Selectivity of coronaridine congeners at nicotinic acetylcholine receptors and inhibitory activity on mouse medial habenula

Abstract

The inhibitory activity of coronaridine congeners on human (h) α4β2 and α7 nicotinic acetylcholine receptors (AChRs) is determined by Ca2+ influx assays, whereas their effects on neurons in the ventral inferior (VI) aspect of the mouse medial habenula (MHb) are determined by patch-clamp recordings. The Ca2+ influx results clearly establish that coronaridine congeners inhibit hα3β4 AChRs with higher selectivity compared to hα4β2 and hα7 subtypes, and with the following potency sequence, for hα4β2: (±)-18-methoxycoronaridine [(±)-18-MC] > (+)-catharanthine > (±)-18-methylaminocoronaridine [(±)-18-MAC] ∼ (±)-18-hydroxycoronaridine [(±)-18-HC]; and for hα7: (+)-catharanthine > (±)-18-MC > (±)-18-HC > (±)-18-MAC. Interestingly, the inhibitory potency of (+)-catharanthine (27 ± 4 μM) and (±)-18-MC (28 ± 6 μM) on MHb (VI) neurons was lower than that observed on hα3β4 AChRs, suggesting that these compounds inhibit a variety of endogenous α3β4* AChRs. In addition, the interaction of bupropion with (−)-ibogaine sites on hα3β4 AChRs is tested by [3H]ibogaine competition binding experiments. The results indicate that bupropion binds to ibogaine sites at desensitized hα3β4 AChRs with 2-fold higher affinity than at resting receptors, suggesting that these compounds share the same binding sites. In conclusion, coronaridine congeners inhibit hα3β4 AChRs with higher selectivity compared to other AChRs, by interacting with the bupropion (luminal) site. Coronaridine congeners also inhibit α3β4*AChRs expressed in MHb (VI) neurons, supporting the notion that these receptors are important endogenous targets for their anti-addictive activities.

Selectivity of (±)-citalopram at nicotinic acetylcholine receptors and different inhibitory mechanisms between habenular α3β4* and α9α10 subtypes

“…The activation potency of (±)-epibatidine on each human AChR was first determined by assessing the fluorescence change in the respective AChR-expressing cells after (±)epibatidine stimulation. The respective EC 50 values for (±)-epibatidine are in the same range as previous determinations (Arias et al, 2018a(Arias et al, , 2018b(Arias et al, , 2017(Arias et al, , 2016(Arias et al, , 2010a(Arias et al, , 2010b(Arias et al, , 2010c).…”

“…Ca 2+ influx measurements were performed on HEK293-hα3β4, HEK293-hα4β2, and GH3-hα7 cells as previously described (Arias et al, 2018a(Arias et al, , 2018b(Arias et al, , 2017(Arias et al, , 2016(Arias et al, , 2010a(Arias et al, , 2010b(Arias et al, , 2010c. Briefly, 5 x 10 4 cells per well were seeded 72 h prior to the experiment on black 96well plates (Costar, New York, USA) and incubated at 37°C in a humidified atmosphere (5% CO 2 /95% air).…”

“…Brain slices were prepared as previously described (Arias et al, 2017;Shih et al, 2014). Mice were anesthetized with Euthasol (sodium pentobarbital, 100 mg/kg; sodium phenytoin, 12.82 mg/kg) before trans-cardiac perfusion with oxygenated (95% O 2 /5% CO 2 ), 4 °C N-methyl-D-glucamine (NMDG)-based recovery solution that contains (in mM): 93 NMDG, 2.5 KCl, 1.2 NaH 2 PO 4 , 30 NaHCO 3 , 20 HEPES, 25 glucose, 5 sodium ascorbate, 2 thiourea, 3 sodium pyruvate, 10 MgSO 4 •7H 2 O, and 0.5 CaCl 2 •2H 2 O; 300-310 mOsm; pH 7.3-7.4).…”

“…Neurons within brain slices were visualized with infrared or visible differential interference contrast (DIC) optics. Neurons in the ventral inferior (VI) aspect of the MHb were targeted for recordings, as previously described (Arias et al, 2017;Shih et al, 2014). Electrophysiology experiments were conducted using a Scientifica SliceScope or Nikon FN-1 upright microscope.…”

“…Habenular α3β4* AChRs are considered important targets for several anti-addictive compounds (Glick et al, 2002;Maisonneuve and Glick, 2003;McCallum et al, 2012). Compounds with relatively higher selectivity for this receptor subtype such as 18-methoxycoronaridine (Arias et al, 2017) have anti-addictive properties and decrease alcohol intake in rodents (Rezvani et al, 2016), whereas bupropion and mecamylamine have antidepressant activity (Arias et al, 2018a(Arias et al, , 2014. Nevertheless, no direct evidence of (±)-citalopram-induced habenular α3β4* AChRs inhibition has been demonstrated so far.…”