MMP2 and MMP9 Activity Is Crucial for Adult Visual Cortex Plasticity in Healthy and Stroke-Affected Mice

Abstract

A fundamental regulator of neuronal network development and plasticity is the extracellular matrix (ECM) of the brain. The ECM provides a scaffold stabilizing synaptic circuits, while the proteolytic cleavage of its components and cell surface proteins are thought to have permissive roles in the regulation of plasticity. The enzymatic proteolysis is thought to be crucial for homeostasis between stability and reorganizational plasticity and facilitated largely by a family of proteinases named matrix metalloproteinases (MMPs). Here, we investigated whether MMP2 and MMP9 play a role in mediating adult primary visual cortex (V1) plasticity as well as stroke-induced impairments of visual cortex plasticity in mice. In healthy adult mice, selective inhibition of MMP2/9 for 7 d suppressed ocular dominance plasticity. In contrast, brief inhibition of MMP2/9 after a cortical stroke rescued compromised plasticity. Our data indicate that the proteolytic activity of MMP2 and MMP9 is critical and required to be within a narrow range to allow adult visual plasticity.SIGNIFICANCE STATEMENT Learning and recovery from injuries depend on the plasticity of neuronal connections. The brain's extracellular matrix (ECM) provides a scaffold for stabilizing synaptic circuits, while its enzymatic proteolysis is hypothesized to regulate homeostasis between stability and reorganizational plasticity. ECM digestion is facilitated by a family of matrix metalloproteinases (MMPs). Here, we show that treatments that inhibit MMP2/9 can either inhibit or rescue cortical plasticity depending on cortical state: in the visual cortex of healthy adult mice, inhibition of MMP2/9 suppressed cortical plasticity. In contrast, brief inhibition of MMP2/9 after a stroke rescued compromised plasticity. Our data provide strong evidence that an optimal level of MMP2/9 proteolytic activity is crucial for adult visual plasticity.

Keywords: MMP; extracellular matrix; ocular dominance plasticity; plasticity; stroke; visual cortex.

Figure 1.

Treatment with the MMP2/9 inhibitor SB3CT during the MD period suppressed OD plasticity in adult mouse V1. A, Experimental timeline testing the effect of selective MMP2 and MMP9 inhibition on visual plasticity in healthy adult mice. Between P70 and P110, a group of mice received MD. From d0 (before MD), mice were checked daily for 7 d in the optomotor setup for the assessment of both spatial frequency and contrast sensitivity thresholds. One hour after MD, mice of all experimental groups were injected intraperitoneally with SB3CT or vehicle solution; injections continued until d7, when V1 activity was visualized using intrinsic signal optical imaging. B–E, Representative activity maps of the left binocular V1 and ODI values of vehicle-injected (B, D) or SB3CT-injected (C, E) mice. Greyscale-coded response magnitude maps, polar maps, two-dimensional OD maps, and histograms of OD scores including the average ODIs are illustrated. In mice without MD, activity patches evoked by stimulation of the contralateral eye were darker than those of the ipsilateral eye, the average ODI was positive, and warm colors prevailed in the OD maps, indicating contralateral dominance (B, C). In vehicle-injected mice, 7-d MD induced a strong OD shift toward the nondeprived (ipsilateral) eye (D): ipsilateral eye stimulation now caused darker V1-activity patches compared with noMD mice, colder colors appeared in the OD map, and average OD scores shifted to the left. In contrast, SB3CT-injected mice did not display OD plasticity (E). After MD, the contralateral eye continued to dominate V1 activation, warm colors continued to prevail in the OD map, and the average ODI was not changed (red) in SB3CT-treated mice (E). Scale bars: 1 mm.

Figure 2.

Quantification of ODIs (A) and V1 activation (B) in vehicle-injected (black) and SB3CT-injected (green) mice with or without MD. A, Administration of SB3CT for 7 d abolished OD plasticity in MD mice. Symbols represent ODI values of individuals; filled circles represent mice without MD and half-filled circles mice with MD; mean values are shown as horizontal lines. B, V1 activation (fractional change in reflectance ×10⁻⁴) elicited by stimulation of the contralateral (C) or ipsilateral (I) eye without and after MD (deprived eye is marked by a black filled circle on the x-axis). In vehicle-treated mice, the OD shift was primarily mediated by an increase of open (ipsi) eye responses in V1, while after SB3CT treatments, there was no significant increase in open eye responses of MD mice receiving injections. Multivariate ANOVA, followed up by Sidak-corrected multiple comparisons. Mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001. Tables comparing the two independent analyses are provided as the Extended Data Figure 2-1.

Figure 3.

Daily injection of the MMP2/9-specific inhibitor SB3CT increased PNNs in V1, stained with WFA. A, WFA-positive staining of PNNs in vehicle-injected (left) and SB3CT-injected (right) mice. PNNs were identified by their shape and bright circular outline throughout the layers of V1. One example of each condition is displayed. On the right, noMD/MD conditions of vehicle-injected mice are shown. On the left, noMD/MD conditions of SB3CT-inejcted mice are shown. Scale bar: 100 µm. B, Quantification of PNNs in V1. Number of WFA-positive PNNs were significantly higher in SB3CT-injected mice, regardless of MD. From each brain, three sections containing V1 were analyzed. ANOVA followed up by Sidak-corrected test for simple effects. Scale bar: 100 µm. Mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001.

Figure 4.

Injection of the MMP2/9 inhibitor SB3CT prevented experience-induced enhancements of both the spatial frequency (A) and contrast sensitivity (B) thresholds of the optomotor reflex of the open eye in adult mice after MD (half-filled circles). A, Spatial frequency threshold values in cyc/° are plotted against days, for mice without (filled circles) and with MD (half-filled circles) for both vehicle-treated (black) and SB3CT-treated (green) mice. After 7 d of MD, spatial frequency thresholds improved significantly in vehicle-injected MD mice when compared with mice without MD and SB3CT-injected MD mice. In contrast, SB3CT-injected MD mice did not significantly increase thresholds until D7, while control MD mice improved significantly each day. Mice without MD did not show any change over days. Three-way mixed ANOVA was used, followed up by Sidak-corrected tests for simple effects. B, In vehicle-treated MD mice, contrast sensitivity thresholds of the open eye increased significantly after 7-d MD compared with both noMD mice (filled circles) and SB3CT-injected noMD mice (blue). SB3CT MD mice (blue, half-filled circles) had significantly lower contrast sensitivity thresholds than vehicle-injected mice (black, half-filled circles), similar to noMD mice (filled circles). Gratings left of the y-axis illustrate the contrast steps for a given (constant) spatial frequency. Four-way mixed ANOVA, followed up by Sidak-corrected tests for simple effects. Mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001.

Figure 5.

Brief treatment with the MMP2 and MMP9-specific inhibitor SB3CT rescued adult OD plasticity in V1 after a stroke in S1. A, The experimental design to test the effect of MMP2 and MMP9 inhibition on OD plasticity after a PT stroke in S1. A small (1 mm in diameter) lesion in S1 was induced on young adult mice (P70–P90) using PT. Shortly after (1 h), half of the PT mice received MD. One hour after MD, mice were injected with SB3CT or vehicle solution and returned to their home cages. Starting from d0 before MD, mice were checked daily in the optomotor setup for the assessment of spatial frequency and contrast sensitivity thresholds of the optomotor reflex for the following 7 d. On d7, optical imaging of intrinsic signals was performed, after which the mice were perfused, and the brains were harvested. After tissue preparation and immunohistochemistry, PT lesions were measured. Representative activity maps of binocular V1 and ODI values of the left hemisphere measured by optical imaging of intrinsic signals in PT-lesioned, one-time vehicle-injected (B, D) or SB3CT-injected (C, E), or sham-treated vehicle-injected (F, H) or SB3CT-injected (G, I), mice are displayed. In PT mice without MD (B, C), activity patches evoked by stimulation of the contralateral eye were darker than those of the ipsilateral eye, the average ODI was positive, and warm colors prevailed in the OD maps, indicating contralateral dominance in the left hemisphere binocular V1. Vehicle-injected PT mice with MD (deprived eye is marked by black filled circles) did not show OD plasticity (D). In contrast, a single injection of the MMP2/9 inhibitor SB3CT after the PT lesion rescued OD plasticity so that MD induced a strong OD shift toward the nondeprived eye (ipsilateral; E). Likewise, in sham-lesioned mice, neither vehicle nor SB3CT injections compromised OD plasticity, nor MD induced an OD shift toward the nondeprived eye (H, I). Data display same as Figure 1. Scale bars: 1 mm. Tables comparing the two independent analyses are provided as the Extended Data Figure 5-1.

Figure 6.

Quantification of OD plasticity on the brief (1-d) inhibition of MMP2/MMP9 with SB3CT after a PT lesion in S1: both ODIs (A) and V1 activation (B) without and with MD in the various treatment groups (vehicle/SB3CT and PT/sham lesion) are illustrated. A, Optically imaged ODIs of mice injected with vehicle (black/gray) or SB3CT (dark green, light green). Symbols represent ODI values of individuals (squares specify PT-lesioned mice, circles sham-lesioned mice). In PT mice with MD, a single systemic administration of SB3CT on d0 rescued the impaired OD shift (impaired by PT in vehicle-treated mice). B, V1 activation elicited by stimulation of the contralateral (C) or ipsilateral (I) eye without and after MD. Multivariate ANOVA, followed up by Sidak-corrected tests for simple effects. Data display as in Figure 2. Mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001.

Figure 7.

Injection of the MMP2/9 inhibitor SB3CT after the induction of a PT lesion in S1 rescued experience-enabled enhancements of both the spatial frequency (A) and contrast sensitivity (B) thresholds of the optomotor reflex in adult mice. A, Spatial frequency threshold values in cyc/° are plotted against days for PT (squares), sham (circles), in mice without (filled symbols) and with MD (half-filled symbols), both for vehicle (black/gray) and SB3CT (dark green, light green) treatment. Seven days of MD induced a significant increase in spatial frequency threshold in SB3CT-injected PT-treated mice each day but not in vehicle-injected PT mice. In contrast, sham-lesioned mice showed significant threshold increases over days after MD (half-filled squares) compared with noMD mice, regardless of the type of injection (vehicle/SB3CT). B, Likewise, SB3CT treatment after PT rescued the experience-induced increase in contrast sensitivity thresholds of the open eye after MD: PT mice receiving a single SB3CT injection (dark green) had higher contrast thresholds, while the contrast thresholds of vehicle-treated PT mice (black) were significantly lower compared with SB3CT-injected PT mice. Sham-lesioned MD mice also had significantly higher contrast thresholds at every spatial frequency compared with noMD mice, regardless of treatment (vehicle and SB3CT). Gratings left of the y-axis illustrate the contrast steps for a given (constant) spatial frequency. Mixed ANOVA followed up by Sidak-corrected test for simple effects. Mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001.