The distribution and heterogeneity o f cholecystokinin (CCK) in different regions of the central nervous systcni (CNS) of 12 adult rats and six cows was determined by means of specific radioimmunoassays. In addition, the biologic activity of immunoreactivc material was evaluatcd in a bioassay that uscd a rabbit gallbladder-strip preparation. High amounts of imtnunochemically detectable CCK-8 (20.2 1.5 pmolig) and CCK-33 (33.7 F 3.3 pmolig) and of biologically active CCK were found in the telenccphalon, although conccntrations decreased progressively from the rostra1 to the caudal portions of the brain. The distribution and biologic activity of CCK was similar in rat and cow brain. Ratios of biologic to immunologic activity ranged from 1.0 to 8.0 and were higher for CCK-8 than for CCK-33. The results indicate that (1) CCK-8 and CCK-33 immunoreactivity are widely distributed throughout the CNS in a unique and differcntial manner: ( 2 ) CCK is biologically active in all the regions of the brain, with a pattern of distribution similar to that found with radioinimunoassay; (3) measurable bioactivity was equal to or greatcr than measurable iinmunoreactivity in all areas of the brain in rat and cow: and (4) CCK i s present in tissues without significant hpccics differences between rat and cow.