Loss of
amino acids into the coronary artery perfusate, which is exacerbated during anoxic stress, may have important implications for the ability of hearts subjected to
ischemia or
anoxia to recover function during reoxygenation. This work investigates the mechanisms underlying the
amino acid efflux. Rat Langendorff heart preparations were used to study
amino acid loss into coronary artery perfusates during
anoxia or
anoxia/reoxygenation sequences. Coronary flow rates, heart rates and intra-aortic pressures were recorded. Changes in myocardial
amino acid concentrations were equated with
amino acid levels in collected anoxic perfusate. With the exception of
taurine, the differences in
amino acid levels between normoxic and anoxic hearts were smaller than the amounts lost into the coronary perfusates, indicating ongoing replenishment of most
amino acids during the anoxic episode. Fifteen-minute periods of exposure to low
oxygen levels (P02 18-20 mmHg) resulted in large percentage increases in perfusate
amino acid levels which returned slowly towards control levels upon reoxygenation.
Anion channel blockers,
anthracene-9-carboxylic acid,
furosemide, and 4-acetamido-4-isothiocyanostilbene-2,2'-disulfonic
acid (
SITS), depressed
anoxia-elicited increases in
amino acid release.
Phospholipase inhibition with
quinacrine,
4-bromophenacyl bromide and 7,7-dimethyl-eicosadenoic
acid (DEDA) depressed the
anoxia-evoked release of
amino acids. Combined applications of
SITS and DEDA exhibited additive effects, virtually abolishing
anoxia-evoked release of all the
amino acids. The
protein kinase C inhibitor,
chelerythrine chloride, and the
protein tyrosine kinase inhibitors,
genistein and
lavendustin A, inhibited
anoxia-evoked
amino acid release.
Polyunsaturated fatty acids, arachidonic and linoleic, reduced
anoxia-evoked
amino acid release whereas monosaturated (oleic) and saturated (
stearic) acids were ineffective. The
glutamate transport inhibitor,
dihydrokainate, depressed
anoxia-evoked
glutamate and
aspartate release. These results suggest that at least three possible mechanisms for the
anoxia-evoked
amino acid efflux including (a) diffusional release through volume activated
anion channels, (b) leakage across myocyte plasma membranes as a consequence of
phospholipase activation and (c) reversal of Na+ dependent high-affinity transporters.