Human Α3β4 Nicotinic Acetylcholine Research Articles

Alkaloids Purified from Aristotelia chilensis Inhibit the Human α3β4 Nicotinic Acetylcholine Receptor with Higher Potencies Compared with the Human α4β2 and α7 Subtypes.

The alkaloids aristoteline (1), aristoquinoline (2), and aristone (3) were purified from the leaves of the Maqui tree Aristotelia chilensis and chemically characterized by NMR spectroscopy. The pharmacological activity of these natural compounds was evaluated on human (h) α3β4, α4β2, and α7 nicotinic acetylcholine receptors (AChRs) by Ca2+ influx measurements. The results suggest that these alkaloids do not have agonistic, but inhibitory, activity on each receptor subtype. The obtained IC50 values indicate the following receptor selectivity: hα3β4 > hα4β2 ≫ hα7. In the particular case of hα3β4 AChRs, 1 (0.40 ± 0.20 μM) and 2 (0.96 ± 0.38 μM) show higher potencies compared with 3 (167 ± 3 μM). Molecular docking and structure-activity relationship results indicate that ligand lipophilicity is important for the interaction with the luminal site located close to the cytoplasmic side of the hα3β4 ion channel between positions -2' and -4'. Compound 1 could be used as a molecular scaffold for the development of more potent noncompetitive inhibitors with higher selectivity for the hα3β4 AChR that could serve for novel addiction and depression therapies.

Bupropion inhibits human α3β4 nicotinic acetylcholine receptors by interacting with luminal and non-luminal sites

The interaction of (±)-bupropion [(±)-BP] with the human (h) α3β4 nicotinic acetylcholine receptor (AChR) is determined by functional and structural approaches. The Ca 2+ influx results indicate that (±)-BP inhibits hα3β4 AChRs with ~2-fold lower potency than that for (±)-2-( N - tert -butylamino)-3’-iodo-4’-azidopropiophenone [(±)-SADU-3-72], indicating that this photoreactive analog can be used to further characterize the (±)-BP binding sites. The competition binding results show that (±)-BP binds to the [ 3 H]imipramine sites at desensitized hα3β4 AChRs with ~4-fold higher affinity compared to the resting state. Molecular docking results indicate that both enantiomers of BP and SADU-3-72, in the protonated state, interact with luminal and non-luminal sites. BP interacts with a luminal site which overlaps that for imipramine, and with non-luminal sites located in the transmembrane domain (TMD) at interfacial (+α3/-β4) and α3 intrasubunit sites, and in the TMD-ECD (extracellular domain) junction at the +β4/-α3 and +β4/-β4 interfaces. Our results are consistent with a hα3β4 model where BP and SADU-3-72 bind to overlapping non-luminal sites as well as non-overlapping luminal sites, probably in physiological conditions.

Coronaridine congeners inhibit human α3β4 nicotinic acetylcholine receptors by interacting with luminal and non-luminal sites

To characterize the interaction of coronaridine congeners with human (h) α3β4 nicotinic acetylcholine receptors (AChRs), structural and functional approaches were used. The Ca2+ influx results established that coronaridine congeners noncompetitively inhibit hα3β4 AChRs with the following potency (IC50's in μM) sequence: (−)-ibogamine (0.62±0.23)∼(+)-catharanthine (0.68±0.10)>(−)-ibogaine (0.95±0.10)>(±)-18-methoxycoronaridine [(±)-18-MC] (1.47±0.21)>(−)-voacangine (2.28±0.33)>(±)-18-methylaminocoronaridine (2.62±0.57μM)∼(±)-18-hydroxycoronaridine (2.81±0.54)>(−)-noribogaine (6.82±0.78). A good linear correlation (r2=0.771) between the calculated IC50 values and their polar surface area was found, suggesting that this is an important structural feature for its activity. The radioligand competition results indicate that (±)-18-MC and (−)-ibogaine partially inhibit [3H]imipramine binding by an allosteric mechanism. Molecular docking, molecular dynamics, and in silico mutation results suggest that protonated (−)-18-MC binds to luminal [i.e., β4-Phe255 (phenylalanine/valine ring; position 13′), and α3-Leu250 and β4-Leu251 (leucine ring; position 9′)], non-luminal, and intersubunit sites. The pharmacophore model suggests that nitrogens from the ibogamine core as well as methylamino, hydroxyl, and methoxyl moieties at position 18 form hydrogen bonds. Collectively our data indicate that coronaridine congeners inhibit hα3β4 AChRs by blocking the ion channel's lumen and probably by additional negative allosteric mechanisms by interacting with a series of non-luminal sites.

Interaction of ibogaine with human α3β4-nicotinic acetylcholine receptors in different conformational states

The interaction of ibogaine and phencyclidine (PCP) with human (h) α3β4-nicotinic acetylcholine receptors (AChRs) in different conformational states was determined by functional and structural approaches including, radioligand binding assays, Ca 2+ influx detections, and thermodynamic and kinetics measurements. The results established that (a) ibogaine inhibits (±)-epibatidine-induced Ca 2+ influx in hα3β4 AChRs with ∼9-fold higher potency than that for PCP, (b) [ 3H]ibogaine binds to a single site in the hα3β4 AChR ion channel with relatively high affinity ( K d = 0.46 ± 0.06 μM), and ibogaine inhibits [ 3H]ibogaine binding to the desensitized hα3β4 AChR with slightly higher affinity compared to the resting AChR. This is explained by a slower dissociation rate from the desensitized ion channel compared to the resting ion channel, and (c) PCP inhibits [ 3H]ibogaine binding to the hα3β4 AChR, suggesting overlapping sites. The experimental results correlate with the docking simulations suggesting that ibogaine and PCP interact with a binding domain located between the serine (position 6′) and valine/phenylalanine (position 13′) rings. This interaction is mediated mainly by van der Waals contacts, which is in agreement with the observed enthalpic contribution determined by non-linear chromatography. However, the calculated entropic contribution also indicates local conformational changes. Collectively our data suggest that ibogaine and PCP bind to overlapping sites located between the serine and valine/phenylalanine rings, to finally block the AChR ion channel, and in the case of ibogaine, to probably maintain the AChR in the desensitized state for longer time.

Different interaction between tricyclic antidepressants and mecamylamine with the human α3β4 nicotinic acetylcholine receptor ion channel

The interaction of tricyclic antidepressants (TCAs) with the human (h)alpha3beta4 nicotinic acetylcholine receptor (AChR) in different conformational states was compared with that for mecamylamine by using functional and structural approaches including, Ca(2+) influx, radioligand binding, and molecular docking. The results established that: (a) [(3)H]imipramine binds to a single site with relatively high affinity (K(d) = 0.41 +/- 0.04 microM), (b) imipramine inhibits [(3)H]imipramine binding to the resting/kappa-bungarotoxin-bound AChR (K(i) = 0.68 +/- 0.08 microM) with practically the same affinity as to the desensitized/epibatidine-bound AChR (K(i) = 0.83 +/- 0.08 microM), suggesting that TCAs do not discriminate between these conformational states, and (c) although TCAs (IC(50) approximately 1.8-2.7 microM) and mecamylamine (IC(50) = 3.3 +/- 0.4 microM) inhibit (+/-)-epibatidine-induced Ca(2+) influx with potencies in the same concentration range, TCAs (K(i) approximately 1-3.6 microM), but not mecamylamine (apparent IC(50) approximately 0.2 mM), inhibit [(3)H]imipramine binding to halpha3beta4 AChRs in different conformational states. This is explained by our docking results where imipramine, in the neutral and protonated states, interacts with the leucine (position 9') and valine/phenylalanine (position 13') rings, whereas protonated mecamylamine (>99% at physiological pH) interacts with the outer ring (position 20'). Our data indicate that TCAs bind to overlapping sites located between the serine and valine/phenylalanine rings in the halpha3beta4 AChR ion channel, whereas protonated mecamylamine can be attracted to the channel mouth before blocking ion flux by interacting with a luminal site in its neutral state.

Interaction of Novel Ibogaine Analogs With The Human α3β4 Nicotinic Receptor

This work is an attempt to characterize the binding site and the inhibitory activity of ibogaine analogs on the human α3β4 nicotinic acetylcholine receptor (hα3β4). In this regard, we used [3H]ibogaine equilibrium binding and Scatchard-plots, [3H]ibogaine and [3H]epibatidine competition binding, and ibogaine-induced inhibition of Ca2+ influx approaches. The results indicate that: (1) there is one high-affinity binding site for [3H]ibogaine, (2) ibogaine inhibits the hα3β4 with higher potency than that for the α1β2γδ AChR, (3) ibogaine interacts with different conformations of the hα3β4 with the indicated affinity (or potency) sequence: Desensitized > Resting > Open, (4) [3H]ibogaine competition experiments indicate that ibogaine and 18-MAC, among ibogaine analogs, and imipramine and dextromethorphan, among other known noncompetitive antagonists, have the highest affinities for the hα3β4 ion channel, and (5) [3H]epibatidine competition experiments indicate that ibogaine analogs interact with the agonist sites with very low affinities. Interaction of 18-MAC with the hα3β4 ion channel could be important for its anti-addictive property.