Sensory deprivation alters aggrecan and perineuronal net expression in the mouse barrel cortex

Abstract

An important role for the neural extracellular matrix in modulating cortical activity-dependent synaptic plasticity has been established by a number of recent studies. However, identification of the critical molecular components of the neural matrix that mediate these processes is far from complete. Of particular interest is the perineuronal net (PN), an extracellular matrix component found surrounding the cell body and proximal neurites of a subset of neurons. Because of the apposition of the PN to synapses and expression of this structure coincident with the close of the critical period, it has been hypothesized that nets could play uniquely important roles in synapse stabilization and maturation. Interestingly, previous work has also shown that expression of PNs is dependent on appropriate sensory stimulation in the visual system. Here, we investigated whether PNs in the mouse barrel cortex are expressed in an activity-dependent manner by manipulating sensory input through whisker trimming. Importantly, this manipulation did not lead to a global loss of PNs but instead led to a specific decrease in PNs, detected with the antibody Cat-315, in layer IV of the barrel cortex. In addition, we identified a key activity-regulated component of PNs is the proteoglycan aggrecan. We also demonstrate that these Cat-315-positive neurons virtually all also express parvalbumin. Together, these data are in support of an important role for aggrecan in the activity-dependent formation of PNs on parvalbumin-expressing cells and suggest a role for expression of these nets in regulating the close of the critical period.

Figure 1.

Monoclonal antibody Cat-315 expression in the mouse somatosensory cortex. Cat-315 marks a subset of cells within the barrel region of the mouse. A, A coronal section shows that Cat-315 is more enriched in layers IV and VI of the somatosensory cortex than in adjacent cortical areas. B, Higher magnification of the boxed region in A demonstrates high expression of Cat-315-reactive nets in the somatosensory cortex (arrow denotes the border). C, A tangential section through layer IV of the barrel cortex showing Cat-315 (green) immunoreactivity is highly expressed within the boundaries of the barrel cortex, defined by fluorescent Nissl (red). Scale bars: A, 1 mm; C, 200 μm.

Figure 2.

Effects of sensory deprivation on Cat-315-positive PNs in the mouse barrel cortex. Tangential sections through layer IV of the barrel cortex of control (A–H) and sensory-deprived (I–P) mice stained with fluorescent Nissl (A, E, I, M) and Cat-315 (B, F, J, N) are shown. C, G, K, and O show the merge of the fluorescent Nissl (red) and Cat-315 (green). D, H, L, and P are higher-magnification images of C, G, K, and O. The right hemisphere (A–D) did not differ from the left hemisphere (E–H) (p = 0.3878). Unilateral whisker trimming from the right whisker pad did not alter barrel morphology (I, M) but did substantially reduce Cat-315 immunoreactivity in the sensory-deprived hemisphere (N–P) compared with the nondeprvied hemisphere (J–L). The level of reduction can also be observed by comparing Cat-315 levels in the deprived barrel cortex with those of control animals (B–D, F–H). Quantification of these data are shown in Table 1. Scale bar, 100 μm.

Figure 3.

Effects of sensory deprivation in adulthood, or sensory deprivation followed by normal activity, on Cat-315-expressing PNs in the mouse barrel cortex. A, B, Tangential sections through layer IV of the nondeprived barrel cortex (A) and sensory-deprived barrel cortex (B) of adult mice after sensory deprivation from P90 to P120 did not alter Cat-315 expression (green) or fluorescent Nissl staining (red). C, D, Sensory deprivation through P30 followed by 30 d of normal activity did not alter barrel morphology, as shown by the fluorescent Nissl staining, but did significantly reduce Cat-315 immunoreactivity in the sensory-deprived hemisphere (D) compared with the nondeprvied hemisphere (C) (see Table 1). Scale bar, 100 μm.

Figure 4.

aggrecan mRNA is expressed by neurons in the mouse barrel cortex and altered by sensory deprivation. A, Tangential section through layer IV of the barrel cortex stained with cresyl violet to outline the barrel structure. B, In situ hybridization, using a ³³P- labeled antisense probe to exon 12, was performed on the same section, and aggrecan mRNA expression is both enriched and localized within the boundaries of the barrel cortex. C, aggrecan mRNA is more enriched in the barrel cortex than neighboring cortical areas. There is a laminar distribution with aggrecan expression more prominent in layers IV and VI of the barrel cortex. D, There is a decrease in aggrecan mRNA in the sensory-deprived barrel cortex relative to the nondeprived barrel cortex of the same animal after 30 d of sensory deprivation during development (the arrow points out the sensory-deprived barrel cortex). The aggrecan expression levels are normal in the nondeprived hemisphere of the trimmed animal. The sensory-deprived barrel cortex shows a decrease in aggrecan message in layer IV, whereas layer VI expression levels appear normal.

Figure 5.

Effects of sensory deprivation on PNs detected with WFA in the mouse barrel cortex. A–C, G–I, In coronal sections, WFA (green) is immunoreactive in layers IV and VI of the mouse somatosensory cortex. A, B, G, H, Trimming the whiskers does not alter WFA expression in either the sensory-deprived barrel cortex (A, G) or the nondeprived barrel cortex (B, H). D, E, G, H, Trimming leads to reduced expression of Cat-315 (red) in the deprived barrel cortex (D, G) but not in the nondeprived barrel cortex (E, H). C, F, I, High-magnification images from the boxed regions show that Cat-315 and WFA have partial levels of overlap but are identifying distinct subsets of cells. Scale bar, 100 μm.

Figure 6.

Effects of sensory deprivation on Cat-315-immunoreactive PNs surrounding parvalbumin-expressing interneurons in the mouse barrel cortex. In tangential sections, parvalbumin (red) and Cat-315 (green) were used to stain layer IV of both the nondeprived barrel cortex (A, C, E, G) and the sensory-deprived barrel cortex (B, D, F, H) of trimmed animals. A, B, There is no change in the level of parvalbumin reactivity in the nondeprived barrel cortex (A) relative to the deprived barrel cortex (B). C, D, Sensory deprivation does not alter Cat-315 expression in the nondeprived barrel cortex (C) but does lead to a reduction in Cat-315 expression in the deprived barrel cortex (D). C, E, G, In the nondeprived barrel cortex, Cat-315 forms PNs in the extracellular space around parvalbumin-immunoreactive cells. D, F, H, In the sensory-deprived hemisphere, there are parvalbumin-positive interneurons not ensheathed by Cat-315. Scale bar, 100 μm.

Figure 7.

WFA-positive PNs are rarely colocalized with parvalbumin-expressing interneurons in layer IV of the barrel cortex. A, Coronal section showing PNs detected with WFA in layer IV of the barrel cortex. B, Parvalbumin-immunoreactive cells found in layer IV of the barrel cortex. C, A merged image shows that the majority of WFA-positive cells do not express the calcium-binding protein parvalbumin. Scale bar, 100 μm.