Studies on lithium transport across the red cell membrane

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Abstract
The reactivity of the SH-group essential for ouabain-resistant Na+−Li+ (and Na+−Na+) exchange and its location within the membrane are studied on human and beef erythrocytes and beef red cell ghosts. N-ethylmalcimide (NEM), 1,6-hexane dimaleimide, and iodoacetamide can induce an irreversible, partial inhibition of Na+−Li+ exchange in erythrocytes of the two species. The development of the inhibition due to the alkylating agents is greatly accelerated by external Na+ and Li+. The inhibition takes 3 min (NEM) and 60 min (iodoacetamide) to come to completion in isotonic Na+ media, but is hardly detectable in choline+, K+ or Mg2+ media. The transport site of the exchange system and the site promoting NEM binding exhibit similar affinities for external Na+. The impermeable, monofunctional glutathione derivative of 1,6-hexane dimaleimide does not inhibit Na+−Li+ exchange. The mercurials PCMBS, PCMB, and Hg2+ inhibit Na+−Li+ exchange in beef, but not in human erythrocytes. The inhibitory action of PCMBS, being slightly accelerated by external Na+, is fully reversed by penetrating thiols such as 2-mercaptoethanol, whilst glutathione, an impermeable thiol, is ineffective. Pretreatment with PCMBS affords partial protection from the irreversible inhibition caused by NEM. Oxidation with copper orthophenanthroline inhibits Na+−Li+ exchange only when performed in the presence of penetrating thiols such as 2-mercaptoethanol. It is concluded that the SH-reagents studied inhibit Na+−Li+ exchange by modifying an essential SH-group of a membrane protein in such a way that the turnover number of the exchange system is reduced. This SH-group is separated from both the red cell exterior and interior by a penetration barrier and seems to be distinct from the cation binding site. The action of external Na+ and Li+ in promoting the reaction of alkylating inhibitors is interpreted to result from a conformational change of the transport protein induced by the binding of external Na+ or Li+.