Changes in [Na⁺]_i, compartmental [Ca²⁺], and NADH with dysfunction after global ischemia in intact hearts

Abstract

We measured the effects of global ischemia and reperfusion on intracellular Na(+), NADH, cytosolic and mitochondrial (subscript mito) Ca(2+), relaxation, metabolism, contractility, and Ca(2+) sensitivity in the intact heart. Langendorff-prepared guinea pig hearts were crystalloid perfused, and the left ventricular (LV) pressure (LVP), first derivative of LVP (LV dP/dt), coronary flow, and O(2) extraction and consumption were measured before, during, and after 30-min global ischemia and 60-min reperfusion. Ca(2+), Na(+), and NADH were measured by luminescence spectrophotometry at the LV free wall using indo 1 and sodium benzofuran isophthalate, respectively, after subtracting changes in tissue autofluorescence (NADH). Mitochondrial Ca(2+) was assessed by quenching cytosolic indo 1 with MnCl(2). Mechanical responses to changes in cytosolic-systolic (subscript sys), diastolic (subscript dia), and mitochondrial Ca(2+) were tested over a range of extracellular [Ca(2+)] before and after ischemia-reperfusion. Both [Ca(2+)](sys) and [Ca(2+)](dia) doubled at 1-min reperfusion but returned to preischemia values within 10 min, whereas [Ca(2+)](mito) was elevated over 60-min reperfusion. Reperfusion dissociated [Ca(2+)](dia) and [Ca(2+)](sys) from contractile function as LVP(sys-dia) and the rise in LV dP/dt (LV dP/dt(max)) were depressed by one-third and the fall in LV dP/dt (LV dP/dt(min)) was depressed by one-half at 30-min reperfusion, whereas LVP(dia) remained markedly elevated. [Ca(2+)](sys-dia) sensitivity at 100% LV dP/dt(max) was not altered after reperfusion, but [Ca(2+)](dia) at 100% LV dP/dt(min) and [Ca(2+)](mito) at 100% LV dP/dt(max) were markedly shifted right on reperfusion (ED(50) +36 and +125 nM [Ca(2+)], respectively) with no change in slope. NADH doubled during ischemia but returned to normal on initial reperfusion. The intracellular [Na(+)] ([Na(+)](i)) increased minimally during ischemia but doubled on reperfusion and remained elevated at 60-min reperfusion. Thus Na(+) and Ca(2+) temporally accumulate during initial reperfusion, and cytosolic Ca(2+) returns toward normal, whereas [Na(+)](i) and [Ca(2+)](mito) remain elevated on later reperfusion. Na(+) loading likely contributes to Ca(2+) overload and contractile dysfunction during reperfusion.

GSK-3.BETA., a Therapeutic Target for Cardiomyocyte Protection

“…7,8 Second, intracellular Na + accumulates due to Na + influx via Na + -H + exchangers and Na + channels and due to reduced Na + efflux via the Na + -K + pump. [9][10][11] This Na + overload predisposes ischemic cardiomyocytes to Ca 2+ influx by the Na + -Ca 2+ exchanger. Third, Ca 2+ influx via the Na + -Ca 2+ exchanger, reduced Ca 2+ uptake into the sarcoplasmic reticulum and reduced Ca 2+ efflux via the sarcolemmal Ca 2+ pump induce Ca 2+ overload in ischemic cardiomyocytes.…”

“…9-12 However, elevation of intracellular Ca 2+ is modest during ischemia because acidosis inhibits the Na + -Ca 2+ exchanger and cytosolic Ca 2+ is taken up by the mitochondria as long as its membrane potential is maintained by use of ATP. [9][10][11]13 Severely ischemic cardiomyocytes ultimately 'starve to death', although sarcolemmal damage by detergent actions of accumulated long-chain Acyl-CoA and by Ca 2+ -activated proteases and phospholipases might also be involved in ischemia-induced necrosis. 7 Nevertheless, reperfusion is necessary to salvage ischemic cardiomyocytes from necrosis.…”

“…[9][10][11] The abrupt increase in intracellular Ca 2+ level induces activation of calpain, a Ca 2+ -activated protease, which compromises Na + -K + pump function 10 and Ca 2+ overload of the mitochondria. 6,7,9,11 At the same time as Ca 2+ overload, cardiomyocytes abruptly develop cell edema after reperfusion as a result of osmotic loading during ischemia. ATP production restituted by re-supply of oxygen in Ca 2+ -overloaded and swelling myocytes with fragile sarcolemma induces hypercontraction and sarcolemmal rupture.…”

Altered NADH and improved function by anesthetic and ischemic preconditioning in guinea pig intact hearts

“…Depending on the duration and magnitude of the ischemia, the injury can be reversible (stunning) or irreversible (infarction) (20). Several interrelated mechanisms are responsible: 1) cytosolic Na ϩ and Ca 2ϩ overload (39) with myofibrillar hypercontracture, cytoskeletal damage, and cell disruption; 2) mitochondrial Ca 2ϩ overload (27,39), inefficient ATP synthesis or utilization, and NADH accumulation (39); 3) reduced maximal Ca 2ϩ -activated force and/or Ca 2ϩ sensitivity (39); 4) impaired myocardial vascular perfusion (32); and 5) reactive oxygen species-induced damage to Na ϩ and Ca 2ϩ pump proteins and membranes by lipid peroxidation (4).…”

“…Hypoxia and ischemia lead to excess accumulation of NADH (19,39) and suggest uncoupled oxidative phosphorylation. Because APC and IPC may trigger mechanisms that induce metabolic protection during ischemia, we hypothesized that APC and IPC cause similar changes in mitochondrial function before, during, and after ischemia as evidenced by online recordings of NADH fluorescence.…”