Peptide hormones and neurotransmitters play crucial roles in the maint
enance of physiological function at both the cellular and organ level.
Although peptide neuropharmaceuticals have enormous potential in the
treatment of disease states, the blood-brain barrier (BBB) generally p
revents the entry of peptides into the brain either by enzyme degradat
ion or by specific properties of the BBB. Peptides that act at opioid
receptors are currently being designed for analgesia and to reduce the
unwanted side effects associated with morphine, such as addiction and
inhibition of gastric motility. It has been the focus of our group to
produce stabile peptide analogues of Metenkephalin, that lead to anal
gesia without side effects. In this paper we present the methodologies
that have been used to elucidate the transport mechanisms of three pe
ptides across the BBB. Using a primary endothelial cell culture model
of the BBB, in situ perfusion, and kinetic analysis we show that D-Phe
-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP) crosses the BBB via diffusio
n, [D-penicillamine(2,5)]-enkephalin uses a combination of diffusion a
nd a saturable transport mechanism, and biphalin ([Tyr-D-Ala-Gly-Phe-N
H](2)) uses diffusion and the large neutral amino acid carrier. Unders
tanding BBB transport mechanisms for peptides will aid in the rational
design of peptides targeted to the brain.