Major glycosaminoglycan species in the developing retina: synthesis, tissue distribution and effects upon cell death Rafael B. Erlich a , Cla ´udio C. Werneck b , Paulo A.S. Moura ˜o b , Rafael Linden a, * a Instituto de Biofı ´sica da UFRJ, Rio de Janeiro, RJ Brazil b Laborato ´rio de Tecido Conjuntivo, Departamento de Bioquı ´mica Me ´dica, Centro de Cie ˆncias da Sau ´de and Hospital Universita ´rio Clementino Fraga Filho, UFRJ, Rio de Janeiro, RJ, Brazil Received 25 February 2003; accepted in revised form 12 May 2003 Abstract Retinal explants mantained in culture medium retain their histotypic structure and develop similarly to the in vivo condition. Extracellular matrix components, particularly the glycosaminoglycans which are not routinely present in dissociated cell cultures are involved in various cellular events. In this work we characterized and determined the localization of sulfated glycosaminoglycans in the extracellular matrix of rat retinal explants at various stages of normal postnatal development and tested whether disruption of the tissue glycosaminoglycan composition may impose either trophic or toxic effects upon distinct retinal cell populations. Our data show that chondroitin sulfate and heparan sulfate glycosaminoglycan chains are synthesized in different proportions during postnatal retinal development. A peak of synthesis of chondroitin sulfates is evident at around P14. Immunohistochemistry showed chondroitin 6-sulfate in the plexiform layers during the earlier stages while later, intense immunoreactivity was found in the outer retina. Heparan sulfate was found in the neuroblastic layer (NBL) at P1, in both nuclear layers from P5 onwards and in the ganglion cell layer (GCL) at all stages. In contrast to chondroitin 6-sulfate, immunoreactivity to heparan sulfate was absent from the outer retina at both P14 and P21. Treatment with heparitinase modulated the rates of cell death in both the GCL and the NBL in P1 retinal explants. Taken together our data show that among the major sulfated glycosaminoglycans, the developing rat retina synthesizes only heparan sulfate and chondroitin sulfates in a spatiotemporally regulated manner, with a peak of chondroitin sulfates at P14, possibly related to photoreceptor differentiation. In addition, our data suggest a role for heparan sulfate as a modulator of sensitivity to cell death in the retina. q 2003 Elsevier Ltd. All rights reserved. Keywords: retina; glycosaminoglycan; chondroitin sulfate; heparan sulfate; development; cell death; rat 1. Introduction Proteoglycans are macromolecules composed by a core protein covalently bound to a variety of glycosaminoglycan side chains (GAGs), located mostly in the extracellular milieu. Distinct classes of proteoglycans are expressed in the nervous system in a developmentally coordinated way (Oohira et al., 2000; Hartmann and Maurer, 2001). These macromolecules affect the correct wiring of neural connections, proliferation and differentiation of neural cells, besides many others cellular events (Margolis and Margolis, 1989; Iozzo, 1998; Hartmann and Maurer, 2001). The great heterogeneity of this class of macromolecules is in part due to various GAG chains, which are polysacharides composed by repetitive dissacharidic units in a linear distribution attached to the core protein. The capability of proteoglycans to bind different growth factors, growth factor binding proteins, protease inhibitors and other ECM molecules are, at least in part, related to their GAGs chains and explain the roles of this class of macromolecules in various cellular events (Lander and Selleck, 2000; Perrimon and Bernfield, 2000; Tumova et al., 2000). Among these, proteoglycans and GAGs have been 0014-4835/03/$ - see front matter q 2003 Elsevier Ltd. All rights reserved. DOI:10.1016/S0014-4835(03)00129-5 Experimental Eye Research 77 (2003) 157–165 www.elsevier.com/locate/yexer * Corresponding author. Dr Rafael Linden, Instituto de Biofı ´sica da UFRJ, CCS bloco G, Cidade Universita ´ria 21949-900, Rio de Janeiro, Brazil. E-mail addresses: erlich@anato.ufrj.br (R.B. Erlich), rlinden@biof. ufrj.br (R. Linden).