A brain Na+, K+-ATPase inhibitor (endobain E) enhances norepinephrine release in rat hypothalamus

We have shown that synaptosomal membrane Na+, K+-ATPase activity is stimulated or inhibited by norepinephrine according to the presence or absence of a brain soluble fraction. Gel filtration of such soluble fraction has allowed the separation of two fractions, peaks I and II, able to stimulate and inhibit Na+, K+-ATPase activity, respectively. Peak II behaves much like ouabain, which has suggested the term endobain. From peak II, a subfraction termed II-E (endobain E), which highly inhibits Na+, K+-ATPase, has been separated by anionic exchange chromatography in a Synchropack AX-300 column. We determined the in vitro effect of endobain E obtained from rat cerebral cortex on neuronal norepinephrine release by incubating rat hypothalamic tissue in the presence of [3H]norepinephrine. Neuronal norepinephrine release was quantified as the factor above basal [3H]norepinephrine released to the medium at experimental and three post-experimental periods. Endobain E was found to increase norepinephrine release in a concentration-dependent fashion, reaching 200%, equivalent to the effect achieved with 400 microM ouabain. Ouabain effect persisted along three post-experimental periods whereas that of endobain E remained only during the first post-experimental period. These results led us to conclude that endobain increases norepinephrine release in hypothalamic neurons at the presynaptic nerve ending level, an effect resembling that of ouabain. It is postulated that endobain E may enhance catecholamine availability in the synaptic gap, leading to an increase in noradrenergic activity.