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Hippocampal extracellular matrix alterations contribute to cognitive impairment associated with a chronic depressive-like state in rats

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Science Translational Medicine  20 Dec 2017:
Vol. 9, Issue 421, eaai8753
DOI: 10.1126/scitranslmed.aai8753
  • Fig. 1 SDPS induces deficits in rat spatial memory that are reversed by imipramine.

    (A) Rats were exposed to the social defeat–induced persistent stress (SDPS) paradigm, consisting of five daily social defeat episodes and ~3 months of individual housing. Pharmacotherapy with imipramine (IMI) or vehicle (H2O) as control was applied during the last 3 weeks of the isolation period in both groups. Rats underwent behavioral assessment using the object place recognition (OPR) test (B) or the novel object recognition (NOR) test (C). (B) Exploration index during the test phase of the OPR task. SDPS impaired memory retention of the object location; imipramine reversed this deficit but had no effect on control animals. (C) Exploration index during the test phase of the NOR task. Neither SDPS nor imipramine treatment affected recognition performance. Dotted line represents exploration at chance level (0.50); n = number of animals; two-way ANOVA; post hoc Fisher’s LSD; *P < 0.05 (see table S2). Significant memory retention (I) for P < 0.05, and #trend for P < 0.2 by unpaired t test.

  • Fig. 2 SDPS induces increased perisynaptic CSPG expression in the dorsal hippocampus that is reversed by imipramine.

    (A and B) Proteomic analysis using iTRAQ of the dorsal hippocampal synaptic membrane fraction at 3 months after the last social defeat episode. The results revealed 37 SDPS-regulated proteins (adjusted P < 0.05) (A). Expression of 18 of these proteins was rescued by treatment with imipramine (IMI; adjusted P < 0.1, SDPS-IMI versus SDPS-H2O) (A). Extracellular matrix (ECM) proteins, in particular, chondroitin sulfate proteoglycans (CSPGs), were overrepresented in both groups of 37 and 18 differentially expressed proteins (B). (C to F) Independent immunoblot analysis revealed that SDPS increased the synaptic expression of several CSPGs, including brevican (C), neurocan (D), phosphacan (E), and the PNN backbone protein hyaluronan and proteoglycan link protein 1 (HPLN1) (F). Imipramine (IMI) treatment reversed this effect. (G and H) Immunoblots for tenascin-R (160 and 180 kDa) (G), aggrecan and versican (H) showed a moderate effect of SDPS on expression (0.05 < P < 0.20). (I) Representative example blots showing the effect of SDPS on protein expression and that imipramine treatment reversed this effect. The apparent molecular mass is indicated for the specific protein band; total protein loading used for normalization can be found in fig. S4. n = number of samples; PLGEM (A and B), one-way ANOVA (C, D, and F to H), Mann-Whitney (E); *P < 0.05 and **P < 0.01 (see table S2).

  • Fig. 3 SDPS increases the number of PNN-coated parvalbumin-expressing interneurons in the hippocampus.

    (A) PNN-coated (PNN+) parvalbumin-positive (PV+) interneurons of the dorsal hippocampal subfields were quantified for control versus SDPS rats at 2 months after the last defeat. Double-immunopositive interneurons (PNN+ PV+) in the hippocampus CA1 region are indicated by white arrows in the 40× magnification images. (B) SDPS increased the number of PNN+ cells in the CA1 region but not in CA2/3 or the dentate gyrus (DG) regions of the hippocampus. (C and D) The increase in PNN number was specific for PV+ interneurons of the hippocampal CA1 stratum pyramidale region (C) and was not accompanied by an alteration in PNN intensity (D). Scale bars (A), 75 or 25 μm (40×); n = number of animals; N = number of sections; one-way ANOVA (B and C); paired t test (D); *P < 0.05 (see table S2).

  • Fig. 4 SDPS alters parvalbumin-positive interneuron properties and decreases inhibitory transmission in the hippocampus.

    (A and B) In the hippocampal CA1 stratum pyramidale, SDPS did not affect the total number of parvalbumin-positive interneurons having a PNN coat (PNN+ PV+) (A) but did cause a moderate (7.8 ± 1.0%) increase in the intensity of parvalbumin immunoreactivity in SDPS versus control animals (B). (C and D) Representative examples of labeling of hippocampal CA1 parvalbumin-positive interneurons in SDPS and control animals (high intensity, white arrowheads; intermediate-low intensity, yellow arrowheads) (C). (D) Within double-immunopositive (PNN+ PV+) interneurons, SDPS decreased the fraction of intermediate-low parvalbumin–expressing cells (control, 22.1%; SDPS, 11.8%) and increased the fraction of high parvalbumin–expressing interneurons (control, 26.2%; SDPS, 44.1%) (D, left). No difference in the fraction of low or intermediate-high parvalbumin–expressing interneurons was observed. No intensity shift was observed in PNN-free parvalbumin-positive (PNN PV+) interneurons (D, right). (E and F) Quantification of bassoon-positive (Bs+) puncta showed no effect of SDPS on perisomatic excitatory input onto PNN+ PV+ interneurons [representative example (E)]. In control and SDPS animals alike, PNN PV+ interneurons showed increased density of Bs+ puncta versus PNN+ PV+ cells (F). AU, arbitrary units. (G) Example traces of whole-cell patch-clamp recordings (5 s) of hippocampal CA1 pyramidal neurons. (H) SDPS reduced sIPSC frequency (left) while leaving amplitude unaffected (right). Scale bars, 50 μm (C) or 20 μm (E); str.or, stratum oriens; str.pyr, stratum pyramidale; str.rad, stratum radiatum; n = number of animals; N = number of sections/slices; Mann-Whitney (A and F); paired t test (B and F); one-way ANOVA (D and F); *P < 0.05 and **P < 0.01 (see table S2).

  • Fig. 5 Intrahippocampal chondroitinase ABC administration restores PNNs, hippocampal function, and memory recall after SDPS.

    (A) After exposure to SDPS or no exposure (control), animals received either intrahippocampal administration of chondroitinase ABC (ChABC) or penicillinase (Peni) as a control. Performance on the object place recognition (OPR) test was assessed 12 days after administration (H and I) and was followed by LTP measurements at 12 to 24 days after treatment (F and G). Immunohistochemistry (B and C) and sIPSC recordings (D and E) were performed at 12 to 14 days after treatment. (B and C) SDPS increased the number of double-immunopositive (PNN+ PV+) interneurons [representative example, (C)], and treatment with chondroitinase ABC reversed this effect. Chondroitinase ABC treatment reduced the number of PNN+ PV+ neurons compared to penicillinase treatment. (D and E) Frequency of sIPSCs [representative example traces, (D)] was reduced after SDPS, and chondroitinase ABC treatment rescued this effect. Chondroitinase ABC treatment had no effect on sIPSC frequency in control rats. (F and G) Maintenance of LTP, expressed as fEPSP slope, was decreased in SDPS rats and restored after chondroitinase ABC treatment. Chondroitinase ABC treatment had no effect on LTP maintenance in control animals. (G) Representative example of placement on the MED-64 grid with fEPSP traces before and after (gray/black, respectively) high-frequency stimulation to induce LTP. (H and I) Rats exposed to the SDPS paradigm showed impaired object location memory on the OPR test, and chondroitinase ABC reversed this effect. (H) Representative example of animal movements during the OPR test. Yellow squares represent the displaced object. Scale bar (C), 25 μm. Dotted line represents baseline fEPSP slope before high-frequency stimulation (F) or exploration at the chance level (0.50) (I); n = number of animals; N = number of cells/sections; one-way ANOVA and post hoc Fisher’s LSD (B, E, and F); two-way ANOVA and post hoc Fisher’s LSD (I); *P < 0.05 and **P < 0.01 (see table S2). Significant memory retention (I) for P < 0.05, and #trend for P < 0.2 by unpaired t test.

Supplementary Materials

  • www.sciencetranslationalmedicine.org/cgi/content/full/9/421/eaai8753/DC1

    Materials and Methods

    Fig. S1. The SDPS paradigm elicits physiological stress responses that subside after several weeks.

    Fig. S2. The SDPS paradigm triggers imipramine-reversible reduction in LTP maintenance.

    Fig. S3. Expression of synaptic proteins after SDPS.

    Fig. S4. Representative immunoblots and corresponding loading control.

    Fig. S5. Overall CSPG expression is not affected by SDPS.

    Fig. S6. Cat-301 recognizes a CSPG-rich PNN population in hippocampus.

    Fig. S7. Cat-301 recognizes aggrecan-rich PNNs, which increase after SDPS.

    Fig. S8. CSPG-rich PNN characterization in dorsal hippocampus CA1 layers.

    Fig. S9. SDPS does not affect PNN number in the perirhinal cortex.

    Fig. S10. The effects of chondroitinase ABC on CSPGs and PNN recovery 2 weeks after administration.

    Fig. S11. Chondroitinase ABC does not affect sIPSC amplitude.

    Fig. S12. Extracellular matrix reorganization rescues SDPS-induced deficits in social recognition and mildly attenuates social withdrawal.

    Table S1. SDPS-induced changes in dorsal hippocampus synaptic protein expression and rescue by the antidepressant imipramine.

    Table S2. Overview of statistical tests used in the main figures.

    References (7790)

  • Supplementary Material for:

    Hippocampal extracellular matrix alterations contribute to cognitive impairment associated with a chronic depressive-like state in rats

    Danai Riga, Ioannis Kramvis, Maija K. Koskinen, Pieter van Bokhoven, Johanneke E. van der Harst, Tim S. Heistek, A. Jaap Timmerman, Pim van Nierop, Roel C. van der Schors, Anton W. Pieneman, Anouk de Weger, Yvar van Mourik, Anton N. M. Schoffelmeer, Huib D. Mansvelder, Rhiannon M. Meredith, Witte J. G. Hoogendijk, August B. Smit, Sabine Spijker*

    *Corresponding author. Email: s.spijker{at}vu.nl

    Published 13 December 2017, Sci. Transl. Med. 9, eaai8753 (2017)
    DOI: 10.1126/scitranslmed.aai8753

    This PDF file includes:

    • Materials and Methods
    • Fig. S1. The SDPS paradigm elicits physiological stress responses that subside after several weeks.
    • Fig. S2. The SDPS paradigm triggers imipramine-reversible reduction in LTP maintenance.
    • Fig. S3. Expression of synaptic proteins after SDPS.
    • Fig. S4. Representative immunoblots and corresponding loading control.
    • Fig. S5. Overall CSPG expression is not affected by SDPS.
    • Fig. S6. Cat-301 recognizes a CSPG-rich PNN population in hippocampus.
    • Fig. S7. Cat-301 recognizes aggrecan-rich PNNs, which increase after SDPS.
    • Fig. S8. CSPG-rich PNN characterization in dorsal hippocampus CA1 layers.
    • Fig. S9. SDPS does not affect PNN number in the perirhinal cortex.
    • Fig. S10. The effects of chondroitinase ABC on CSPGs and PNN recovery 2 weeks after administration.
    • Fig. S11. Chondroitinase ABC does not affect sIPSC amplitude.
    • Fig. S12. Extracellular matrix reorganization rescues SDPS-induced deficits in social recognition and mildly attenuates social withdrawal.
    • Table S1. SDPS-induced changes in dorsal hippocampus synaptic protein expression and rescue by the antidepressant imipramine.
    • Table S2. Overview of statistical tests used in the main figures.
    • References (7790)

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