We studied molecular variability to clarify intraspecific differentiation and phylogenetic relationships in three pine vole species (genus Microtus, subgenus Terricola): Microtus subterraneus, Microtus daghestanicus, and Microtus majori. Multilocus analysis was performed using the entire mitochondrial cytb gene and fragments of nuclear BRCA1, IRBP, and XIST genes. Results confirmed separation of the species, especially M. majori compared with M. daghestanicus and M. subterraneus. These species showed different molecular polymorphism in the genetic markers. We identified two close forms of M. majori, differing in cytb gene and the nuclear gene XIST; one form inhabits the northern slopes of the Greater Caucasus, another the Transcaucasia. Separation of M. daghestanicus populations from North Ossetia and the others was clear. Microtus subterraneus populations from southern Europe and Asia Minor were characterized by maximal genetic heterogeneity; the specimen from Samsun (northern Asia Minor) appeared to be most distant from the others. Despite polymorphism in the chromosome number in M. subterraneus populations from the East European Plain, they possess a depleted gene pool. Results indicated that M. subterraneus colonized the East European Plain in the Holocene, and chromosome variability originated in this part of the species' range as a result of chromosomal fission and quick fixation of the arrangement in northern populations. We argue that differences in the genetic differentiation patterns of Terricola species are mainly due to their ecological peculiarities.