Complement and microglia mediate early synapse loss in Alzheimer mouse models

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Science  06 May 2016:
Vol. 352, Issue 6286, pp. 712-716
DOI: 10.1126/science.aad8373
  • Fig. 1 C1q up-regulation and deposition onto synapses precede preplaque synapse loss in J20 mice.

    (A) Superresolution SIM images of synaptophysin (green)– and PSD95 (red)–immunoreactive puncta in stratum radiatum of 3 mo J20 or WT hippocampus (CA1). Quantification of synaptic puncta or their apposition using Imaris indicates selective loss of PSD95 in J20 hippocampus as compared to their WT littermate controls. See fig. S1. (B) Region-specific up-regulation of C1q (green) in 1 mo J20; DG, dentate gyrus; FC, frontal cortex; STR, striatum; CRB, cerebellum; DAPI, 4′,6-diamidino-2-phenylindole. See fig. S2. (C) Orthogonal view of SIM image showing colocalization of C1q (green) and PSD95 (red). (D) Higher percentage of PSD95 colocalized with C1q in 1 mo J20 dentate gyrus versus WT. (E) Compound E reduces deposited soluble Aβ (red) and C1q (green) in 1 mo J20 dentate gyrus, with minimal effect on C1q levels in WT mice. Scale bar, 2 μm (A, C, and D) or 10 μm (B and E). Means ± SEM; n = 3 or 4 mice per genotype or per treatment group per genotype. *P < 0.05 , **P < 0.01, or ***P < 0.001 using two-way analysis of variance (ANOVA) followed by Bonferroni posttest (A and B), two-tailed one-sample t test (D), or two-tailed unpaired t test (E).

  • Fig. 2 Oligomeric Aβ increases C1q and microglial phagocytic activity.

    (A and B) Soluble Aβ oligomers in WT mice led to elevation of C1q (green) (A) and a higher percentage of PSD95 (red) colocalization with C1q versus monomers (B). (C and D) oAβ induced high levels of CD68 (green) immunoreactivity in Iba1-positive (red) microglia in WT mice (C), but not in those of C1qa KO mice (D). Both had negligible changes in morphology. See fig. S10. Scale bar, 10 μm (A), 5 μm (B), or 20 μm (C). Means ± SEM; n = 3 to 5 mice per treatment group per genotype. *P < 0.05 using two-tailed t test (B) or *P < 0.05, **P < 0.01 versus control-treated or ##P < 0.01 versus Aβ monomer–treated using two-way ANOVA followed by Bonferroni posttest (C).

  • Fig. 3 Complement is necessary for synapse loss and dysfunction in AD models.

    (A) Aβ oligomers induced loss of colocalized synapsin- and PSD95-immunoreactive puncta in the contralateral hippocampus of 3 mo WT mice (left panel); however, they failed to do so in C1qa KO mice (right panel). (B) Coinjection of Aβ oligomers with the function-blocking antibody against C1q, ANX-M1, but not with its IgG isotype control, prevented synapse loss in WT mice. (C) Pretreatment of hippocampal slices with the anti-C1q antibody, ANX-M1, prevented Aβ-mediated LTP inhibition (green) versus IgG (red). IgG alone had a minimal effect (blue) versus artificial cerebrospinal fluid (aCSF) vehicle (black). n = 6 to 11 slices per group. (D) Percentage of PSD95 colocalized with C3 is increased in APP/PS1 hippocampus versus that of WT mice. (E and F) Genetic deletion of C3 prevents synapse loss in 4 mo APP/PS1 mice. Quantification of colocalized immunoreactive puncta for synaptotagmin and homer in dentate gyrus (E) or synaptophysin and PSD95 in CA1 stratum radiatum (F) of WT, APP/PS1, APP/PS1xC3 KO, and C3 KO hippocampi. Means ± SEM; n = 3 to 5 mice per genotype or per treatment group per genotype. *P < 0.05, **P < 0.01, or ***P < 0.001 using two-tailed one-sample t test (D), one-way (A, C, E, F) or two-way (B) ANOVA followed by Bonferroni posttest. ns, not significant.

  • Fig. 4 Microglia engulf synapses via CR3 upon oligomeric Aβ challenge.

    (A) Orthogonal view of high-resolution confocal image shows colocalization of homer-GFP and Iba1 (red). (B) Three-dimensional reconstruction and surface rendering using Imaris demonstrate larger volumes of homer-GFP puncta inside microglia of oAβ-injected contralateral hippocampus versus those of monomer-injected. (C) Microglia of homer-GFPxCR3 KO mice (right panel) show less engulfment of homer-GFP when challenged with oAβ versus those of homer-GFP mice (left panel). (D) Aβ oligomers failed to induce synapse loss in the contralateral hippocampus of CR3 KO mice (right panel) as they did in WT mice (left panel). Scale bar, 5 μm (A and B). Means ± SEM; n = 3 mice per treatment group per genotype (n = 6 to 17 microglia analyzed per mouse). *P < 0.05, **P < 0.01, or ***P < 0.0001 using two-tailed t test (B) or two-tailed one-sample t test (C and D). ns, not significant.

Supplementary Materials

  • Complement and microglia mediate early synapse loss in Alzheimer mouse models

    Soyon Hong, Victoria F. Beja-Glasser, Bianca M. Nfonoyim, Arnaud Frouin, Shaomin Li, Saranya Ramakrishnan, Katherine M. Merry, Qiaoqiao Shi, Arnon Rosenthal, Ben A. Barres, Cynthia A. Lemere, Dennis J. Selkoe, Beth Stevens

    Materials/Methods, Supplementary Text, Tables, Figures, and/or References

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    • Materials and Methods
    • Figs. S1 to S12

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