The Brain Chondroitin Sulfate Proteoglycan Brevican Associates with Astrocytes Ensheathing Cerebellar Glomeruli and Inhibits Neurite Outgrowth from Granule Neurons

Abstract

Brevican is a nervous system-specific chondroitin sulfate proteoglycan that belongs to the aggrecan family and is one of the most abundant chondroitin sulfate proteoglycans in adult brain. To gain insights into the role of brevican in brain development, we investigated its spatiotemporal expression, cell surface binding, and effects on neurite outgrowth, using rat cerebellar cortex as a model system. Immunoreactivity of brevican occurs predominantly in the protoplasmic islet in the internal granular layer after the third postnatal week. Immunoelectron microscopy revealed that brevican is localized in close association with the surface of astrocytes that form neuroglial sheaths of cerebellar glomeruli where incoming mossy fibers interact with dendrites and axons from resident neurons. In situ hybridization showed that brevican is synthesized by these astrocytes themselves. In primary cultures of cerebellar astrocytes, brevican is detected on the surface of these cells. Binding assays with exogenously added brevican revealed that primary astrocytes and several immortalized neural cell lines have cell surface binding sites for brevican core protein. These cell surface brevican binding sites recognize the C-terminal portion of the core protein and are independent of cell surface hyaluronan. These results indicate that brevican is synthesized by astrocytes and retained on their surface by an interaction involving its core protein. Purified brevican inhibits neurite outgrowth from cerebellar granule neurons in vitro, an activity that requires chondroitin sulfate chains. We suggest that brevican presented on the surface of neuroglial sheaths may be controlling the infiltration of axons and dendrites into maturing glomeruli.

Degradation of Chondroitin Sulfate Proteoglycans Induces Sprouting of Intact Purkinje Axons in the Cerebellum of the Adult Rat

“…1 A-E), which binds to the GAG chains of different types of CSPGs (Hartig et al, 1992). In line with previous reports (Seeger et al, 1994;Yamada et al, 1997), WFA staining was the most intense in the granular layer, which displayed a latex-like labeling pattern, with patches of different intensity and prominent pericellular nets around Golgi neurons ( Fig. 1 A, B).…”

“…This indicates that the peculiar distribution pattern of the intracortical plexus is maintained, at least in part, by inhibitory signals issued by myelin and/or associated proteins (Buffo et al, 2000;Gianola et al, 2003). Because different types of CSPGs are strongly expressed in the adult cerebellum (Rauch et al, 1991;Yamada et al, 1997), here we asked whether these molecules also participate in this regulation. Thus, we induced degradation of CSPGs by intraparenchymal injection of chondroitinase ABC (ChABC) and examined the morphology and distribution of intracortical Purkinje axons.…”

Brevican isoforms associate with neural membranes

“…The observation that chondroitinase ABC treatment before loading the gradient did not influence the flotation behavior suggests that CS chains are not directly involved in brevican linkage to the cell membrane. Plasmalemmal binding was observed in in vitro assays previously (Yamada et al 1997) and is thought to be mediated via a direct interaction of the C-type lectin domain with sulfated membrane glycolipids, such as sulfatides and HNK-1-reactive sulfoglucuronylglycolipids Miura et al 2001), which in turn could provide a cell-substrate recognition system. A further mechanism of membrane anchoring for extracellular proteins is glypiation.…”

“…As a conditional proteoglycan, brevican occurs both as a CSPG and in a CS-free non-proteoglycan form. The presence of CS chains has been reported to be necessary for the neurite outgrowth-inhibiting action of the molecule (Yamada et al 1997). The N-terminal brevican structure harbors an Ig-like domain and a so-called proteoglycan tandem repeat, which is capable of binding hyaluronic acid (HA).…”

Conditional Sox9 ablation reduces chondroitin sulfate proteoglycan levels and improves motor function following spinal cord injury

“…Chief of the many ECM molecules that serve to inhibit axonal regeneration are the chondroitin sulfate proteoglycans (CSPGs) (Eddleston and Mucke, 1993;Silver and Miller, 2004) that experience a great increase in expression following SCI (Lemons et al, 1999;McKeon et al, 1991). Both in vitro and in vivo studies have shown that axons do not extend into CSPG-rich ECM (Davies et al, 1997(Davies et al, , 1999McKeon et al, 1991;Meiners et al, 1995;Zuo et al, 1998), and specific CSPGs that inhibit neurite outgrowth have been identified including: aggrecan (Condic et al, 1999), neurocan , phosphocan , brevican (Yamada et al, 1997), versican (Schmalfeldt et al, 2000), and NG2 (Dou and Levine, 1994).…”