport was depressed after a single exhaustive exercise period. After three exercise bouts SR Ca2+ accumulation remained depressed; however, mitochondrial Ca2+ transport was found to be augmented. If the exhaustive exercise protocol was continued up to seven days, only mitochondrial Ca2+ accumulation was depressed. Various parameters of sarcolemmal membrane function were observed to be unaltered after exhaustive exercise. These findings demonstrate that exhaustive swimming exercise in rats is capable of producing significant alterations in the Ca2+ transport capacity of the SR and mitochondrial membrane systems of the heart but is without apparent effect on the sarcolemmal membrane.

Animals↗

PubMed1984

Sarcolemmal phosphatidylethanolamine N-methylation in diabetic cardiomyopathy.

Phosphatidylethanolamine N-methylation was studied in cardiac sarcolemma 8 weeks after the induction of chronic experimental diabetes in rats by a streptozotocin injection (65 mg/kg, iv). Incorporation of radiolabeled methyl groups from S-adenosyl-L-methionine into intramembranal phosphatidylethanolamine, assayed under optimal conditions, confirmed the existence of three catalytic sites involved in the sequential methyl transfer reactions. Total methyl group incorporation at all three sites was significantly depressed in diabetic myocardium, but this change was reversible by a 14-day insulin therapy to the diabetic animals. Measurements of phospholipid N-methylation activity at different times indicated that the depression was evident at 6 weeks after the induction of diabetes. This defect was also seen for the individual methylated lipid products (monomethyl-, dimethylphosphatidylethanolamine, and phosphatidylcholine) specifically formed at each catalytic site. Experiments with different concentrations of S-adenosyl-L-methionine revealed that, for all three sites, the apparent affinity for the methyl donor did not change, whereas the apparent Vmax values were significantly lowered in diabetes. The results of this study identify a defect in the sarcolemmal phosphatidylethanolamine N-methylation in diabetic cardiomyopathy.

Animals↗

PubMed1984

Influence of myosin antiserum on heart sarcolemmal Ca2+ or Mg2+ ATPase activity.

Sarcolemmal Ca2+/Mg2+ ATPase was inhibited only about 30% by myosin antiserum that decreased myofibrillar ATPase activity by about 80%. There was a remarkable difference in the effect of myosin antiserum on sarcolemmal Ca2+/Mg2+ ATPase and myofibrillar ATPase with regard to its pre-incubation time with these organelles. Tryptic digestion of the sarcolemmal membrane did not show any change in its inhibitory effect of the myosin antiserum on Ca2+ ATPase. The data distinguish the Ca2+/Mg2+ ATPase from myosin ATPase and suggest that it is an enzyme of the sarcolemmal origin.

Adenosine Triphosphatases↗

PubMed1984

Cardiac cell damage: a primary myocardial disease in streptozotocin-induced chronic diabetes.

Ultrastructural changes in heart muscle due to chronic diabetes subsequent to a single injection of streptozotocin (65 mg/kg body wt, i.v.) were studied in rats. Presence of diabetes was indicated by hyperglycaemia (plasma glucose, control, 120 +/- 7; diabetic, 448 +/- 21 mg/dl) as well as hypo-insulinaemia (plasma insulin, control, 25.6 +/- 5.2; diabetic, 11.2 +/- 0.5 microU/ml). After 8 weeks of diabetes, the hearts were processed for electron microscopic examination. Cardiac muscle cells in diabetic hearts showed condensation of nuclear chromatin and folding of nuclear membranes. Swelling of mitochondria, clearing of mitochondrial matrix and incorporation of lysosomal membranes into mitochondrial matrix was also noted. A marked increase in both lysosomes and lipid droplets was apparent. Focal areas in diabetic hearts showed contracted sarcomeres, myofibrillar degeneration and separation of the intercalated disc. Atherosclerotic plaque formation as well as structural changes in the smooth muscle or endothelial cells in the small arteries, arterioles or capillaries were not seen to accompany the structural changes in the cardiac muscle cells of the diabetic hearts. This study provides strong evidence for the occurrence of primary myocardial disease in streptozotocin-induced chronic diabetes.

Animals↗

PubMed1983

Association of adenylate cyclase inhibition by NaF with loss of a factor in rat heart sarcolemma.

Heart sarcolemma from different animals was isolated for studying the effect of NaF on membrane-bound adenylate cyclase activity. Unlike dog and rabbit heart, a depression of adenylate cyclase by NaF was observed in sarcolemma from rat heart. There was a progressive attenuation of the NaF ability to stimulate the enzyme at different steps of the sarcolemmal isolation procedure. The activation by epinephrine in the presence of Gpp(NH)p also decreased progressively but unlike NaF, this agent did not show an inhibition of the enzyme. The inhibitory action of NaF was not reversed upon the treatment of heart membranes with deoxycholate or by Ca2+. Lubrol extract (supernatant) of a particulate fraction from rat heart, which showed NaF activation, returned the stimulatory response of the sarcolemmal adenylate cyclase to NaF. These results suggest that some regulatory factor is required for the stimulation of adenylate cyclase by NaF in myocardium and rat heart is susceptible for the loss of such a factor during the sarcolemmal isolation by the hypotonic shock-LiBr treatment method.

Adenylyl Cyclase Inhibitors↗

PubMed1983

Properties of 5'-nucleotidase in rat heart sarcolemma.

The activity of 5'-nucleotidase (5'-ribonucleotide phosphohydrolase, EC 3.1.3.5) was examined in membrane fractions isolated by hypotonic shock-LiBr treatment (fraction HL) and sucrose gradient separation (fraction S) of rat ventricle homogenate. The enzyme activity in these two fractions differed significantly in several respects. In fraction HL, 5'-nucleotidase had a high affinity for AMP (Km 35 microM), and ATP was a potent competitive inhibitor. In contrast, the 5'-nucleotidase displayed by fraction S showed a low substrate affinity (Km 130 microM) and less sensitivity to ATP. Treatment of membranes with trypsin and neuraminidase markedly stimulated 5'-nucleotidase in fraction HL, whereas only a modest effect was observed in fraction S. Exposure of the membranes to Triton X-100 resulted in a 60% and 10% increase in the enzyme activity in fractions HL and S, respectively. The characteristic activity ratios of 5'-nucleotidase at 200 microM relative to 50 microM AMP in fractions HL and S were modified by alamethicin in an opposite way and became identical. Although concanavalin A almost completely inhibited the 5'-nucleotidase activity in both membrane preparations at a concentration of 2 microM, Hill plots of the data on concanavalin A inhibition revealed a coefficient of 2.2 for fraction S and 1.1 for fraction HL. The differences in 5'-nucleotidase activity of the two membrane fractions are considered to be due to differences in the orientation of the vesicles of the sarcolemmal preparations. These results suggest that two distinct catalytic sites for 5'-nucleotidase are present at the intra- and extracellular surface of the rat heart sarcolemma.

Alamethicin↗

PubMed1983

Energy production and utilization in contractile failure due to intracellular calcium overload.

Intracellular calcium overload was produced by perfusing the Ca2+- deprived rat hearts for 5 or 10 min with normal medium containing 1.25 mM Ca2+ for 10 min and changes in the myofibrillar and membrane ATPase, sarcolemmal adenylate cyclase, mitochondrial oxidative phosphorylation and high energy phosphate stores in failing hearts were examined. Myocardial creatine phosphate and ATP were decreased by the intracellular calcium overload whereas the myofibrillar, mitochondrial and microsomal ATPase activities were not altered. The intracellular calcium overload markedly depressed the mitochondrial oxidative phosphorylation as well as sarcolemmal Ca2+ ATPase, Mg2+ ATPase, Na+ - K+ ATPase and adenylate cyclase. These results suggest that abnormalities in the process of energy production rather than energy utilization may primarily account for the depressed energy state of hearts failing due to an intracellular calcium overload.

Adenosine Diphosphate↗

PubMed1983

Potential oxidative pathways of catecholamines in the formation of lipid peroxides and genesis of heart disease.

Effects of vitamin E, a fat soluble antioxidant, on the isoproterenol-induced changes in the lipid peroxide activity as determined by a quantitation of malondialdehyde (MDA) content in the myocardium were examined. Isoproterenol treatment (80 mg/kg given over two days in two equal doses) caused more than 100 percent increase in the MDA content which was prevented by pretreatment of the animals with vitamin E (alpha-tocopherol acetate, 10 mg/kg) for two weeks. Animals maintained on vitamin E deficient diet for 8 weeks were found to be more sensitive to isoproterenol-induced increase in the MDA content. A small increase in MDA content was also seen due to vitamin E deficiency alone. These changes were found to be reversible upon a 2 week feeding of the animals on the normal diet coupled with vitamin E treatment. Based on these data it is proposed that free radical mediated increase in lipid peroxide activity may have a role in catecholamine-induced heart disease.

Adenosine Triphosphate↗

PubMed1983

Alterations in Ca2+ binding by and composition of the cardiac sarcolemmal membrane in chronic diabetes.

Chronic streptozotocin-induced diabetes in rats was associated with a significant loss in the ability of isolated cardiac sarcolemmal membranes to bind Ca2+. Administration of insulin to the diabetic rats normalized the sarcolemmal Ca2+ binding capacity. The content of sialic acid residues, which are considered to represent a superficial Ca2+ pool in sarcolemma, was decreased in preparations from diabetic rats, and this change also was reversible upon insulin treatment of the diabetic rats. Treatment of sarcolemma with neuraminidase decreased Ca2+ binding by 37% in control preparations but had no effect on diabetic preparations. Diphosphatidylglycerol content was decreased but other acidic phospholipids such as phosphatidylinositol and phosphatidylserine, which also bind Ca2+, were not altered during diabetes. An increase in lysophosphatidylcholine and a decrease in phosphatidylethanolamine contents were observed in membranes isolated from diabetic rats. These results suggest that some alterations occur in Ca2+ binding and composition of heart sarcolemma in chronically diabetic rats and may provide further insight into the pathogenesis of diabetic cardiomyopathy.

Animals↗

PubMed1983

Role of Na+-Ca2+ exchange in the development of cardiac abnormalities due to calcium paradox.

Reperfusion of rat heart with Ca2+-containing medium for 1 to 10 min after a 5 min perfusion with Ca2+-free medium resulted in a generalized disruption of myocardial ultrastructure including swelling of sarcoplasmic reticulum and mitochondria, depletion of creatine phosphate and adenosine triphosphate stores, reduction of the microsomal but augmentation of the mitochondrial Ca2+ uptake activities and elevation of the cardiac Na+ as well as Ca2+ contents. These hearts developed contracture and were unable to generate contractile force. Lowering the concentration of Na+ from 145 to 35 mmol l-1 in the medium during Ca2+-free perfusion was observed to markedly reduce or prevent the reperfusion induced changes in myocardium. On the other hand, lowering the concentration of Na+ in the medium during the reperfusion phase following a 5 min perfusion with Ca2+-free medium enhanced the rate of depletion of the high energy phosphate stores and the rate of elevation of myocardial Ca2+ contents but markedly depressed the rise in Na+ contents. These results suggest that Ca2+-paradoxical changes in myocardium occur as a consequence of the intracellular Ca2+ overload in which Ca2+ entry through Na+-Ca2+ exchange mechanism at the cell membrane may be an important contributory factor.

Animals↗