Subunit-specific determinants of function and pharmacology of nicotinic acetylcholine receptors.

摘要

Some noncompetitive inhibitors (e.g., ganglionic blockers) exhibit selectivity for the inhibition of neuronal nicotinic acetylcholine receptors (nAChRs). The main goal of the present study is to characterize the mechanism of selective long-term inhibition of neuronal and muscle-neuronal chimeric nAChRs by bis-TMP-10 (bis (2,2,6,6-tetramethyl-4-piperidinyl) sebacate or BTMPS), a bifunctional form of the potent ganglionic blocker tetramethylpiperidine. Long-term inhibition of neuronal nAChRs by bis-TMP-10 has been previously demonstrated to arise, at least in part, from the binding of the bis-compound to neuronal beta subunits. In this study, long-term inhibition is demonstrated to be dependent upon the presence of sequence element(s) within the pore-lining second transmembrane domain (TM2) of either neuronal beta subunits or muscle delta subunits; however, for either class of subunit, because the onset of inhibition does not appear to be strongly voltage-dependent, the inhibitor binding site itself does not appear to be contained within the segment of the channel pore influenced by the membrane electric field. In the case of the neuronal beta subunits, long-term inhibition is also not affected by preapplication of the open-channel blocker QX-314. Furthermore, we demonstrate a compound length requirement for long-term inhibition which would be consistent with binding to multiple sites contributed by separate subunits (either beta or delta) located on the extracellular portion of the receptor. Our results may imply that bis-TMP-10 interacts with an activation-sensitive element, the availability of which may be regulated by sequence in the TM2 domain. It is interesting to note that the mechanism of inhibition by bis-TMP-10 appears to be distinct from that of other ganglionic blockers such as mecamylamine. Knowledge of the mechanism underlying the basis for inhibition by pure antagonists may be useful for consideration of observations of mixed agonist/antagonist properties of certain experimental therapeutics for nicotinic receptors. Furthermore, if bis-TMP-10 is binding to an activation-sensitive element distinct from TM2, it may be possible to use sensitivity to inhibition by this compound to investigate the structural changes which take place with channel gating.