The clade was conceived largely on the basis of the Hyperoartia (lampreys and kin) being more closely related to the Gnathostomata (jawed vertebrates) than the Myxini (hagfishes).

Molecular-genetic analysis of craniates reveals that, compared to less complex animals, they developed duplicate sets of many gene families that are involved in cell signaling, transcription, and morphogenesis (see homeobox).

Craniates have cardiovascular systems that include a heart with at least two chambers, red blood cells, oxygen transporting hemoglobin as well as myoglobin, livers and kidneys.

[10] Dumeril (1806)[11] grouped hagfishes and lampreys in the taxon Cyclostomi, characterized by horny teeth borne on a tongue-like apparatus, a large notochord as adults, and pouch-shaped gills (Marspibranchii).

Løvtrup (1977)[11] argued that lampreys are more closely related to gnathostomes based on a number of uniquely derived characters, including: In other words, the cyclostome characteristics (e.g. horny teeth on a "tongue", gill pouches) are either instances of convergent evolution for feeding and gill ventilation in animals with an eel-like body shape, or represent primitive craniate characteristics subsequently lost or modified in gnathostomes.

The validity of the taxon "Craniata" was recently examined by Delarbre et al. (2002) using mtDNA sequence data, concluding that Myxini is more closely related to Hyperoartia than to Gnathostomata - i.e., that modern jawless fishes form a clade called Cyclostomata.

In 2010, Philippe Janvier stated: Although I was among the early supporters of vertebrate paraphyly, I am impressed by the evidence provided by Heimberg et al. and prepared to admit that cyclostomes are, in fact, monophyletic.