Since 1983, when it was discovered that inositol 1,4,5-trisphosphate can act as second messenger to release Ca2+ from the endoplasmic reticulum, widespread research has focused on the phosphatidylinositol signalling transduction pathway and the host of inositolphosphates formed intracellularly after stimulation therof. Although the polyphosphates, inositoltetrakisphosphate (InsP4) and inositolhexakisphosphate (InsP6), have received their share of attention, a definite physiological role has not been ascribed to them as yet. Different binding proteins for these two polyphosphates have been demonstrated, especially in brain tissue, indicating their possible importance in the cell. InsP6 is known as one of nature's most powerful antioxidants and has already been demonstrated to possess the abilities to be of use in the industry as well as in the medical profession. As its natural actions are poorly understood and its possible side-effects have not been widely investigated, basic research regarding its cellular and subcellular activities is urgently called for.

Animals↗

PubMedDate not supplied

Signal transduction in myocardial ischaemia and reperfusion.

Recent studies in the non-ischaemic myocardium indicated that drugs stimulating cAMP formation inhibit alpha 1-mediated inositol phosphate generation, while alpha 1-adrenergic stimulation lowered tissue cAMP levels, implicating cross-talk between alpha 1- and beta-adrenergic signalling pathways in normal physiological conditions. Massive amounts of endogenous catecholamines, predominantly noradrenaline, are released during myocardial ischaemia and reperfusion, causing stimulation of both alpha 1- and beta-adrenergic receptors which, in turn, may contribute to intracellular Ca2+ overload and subsequent cell damage. Since no information is available regarding cross-talk in pathophysiological conditions, the aim of this study was to evaluate the interactions between alpha 1- and beta-adrenergic signalling pathways during different periods of ischaemia and reperfusion. Isolated rat hearts were perfused retrogradely for 30 min before being subjected to (i) 5-25 min global ischaemia and (ii) 1-5 min of reperfusion after 20 min global ischaemia. Drugs (prazosin, 10(-7) M; propranolol, 10(-6) M; phenylephrine 3 x 10(-5) M; isoproterenol 10(-9) M) were added 10 min before the onset of ischaemia and were present during reperfusion. Increasing periods of ischaemia caused an immediate rise and progressive lowering in tissue cAMP and Ins(1,4,5)P3 levels respectively. In contrast, reperfusion caused an elevation in Ins(1,4,5)P3 levels and reduced cAMP. Prazosin elevated cAMP levels during both ischaemia and reperfusion, while propranolol had no effects on tissue Ins(1,4,5)P3. The activity of the alpha 1-adrenergic signal transduction pathway appears to have an inhibitory effect on the activity of the beta-adrenergic system during ischaemia and reperfusion.

Adrenergic alpha-Antagonists↗

PubMedDate not supplied

Substrate effects on sarcolemmal permeability in the normoxic and hypoxic perfused rat heart.

OBJECTIVES: Based on the hypothesis that provision of glucose is good and fatty acids are bad for the ischaemic myocardium, the aims of this study were to determine i) the effects of different substrates on sarcolemmal permeability during normoxia, low-flow hypoxia (HLF) and reperfusion, ii) whether increased membrane permeability is associated with ultrastructural damage and increased influx of Ca2+ into cells and iii) whether changes in membrane permeability correlate with myocardial function and high energy phosphate metabolism. METHODS: The isolated rat heart subjected to HLF was used as model of global ischaemia, and sarcolemmal permeability assessed by release of LDH from and influx of lanthanum and Ca2+ into myocardial tissue. Myocyte structural injury was also evaluated quantitatively, and mechanical activity was monitored throughout the experimental protocol. RESULTS: Regardless of the substrate used, HLF caused a 80-90% and 20-40% reduction in myocardial oxygen uptake and coronary flow rate, respectively. Palmitate (0.5 mM conjugated to 0.1 mM albumin) or substrate-free perfusion caused ultrastructural damage and loss of normal sarcolemmal integrity during both normoxia and HLF. Although reperfusion reversed injury in some cells, in general, myocytes exhibited myofibrillar contracture, while membrane integrity recovered to some extent, as indicated by reduced lanthanum influx. Intracellular Ca2+ increased significantly upon reperfusion. Mechanical function as well as tissue high energy phosphates were significantly depressed during both HLF and reperfusion. Glucose, on the other hand, protected against ischaemia-induced structural damage and loss of sarcolemmal integrity. Reperfusion in these experiments resulted in almost complete recovery of normal morphology, ultrastructure and sarcolemmal integrity, while intracellular Ca2+ remained unchanged. Mechanical function and tissue high energy phosphates were significantly higher in glucose-perfused hearts than in palmitate-perfused or substrate-free hearts. Glucose was also able to attenuate the harmful effects of palmitate on myocardial ultrastructure, membrane integrity, mechanical function, energy metabolism and prevented Ca2+ overloading during reperfusion. CONCLUSION: The results provide new evidence for the protective role of glucose during myocardial ischaemia and reperfusion. Although the exact mechanism of the beneficial actions of glucose remains to be established, the results suggest that glycolytic flux and thus glycolytically derived ATP protect against ischaemic damage via preservation of membrane integrity.

Adenine Nucleotides↗

PubMedDate not supplied

A comparison between ischemic preconditioning and anti-adrenergic interventions: cAMP, energy metabolism and functional recovery.

OBJECTIVES: The postulate that ischemic preconditioning caused an attenuation in ischemia induced increases in tissue cAMP, and that this may pertain to the mechanism of ischemic preconditioning, was investigated in the isolated rat heart. A significant reduction in tissue cAMP in preconditioned hearts was observed for all time periods of global ischemia studied. The significance of this observation was evaluated by comparing the effect of anti-adrenergic interventions on energy metabolism and post-ischemic functional recovery of both non-preconditioned and preconditioned hearts. METHODS: The isolated perfused rat heart was used as experimental model. Six groups were studied: Non-preconditioned rat hearts: i) untreated controls (Non-PC), ii) reserpinised (Non-PC Res), iii) propranolol treated (10(-7) M) (Non-PC Prop); Preconditioned rat hearts: iv) preconditioned controls (PC), v) reserpinised (PC Res) and vi) propranolol (10(-7) M) treated (PC Prop). RESULTS: After 25 min global ischemia the concentration of cAMP was increased by 79.6% in the Non-PC group. This increase was attenuated in all of the treated groups, although in varying degrees. Energy utilization in these hearts also differed markedly between the groups. Functional recovery was however similar in all Non-PC and PC treated groups and significantly superior to that of Non-PC control hearts. Prior reserpinisation mimicked the protective effect of preconditioning on energy metabolism and functional recovery. To determine the significance of attenuation of the increase in cAMP in the protection conferred by preconditioning, hearts were pretreated with forskolin (10(-6) M). This caused an accumulation of tissue cAMP in preconditioned hearts to similar absolute values as seen in untreated non-preconditioned hearts during 25 min global ischemia. However, the percentage increase in forskolin-pretreated preconditioned hearts during sustained ischemia was only 50% vs. 71% in non-preconditioned hearts treated with forskolin, confirming an attenuated beta-response induced by preconditioning. Forskolin treatment of preconditioned hearts did not abolish the protective effect. CONCLUSIONS: The findings suggest that the protection against ischemic damage conferred by preconditioning is associated with an attenuated beta-adrenergic response. However, whether the changes in cAMP occurring during sustained global ischemia is the cause of consequence of the elicited protection, remains to be established.

Adenine Nucleotides↗

PubMedDate not supplied

The hypoxic, low-flow perfused rat heart: characterization as a model of global ischaemia.

In the study of the mechanism of mitochondrial damage in myocardial ischaemia, production of a small animal model of global ischaemia proved to be necessary. Thus, the effects of coronary flow rate, oxygen availability and substrate availability were studied on the functional capacity of mitochondria isolated from perfused rat hearts. The hypoxic, low-flow perfused heart eventually chosen was evaluated as a model of myocardial ischaemia by measuring tissue contents of high energy phosphate compounds and glycolytic intermediates and by assessing the effects of exogenous substrates on the above parameters.

Adenosine Diphosphate↗

PubMedDate not supplied

Inhibition of contractility during the early phase of total ischaemia in the working heart. Recovery during reperfusion.

Total global ischaemia of the normothermic working rat heart caused an initial positive inotropic response characterized by vigorous contractions. After +/- 15 s this response reached a peak whereafter the isotonic contraction amplitude started to decline. After +/- 3.5 min the heart ceased to beat. The low level of high energy phosphates (HEP), determined 3 min after the onset of ischaemia, indicated that these phases of contractility during ischaemia might play a significant role in depleting HEP. This was substantiated by the observation that inhibition of the contractions during ischaemia by low calcium or high potassium solutions resulted in conservation of myocardial adenosine triphosphate (ATP) and creatine phosphate (CP) stores. It also resulted in the prevention of contracture development during ischaemia and improved mechanical recovery during reperfusion. It was therefore concluded that inhibition of contractility immediately after the onset of total global ischaemia of the normothermic working rat heart is of prime importance in mechanical recovery during reperfusion.

Adenosine Triphosphate↗

PubMedDate not supplied

Substrate effects on mitochondrial function and tissue lipids in low-flow hypoxia of isolated perfused rat hearts.

A possible causal relationship between tissue FFA contents and the depression in mitochondrial oxidative phosphorylation in myocardial ischaemia has been suggested. To test this hypothesis, the effects of different substrates added to the perfusates of hypoxic, low-flow perfused hearts were examined on oxidative phosphorylation catalysed by mitochondria isolated from such tissue. In an additional series of experiments tissue neutral glyceride and FFA levels were analysed and correlated with changes in mitochondrial function. Mitochondria isolated from hearts with a high tissue FFA content exhibited the lowest ADP/O ratios, RCI and QO2 values. On the other hand, mitochondria isolated from hearts with reduced FFA contents, performed significantly better with respect to these parameters of mitochondrial function studied.

Animals↗

PubMedDate not supplied

Sarcolemmal integrity during ischaemia and reperfusion of the isolated rat heart.

The effects of ischaemia and reperfusion on sarcolemmal permeability were studied in the isolated rat heart using an ionic lanthanum probe technique. Detection of increased permeability after exposure to short periods of ischaemia without accompanying reperfusion effects was achieved by employing a substrate-free, low calcium saline solution as vehicle for ionic lanthanum. Quantifiable results showed a rapid increase in sarcolemmal permeability in the subendocardium after 15 min of ischaemia, with the subepicardium showing similar alterations after longer periods. Reperfusion after 15 min of ischaemia led to recovery of normal permeability characteristics, correlating well with ultrastructural and functional recovery. Reperfusion after 25 min of ischaemia did not allow recovery of sarcolemmal permeability, ultrastructure, or function. These results show a clear correlation between the development of irreversible myocardial damage and increased sarcolemmal membrane permeability.

Animals↗

PubMedDate not supplied

Normothermic ischaemic cardiac arrest of the isolated perfused rat heart: effects of trifluoperazine and lysolecithin on mechanical and metabolic recovery.

To evaluate the hypothesis that maintenance of the integrity of myocardial membrane systems and prevention of Ca2+ influx into the cell are significant in the survival of ischaemic tissue, the effect of trifluoperazine and lysolecithin, were tested on the recovery of globally ischaemic rat hearts. Trifluoperazine increases membrane stabilization, inhibits calmodulin and binds to other Ca2+-dependent proteins. Lysolecithin, on the other hand, has a detergent action on myocardial cell membranes and facilitates Ca2+ ingress in ischaemic tissue. With trifluoperazine (2.45 microM), added before induction of ischaemia or during reperfusion only, hearts subjected to 40 min normothermic ischaemic cardiac arrest recovered mechanically. Untreated hearts failed after 20 min of ischaemia. The drug had no effect on tissue high energy phosphate levels or mitochondrial oxidative phosphorylation. Conversely, lysolecithin (2.5-10 microM) caused all hearts to fail after being subjected to 15 min ischaemia. Mechanical failure during reperfusion of such hearts was associated with a significant reduction in tissue ATP and CrP levels. Trifluoperazine counteracted the harmful effects of lysolecithin to a limited extent.

Adenosine Diphosphate↗

PubMedDate not supplied

Oxidative phosphorylation function of two mitochondrial preparations from heart: effects of ischaemia and cytochrome C.

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Animals↗