Radioactive Labeling of Phospholipids and Proteins by Cochlear Perfusion in the Guinea Pig and the Effect of Neomycin

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Abstract
Phospholipids and proteins of guinea pig stria vascularis, spiral ligament and organ of Corti were radio-actively labeled by perilymphatic perfusion with artificial perilymph containing [32P]orthophosphate or radioactive amino acids. Phospholipids were separated by thin-layer chromatography, proteins by disc gel electropho-resis and quantitated by liquid scintillation counting. The addition of 10-4M to 10-2M neomycin to the perfusion fluid resulted in a dose-dependent increase of tissue permeability to the radioactive precursors, and a specific decrease in the 32P-incorporation into phosphatidylinositol diphosphate in stria vascularis and organ of Corti. No effect of neomycin on protein labeling was observed using a double label approach with [3H]methionine and [35S]-methionine. In vitro, low concentrations of neomycin led to the formation of a complex with polyphosphoinositides. Much higher concentrations of the drug were needed for a comparable reaction with the acid mucopolysaccharide, chondroitin sulfate A. The implications of these findings for the mechanism of neomycin ototoxicity are discussed.