Lysosomes are reliably labeled by LAMP1-GFP and are found in distal dendrites and dendritic spines. (A) Endogenous LAMP1 distribution is similar to LAMP1-GFP, as both labeled structures were found in somatic and dendritic compartments. Representative images of dissociated hippocampal neurons (DIV16) either stained for endogenous LAMP1 (left) or transfected with LAMP1-GFP (right) and allowed to express for ≤24 h. (B) Representative image of LysoTracker red–labeled acidic vesicles found in dendritic compartments (DIV16). Inset shows entire cell image of neuron expressing GFP (Sindbis virus). (C) LAMP1-GFP marks acidic vesicles. Representative image of a dendrite from a dissociated hippocampal neuron transfected with LAMP1-GFP and costained with LysoTracker red, which marks acidic compartments. Neurons were imaged live at DIV16. (D) LAMP1-GFP does not colocalize with early endosomes. Representative image of a dendrite from dissociated hippocampal neuron (DIV16) transfected with mCherry and LAMP1-GFP and stained for the endogenous early endosomal protein marker EEA1. (E and F) LAMP1-GFP–positive vesicles are found in distal dendrites. A straightened secondary distal dendrite from a dissociated hippocampal neuron (DIV16) with LAMP1-GFP present throughout the dendrite. Depicted are a representative whole-cell image (E) and straightened distal secondary dendrite (F). (G) LAMP1-GFP is present at the base, neck, and head of dendritic spines. Representative images of dendritic spines in cultured hippocampal neurons (DIV16) transfected with mCherry and LAMP1-GFP. (H) Quantification the percentage of spines with a lysosome either in the head, neck, or base of the spine. Data represent means ± SEM. (I) Representative two-photon image of a CA1 pyramidal neuron from a hippocampal organotypic (DIV8) slice biolistically transfected with LAMP1-GFP and dsRed. LAMP1-GFP–labeled lysosomes are found in dendritic spines.