The binding and internalization of a novel adrenocorticotropic hormone (ACTH) analog having a potent neuromodulating effect, ebiratide (H-Met(O2)-Glu-His-Phe-D-Lys-Phe-NH(CH2)8NH2), by isolated bovine brain capillaries, were examined. Metabolism of [5-125I-His]ebiratide occurred during a 30-min incubation with bovine brain capillaries at 37°C. In the presence of 20 mM EDTA, added to inhibit this metabolism, the medium, after 30 min of incubation, contained 82.3 ± 0.5% of the unchanged ebiratide. The total binding and acid-resistant binding of [125I]ebiratide increased with time and reached an equilibrium at about 15 min. The total binding and acid-resistant binding at 30 min (as the cell/medium ratios corrected with [14C]sucrose) were 13.07 ± 0.86 and 5.00 ± 0.18 μl/mg of protein, respectively. The acid-resistant binding showed significant dependence on temperature and medium osmolarity. The [125I]ebiratide binding was significantly inhibited by dansylcadaverine, an endocytosis inhibitor. The saturable acid-resistant binding was obtained by the addition of unlabeled ebiratide (100 nM-5 mM), and the maximal internalization capacity (B(max)) at 30 min was 144.2 pmol/mg of protein, with the half-saturation constant (K(D)) of 62.1 μM. The nonsaturable acid-resistant binding [cell/medium ratio in the presence of the unlabeled compound (1 mM or more)] was 2.2 μl/mg of protein. The acid-resistant binding was significantly inhibited by human ACTH, poly-L-lysine, protamine and E-2078, a basic peptide, but was not inhibited by poly-L-glutamate, insulin or transferrin. These results demonstrate that ebiratide is transported through the blood-brain barrier via a basic peptide-specific absorptive-mediated endocytosis.
|ジャーナル||Journal of Pharmacology and Experimental Therapeutics|
|出版ステータス||Published - 1991 1 1|
ASJC Scopus subject areas