BRAIN RESEARCH ELSEVIER Brain Research 736 (1996) 329-337 Research report Evidence for receptors for hyaluronan in discrete nerve cell populations of the brain K. Fuxe a,*, L.F. Agnati b, B. Tinner a, N. Forsberg c, P. McCourt c, S. Gustafson c a Department ofNeuroscience, Karolinska Institutet, Stockholm 171 77, Sweden b Department of Human Physiology, University ofModena, Modena, Baly c Department of Medical and Physiological Chemistry, Biomedical Centre, Uppsala Unir'ersity, Uppsala, Sweden Accepted 11 June 1996 Abstract Evidence is presented, based on immunoblotting, immunohistochemistry and double immunolabelling procedures, for the existence of hyaluronan receptor immunoreactivity in discrete nerve cell populations of the rat brain, present within the zona compacta and the zona reticulata of the substantia nigra, the ventral tegmental area, the locus coeruleus, the mesencephalic trigeminal nucleus, the nucleus of the trapezoid body, the motor trigeminal nucleus and the lateral cerebellar nucleus. With preimmune serum control, this hyaluronan receptor immunoreactivity could not be demonstrated. Double immunofluorescence immunocytochemistry, using a well-characterized hyaluronan receptor antiserum, together with the tyrosine hydroxylase antiserum, in the presence or absence of detergent, demonstrated the existence of hyaluronan receptor immunoreactivity in dopamine nerve cells of the substantia nigra and the ventral tegmental area and in noradrenaline nerve cells of the locus coeruleus, previously shown not to stain for hyaluronan. In all the nerve cells, the immunoreactivity had the appearance of punctate bodies mainly located in the cytoplasm of the perikarya of the above nerve cell populations as also shown by confocal laser microscopy in the mesencephalic trigeminal nucleus. Based on these observations, it is concluded that hyaluronan receptors exist in discrete nerve cell populations of the brain, including many noradrenaline and dopamine neurones. In all nerve cells, it is located intracellularly in bodies possibly representing clustered hyaluronan receptors undergoing endocytosis. The results open up the possibility that hyaluronan receptors may reduce high concentrations of hyaluronic acid in the surrounding matrix, thereby facilitating communication between adjacent neurones. Intracytoplasmatic hyaluronic acid may also be of special importance for neuronal plasticity, in view of the ability of hyaluronic acid to activate protein kinase activity and/or by influencing the architecture of the cytoskeleton. Keywords: Hyaluronan receptor; Nerve cell; Brain; Plasticity 1. Introduction Recently, evidence was obtained for a regional distribu- tion of hyaluronan in the rat brain using a highly specific hyaluronan-recognizing protein [8,15,22]. Histochemistry performed without permeabilization, revealed labelling as- sociated with the plasma membrane in highly discrete nerve cell bodies of the frontoparietal cortex, the red Abbreviations: CEC, corneal endothelial cell; FITC, fluorescein iso- thiocyanate; HA, hyaluronan; HARLEC, hyaluronan receptor on liver endothelial cell; ICAM-1, intercellular adhesion molecule-1; IR, immuno- reactivity; LEC, liver endothelial cell; PB, phosphate buffer; PBS, phos- phate-buffered saline; RHAMM, receptor for hyaluronic acid-mediated motility; TH, tyrosine hydroxylase. * Corresponding author. Fax: +46 (8) 33-7941. nucleus, the zona reticulata of the substantia nigra and the oculomotor nucleus. In view of these findings, it became of interest to analyze for the possible presence of a hyaluronan (HA) receptor in the brain and especially in the above-described discrete nerve cell populations. It has recently been demonstrated that the receptor for HA-medi- ated motility plays a role for neurite extension and motility in primary neurones and neuronal cell lines as well as for astroglial and microglial motility [21,24]. Immunohisto- chemistry indicates the presence of this receptor at the periphery of cells, along processes and at growth cones as analyzed in vitro. The antibodies raised against the hyaluronan receptor on liver endothelial cells (HARLEC) and used in this study, have been shown to inhibit HA binding to (liver endothelial cells) LEC and LEC membranes [4]. Previous 0006-8993/96/$15.00 Copyright © 1996 Elsevier Science B.V. All rights reserved. PH S0006-8993(96)00720-2