Abstract This is a preliminary and exploratory study of cranial variation in European populations of grayling. We investigated the correspondence between size/shape variation of the dorsal (dc), ventral (vc) and occipital (oc) cranium and phylogenetic relationships (inferred from mitochondrial control region – mtDNA cr and microsatellite dna data) of six grayling populations: three from Balkan phylogenetic clade and two from Caspian phylogenetic clade of the European grayling Thymallus thymallus and one population of the Adriatic grayling Thymallus aeliani, which until recently was considered the Adriatic phylogenetic clade of T. thymallus. Significant size and shape differences were found between populations in all three cranial views. However, significant size-related shape variation (allometry) was found for dc and vc, but not for oc. The size variation of each cranial view does not contain phylogenetic signal, but size variation of oc is consistent with genetic variation inferred from microsatellite dna. Regarding shape variation, a significant phylogenetic signal was detected only for oc, and only the shape variation of oc is consistent with the genetic variation inferred from the mtDNA cr. Moreover, the Adriatic grayling T. aeliani (Soča population) was clearly separated from the three T. thymallus populations of the Balkan phylogenetic clade and the two T. thymallus populations of the Caspian phylogenetic clade only at the level of oc. Thus, our results suggest that different cranial regions differ in allometry, reflect phylo(genetic) relationships differently, and exhibit differences in ecophenotypic plasticity, with oc seeming best suited to represent the phylogenetic relationships of the grayling populations studied.