Toxicity of glucosylsphingosine (glucopsychosine) to cultured neuronal cells: a model system for assessing neuronal damage in Gaucher disease type 2 and 3 U.H. Schueler, a,b,c, * T. Kolter, b C.R. Kaneski, a J.K. Blusztajn, d M. Herkenham, c K. Sandhoff, b and R.O. Brady a a Developmental and Metabolic Neurology Branch, NINDS, NIH, DHHS, Bethesda, MD, USA b Kekule ´-Institut fu ¨r Organische Chemi und Biochemie der Universita ¨t, Bonn, Germany c Section on Functional Neuroanatomy, Laboratory of Cellular and Molecular Regulation, NIMH, NIH, DHHS, Bethesda, MD, USA d Department of Pathology and Laboratory Medicine, Boston University School of Medicine, Boston, MA, USA Received 5 December 2002; revised 1 May 2003; accepted 13 August 2003 Abstract Patients with Gaucher disease have been classified as type 1 nonneuronopathic, type 2 acute neuronopathic, and type 3 chronic neuronopathic phenotypes. Increased quantities of glucocerebroside and glucosylsphingosine (glucopsychosine) are present in the brain of type 2 and type 3 Gaucher patients. Galactosylsphingosine has previously been shown to be neurotoxic in globoid cell leukodystrophy (Krabbe disease). To determine whether glucosylsphingosine is also neurotoxic, we examined its effect on cultured cholinergic neuron-like LA-N-2 cells. When these cells were exposed to 1, 5, or 10 M glucosylsphingosine for a period of 18 h, they became shriveled, neurite outgrowth was suppressed, and the activities of the lysosomal enzymes glucocerebrosidase, sphingomyelinase, and -galactosidase were reduced in a dose-dependent manner. Acetylcholine in cells exposed to glucosylsphingosine also declined. Cells switched to glucosyl- sphingosine-free medium partially recovered. The data suggest that accumulation of glucosylsphingosine contributes to neuronal dysfunc- tion and destruction in patients with neuronopathic Gaucher disease. © 2003 Elsevier Inc. All rights reserved. Keywords: Neuronopathic Gaucher disease; Glucosylsphingosine; Glucopsychosine; Acetylcholine; Cholinergic LA-N-2 cells Introduction Gaucher disease is an autosomal recessive metabolic disorder caused by the accumulation of glucosylceramide (glucocerebroside) due to a deficiency of the enzyme glu- cosylceramide--glucosidase (glucocerebrosidase; EC 3.2.1.45) (Brady et al., 1965). Glucocerebroside arises from the turnover of senescent white and red blood cells and is a metabolic intermediate in both the synthesis and the degra- dation of complex glycosphingolipids such as gangliosides and globoside (Kolter et al., 2002; Kolter and Sandhoff, 1998). Patients with Gaucher disease have been subclassified into three principal clinical phenotypes: non-neuronopathic (type 1), acute neuronopathic (type 2), and chronic neurono- pathic (type 3) forms. Patients with type 1 Gaucher disease are successfully treated by enzyme replacement therapy (Barton et al., 1991). In addition to systemic manifestations, patients with type 2 and 3 Gaucher disease are characterized by neuronopathic effects leading to loss of neurons and the presence of lipid-storing cells in the Virchow-Robin spaces of the brain. In these phenotypes, visceral signs improve with enzyme therapy; but neurological signs inexorably progress in the type 2 patients (Erikson et al., 1993; Prows et al., 1997), and may or may not worsen in type 3 patients * Corresponding author. NIH, NIMH, LCMR, Section on Functional Neuroanatomy, Building 36, Room 2D15, 36 Convent Drive, Bethesda, MD 20892-4070. Fax: +1-301-402-2200. E-mail address: SchueleU@ninds.nih.gov (U.H. Schueler). R Available online at www.sciencedirect.com Neurobiology of Disease 14 (2003) 595– 601 www.elsevier.com/locate/ynbdi 0969-9961/$ – see front matter © 2003 Elsevier Inc. All rights reserved. doi:10.1016/j.nbd.2003.08.016