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Spine dynamics of PSD-95-deficient neurons in the visual cortex link silent synapses to structural cortical plasticity - PubMed

  • ️Fri Jan 01 2021

Spine dynamics of PSD-95-deficient neurons in the visual cortex link silent synapses to structural cortical plasticity

Rashad Yusifov et al. Proc Natl Acad Sci U S A. 2021.

Abstract

Critical periods (CPs) are time windows of heightened brain plasticity during which experience refines synaptic connections to achieve mature functionality. At glutamatergic synapses on dendritic spines of principal cortical neurons, the maturation is largely governed by postsynaptic density protein-95 (PSD-95)-dependent synaptic incorporation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors into nascent AMPA-receptor silent synapses. Consequently, in mouse primary visual cortex (V1), impaired silent synapse maturation in PSD-95-deficient neurons prevents the closure of the CP for juvenile ocular dominance plasticity (jODP). A structural hallmark of jODP is increased spine elimination, induced by brief monocular deprivation (MD). However, it is unknown whether impaired silent synapse maturation facilitates spine elimination and also preserves juvenile structural plasticity. Using two-photon microscopy, we assessed spine dynamics in apical dendrites of layer 2/3 pyramidal neurons (PNs) in binocular V1 during ODP in awake adult mice. Under basal conditions, spine formation and elimination ratios were similar between PSD-95 knockout (KO) and wild-type (WT) mice. However, a brief MD affected spine dynamics only in KO mice, where MD doubled spine elimination, primarily affecting newly formed spines, and caused a net reduction in spine density similar to what has been observed during jODP in WT mice. A similar increase in spine elimination after MD occurred if PSD-95 was knocked down in single PNs of layer 2/3. Thus, structural plasticity is dictated cell autonomously by PSD-95 in vivo in awake mice. Loss of PSD-95 preserves hallmark features of spine dynamics in jODP into adulthood, revealing a functional link of PSD-95 for experience-dependent synapse maturation and stabilization during CPs.

Keywords: awake; plasticity; silent synapses; spine dynamics; visual cortex.

Copyright © 2021 the Author(s). Published by PNAS.

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Conflict of interest statement

The authors declare no competing interest.

Figures

Fig. 1.
Fig. 1.

Repeated two-photon imaging of apical dendrites of L2/3 pyramidal neurons in the primary visual cortex (V1) of awake head-fixed PSD-95 WT and KO mice during periods of normal vision (NV), monocular deprivation (MD), and after reopening the formerly deprived eye (RO). (A) Experimental timeline: a plasmid expressing eGFP or eGFP/sh95 was delivered into the left ventricle of E15.5 PSD-95 WT and KO pups and electroporated targeting V1. After implanting a chronic cranial window (∼P45), optical imaging of intrinsic signals was used to locate binocular V1 at ∼P50. Mice were then screened for sparse neuronal labeling and habituated to head restraint for 15 to 20 d. After baseline measurements (d0 to d4), a 4-d MD of the contralateral eye was started on d4, and the eye was reopened on d8. Awake two-photon imaging of dendritic spines was performed on all days indicated by the black arrows. (B) Coronal slice of an electroporated brain illustrating L2/3 pyramidal neuron-specific eGFP expression in V1. (Scale bar, 400 μm.) (C) Cortical blood vessel pattern imaged through the cranial window with an outline of binocular V1 (dashed ellipse), located by (D) retinotopic mapping using intrinsic signal optical imaging (Scale bar, 1 mm). (E) Low-magnification top view of an imaged region with sparsely eGFP-labeled L2/3 pyramidal neurons. (Scale bar, 50 μm.) (F) Average baseline spine densities (±SEM) of apical dendrites in PSD-95 WT and KO mice on d0 to d4. (G) The same dendrites were repeatedly imaged with 1-d (5 WT/4 KO: 25/21 dendrites) and 4-d (6 WT/6 KO: 40/36 dendrites) intervals during NV and MD periods. Imaging was continued 2 d (6 WT/6 KO: 33/31 dendrites) and 4 d (6 WT/6 KO: 33/30 dendrites) after RO. Spine density (normalized to average baseline) was significantly reduced in PSD-95 KO but not WT mice after 4-d MD, and the reduction persisted after reopening the eye. *P < 0.05; **P < 0.01; ***P < 0.001; ns, P > 0.05.

Fig. 2.
Fig. 2.

In V1 of adult PSD-95 KO mice, monocular deprivation induces increased spine elimination and decreased spine formation on apical dendrites of L2/3 pyramidal neurons. Experience-dependent spine dynamics of L2/3 pyramidal neurons and their quantification. (A) Representative examples of dendrites from PSD-95 WT (n = 6) and (B) KO mice (n = 6) on d0 (Left), d4 (Middle), and d8 (Right): red arrows mark eliminated and green arrows mark newly formed spines during 4-d intervals. (Scale bar, 5 μm). (C and D) Mean (±SD) spine elimination and (F and G) spine formation ratios during 1-d (C and F; 5 WT/4 KO: 25/21 dendrites) and 4-d (D and G; 6 WT/6 KO: 40/36 dendrites) intervals, calculated as R = Nelmn or Nform/(Ninitial + Nfinal). (E) MD-induced change Δ = RMD - RNV in spine elimination and (H) formation ratios (mean ± SEM). *P < 0.05; **P < 0.01; ***P < 0.001; ns, P > 0.05.

Fig. 3.
Fig. 3.

Newly formed spines in PSD-95 KO mice are more likely to be eliminated during MD. MD-induced changes in persistent (lifetime > 4 d) and newly formed spine populations in PSD-95 KO and WT mice. (A) Percentage of persistent spines (mean ± SD) present from d0 throughout NV and MD periods. (B) Histogram depicts relative frequency of PSD-95 WT and KO dendrites binned according to the fractions of newly formed spines that were eliminated during 4-d MD (0.00 = none eliminated; 1.00 = all new spines eliminated during MD). Note that in PSD-95 KO mice, the frequency histogram is skewed toward more spine elimination during MD. *P < 0.05.

Fig. 4.
Fig. 4.

Dendritic spine formation of L2/3 pyramidal neurons returns to baseline values in PSD-95 KO mice, while no significant changes happen in WT mice after reopening. Imaging was continued in a subset of dendrites described in Fig. 2, 4 d (6 WT/6 KO: 33/30 dendrites) after reopening the MD eye. (A) Representative examples of dendrites from PSD-95 WT and (B) PSD-95 KO mice on d8 (Left) and d12 (Right): green arrows mark formed and red arrows mark eliminated spines compared to d8 (Scale bar, 5 μm). (C) Comparison of mean (±SD) spine elimination and (D) formation ratios during 4-d reopening to NV and MD periods (gray rectangle, data replotted from Fig. 2 D and G for comparison). *P < 0.05; **P < 0.01; ***P < 0.001; ns, P > 0.05.

Fig. 5.
Fig. 5.

In V1 of WT mice, MD induces increased spine elimination on apical dendrites of PSD-95 knockdown L2/3 pyramidal neurons. (A) Apical dendrites of L2/3 pyramidal neurons expressing either eGFP-tagged control (gfp: 4 mice/n = 19) or (B) PSD-95 knockdown plasmids (sh95: 3 mice/n = 21) were repeatedly imaged with 1- and 4-d intervals during NV and MD periods: red arrows mark eliminated, green arrows mark formed spines during 4-d intervals. (Scale bar, 5 μm.) (C) Average baseline spine density (±SEM) during NV and net change of spine density during NV and MD periods (gray shaded area) relative to baseline (d0 to d4). (D and E) Mean (±SD) spine elimination and (G and H) spine formation ratios during (D and G) 1-d and (E and H) 4-d intervals. (F) MD-induced change (Δ = RMD - RNV) in spine elimination and (I) formation ratios (mean ± SEM). *P < 0.05; **P < 0.01; ns, P > 0.05.

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