The transport and metabolism of ceftriaxone was studied in vitro in the isolated choroid plexus and in vivo in New Zealand White rabbits. In vitro, [14C]ceftriaxone was accumulated by a saturable, probenecid-sensitive system in choroid plexus, although much less readily than [14C]penicillin G. Ceftriaxone was also a much less potent inhibitor of [14C]penicillin G accumulation by the isolated choroid plexus than penicillin G itself (IC50 = 1.6 vs. 0.07 mM, respectively). In vivo, 2 hr after intraventricular injection, [14C]ceftriaxone was not metabolized or cleared from the cerebrospinal fluid more rapidly than [3H]mannitol, a molecule transported in the central nervous system by simple diffusion. These in vitro and in vivo results show that ceftriaxone, unlike penicillin G, has minimal affinity for the choroid plexus active transport system that transfers most penicillins and cephalosporins from cerebrospinal fluid to blood.