Characterization and partial purification of the sodium-potassium-ATPase inhibitor released from cultured rat hypothalamic cells.
An inhibitor of sodium-potassium-ATPase has been partially purified from the culture medium obtained from hypothalamic cells maintained in a capillary membrane perfusion system, and some of the properties of this inhibitory factor have been investigated. Gel filtration (Sephadex G-25 Superfine) of heat-treated medium (80 degrees C for 10 min) resulted in elution of inhibitory activity in the post-salt fraction. These fractions inhibited active (i.e. sodium-potassium-ATPase-mediated) sodium transport in intact human erythrocytes, displaced [3H]ouabain from its binding site, and directly inhibited canine kidney sodium-potassium-ATPase as measured by NADH oxidation. High-performance liquid chromatography (on Hypersil ODS) of these fractions after desalting yielded one region which showed inhibitory activity on all three assays. Inhibition of sodium-potassium-ATPase was dose-related and filtered through an Amicon UM10 membrane. Incubation of this material with dispase, carboxypeptidase A, chymotrypsin, and prolidase destroyed inhibitory activity, whereas trypsin and leucine aminopeptidase were ineffective. These studies show that hypothalamic neurones release a low molecular weight heat-stable peptide which inhibits active sodium transport, ouabain binding, and sodium-potassium-ATPase.
D000818:Animals; D002478:Cells, Cultured; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D004305:Dose-Response Relationship, Drug; D005260:Female; D007031:Hypothalamus / Q000032:analysis*; D011247:Pregnancy; D051381:Rats; D011919:Rats, Inbred Strains; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors*
D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase