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 stimula ted or inhibited by norepinephrine according to the presence or absence of a brain soluble fraction. Gel filtration of such soluble fraction has allow ed the separation of two fractions, peaks I and II, able to stimulate and i nhibit Na+, K+-ATPase activity, respectively. Peak II behaves much like oua bain, which has suggested the term endobain. From peak II, a subfraction te rmed II-E (endobain E), which highly inhibits Na+, K+-ATPase, has been sepa rated by anionic exchange chromatography in a Synchropack AX-300 column. We determined the in vitro effect of endobain E obtained from rat cerebral co rtex on neuronal norepinephrine release by incubating rat hypothalamic tiss ue in the presence of [H-3]norepinephrine. Neuronal norepinephrine release was quantified as the factor above basal [H-3]norepinephrine released to th e medium at experimental and three post-experimental periods. Endobain E wa s found to increase norepinephrine release in a concentration-dependent fas hion, reaching 200%, equivalent to the effect achieved with 400 mu M ouabai n. Ouabain effect persisted along three post-experimental periods whereas t hat of endobain E remained only during the first post-experimental period. These results led us to conclude that endobain increases norepinephrine rel ease in hypothalamic neurons at the presynaptic nerve ending level, an effe ct 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. (C) 1999 IBRO. Published by Elsevier Science Ltd.