Different studies point to the implication of the endocannabinoid system in multiple sclerosis (MS) and animal models of MS. The purpose of this study was to evaluate a possible association of MS with polymorphic markers at the CNR1 gene, encoding the cannabinoid 1 (CB(1)) receptor. We have performed a genetic analysis of an AAT repeat microsatellite localized in the downstream region of the CNR1 gene, in two case-control groups of MS patients and healthy controls (HC) from Spain (Madrid and Bilbao). MS patients with primary progressive MS (PPMS) had more commonly long ((AAT) > or = (13)) alleles and genotypes with a significant difference for genotype 7/8 in Madrid (p = 0.043) and in the sum of both groups (p = 0.016); short alleles were less frequently found in PPMS with a significant difference for allele 5 in the analysis of both groups together (p = 0.039). In patients with relapsing MS, no consistent differences in allele and genotype distribution were found. Disease severity and progression was unrelated to AAT repeat variations. In conclusion, long (AAT) > or = (13) CNR1 genotypes could behave as risk factors for PPMS.