Oligodendrocyte precursor cells facilitate neuronal lysosome release

Abstract

Oligodendrocyte precursor cells (OPCs) shape brain function through many non-canonical regulatory mechanisms beyond myelination. Here we show that OPCs form contacts with their processes on neuronal somata in a neuronal activity-dependent manner. These contacts facilitate exocytosis of neuronal lysosomes. A reduction in the number or branching of OPCs reduces these contacts, which is associated with lysosome accumulation and altered metabolism in neurons and more senescent neurons with age. A similar reduction in OPC branching and neuronal lysosome accumulation is seen in an early-stage mouse model of Alzheimer's disease. Our findings have implications for the prevention of age-related pathologies and the treatment of neurodegenerative diseases.

Fig. 1. OPC processes contact neuronal somata with a preference for active neurons.

A, B Immunostaining and 3D reconstruction of OPCs and neurons with GFP, PDGFRα and NeuN antibodies in coronal brain slices of NG2^EYFP mice. B₁ Exemplary images of process-somata contact (PSC) between OPCs and neurons from the boxed area of B, shown in three directions. C Percentage of neurons with PSCs in the cortex (ctx), hippocampal CA1 region (ca1), thalamus (th), hypothalamus (hy) and amygdala (amg) (N = 3 mice). D–F Ultrastructural images (D, E) and quantification (F) of PSCs in mouse (D) and human cortex (E). G, H Immunostaining and quantification of OPC (GFP⁺) contacts on GABAergic (GABA⁺) and non-GABAergic (GABA^-) neurons in NG2^EYFP mice (127 GABA^- neurons and 109 GABA⁺ neurons from 5 mice, normalized to GABA^- neurons, two-sided unpaired t test). I Immunostaining of OPCs and neurons with GFP and NeuN in the cortex of NG2^EYFP mice. Neuronal activity was indicated with cFos (magenta) immunoreactivity and contacting processes were indicated in yellow. I₁ 3D-reconstruction of neuronal somata and OPCs. OPC processes contacting neuronal somata were indicated in yellow. I₂ Exemplary cFos⁺ and cFos^- neurons with OPC contacts. J Quantification of relative volume of OPC processes (GFP⁺) contacting cFos⁺ and cFos^- neurons (normalized to cFos^- cells). (117 cFos^- cells and 118 cFos⁺ cells from 8 mice, two-sided unpaired t-test for cells (small dots) and two-sided paired t-test for mice (large dots)). K Live-imaging of neuron (hM3D(Gq)-mCherry⁺)-OPC (CellTrace⁺) interaction before and after CNO application. L The relative number of processes contacting (0–0.25 µm), near (0.25–0.5 µm) and far from (0.5–1.0 µm) the neurons were quantified and compared before and after CNO application (5 cells/5 independent experiments, one-way ANOVA, Multiple comparison). R.U.=relative units, normalized to the controls (5 cells/5 independent experiments, in each experiment OPCs are prepared from 4 pups). Data are shown as mean ± SEM in C, H, J, and L. Source data are provided as a Source Data file. Created in BioRender. Fang, L. (2025) https://BioRender.com/n88b804.

Fig. 2. OPC contact facilitates neuronal lysosome release at the contact site.

A–C Immunostaining and 3D-reconstruction of neurons, OPCs and lysosomes in NG2^EYFP mouse cortex using NeuN, GFP and Lamp1 antibodies. B OPC processes contacting neuronal somata were indicated in yellow. C Neuronal lysosomes shown in colors based on their volume. D Display of lysosome volume (indicated by circle size) and their proximity to OPC and neuronal surfaces (5925 lysosomes from 17 cells/3 mice). The color indicates the frequency of lysosomal appearance at corresponding location. E Correlation of lysosome volume and proximity to OPC surface (2077 lysosomes from 17 cells/3 mice). F Live-imaging of a lysosome (dotted circle) trafficking in neuron-OPC co-culture and analysis of maximum fluorescence intensity (MAX FI) of lysosomes over the period. Dotted line indicates neuronal membrane. G Live-imaging and quantification of lysosome exocytosis in neurons from pure neuronal culture and neurons with (neuron^OPC-contact) or without OPC contact (neurons^{w/o-OPC-contact}) from co-culture. In neurons^OPC-contact, the Alexa647⁺ puncta were classified into proximal (<1 µm, magenta) and distal (>1 µm, green) based on their distance to OPC contact site (Pure neuron=9 cells/4 independent experiments; neurons^OPC-contact = 9 cells/4 independent experiments, neurons^{w/o-OPC-contact} = 3 cells/3 independent experiments; 4 pups/experiment, two-way ANOVA, multiple comparisons). H Lysosome exocytosis in acute brain slices was imaged and the number of Alexa647⁺ puncta in each neuron was quantified (ctl = 10 cells/3 mice; dKO=6 cells/3 mice). I Immunostaining and 3D-reconstruction of OPCs (PDGFRα⁺), neurons (NeuN⁺) and lysosomes (Cathepsin D, CTSD⁺). I₁ Quantification of relative volume of OPC processes contacting neuronal somata (ctl = 190 cells/5 mice, dKO = 116 cells/5 mice). I₂, I₃ Analysis of the relative number and volume of neuronal lysosomes. (ctl = 112 cells/7 mice, dKO = 74 cells/4 mice). J Immunolabeling and 3D reconstruction of neurons, OPCs and lysosomes with NeuN, PDGFRα and Lamp1 antibodies. The relative number and volume of neuronal lysosomes were quantified. (ctl = 80 cells/5 mice, DTA = 52 cells/4 mice). In (H–J), small dots represent cells while big dots represent mice, two-sided unpaired t-tests were used. R.U. = relative units, normalized to the controls. Data are shown as mean ± SEM in (G–J). Source data are provided as a Source Data file. Created in BioRender. Fang, L. (2025) https://BioRender.com/a00p319 and https://BioRender.com/v85j019.

Fig. 3. Reduced OPC-neuron contacts induce an accumulation of lysosomes and functional impairment in neurons.

A Immunostaining and quantification of lipid droplets in cortical neurons of control (ctl) and double knockout (dKO) mice with NeuN and BODIPY antibodies, respectively (4 mice per group). B Immunostaining and quantification of cFos⁺ cells in the whole brain of ctl and dKO mice (4 mice per group). C, D Dot plot of enriched gene ontology terms for biological processes (BP), cellular components (CC), molecular functions (MF), and KEGG pathways in genes from OPCs and neurons of dKO mice. E List of differentially expressed genes in OPCs and neurons of the dKO mice (M, bulkRNA-seq) and AD patients in early pathology (H, scRNA-seq). F, G Immunostaining and quantification of senescent neurons with NeuN and p16^INK4A (P16) at 12 months of age (ctl = 3 mice, dKO = 4 mice). H 3D reconstruction of OPCs immunolabelled with PDGFRα (Pα) at 6 months of age. I, J Morphological analysis of OPCs for the number of process intersections (I) and total process length (J) (ctl = 51 cells/3 mice, Tg2576 = 52 cells/3 mice). K Immunostaining and 3D reconstruction of OPCs (Pα), neurons (NeuN) and lysosomes (Cathepsin D, CSTD). L Quantification of the relative volume of OPC processes in contact with neuronal soma (ctl = 91 cells/3 mice, Tg2576 = 117 cells/3 mice) and the number of neuronal lysosomes (ctl=87 neurons/5 mice, Tg2576 = 115 neurons/5 mice). M, N Quantification of the relative mean distance between neuronal lysosomes and OPC surface (ctl = 91 cells/3 mice, Tg2576 = 117 cells/3 mice). M The size of the lysosomes is represented by the size of the filled circles, while the color indicates the frequency of each size in the total analysis. R.U. = relative units, normalized to the controls. Data are shown as mean ± SEM in (A, G, I, J, L, and N). Two-sided unpaired t test was used in (A, B, G, I, J, L, and N). Source data are provided as a Source Data file. Created in BioRender. Fang, L. (2025) https://BioRender.com/f32o880.