Effects of 4-Aminopyridine on Organelle Movement in Cultured Mouse Dorsal Root Ganglion Neurites

Abstract

Aminopyridines, widely used as a K⁺ channel blocker, are membrane-permeable weak bases and have the ability to form vacuoles in the cytoplasm. The vacuoles originate from acidic organelles such as lysosomes. Here, we investigated the effects of 4-aminopyridine (4-AP) on organelle movement in neurites of cultured mouse dorsal root ganglion (DRG) neurons by using video-enhanced microscopy. Some experiments were carried out using fluorescent dyes for lysosomes and mitochondria and confocal microscopy. Treatment of DRG neurons with 4 mM 4-AP caused Brownian movement of some lysosomes within 5 min. The Brownian movement gradually became rapid and vacuoles were formed around individual lysosomes 10–20 min after the start of treatment. Axonal transport of organelles was inhibited by 4-AP. Lysosomes showing Brownian movement were not transported in longitudinal direction of the neurite and the transport of mitochondria was interrupted by vacuoles. The 4-AP-induced Brownian movement of lysosomes with vacuole formation and inhibition of axonal transport were prevented by the simultaneous treatment with vacuolar H⁺ ATPase inhibitor bafilomycin A1 or in Cl⁻-free SO ²⁻₄ medium. These results indicate that changes in organelle movement by 4-AP are related to vacuole formation and the vacuolar H⁺ ATPase and Cl⁻ are required for the effects of 4-AP.