Study of endogenous Tyr-Gly-Gly, a putative enkephalin metabolite, in mouse brain: validation of a radioimmunoassay, localisation and effects of peptidase inhibitors.
The tripeptide Tyr-Gly-Gly, a hydrolysis product of enkephalins and related opioid peptides obtained with 'enkephalinase', was identified and quantified in various regions of mouse brain by means of HPLC and a sensitive and specific radioimmunoassay. Similar levels i.e. about 8 pmol/brain were found after the animals were killed by various procedures, including microwave irradiation, suggesting its pre-mortem formation. The distribution of Tyr-Gly-Gly immunoreactivity among brain regions was highly heterogeneous and paralleled to a certain extent the [Met5]enkephalin distribution, molar levels of Tyr-Gly-Gly representing 10-30% of those of the enkephalin. Following gentle homogeneisation of striata in 0.32 M sucrose and centrifugation, 73% of Tyr-Gly-Gly immunoreactivity was recovered in the supernatant, a result consistent with its extracellular localisation in vivo. Administration of enkephalinase inhibitors rapidly elicited marked decrease in Tyr-Gly-Gly immunoreactivity whereas bestatin, an aminopeptidase inhibitor, elicited 100% increase and captopril, an ACE inhibitor, was without significant effect. These data indicate that the tripeptide is in a dynamic state in the brain and that its levels might reflect the release of endogenous enkephalins or related opioid peptides and their subsequent metabolism by enkephalinase.
D000818:Animals; D001921:Brain / Q000276:immunology / Q000378:metabolism* / Q000528:radiation effects; D002216:Captopril / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D004743:Enkephalin, Leucine / Q000276:immunology / Q000378:metabolism; D004744:Enkephalin, Methionine / Q000276:immunology / Q000378:metabolism; D007930:Leucine / Q000031:analogs & derivatives / Q000494:pharmacology; D008297:Male; D051379:Mice; D008872:Microwaves; D009269:Nalorphine / Q000378:metabolism; D015260:Neprilysin; D009842:Oligopeptides / Q000032:analysis / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology*; D011863:Radioimmunoassay; D015244:Thiorphan; D008625:Tiopronin / Q000031:analogs & derivatives / Q000494:pharmacology
D009842:Oligopeptides; D011480:Protease Inhibitors; C044757:tyrosyl-glycyl-glycine; D004744:Enkephalin, Methionine; D004743:Enkephalin, Leucine; D002216:Captopril; D015244:Thiorphan; D008625:Tiopronin; D015260:Neprilysin; D007930:Leucine; C012211:ubenimex; D009269:Nalorphine