Ca²⁺ Enhancement of Hemolysis Induced by the Topical Anesthetic Oxethazaine in Vitro

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Oxethazaine (OXZ), a potent topical anesthetic, was found to induce red blood cell (RBC) lysis in vitro, depending upon concentrations of OXZ, RBC and Ca²⁺. In a 2% RBC suspension, 100 μM OXZ caused almost complete hemolysis in the presence of 1.3 mM Ca²⁺ with only a minimal effect in its absence, while higher concentrations of OXZ (400 μM<) produced hemolysis without Ca²⁺. The hemolysis induced by OXZ plus Ca²⁺ was poreceded by a rapid increase in ⁴⁵Ca²⁺ uptake by RBCs, with both the hemolysis and Ca²⁺ uptake being inhibited by 1 mM CoCl₂, NiCl₂, and quinine. Together with the Ca²⁺ influx, rapid influx of Na⁺ and efflux of K⁺ occurred, and an increasing external K⁺/Na⁺ concentration ratio inhibited both hemolysis and Ca²⁺ influx. Morphologically, OXZ plus Ca²⁺ caused rapid transformation to spheroechinocytes, the formation of blebs and the pinching-off of blebs, whereas OXZ alone produced membrane invagination. SDS-PAGE analysis of the ghosts prepared from the RBCs treated with OXZ plus Ca²⁺ revealed derangement of cytoskeletal components. OXZ is a rate drug that exhibits a Ca²⁺ ionophore-like action, increasing the Ca²⁺ permeability of plasma membranes.