EFFECT OF PEPTIDASES AT THE BLOOD-BRAIN-BARRIER ON THE PERMEABILITY OF ENKEPHALIN

The blood brain barrier (BBB) presents an enzymatic barrier to the pas sage of peptides from blood to brain. The studies presented here used a well established in vitro model of the BBB to measure the presence o f peptidases and the permeability of two opioid peptides. The in vitro BBB model consisted of confluent monolayers of bovine brain microvess el endothelial cells (BMECs). Enkephalin metabolizing enzymes, total a minopeptidase, aminopeptidase M (APM), angiotensin converting enzyme ( ACE) and neutral endopeptidase (NEP) activities were measured in BMEC monolayers. The effect of specific inhibitors of APM, ACE and NEP on t he permeability of [Met(5)]enkephalin (Met-Enk) and a conformationally constrained and enzymatically stable analog, DPDPE, also was determin ed. High levels of membrane-associated enzyme activity were measured f or total aminopeptidase, APM and ACE. Interestingly, the permeability coefficient of Met-Enk was increased 4-fold in the presence of specifi c inhibitors of APM and ACE. Low levels of NEP activity were measured in BMEC monolayers and inhibition of NEP had no effect on Met-Enk perm eability. The permeability coefficient for DPDPE was not increased wit h enzyme inhibitors but was 4-fold greater than Met-Enk alone. In the presence of APM or ACE inhibitors, there was no difference in the perm eability of DPDPE and Met-Enk. These experiments demonstrate the prese nce of specific peptidases in BMECs and that the presence of inhibitor s to Met-Enk inactivating peptidases significantly increased permeabil ity of this biologically active peptide.