Investigations into mechanisms of magnesium homeostasis in the isolated rat heart

Abstract

Magnesium (Mg24) is essential for normal functioning of the heart. It is the second most abundant divalent cation in living cells and plays vital roles in cellular regulation. Preservation of intracellular Mg 24 concentration ([Mg24]) is essential in the heart. The concentration of ionized Mg 24 is similar in both the extracellular and intracellular milieu of cardiac myocytes although a large electrochemical gradient exists which will tend to promote Mg24 influx across the sarcolemmal membrane. Therefore homeostatic mechanism(s) must exist which will extrude Mg 24 against this gradient. The existence of a Na/Mg2 counterport mechanism or an ATP driven pump for the extrusion of Mg 2 from the cell has been suggested.
This work examines various possible mechanisms for Mg 24 extrusion. In a series of experiments, the effects on Mg 24 efflux and contractile force, of increased and decreased extracellular sodium [Na *], lithium chloride, specific ion channel inhibitors, osmotic shock, metabolic inhibition and global ischaemia, in the isolated, perfused (5 ml mm", 37°C) and paced (20V, I msec, 4-6 Hz) rat-heart were investigated in an attempt to characterize further, the mechanism of Mg 24 transport. Following a recovery period of 20 mm, the heart was perfused with a nominally Mg2tfree Krebs-Henseleit (1(H) solution, which facilitates
the measurement of Mg24 in effluent samples using a spectroscopic method.
Perfusion of the isolated heart with Mg 2 -free KH containing increased [Na], in chloride, nitrate, isethionate or sulphate forms, all promoted a large and significant (P C 0.05) Mg2t efflux. This efflux was, in general, accompanied by an increase in contractile force.
Typically, net total Mg 24 efflux in elevated [Na']. (as NaCl), measured over a period of 10 min was 590.1 ± 26.7 ng (100 mg tissue)" (mean and SEM, n = 6) compared to 376.1 ± 17.9 ng (100 mg tissue)" (n = 7) in controls. However, lithium chloride also promoted significant (P C 0.05) increases in Mg 2 efflux and contractile force. Mg 2 efflux in response to elevated (Na*], was significantly inhibited by 10" M and 10 M amiloride, but not by 10 M ouabain. Propranolol (10" M) and 10 7 M verapamil inhibited Mg2t efflux under control conditions. Niflumic acid (10' M) produced a small but delayed response in Mg2t efflux, under both control conditions and in response to increased [Na']... whereas 103 M Ibrosemide elicited a large and significant increase under these conditions. Both niflumic acid and ifirosemide inhibited contractile force. Osmotic shock induced by either sucrose, glucose or fructose resulted in a significant (P < 0.01) increase in Mg2t efilux and a concomitant increase in contractile force, both of which were found to be dose related.
Metabolic inhibition with either rotenone, potassium cyanide (KCN) or oligomycin B promoted small but not significant increases in Mg 2 efflux and these were accompanied by significant decreases in force of contraction of the heart. Uncoupling phosphorylation and oxidation using dinitrophenol (DNP) elicited a highly significant increase in Mg 2t efflux and
a concomitant decrease in force. Lactate dehydrogenase activity did not significantly increase during perfusion of the heart with either increased [Na t] 0 or DMP, suggesting that cell damage was not responsible for the Mg2t effiux. When electrically paced or spontaneously beating hearts were subjected to 10 min of global ischaemia, a significant increase in Mg2 efilux occurred together with a concomitant decrease in force of contraction. The responses in electrically paced hearts were more pronounced than in spontaneously beating hearts. These results suggest the possibility of a small amiloride and
propranolol sensitive component to Mg 2 efflux, which could be a NaVMg2t counterport mechanism. However, since lithium chloride, sucrose, glucose and fructose also elicit Mg2t efflux, it is concluded that the response to these and to elevated [Na - ],, may be osmotic in origin. A further mechanism involving chloride transport and/or intracellular Ca2t may also exist and it appears that Mg 2t efflux is linked either to the proton gradient in the mitochondria, or to intracellular pH or to ATP production.
Dietary studies showed that the heart is able to conserve Mg 2 at the expense of other organs, but that young rats are more adversely affected by low dietary Mg2t intake than older rats.