Postnatal development of perineuronal nets in wild-type mice and in a mutant deficient in tenascin-R

The extracellular matrix glycoprotein tenascin-R (TN-R), colocalizing with hyaluronan, phosphacan, and aggregating chondroitin sulphate proteoglycans in the white and grey matter, is accumulated in perineuronal nets that surround different types of neurons in many brain regions. To characterize the role of TN-R in the formation of perineuronal nets, we studied their postnatal development in wild-type mice and in a TN-R knock-out mutant by using the lectin Wisteria floribunda agglutinin and an antibody to nonspecified chondroitin sulphate proteoglycans as established cytochemical markers. We detected the matrix components TN-R, hyaluronan, phosphacan, neurocan, and brevican in the perineuronal nets of cortical and subcortical regions. In wild-type mice, lectin-stained, immature perineuronal nets were first seen on postnatal day 4 in the brainstem and on day 14 in the cerebral cortex. The staining intensity of these nets for TN-R, hyaluronan, phosphacan, neurocan, and brevican was extremely weak or not distinguishable from that of the surrounding neuropil. However, all markers showed an increase in staining intensity of perineuronal nets reaching maximal levels between postnatal days 21 and 40. In TN-R-deficient animals, the perineuronal nets tended to show a granular component within their lattice-like structure at early stages of development. Additionally, the staining intensity in perineuronal nets was reduced for brevican, extremely low for hyaluronan and neurocan, and virtually no immunoreactivity was detectable for phosphacan. The granular configuration of perineuronal nets became more predominant with advancing age of the mutant animals, indicating the continued abnormal aggregation of chondroitin sulphate proteoglycans complexed with hyaluronan. As shown by electron microscopy in the cerebral cortex, the disruption of perineuronal nets was not accompanied by apparent changes in the synaptic structure on net-bearing neurons. The regional distribution patterns and the temporal course of development of perineuronal nets were not obviously changed in the mutant. We conclude that the lack of TN-R initially and continuously disturbs the molecular scaffolding of extracellular matrix components in perineuronal nets. This may interfere with the development of the specific micromilieu of the ensheathed neurons and adjacent glial cells and may also permanently change their functional properties.