Biomedical subjects

N S Dhalla

Publications and source records attributed to N S Dhalla.

At least 55 records · Page 3Linked to original sources

PubMed1999

Changes in cardiac protein kinase C activities and isozymes in streptozotocin-induced diabetes.

To understand cardiac dysfunction in diabetes, the activity of protein kinase C (PKC) and protein contents of its isozymes (PKC-alpha, -beta, -epsilon, and -zeta) were examined in diabetic rats upon injection of streptozotocin (65 mg/kg iv). The hearts were removed at 1, 2, 4, and 8 wk, and some of the 6-wk diabetic animals had been injected with insulin (3 U/day) for 2 wk. The Ca(2+)-dependent PKC activity was increased by 43 and 51% in the homogenate fraction and 31 and 70% in the cytosolic fraction from the 4- and 8-wk diabetic hearts, respectively, in comparison with control values. The Ca(2+)-independent PKC activity was increased by 24 and 32% in the homogenate fraction and 52 and 89% in the cytosolic fraction from the 4- and 8-wk diabetic hearts, respectively, in comparison with control values. The relative protein contents of PKC-alpha, -beta, -epsilon, and -zeta isozymes were increased by 43, 31, 48, and 38%, respectively, in the homogenate fraction and by 126, 119, 148, and 129%, respectively, in the cytosolic fraction of the 8-wk diabetic heart. The observed changes in heart homogenate and cytosolic fractions were partially reversible upon treatment of the diabetic rats with insulin. The results suggest that the increased myocardial PKC activity and increased protein contents of the cytosolic PKC isozymes are associated with subcellular alterations and cardiac dysfunction in the diabetic heart.

Animals↗

PubMed1999

Changes in sarcolemmal PLC isoenzymes in postinfarct congestive heart failure: partial correction by imidapril.

We have examined the changes in quantity and activity of cardiac sarcolemmal (SL) phosphoinositide-phospholipase C (PLC)-beta(1), -gamma(1), and -delta(1) in a model of congestive heart failure (CHF) secondary to large transmural myocardial infarction (MI). We also instituted a late in vivo monotherapy with imidapril, an ANG-converting enzyme (ACE) inhibitor, to test the hypothesis that its therapeutic action is associated with the functional correction of PLC isoenzymes. SL membranes were purified from the surviving left ventricle of rats in a moderate stage of CHF at 8 wk after occlusion of the left anterior descending coronary artery. SL PLC isoenzymes were examined in terms of protein mass and hydrolytic activity. CHF resulted in a striking reduction (to 6-17% of controls) of the mass and activity of gamma(1)- and delta(1)-isoforms in combination with a significant increase of both PLC beta(1) parameters. In vivo treatment with imidapril (1 mg/kg body wt, daily, initiated 4 wk after coronary occlusion) improved the contractile function and induced a partial correction of PLCs. The mass of SL phosphatidylinositol 4,5-bisphosphate and the activities of the enzymes responsible for its synthesis were significantly reduced in post-MI CHF and partially corrected by imidapril. The results indicate that profound changes in the profile of heart SL PLC-beta(1), -gamma(1), and -delta(1) occur in CHF, which could alter the complex second messenger responses of these isoforms, whereas their partial correction by imidapril may be related to the mechanism of action of this ACE inhibitor.

Angiotensin-Converting Enzyme Inhibitors↗

PubMed1999

Alterations in sarcoplasmic reticulum function and gene expression in ischemic-reperfused rat heart.

In view of the critical role of sarcoplasmic reticular (SR) Ca(2+) release and the Ca(2+) pump in cardiac contraction-relaxation, this study was undertaken to assess the status of SR function, protein content, and gene expression in isolated rat hearts subjected to global ischemia for 30 min followed by 60 min of reperfusion (I/R). Attenuated recovery of contractile function in the I/R hearts was associated with reduced SR Ca(2+) uptake, Ca(2+) release, and ryanodine-binding activities. mRNA levels and protein contents for SR Ca(2+) pump ATPase and Ca(2+) release channels were markedly depressed in the I/R hearts. Perfusion of hearts with superoxide dismutase plus catalase, well-known scavengers of oxyradicals, prevented the I/R-induced alterations in cardiac function and partially prevented SR Ca(2+) transport activities and mRNA abundance. In hearts perfused with xanthine plus xanthine oxidase or H(2)O(2), changes similar to those in the I/R hearts were observed. These results indicate that oxyradicals may participate in depressing the SR Ca(2+) handling and gene expression in the I/R heart. It is suggested that treatment of hearts with antioxidants may improve the recovery of cardiac function by preserving the SR function and partially protecting the SR gene expression.

Animals↗

PubMed1999

Alterations of heart function and Na+-K+-ATPase activity by etomoxir in diabetic rats.

To examine the role of changes in myocardial metabolism in cardiac dysfunction in diabetes mellitus, rats were injected with streptozotocin (65 mg/kg body wt) to induce diabetes and were treated 2 wk later with the carnitine palmitoyltransferase inhibitor (carnitine palmitoyltransferase I) etomoxir (8 mg/kg body wt) for 4 wk. Untreated diabetic rats exhibited a reduction in heart rate, left ventricular systolic pressure, and positive and negative rate of pressure development and an increase in end-diastolic pressure. The sarcolemmal Na+-K+-ATPase activity was depressed and was associated with a decrease in maximal density of binding sites (Bmax) value for high-affinity sites for [3H]ouabain, whereas Bmax for low-affinity sites was unaffected. Treatment of diabetic animals with etomoxir partially reversed the depressed cardiac function with the exception of heart rate. The high serum triglyceride and free fatty acid levels were reduced, whereas the levels of glucose, insulin, and 3,3',-5-triiodo-L-thyronine were not affected by etomoxir in diabetic animals. The activity of Na+-K+-ATPase expressed per gram heart weight, but not per milligram sarcolemmal protein, was increased by etomoxir in diabetic animals. Furthermore, Bmax (per g heart wt) for both low-affinity and high-affinity binding sites in control and diabetic animals was increased by etomoxir treatment. Etomoxir treatment also increased the depressed left ventricular weight of diabetic rats and appeared to increase the density of the sarcolemma and transverse tubular system to normalize Na+-K+-ATPase activity. Therefore, a shift in myocardial substrate utilization may represent an important signal for improving the depressed cardiac function and Na+-K+-ATPase activity in diabetic rat hearts with impaired glucose utilization.

Animals↗

PubMed1999

Biphasic changes in heart performance with food restriction in rats.

To examine effects of food restriction resembling very-low-calorie dieting on heart performance, normal rats were fed 25% of ad libitum food intake for 14 days. Although heart weight decreased (P < 0.05) after 5 days, left ventricular systolic pressure as well as rates of pressure development and fall were increased (P < 0.05) at 7 days and decreased (P < 0.05) after 14 days. Systolic and diastolic blood pressures were also increased from 5 to 7 days and decreased after 14 days. The increased hemodynamic performance of heart was associated with a raised plasma norepinephrine concentration, which peaked at day 7 of food restriction; epinephrine concentration was increased (P < 0.05) also at day 7. An increased catecholamine synthesis was indicated by the raised (P < 0.05) plasma dopamine beta-hydroxylase activity at 3 days, but this was decreased (P < 0. 05) at 14 days. The concentration of dopamine in the heart was increased (P < 0.05) at 2-14 days, of norepinephrine at 7-14 days, and of epinephrine at 10 and 14 days. Food restriction thus appears initially to be associated with an enhanced catecholamine influence on the heart and is followed by a depressed cardiac performance.

Animals↗

PubMed1999

Development of cardiac sensitivity to oxygen deficiency: comparative and ontogenetic aspects.

Hypoxic states of the cardiovascular system are undoubtedly associated with the most frequent diseases of modern times. They originate as a result of disproportion between the amount of oxygen supplied to the cardiac cell and the amount actually required by the cell. The degree of hypoxic injury depends not only on the intensity and duration of the hypoxic stimulus, but also on the level of cardiac tolerance to oxygen deprivation. This variable changes significantly during phylogenetic and ontogenetic development. The heart of an adult poikilotherm is significantly more resistant as compared with that of the homeotherms. Similarly, the immature homeothermic heart is more resistant than the adult, possibly as a consequence of its greater capability for anaerobic glycolysis. Tolerance of the adult myocardium to oxygen deprivation may be increased by pharmacological intervention, adaptation to chronic hypoxia, or preconditioning. Because the immature heart is significantly more dependent on transsarcolemmal calcium entry to support contraction, the pharmacological protection achieved with drugs that interfere with calcium handling is markedly altered. Developing hearts demonstrated a greater sensitivity to calcium channel antagonists; a dose that induces only a small negative inotropic effect in adult rats stops the neonatal heart completely. Adaptation to chronic hypoxia results in similarly enhanced cardiac resistance in animals exposed to hypoxia either immediately after birth or in adulthood. Moreover, decreasing tolerance to ischemia during early postnatal life is counteracted by the development of endogenous protection; preconditioning failed to improve ischemic tolerance just after birth, but it developed during the early postnatal period. Basic knowledge of the possible improvements of immature heart tolerance to oxygen deprivation may contribute to the design of therapeutic strategies for both pediatric cardiology and cardiac surgery.

Aging↗

PubMed1999

Evidence for the role of oxidative stress in acute ischemic heart disease: a brief review.

BACKGROUND: Although contractile performance of the acutely damaged ischemic heart is invariably depressed on reperfusion (myocardial stunning), the mechanisms of cardiac dysfunction in stunned myocardium are poorly understood. OBJECTIVES: To review briefly the current state of knowledge and to provide further experimental evidence of whether oxidative stress plays a crucial role in cardiac dysfunction and membrane abnormalities due to ischemia-reperfusion. MATERIALS AND METHODS: Isolated rat hearts perfused in the absence or presence of superoxide dismutase (SOD) plus catalase, a well known oxyradical scavenger system, were subjected to 30 mins of global ischemia and 60 mins of reperfusion. Cardiac performance, hydrogen peroxide and calcium contents, lipid peroxidation and membrane activities associated with calcium transport were measured in the control, ischemic and ischemic- reperfused hearts. RESULTS: Cardiac dysfunction, as reflected by depressed left ventricular developed pressure, +dP/dt and -dP/dt as well as elevated left ventricular end-diastolic pressure, in the ischemic-reperfused heart was associated with increased hydrogen peroxide, calcium and malondialdehyde contents as well as increased formation of conjugated dienes. These changes due to ischemia-reperfusion were attenuated in hearts treated with SOD plus catalase. Both ischemia and ischemic-reperfused hearts showed depressions in sarcolemmal Na+/K+-ATPase and sodium-calcium exchange, as well as sarcoplasmic reticular calcium uptake and calcium release activities; these membrane abnormalities were also partially prevented by the presence of SOD plus catalase. CONCLUSIONS: Oxidative stress due to the formation of hydrogen peroxide leading to lipid peroxidation and sulfhydryl group oxidation during ischemia-reperfusion seems to be one of the mechanisms that may produce membrane defects and result in intracellular calcium overload and cardiac contractile dysfunction in the stunned myocardium.

Animals↗

PubMed1999

Alterations in protein kinase A and protein kinase C levels in heart failure due to genetic cardiomyopathy.

BACKGROUND: It is becoming evident that both cardiac and skeletal muscles are affected in congestive heart failure. Although protein kinases are known to regulate cardiac function, very little is known about their status in cardiac and skeletal muscles during the development of congestive heart failure. OBJECTIVE: To determine changes in the activities and protein levels of protein kinase A (PKA) and protein kinase C (PKC) in cardiac and skeletal muscles in congestive heart failure due to genetic cardiomyopathy on the basis that PKA and PKC are crucial for protein phosphorylation. ANIMALS AND METHODS: Genetically cardiomyopathic UM-X7.1 hamsters (250 to 300 days old) and age-matched Syrian hamsters were used in this study. PKA and PKC activities were assayed by measuring 32P from [gamma-32P]ATP incorporated into synthetic substrates. Relative protein contents of these protein kinases were obtained by using immunoblot analysis in control and failing hamster hearts and skeletal muscles. RESULTS: PKC activity was significantly increased in the failing hearts compared with control preparations. The relative protein contents of cytosolic PKC-alpha and -epsilon , and of particulate PKC-epsilon isozymes were significantly increased in failing hearts. PKC activity was also markedly increased in cardiomyopathic skeletal muscle. Furthermore, PKA activity and protein level in both cardiac and skeletal muscles were significantly increased in the failing heart group compared with control values. CONCLUSIONS: Increased PKC activity in heart failure may be due to changes in PKC-alpha and -epsilon isozymes in cardiomyopathic hearts. Alterations of PKA and PKC in congestive heart failure were not limited to the heart because similar changes in enzyme activities were evident in skeletal muscle.

Animals↗

PubMed1999

Inhibition of serotonin-induced vascular smooth muscle cell proliferation by sarpogrelate.

Antiproliferative behavior of sarpogrelate (Anplag, MCI-9042, (+/-)-1-[2-[2-(3-methoxyphenyl)ethyl]phenoxy]-3-(dimethylamino)-2-pro pyl hydrogen succinate hydrochloride), a serotonin 2A (5-HT2A) receptor antagonist, was established using radioactive incorporation of [(3)H]thymidine, [(3)H]uridine, and [(3)H]phenylalanine in cultured rat aortic smooth muscle cells in response to a 5-HT-induced cytokine trigger. Fluorescence-activated cell sorting was used to confirm these observations. 5-HT-induced DNA, RNA, and protein synthesis were inhibited maximally at a concentration of 1 microM sarpogrelate. Although other cytokines such as platelet-derived growth factor and endothelin also induced DNA, RNA, and protein synthesis in rat aortic smooth muscle cells, cell proliferation was not influenced by sarpogrelate, even at large pharmacological concentrations (10 microM). Sarpogrelate's antiproliferative actions were found to be more potent than ketanserin. These data indicate that sarpogrelate operates as a specific inhibitor of 5-HT-mediated cell proliferation and is a good candidate for preventing serotonin-induced neointimal hyperplasia.

Animals↗

PubMed1998

Ca2+ mobilization in adult rat cardiomyocytes by angiotensin type 1 and 2 receptors.

The role of angiotensin II (AngII) in the regulation of heart function under normal and pathological conditions has been well documented. Although two types of AngII receptors (AT1 and AT2 receptors) are found in equal proportions in the rat heart, most studies have focused primarily on AT1 receptor-coupled events. In this study, the contribution of both types of AngII receptors to cardiac function was evaluated by measuring intracellular calcium ([Ca2+]i) levels at ambient temperature in freshly isolated adult rat ventricular cardiomyocytes. Exposure of cardiomyocytes to AngII (0.01 to 10 microM) resulted in an immediate and sustained increase in [Ca2+]i in a concentration-dependent manner. The increase in [Ca2+]i in cardiomyocytes by AngII was blocked by either losartan or compound PD123319 (1-[[4-(dimethylamino)-3-methylphenyl]methyl]-5-(diphenylacetyl)- 4,5,6,7-tetrahydro-1H-imidazo[4,5-c]pyridine-6-carboxylic acid), non-peptide antagonists of the AT1 and AT2 receptors, respectively. The specificity of the action of these antagonists was verified by their inability to alter the basal levels of [Ca2+]i as well as KCl- or ATP-induced increases in [Ca2+]i. AngII was also observed to initiate spontaneous beating activity in cardiomyocytes, which was prevented by both losartan and compound PD123319 in a concentration-dependent manner (0.01 to 10 microM). These data indicate that the activation of both AT1 and AT2 receptors may stimulate a signalling pathway that influences [Ca2+]i and spontaneous beating activity in cardiomyocytes.

Adenosine Triphosphate↗

PubMed1998

Differential alterations in left and right ventricular G-proteins in congestive heart failure due to myocardial infarction.

In order to examine the status of G-proteins in congestive heart failure due to myocardial infarction, the left coronary artery in rats was ligated and animals assessed after 4, 8 and 16 weeks. Sham-operated control and experimental animals were used for the preparation of membranes from the viable (uninfarcted) left and right ventricles. Adenylyl cyclase activities in the presence of pertussis toxin and cholera toxin were increased and decreased in left ventricles from all groups, respectively. On the other hand, adenylyl cyclase activities in 8 and 16-week experimental right ventricles were unaltered in the presence of pertussis toxin and increased in the presence of cholera toxin. Depression of adenylyl cyclase activities in left ventricles from all groups as well as in the right ventricle at 4 weeks were not evident when enzyme activity was determined in the pertussis toxin-treated membranes in the absence or presence of Gpp(NH)p. Cholera toxin-catalyzed ADP ribosylation was decreased in left ventricles from all infarcted groups and increased in the right ventricles at 8 and 16 weeks whereas the pertussis toxin-catalyzed ADP ribosylation was increased in all experimental tissues except in the right ventricles at 8 and 16 weeks. G(s alpha)-protein content was decreased in the left ventricle at 16 weeks and increased in the right ventricles at 8 and 16 weeks of myocardial infarction. On the other hand, G(i alpha)-protein content was increased in left ventricles from all infarcted groups and the 4-week right ventricle but was unaltered in 8 and 16-week right ventricles. An increase in mRNA abundance for G(i alpha)-protein was seen in both left and right ventricles following myocardial infarction. A significant increase in mRNA level for G(s alpha)-protein was observed in all left ventricles and 8-week right ventricle following the coronary occlusion. These results suggest that changes in Gs- and Gi-proteins in the failing heart due to myocardial infarction are chamber-specific and are dependent upon the stage of congestive heart failure.

Adenylyl Cyclases↗

PubMed1998

Molecular cloning of rat cardiac sarcolemmal Ca2+/Mg2+ ectoATPase (Myoglein).

Rat cardiac sarcolemmal Ca2+/Mg2+ ectoATPase (Myoglein), a membrane-bound enzyme requiring millimolar concentrations of Ca2+ or Mg2+ for maximal hydrolysis of ATP, has been purified to apparent homogeneity. Tryptic digestion and amino acid sequencing was used to design an oligonucleotide probe for screening a rat heart cDNA library; this produced a partial cDNA clone (pND2.1), and sequencing of a 400 base pair portion revealed a 100% homology to human platelet CD36. Northern blotting with pND2.1 detected a 3.1 kb transcript in rat heart but not in other tissues. Interspecies expression analysis (cardiac tissue total RNA blot probed with pND2.1) detected a approximately 2.0 kb transcript in canine, rabbit and porcine heart, whereas transcripts of a 4.1 kb, approximately 3.0 kb and 2.1 kb were observed in human cardiac tissue. A rat genomic DNA Southern blot, probed with pND2.1, indicated that there was a single copy of the gene in the rat genome. Expression of the pND2.1 cDNA in E. coli produced an 89 kDa polypeptide recognized by anti-human CD36 antibody but not by anti-rat Ca2+/Mg2+ ectoATPase antibody. It is concluded that rat cardiac Ca2+/Mg2+ ectoATPase is tightly associated with a protein highly homologous to the adhesion molecule CD36.

Adenosine Triphosphatases↗

PubMed1998

Subcellular remodeling and heart dysfunction in chronic diabetes.

Heart dysfunction in chronic diabetes has been observed to be associated with depressed myofibrillar adenosine triphosphatase activities as well as abnormalities in the sarcoplasmic reticular and sarcolemmal calcium transport processes. The evidence has been presented to show that alterations in the expression of myosin isozymes and regulatory proteins as well as myosin phosphorylation contribute to the development of myofibrillar remodeling in the diabetic heart. Defects in sarcoplasmic reticular and sarcolemmal calcium transport appear to be due to the accumulation of lipid metabolites in the membrane. Different agents, such as calcium-antagonists, beta-adrenoceptor blockers, angiotensin converting enzyme inhibitors, metabolic interventions and antioxidants, have been reported to exert beneficial effects in preventing subcellular remodeling and cardiac dysfunction in chronic diabetes. Clinical and experimental investigations have suggested that increased sympathetic activity, activated cardiac renin-angiotensin system, myocardial ischemia/functional hypoxia and elevated levels of glucose for a prolonged period, due to insulin deficiency, result in oxidative stress. It is proposed that oxidative stress associated with a deficit in the status of the antioxidant defense system may play a critical role in subcellular remodeling, calcium-handling abnormalities and subsequent diabetic cardiomyopathy.

Calcium↗

PubMed1998

Beneficial effect of ischemic preconditioning on Ca2+ paradox in the rat heart.

The effect of ischemic preconditioning (IP; 3 min ischemia plus 3 min reperfusion) on the recovery of cardiac function after Ca2+ depletion was investigated. Isolated rat hearts were subjected to different cycles of IP episodes followed by Ca2+ free perfusion and repletion. Perfusion of control hearts with Ca2+ free medium for 5 min followed by repletion of Ca2+ for 30 min resulted in a marked decrease in the left ventricular (LV) developed pressure and an increase in LV end-diastolic pressure (Ca2+ paradox). The depressed function due to Ca2+ paradox recovered with three cycles of IP. Myoglobin release during Ca2+ repletion also decreased significantly by three cycles of IP. The beneficial effects of IP were also evident when the hearts were subjected to a mild form of Ca2+ paradox involving 3 min Ca2+ depletion. The protective effect rendered by IP disappeared when 10 microM of 8-(p-sulfophenyl)-theophylline, adenosine antagonist was perfused for 10 min before IP. These results suggest that IP exerts beneficial effects on Ca2+ paradox which may be mediated by adenosine.

Animals↗

PubMed1998

Biophysical characterization of rat cardiac Ca2+/Mg2+ ecto-ATPase (myoglein).

Sarcolemmal Ca2+/Mg2+ ecto-ATPase (Myoglein; MW 180 kD) is a membrane bound enzyme which requires a millimolar concentration of either Ca2+ or Mg2+ for maximal hydrolysis of ATP. The isoelectric point (pI) of the cardiac ecto-ATPase was 5.7. The purified Ca2+/Mg2+ ecto-ATPase from the rat heart sarcolemmal appeared as a single band with MW approximately 90 kD in the SDS-PAGE. In order to understand the nature of this enzyme, the 90 kD band in the SDS-PAGE was electroeluted; the analysis of the eluate showed 2 prominent bands with MW approximately 90 and 85 kD. The presence of 2 bands was further confirmed by gradient gel (10-20%) electrophoresis in 0.375 M Tris-HCl buffer, pH 8.8. Analysis of the purified Ca2+/Mg2+ ecto-ATPase as well as the electroeluted protein in a non-equilibrium linear two dimensional electrophoresis (Ampholyte pI 3.0-10.0) also showed two distinct bands. Mass spectroscopic analysis of the enzyme using different matrix combinations revealed the presence of multi-components indicating microheterogeneity in the protein structure. Treatment of the ecto-ATPase with DL-dithiothreitol did not alter the pattern of mass spectroscopic analysis and this indicated that the microheterogeneity may be due to some posttranslational modifications. It is concluded that rat cardiac Ca2+/Mg2+ ecto-ATPase is an acidic protein having two subunits. Furthermore, the enzyme shows microheterogeneity in its molecular structure.

Adenosine Triphosphatases↗

PubMed1998

Alterations of KCl- and ATP-induced increase in [Ca2+]i in cardiomyocytes from vitamin B6 deficient rats.

Although vitamin B6 deficiency is related to coronary heart disease, no information regarding changes in myocardium due to vitamin B6 deficiency is available in the literature. In view of the critical role played by Ca2+ in cellular function, we investigated alterations in [Ca2+]i induced by KCI or ATP in vitamin B6 deficient and age-matched control rats. [Ca2+]i was measured in isolated cardiomyocytes by using the Fura-2 fluorescence technique. The KC1-induced increase in [Ca2+]i was augmented in vitamin B6 deficient cardiomyocytes, whereas the ATP-induced increase in [Ca2+]i was attenuated. The specific ATP binding to sarcolemma from hearts of vitamin B6 deficient rats was decreased. A single injection of vitamin B6 (10 mg/kg) to vitamin B6 deficient animals completely reversed the KC1- or ATP-induced changes in [Ca2+]i in cardiomyocytes as well as ATP binding with sarcolemma. These results regarding altered regulation of [Ca2+]i in cardiomyocytes and sarcolemmal ATP receptors indicate myocardial abnormalities due to vitamin B6 deficiency.

Adenosine Triphosphate↗

PubMed1998

Mechanism of depression in cardiac sarcolemmal Na+-K+-ATPase by hypochlorous acid.

Oxidative stress during pathological conditions such as ischemia-reperfusion is known to promote the formation of hypochlorous acid (HOCl) in the heart and to result in depression of cardiac sarcolemmal (SL) Na+-K+-ATPase activity. In this study, we examined the direct effects of HOCl on SL Na+-K+-ATPase from porcine heart. HOCl decreased SL Na+-K+-ATPase activity in a concentration- and time-dependent manner. Characterization of Na+-K+-ATPase activity in the presence of different concentrations of MgATP revealed a decrease in the maximal velocity (Vmax) value, without a change in affinity for MgATP on treatment of SL membranes with 0.1 mM HOCl. The Vmax value of Na+-K+-ATPase, when determined in the presence of different concentrations of Na+, was also decreased, but affinity for Na+ was increased when treated with HOCl. Formation of acylphosphate by SL Na+-K+-ATPase was not affected by HOCl. Scatchard plot analysis of [3H]ouabain binding data indicated no significant change in the affinity or maximum binding capacity value for ouabain binding following treatment of SL membranes with HOCl. Western blot analysis of Na+-K+-ATPase subunits in HOCl-treated SL membranes showed a decrease (34 +/- 9% of control) in the beta1-subunit without any change in the alpha1- or alpha2-subunits. These data suggest that the HOCl-induced decrease in SL Na+-K+-ATPase activity may be due to a depression in the beta1-subunit of the enzyme.

Adenosine Triphosphate↗

PubMed1998

Modification of cardiac beta-adrenoceptor mechanisms by H2O2.

From the role of oxidative stress in cardiac dysfunction, we investigated the effect of H2O2, an activated species of oxygen, on beta-adrenoceptors, G proteins, and adenylyl cyclase activities. Rat heart membranes were incubated with different concentrations of H2O2 before the biochemical parameters were measured. Both the affinity and density of beta 1-adrenoceptors were decreased, whereas the density of the beta 2-adrenoceptors was decreased and the affinity was increased by 1 mM H2O2. Time- and concentration-dependent biphasic changes in adenylyl cyclase activities in the absence or presence of isoproterenol were observed when membranes were incubated with H2O2; however, activation of the enzyme by isoproterenol was increased or unaltered. The adenylyl cyclase activities in the absence or presence of forskolin, NaF, and Gpp(NH)p were depressed by H2O2. Catalase alone or in combination with mannitol was able to significantly decrease the magnitude of alterations due to H2O2. The cholera toxin-stimulated adenylyl cyclase activity and ADP ribose labeling of Gs proteins were decreased by treatment with 1 mM H2O2, whereas Gi protein activities, as reflected by pertussis toxin-stimulation of adenylyl cyclase and ADP ribosylation, were unaltered. The Gs and Gi protein immunoreactivities, estimated by labeling with respective antibodies, indicate a decrease in binding to the 45-kDa band of Gs protein, whereas no change in the binding of antibodies to the 52-kDa band of Gs protein or the 40-kDa subunit of Gi protein was evident when the membranes were treated with 1 mM H2O2. These results suggest that H2O2 in high concentrations may attenuate the beta-adrenoceptor-linked signal transduction in the heart by changing the functions of Gs proteins and the catalytic subunit of the adenylyl cyclase enzyme.

Adenosine Diphosphate Ribose↗