Distinct expression and regulation of the glutamate transporter isoforms GLT-1a and GLT-1b in cultured astrocytes from a rat model of amyotrophic lateral sclerosis (hSOD1 G93A ) Ste´ phanie Goursaud, Jean-Marie Maloteaux, Emmanuel Hermans * Laboratoire de Pharmacologie Expe´rimentale, Universite´ catholique de Louvain, 54.10, Av. Hippocrate 54, 1200 Brussels, Belgium 1. Introduction Amyotrophic lateral sclerosis (ALS) is an adult neurodegenera- tive disease characterised by a progressive and massive loss of motor neurons in the spinal cord and brainstem. Together with neurodegeneration in the motor cortex, these lesions lead to muscular atrophy and progressive paralysis. ALS occurs in both sporadic (sALS) and familial forms (fALS), the latter representing 5– 10% of reported cases (Siddique and Deng, 1996). Mutations in the gene encoding for superoxide dismutase 1 (SOD1) constitute the most frequent cause of fALS, and transgenic rodents expressing mutated forms of human SOD1 (hSOD1) have been generated. These animals exhibit typical features of ALS and are commonly used experimental models for the study of this human disease (Gurney et al., 1994; Nagai et al., 2001; Howland et al., 2002). Several theories have been proposed to explain the targeted loss of motor neurons in ALS and there is convincing evidence that excitotoxic processes participate in neurodegeneration. Thus, impaired glutamate handling by astrocytes was demonstrated both in patients with sALS and in transgenic animal models during the development and the progression of the disease (Doble, 1999; Maragakis and Rothstein, 2004). Moreover, riluzole which is the single FDA approved drug for ALS treatment is documented to inhibit glutamate release (Van den Bosch et al., 2006) and to increase the activity of glutamate transporters (Fumagalli et al., 2008). Indeed, astrocytes achieve an efficient control of extra- cellular glutamate levels in the central nervous system via the activity of Na + -dependent high-affinity glutamate transporters, namely the glutamate-aspartate transporter (GLAST) and the glutamate transporter 1 (GLT-1) (Danbolt, 2001). Both transporter subtypes have been shown to contribute to the neuronal protection against glutamate-mediated excitotoxicity. Neverthe- less, ALS is mainly associated with a decreased expression and Neurochemistry International 55 (2009) 28–34 ARTICLE INFO Article history: Received 25 November 2008 Received in revised form 10 February 2009 Accepted 12 February 2009 Available online 24 February 2009 Keywords: Amyotrophic lateral sclerosis hSOD1 G93A rats GLT-1a and GL-1b Peptide Histidine Isoleucine Cortical astrocytes ABSTRACT Impaired glutamate uptake associated with accumulation of extracellular glutamate is a well- documented feature of amyotrophic lateral sclerosis (ALS) and related excitotoxicity is frequently proposed to participate in the progression of the disease. We herein characterised the expression and activity of the glutamate transporter glutamate transporter 1 (GLT-1) in cultured cortical astrocytes derived from a transgenic rat strain expressing an ALS-related mutated form of human superoxide dismutase 1 (hSOD1 G93A ). Measurements of D-[ 3 H]-aspartate uptake velocity in the presence of selective glutamate transporter blockers demonstrated that astrocytes from the transgenic rats showed an impaired GLT-1 activity as compared to cells from wild-type animals. In addition, the density of GLT-1a mRNA in cells from hSOD1 G93A animals appeared nearly 2-fold lower while the density of GLT-1b mRNA was nearly 2-fold higher. Besides, we observed that exposing the astrocytes from hSOD1 G93A rats to the neuroprotective transmitter Peptide Histidine Isoleucine (PHI) for 24 h caused a 4.5-fold increase in the GLT-1b mRNA level without influencing the expression of the other key isoform GLT-1a. This selective upregulation of GLT-1b by the neuropeptide was correlated with a significant increase in D-[ 3 H]- aspartate uptake activity. The possibility to specifically regulate a single isoform of the high-affinity transporter GLT-1 is an unprecedented observation which sheds light on new perspectives for the pharmacological manipulation of glutamate transmission in diseases such as ALS. ß 2009 Elsevier Ltd. All rights reserved. * Corresponding author. Tel.: +32 2 7649339; fax: +32 2 7645460. E-mail address: emmanuel.hermans@uclouvain.be (E. Hermans). Abbreviations: ALS, amyotrophic lateral sclerosis; BSA, bovine serum albumin fatty acid free from fraction V; DHK, dihydrokainic acid; EAAT2, excitatory amino acid transporter 2; fALS and sALS, familial and sporadic forms of ALS; GAPDH, glyceraldehyde phosphate deshydrogenase; GLAST, glutamate-aspartate transpor- ter; GLT-1, glutamate transporter 1; hSOD1, human SOD1; L-SOS, L-serine O-sulfate potassium salt; LTHA, L-(À)-threo-3-hydroxyaspartic acid; PHI, Peptide Histidine Isoleucine; PACAP, pituitary adenylyl cyclase-activating polypeptide; RT, reverse transcription; SOD1, superoxide dismutase 1; VIP, vasoactive intestinal peptide; VPAC 1 and VPAC 2 , polyvalent VIP/PACAP type 1 and 2 receptors. Contents lists available at ScienceDirect Neurochemistry International journal homepage: www.elsevier.com/locate/neuint 0197-0186/$ – see front matter ß 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.neuint.2009.02.003