Unidirectional uptake of enkephalins at the blood-tissue interface of the blood-cerebrospinal fluid barrier: a saturable mechanism

The cellular uptake at the blood-tissue interface of the blood-cerebrospinal fluid (CSF) barrier to tyrosyl-3,5-[³H]enkephalin-[5-l-leucine] (abbreviated to Leu-enkephalin) and of its synthetic analogue d-alanine²-tyrosyl-3,5-[³H]enkephalin-[5-d-leucine] (abbreviated to d-Ala²-d-Leu⁵-enkephalin) was studied in the isolated perfused choroid plexuses from the lateral ventricles of the sheep, using the rapid (<30 s), single circulation, paired-tracer dilution technique, in which d-[¹⁴C]-mannitol serves as an extracellular marker. Cellular uptake of peptides was estimated by directly comparing venous dilution profiles of [³H] and [¹⁴C] radioactivities in the absence and presence of unlabelled peptide, the N-terminal amino acid (l-tyrosine), the typical l-transport system substrate, 2-aminobicyclo(2,2,1)heptane-2-carboxylic acid (BCH) and the inhibitor of aminopeptidase activity, bacitracin. The cellular uptake of both enkephalins was strongly (65–76%) but not completely inhibited by the addition of 5 mM unlabelled peptide to the bolus; the self-inhibition was significantly higher for d-Ala²-d-Leu⁵-enkephalin than for Leu-enkephalin. The addition to the bolus of l-tyrosine (5 mM), BCH (10 mM) or bacitracin (2 mM) reduced the ³H-radioactivity uptake by the choroid plexus of both enkephalins by 20–40%, the degree of inhibition being greater for [³H]-Leu-enkephalin than for its analogue.It is concluded that during single passage of enkephalins through the choroid plexus circulation, unidirectional uptake at the blood-tissue interface of the blood-CSF barrier consists of two components; a saturable component, which represents uptake of the intact peptide by the choroid epithelium, and a non-saturable component, which reflects enzymatic degradation of peptide in the blood and/or at the barrier, with a liberation of the N-terminal tyrosyl residue. Higher penetration of the blood-CSF barrier by d-Ala²-d-Leu⁵-enkephalin can be attributed to its greater resistance to hydrolysis.