Primaquine interferes with membrane recycling from endosomes to the plasmamembrane through a direct interaction with endosomes which does not involve neutralisation of endosomal pH nor osmotic swelling of endosomes

The anti-malaria drug primaquine is a weak base which accumulates in endoso mes in a protonated form and consequently neutralises the endosomal pH, Baf ilomycin Al prevents endosome acidification by inhibiting the vacuolar prot on pump. Although both agents neutralise the endosomal pH, only primaquine has a strong inhibitory effect on recycling of endocytosed proteins to the plasma membrane (Van Weert et al, (1995), J, Cell Biol, 130, 821-834), This suggests that primaquine interferes with a parameter, other than endosomal pH, that is essential for membrane recycling, In the presence of 0.3 mM pr imaquine, endocytosed transferrin-receptors accumulated intracellularly, bu t not in the additional presence of bafilomycin Al. Thus, at relative low c oncentrations proton pump-driven accumulation of primaquine in endosomes wa s required to inhibit membrane recycling, suggesting that the target of pri maquine is associated with endosomes, The inhibitory effect of 1 mM primaqu ine on transferrin receptor recycling was not reversed by the additional pr esence of bafilomycin Al, indicating that osmotic swelling of endosomes due to accumulation of protonated primaquine could also not explain its effect . To study endosome swelling morphologically, we introduce a novel techniqu e for fluorescent labelling of endosomes involving HRP-catalysed biotinylat ion. In the presence of 0.2 mM primaquine endosomal vacuoles with diameters up to 2 mu m were observed. Endosome swelling was not observed when in add ition to primaquine also bafilomycin Al was present, supporting the notion that vacuolar proton pump activity lowers the dose response for primaquine, Factors that are crucial for membrane recycling and may be affected by pri maquine are discussed.