THREONINE-FOR-LEUCINE MUTATION WITHIN DOMAIN M2 OF THE NEURONAL ALPHA(7) NICOTINIC RECEPTOR CONVERTS 5-HYDROXYTRYPTAMINE FROM ANTAGONIST TOAGONIST

A study was made of the effects of 5-hydroxy-tryptamine (5HT) on homom eric neuronal nicotinic receptors (nAcChoR) expressed in Xenopus oocyt es after injection of cDNA encoding the wild-type chicken alpha(7) sub unit. Acetylcholine (AcCho) elicited large currents (I-AcCho) that wer e reduced by 5HT in a reversible and dose-dependent manner, with a hal f-inhibitory concentration (IC50) of 56 mu M and a Hill coefficient (n (H)) of 1.2. The inhibition of I-AcCho by 5HT was noncompetitive and v oltage independent, a behavior incompatible with a channel blockade me chanism, 5HT alone did not elicit membrane currents in oocytes injecte d with the wild-type alpha(7) subunit cDNA, In contrast, 5HT elicited membrane currents (I-5HT) in oocytes injected with cDNA encoding an cv mutant subunit with a threonine-for-leucine-247 substitution (L247T a lpha(7)), I-5HT was inhibited by the potent nicotinic receptor blocker s alpha-bungarotoxin (100 nM) and methyllycaconitine (1 mu M). Further more, the characteristics of I-5HT, including its voltage dependence, were similar to those of I-AcCho. The 5HT dose-I-5HT response gave an apparent dissociation constant EC(50) of 23.5 mu M and a Hill coeffici ent n(H) of 1.7, which were not modified by the presence of AcCho, Sim ilarly, the apparent affinity of L247T alpha(7) for AcCho as well as i ts cooperativity were not influenced by 5HT, indicating a lack of mutu al interactions between 5HT and AcCho. These results show that 5HT is a potent noncompetitive antagonist of neuronal alpha(7) nAcChoR, but i t becomes a noncompetitive agonist following mutation of the highly co nserved leucine residue 247 located in the channel domain M2.