Amyotrophic lateral sclerosis (ALS) is typically characterized by a dramatic loss of lower motor neurons in spinal cord and brainstem. Nevertheless, upper motor neuron dysfunction is also reported in ALS and frequently related to a deficit of transcallosal connections associated to a reduced volume of the corpus callosum. Besides, impairment of the astroglial glutamate transporter GLT-1 associated with accumulation of extracellular glutamate is demonstrated in ALS and related excitotoxicity is likely to participate in the progression of the disease. At the molecular level, the caspase-3-mediated cleavage of GLT-1 leading to a selective and functional inhibition of the transporter was evidenced in spinal cord samples from a transgenic mice model of ALS. We herein characterised the expression and activity of GLT-1 and caspase-3 in cultured callosal astrocytes isolated from a transgenic rat strain expressing an ALS-related mutated form of human superoxide dismutase 1 (hSOD1^G93A). Quantitative RT-PCR and Western-blotting studies revealed that the expression of GLT-1 was higher in the cells prepared from the transgenic animals in comparison to the wild-type rats. However, specific measurements of D-[³H]-aspartate uptake velocity failed to evidence differences in the activity of this transporter. Measures of uptake were also performed in the presence of a selective caspase-3 inhibitor (Ac-Asp-Met-Gln-Asp-aldehyde) or in cells exposed to the Peptide Histidine Isoleucine (PHI) which belongs to VIP/PACAP neuropeptide family and which decreases caspase-3 activity. Reducing the activity of this apoptotic enzyme, which is highly detected in callosal astrocytes from hSOD1^G93Arats, was found to upregulate the GLT-1 activity, exclusively in cells from transgenic animals. Together, these findings reinforce the hypothesis of an involvement of caspase-3-mediated impairment of glutamate uptake in the pathogenesis of ALS.