Extracellular biotransformation of beta-endorphin in rat striatum and cerebrospinal fluid.
Numerous studies have investigated the behavioural effects of beta-endorphin, both endogenous and exogenously applied. However, the potential for biotransformation of beta-endorphin in the extracellular space of the brain has not been previously directly addressed in vivo. Utilising microinfusion/microdialysis and matrix-assisted laser desorption/ionisation mass spectrometry, we investigated beta-endorphin biotransformation in the striatum of rats. We infused 1.0 nmol beta-endorphin into the striatum of adult male Fischer rats and observed rapid cleavage resulting in beta-endorphin 1-18, as well as several fragments resulting from further N-terminal degradation. In vitro studies with incubation of full-length beta-endorphin, with and without protease inhibitors, in the incubation fluid of isolated striatal slices indicate that beta-endorphin is initially cleaved predominantly at the Phe(18)-Lys(19), position, as well as at the Leu(17)-Phe(18) position. Investigations of cerebrospinal fluid revealed similar enzymatic cleavage of beta-endorphin. The observed pattern of cleavage sites (Phe(18)-Lys(19) and Leu(17)-Phe(18)) is consistent with published in vitro studies of purified insulin-degrading enzyme cleavage of beta-endorphin. The binding affinities of full-length beta-endorphin, as well as previously identified beta-endorphin fragments alpha-endorphin (beta-endorphin 1-16) and gamma-endorphin (beta-endorphin 1-17), and the fragment identified in the present study, beta-endorphin 1-18, at heterologously expressed mu, delta and kappa-opioid receptors, respectively, were determined; the affinity of the truncation fragments is reduced at each of the receptors compared to the affinity of full length beta-endorphin.
Reed
B
B
Laboratory of the Biology of Addictive Diseases, The Rockefeller University, New York, NY 10021, USA. reedb@rockefeller.edu
Bidlack
J M
JM
Chait
B T
BT
Kreek
M J
MJ
D000595:Amino Acid Sequence; D000818:Animals; D001479:Basal Ganglia / Q000187:drug effects / Q000378:metabolism*; D001711:Biotransformation; D016466:CHO Cells; D002555:Cerebrospinal Fluid / Q000737:chemistry; D006224:Cricetinae; D003412:Cricetulus; D005110:Extracellular Space / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D013058:Mass Spectrometry; D017551:Microdialysis; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D051381:Rats; D011916:Rats, Inbred F344; D011957:Receptors, Opioid / Q000378:metabolism; D001615:beta-Endorphin / Q000134:cerebrospinal fluid* / Q000493:pharmacokinetics*
D010446:Peptide Fragments; D011957:Receptors, Opioid; D001615:beta-Endorphin