Enhanced Inhibition of a Mutant Neuronal Nicotinic Acetylcholine Receptor by Agonists: Protection of Function by (E)-N-Methyl-4-(3-pyridinyl)-3-butene-1-amine (TC-2403)

Abstract

Inhibition of neuronal nicotinic receptors can be regulated by sequence in the ␤ subunit second transmembrane domain (TM2). The incorporation of a ␤4(6ЈF10ЈT) subunit, which contains sequence from the muscle ␤ subunit at the TM2 6Ј and 10Ј positions of the neuronal ␤4 subunit, increases the loss of receptor responsiveness after the application of acetylcholine (ACh), nicotine, or 3-(2,4-dimethoxybenzylidene)-anabaseine (DMXB), an ␣7-selective partial agonist. Inhibition of receptor responsiveness following agonist exposure may occur through either an enhancement of desensitization, increased channel block by an agonist, or alternatively via allosteric modulation. Although DMXB produces very little activation of either ␣3␤4 or ␣3␤4(6ЈF10ЈT) receptors, DMXB shows an enhanced use-and voltage-dependent inhibition of ␣3␤4(6ЈF10ЈT) receptors compared with wild-type. In contrast, the ␣4␤2-selective agonist (E)-N-methyl-4-(3-pyridinyl)-3-butene-1-amine (TC-2403, previously identified as RJR-2403) shows increased activation of ␣3␤4(6ЈF10ЈT) receptors compared with ␣3␤4 receptors (as related to ACh activation) but with no significant increase in antagonist activity. The interaction between the binding of local anesthetics and the functional inhibition produced by these agonists was evaluated. The binding of the local anesthetics to their inhibitory sites does not affect inhibitory effects of DMXB and nicotine. However, TC-2403 can protect receptor function from the inhibitory effects of other agonists, suggesting that TC-2403, as well as agonists that cause inhibition, may be binding to an allosteric site, either promoting or inhibiting channel opening. The ability of TC-2403 to protect receptor function from agonist-induced inhibition may point toward valuable new combination drug therapies.Our current understanding of the function of the ligandgated ion channels that mediate synaptic function is founded on early studies of the nicotinic receptor of the neuromuscular junction. However, the analysis of nicotinic acetylcholine receptor function is complicated by the fact that, in addition to activating receptors, exposure to agonists ultimately decreases receptor responsiveness. The classical concept of desensitization, originally put forth by Katz and Thesleff (1957), is that it represents conversion of the receptor to an alternative nonfunctional conformational state and that this conversion is promoted by the binding of agonist to the same sites that produce channel activation.Although it is clear that after an episode of strong activation, receptor responsiveness will decline in the continued presence of an agonist, producing a decrease in function that can persist after the agonist is removed, it is difficult to distinguish classical (i.e., Katz and Thesleff) desensitization from other processes, such as channel block by agonist or allosteric modulation, which might also lead to a decrease in receptor responsiveness. Moreover, the degree to which receptor responsiveness is decreased subsequent to agonist-evoke...

Interaction of benzylidene‐anabaseine analogues with agonist and allosteric sites on muscle nicotinic acetylcholine receptors

“…These results rule out the possibility that the BA locus overlaps the tetracaine‐binding site. Other experimental data (Papke, 2002) indicating that tetracaine does not protect the receptor from the inhibition produced by DMXBA is in accord with our results. In addition, both BAs partially inhibit [ 14 C]amobarbital and inhibit [ 3 H]TCP binding to nAChRs arrested in the resting state (in the presence of α‐BTx) in an allosteric manner.…”

“…Consequently, our data are in accord with a model where the desensitized Torpedo ion channel bears overlapping binding sites for BAs, PCP and dizocilpine. The fact that different mutations in the ion channel increase the self‐inhibitory effect of DMXBA on nAChRs (Papke, 2002), supports the idea that certain amino acids in the ion channel are possible sites for the antagonist action of DMXBA.…”

The Structural Basis for GTS-21 Selectivity between Human and Rat Nicotinic α7 Receptors

“…In particular, differences in equilibrium desensitization may manifest as apparent differences in I max . Another factor influencing the concentration response relationships of ␣7 receptors to various agonists is the phenomenon of residual inhibition, which may represent a particularly long-lived form of desensitization or noncompetitive inhibition (Papke, 2002;Uteshev et al, 2002). However, although some mutations in ␣7 that affect desensitization rates have been described (Revah et al, 1991), all of the mutants and chimeras in this study showed similar concentration-dependent rapid desensitization and similar sensitivity to the residual inhibition produced by GTS-21.…”

Heterogeneity of nicotinic acetylcholine receptor expression in the caudal nucleus of the solitary tract

“…In contrast, 0.2 mM RJR-2403 applied via a picospritzer ( Figure 3D, bottom trace) or iontophoretically ( Figure 5B, top trace) failed to elicit mEPSC facilitation. RJR-2403 is a relatively potent and effective agonist of a number of β2* and β4* nAChRs (Papke et al, 2000): α4β2 (EC 50 =16 μM), α3β2 (EC 50 =150 μM) and α4β4 (EC 50 =50 μM), but not α3β4 (EC 50 =347 μM) or α3β2α5 (EC 50 =360 μM) (Papke, 2002;Papke et al, 2000). Moreover, RJR-2403 has a significantly lower efficacy for α3β4 than for other non-α7 nAChRs (Papke et al, 2000).…”