HEME FROM ALZHEIMERS BRAIN INHIBITS MUSCARINIC RECEPTOR-BINDING VIA THIYL RADICAL GENERATION

An endogenous inhibitor (< 3500 Da) of antagonist binding to the musca rinic acetylcholine receptor (mAChR) has been reported to be elevated 3-fold in Alzheimer's disease (AD) brain, This endogenous inhibitor wa s found to require the presence of reducing agents such as reduced glu tathione (GSH) for optimal activity. In the presence of GSH, the inhib itor was shown to generate thiyl radicals which irreversibly inhibited the mAChR. We now report that the inhibitor contains free heme, a wel l-established source of oxidative stress capable of generating free ra dicals and causing neurotoxicity. While FeSO4, microperoxidase and hem in all inhibited antagonist binding to the mAChR, only hemin shared th e inhibitor's requirement for GSH. Both the free radical scavengers Tr olox and Mn2+, and the metal chelator, EDTA, blocked the activity of t he endogenous AD inhibitor and of hemin. Heme oxygenase-1 (HO-1) marke dly reduced the activity of both the endogenous AD inhibitor and hemin , indicating that the endogenous inhibitor contains heme. Mass spectro metric analysis confirmed the presence of free heme and heme fragments in fractions of the endogenous AD inhibitor. The antioxidants estroge n, vitamin E and vitamin C all protected the mAChR from irreversible i nhibition by the endogenous inhibitor or hemin. These antioxidants may function to protect the integrity of the mAChR in vivo and may have t herapeutic potential in AD where free heme could be a source of oxidat ive stress. (C) 1997 Elsevier Science B.V.