Soluble polycations and cationic amphiphiles inhibit volume-sensitive K-Cl cotransport in human red cell ghosts

We have measured the effect of soluble polycations (spermine and methylglyoxal) and cationic amphiphiles (sphingosine and tetracaine) on K-Cl cotransport in shrunken and swollen red cell ghosts. All substances inhibited cotransport, and for each agent, the concentration at which inhibition was half-maximal was about the same for swollen and shrunken ghosts. Acetylspermine was a much less effective inhibitor than spermine, which demonstrates that inhibition depends on the cationic groups of spermine. Spermine was a more effective inhibitor in ATP-free ghosts than in ghosts containing ATP, which eliminates the possibility that inhibition of cotransport activity results from inhibition of protein kinase activity. Inhibition by spermine is as effective in K-free ghosts as in high-K ghosts; spermine does not inhibit cotransport by reducing the effective K concentration at the inner membrane surface. We conclude that regulation of K-Cl cotransport involves negative charges (phosphatidylserine or phosphatidylinositides) at the inner membrane surface and suggest a model that accounts for our findings.