a-85380 and cytisine

Nicotinic acetylcholine receptors (nAChRs) have been investigated for developing drugs that can potentially treat various central nervous system disorders. Considerable evidence supports the hypothesis that modulation of the cholinergic system through activation and/or desensitization/inactivation of nAChR holds promise for the development of new antidepressants. The introductory portion of this Miniperspective discusses the basic pharmacology that underpins the involvement of α4β2-nAChRs in depression, along with the structural features that are essential to ligand recognition by the α4β2-nAChRs. The remainder of this Miniperspective analyzes reported nicotinic ligands in terms of drug design considerations and their potency and selectivity, with a particular focus on compounds exhibiting antidepressant-like effects in preclinical or clinical studies. This Miniperspective aims to provide an in-depth analysis of the potential for using nicotinic ligands in the treatment of depression, which may hold some promise in addressing an unmet clinical need by providing relief from depressive symptoms in refractory patients.

Topics: Alkaloids; Antidepressive Agents; Azetidines; Azocines; Depression; Humans; Ligands; Molecular Targeted Therapy; Nicotinic Agonists; Nicotinic Antagonists; Quinolizines; Receptors, Nicotinic

Nicotine elicited membrane depolarization, elevation of intracellular calcium, rubidium efflux, and release of insulin from mouse beta-TC6 insulinoma cells. Such responses were blocked by the nicotinic antagonist mecamylamine but not by the muscarinic antagonist atropine. Neither the selective alpha4beta2 antagonist dihydro-beta-erythroidine nor the selective alpha7 antagonist methyllycaconitine significantly blocked the nicotine-elicited depolarization or the calcium response. The elevation of intracellular calcium did not occur in calcium-free media, indicating that the increase in intracellular calcium was due to the influx of calcium. The rank order of potency for nicotinic agonists was as follows: epibatidine > nicotine = 3-(azetidinylmethoxy)pyridine (A-85380), cytisine, dimethylphenylpiperazinium (DMPP). Cytisine and DMPP seemed to be partial agonists. The density of nicotinic receptors measured by [3H]epibatidine binding was 7-fold higher in membranes from beta-TC6 cells than in rat brain membranes. No binding of 125I-A-85380 was detected, indicating the absence of beta2-containing receptors. Reverse transcription-polymerase chain reaction analyses indicated the presence of mRNA for alpha3 and alpha4 subunits and beta2 and beta4 subunits in beta-TC6 cells. The binding and functional data suggest that the major nicotinic receptor is composed of alpha3 and beta4 subunits. The beta-TC6 cells thus provide a model system for pharmacological study of such nicotinic receptors.

Topics: Alkaloids; Animals; Azetidines; Azocines; Bridged Bicyclo Compounds, Heterocyclic; Calcium; Calcium Signaling; Cell Membrane; Dimethylphenylpiperazinium Iodide; Insulin; Insulin-Secreting Cells; Insulinoma; Membrane Potentials; Mice; Nicotine; Nicotinic Agonists; Pyridines; Quinolizines; Receptors, Nicotinic; Tumor Cells, Cultured

The subunit composition and pharmacology of alpha-Conotoxin MII-binding (alpha-CtxMII) nicotinic acetylcholine receptors (nAChR) was studied by an improved [(125)I]-alpha-CtxMII membrane binding method. This binding method facilitates pharmacological studies that have been difficult to accomplish with [(125)I]-alpha-CtxMII autoradiography or alpha-CtxMII inhibition of [(125)I]-epibatidine binding. Binding densities and K(d)-values obtained by this [(125)I]-alpha-CtxMII membrane binding were similar to the values obtained by autoradiography or alpha-CtxMII inhibition of [(125)I]-epibatidine binding, verifying that each of these approaches measures the same nAChR population. Binding results with nAChR subunit-null mutant mice confirm and extend observations from earlier studies: [(125)I]-alpha-CtxMII binding measures two sets of alpha6beta2* nAChR (alpha4alpha6beta2beta3 or alpha6beta2beta3). Most nicotinic agonists and antagonists show monophasic inhibition of [(125)I]-alpha-CtxMII binding, indicating that alpha4alpha6beta2beta3 and alpha6beta2beta3 have similar binding properties. Comparison of the binding and activation profiles of alpha6beta2* nAChR to those of other nAChR subtypes (alpha4beta2* and beta4*) indicates that these receptors have distinctly different pharmacology indicating that it may be possible to target alpha6beta2* nAChR selectively to develop compounds that might be therapeutically useful.

Topics: Acetylcholine; Alkaloids; Animals; Azetidines; Azocines; Binding, Competitive; Brain; Bridged Bicyclo Compounds, Heterocyclic; Cell Membrane; Conotoxins; Dose-Response Relationship, Drug; Drug Interactions; Hydrogen-Ion Concentration; Iodine Isotopes; Male; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Nicotine; Nicotinic Agonists; Nicotinic Antagonists; Protein Binding; Protein Subunits; Pyridines; Quinolizines; Radioligand Assay; Receptors, Nicotinic; Time Factors

Comparison of [125I]epibatidine and 5-[125I]iodo-3-(2-azetidinylmethoxy)pyridine ([125I]A-85380) autoradiography showed evidence for nicotinic receptor heterogeneity. To identify the receptor subtypes, we performed [125I]epibatidine autoradiography in the presence of cytisine or A-85380. By comparing these results with binding data from human embryonic kidney (HEK) 293 cells stably transfected with different combinations of rat nicotinic receptor subunits, we were able to quantify three distinct populations of [125I]epibatidine binding sites with characteristics of alpha4beta2, alpha3beta2 and alpha3beta4 receptors. Although the predominant subtype in rat brain was alpha4beta2, non-alpha4beta2 binding sites were prominent in many regions. In the habenulo-peduncular system, cerebellum, substantia gelatinosa, and many medullary nuclei, alpha3beta4-like binding accounted for more than 40% of [125I]epibatidine binding, and nearly all binding in superior cervical ganglion and pineal gland. Other regions enriched in alpha3beta4-like binding included locus ceruleus, dorsal tegmentum, subiculum and anteroventral thalamic nucleus. Regions enriched in alpha3beta2-like binding included the habenulo-peduncular system, many visual system structures, certain geniculate nuclei, and dopaminergic regions. The combination of autoradiography using a broad spectrum radioligand in the presence of selective competitors, and data from binding to defined receptor subtypes in expression systems, allowed us to quantify the relative populations of these three subtypes.

Topics: Alkaloids; Animals; Autoradiography; Azetidines; Azocines; Binding, Competitive; Brain; Bridged Bicyclo Compounds, Heterocyclic; Cell Line; Humans; Iodine Radioisotopes; Kidney; Ligands; Male; Nicotinic Antagonists; Organ Specificity; Pineal Gland; Protein Subunits; Pyridines; Quinolizines; Rats; Rats, Sprague-Dawley; Receptors, Nicotinic; Spinal Cord; Superior Cervical Ganglion; Tissue Distribution

In the past few years the focus on central acetylcholine receptors has shifted from compounds with affinity for muscarinic acetylcholine receptors (mAChR) to compounds with affinity for nicotinic acetylcholine receptors (nAChR). The therapeutic potential includes treatment of a variety of diseases, e.g., Alzheimer's disease, Parkinson's disease, and Tourette's syndrome. This work describes the synthesis of six novel series of potent ligands with nanomolar affinity for the alpha4beta2 nAChR subtype. Structure-activity relationship (SAR) was evaluated by the calculation of a 3D-QSAR model. 3D-QSAR analysis of the compounds using the GRID/GOLPE methodology resulted in a model of high quality (R(2) = 0.97, Q(2) = 0.81). The coefficient plots reveal that the steric interactions between the target and our compounds are of major importance for the affinity. Bulky substituents in the 6-position of the pyridine ring will reduce the affinity of the compounds, whereas bulky ring systems including a sp(3)-nitrogen will increase the affinity of the compounds.

Topics: Alkaloids; Animals; Azocines; Brain; Cholinergic Agents; Ligands; Male; Molecular Conformation; Protein Isoforms; Quinolizines; Rats; Rats, Wistar; Receptors, Nicotinic; Reproducibility of Results; Structure-Activity Relationship; Tritium

Nicotinic acetylcholine receptors (nAChRs) play an important role in tobacco dependence and a potential therapeutic role in neuropsychiatric disorders such as Alzheimer's disease. [123I]5-iodo-3-[2(S)-2-azetidinylmethoxy]pyridine (5-I-A-85380) is a new SPECT tracer that labels alpha4beta2 nAChRs. The purpose of this study was to assess the usefulness of this tracer to measure regional nAChR binding in baboon brain using both a bolus/kinetic paradigm and also a bolus plus constant infusion/equilibrium paradigm.. A pair of bolus/kinetic and bolus plus constant infusion/equilibrium studies was performed in each of 3 isoflurane-anesthetized baboons. Bolus studies were performed by intravenous injection of 191-226 MBq [123I]5-I-A-85380 and image acquisition for 289-367 min. The data were analyzed with 1- and 2-tissue compartment models. Bolus plus constant infusion/equilibrium studies were performed by a bolus injection (74-132 MBq) followed by a 468- to 495-min infusion with a bolus/infusion ratio (B/I) of 4.8-5.0 h. The distribution volumes in the thalamus were measured in these 2 paradigms. To study whether the cerebellum was appropriate as a receptor-poor region, displacement studies were done in 2 baboons using the B/I paradigm with subcutaneous injection of (-)-cytisine (0.8 and 1.0 mg/kg).. The kinetics of this tracer was best described by the 1-tissue compartment model. The 2-compartment model showed poor identifiability of rate constants. The total (specific plus nondisplaceable compartments) distribution volumes (V(T)') agreed between bolus and B/I paradigms (average percentage difference in V(T)', 16.8%). (-)-Cytisine (0.8 and 1.0 mg/kg) displaced 70% and 72% of the radioactivity in the thalamus and 36% and 55% in the cerebellum, respectively, indicating that the latter was not appropriate as a receptor-poor region.. These results show the feasibility of quantifying alpha4beta2 nAChRs using [123I]5-I-A-85380 and support the use of V(T)' as an appropriate outcome measure.

Topics: Alkaloids; Animals; Azetidines; Azocines; Binding, Competitive; Brain; Cerebellum; Image Processing, Computer-Assisted; Infusions, Intravenous; Injections, Intravenous; Iodine Radioisotopes; Kinetics; Least-Squares Analysis; Magnetic Resonance Imaging; Papio; Quinolizines; Receptors, Nicotinic; Thalamus; Tomography, Emission-Computed, Single-Photon

[(125)I]-Epibatidine binds to multiple nicotinic acetylcholine receptor (nAChR) subtypes with high affinity. In this study, [(125)I]-epibatidine was used to label and characterize a novel nAChR subtype found in mouse brain inferior colliculus, interpeduncular nucleus, and olfactory bulb homogenates. Binding of [(125)I]-epibatidine was saturable and apparently monophasic in each brain region (K:(D:)=71+/-12 pM mean+/-s.e.mean across regions) but inhibition of [(125)I]-epibatidine binding (200 pM) by A85380, cytisine and (-)-nicotine was biphasic, indicating the presence of multiple binding sites. The sites with lower agonist affinity comprised 30.0+/-2.2, 58.6+/-0.1 and 48.7+/-3.3% of specific [(125)I]-epibatidine (200 pM) binding in inferior colliculus, interpeduncular nucleus, and olfactory bulb homogenates, respectively. The affinity difference between A85380-sensitive and -resistant binding sites was particularly marked (approximately 1000 fold). Thus A85380 was used to differentiate agonist-sensitive and -resistant sites. The pharmacological profiles of the A85380-resistant sites in each region were assessed with inhibition binding experiments, using 14 agonists and five antagonists. The profiles were indistinguishable across regions, implying that A85380-resistant [(125)I]-epibatidine binding sites in inferior colliculus, interpeduncular nucleus, and olfactory bulb represent a single nAChR subtype. The pharmacological profile of the A85380-resistant sites is very different from that previously reported for high affinity (-)-[(3)H]-nicotine-, [(125)I]-alpha-bungarotoxin-, or [(125)I]-alpha-conotoxin MII-binding sites, suggesting that they represent a novel nAChR population in mouse brain.

Topics: Alkaloids; Animals; Autoradiography; Azetidines; Azocines; Binding Sites; Brain; Bridged Bicyclo Compounds, Heterocyclic; Iodine Radioisotopes; Male; Mice; Mice, Inbred C57BL; Nicotine; Nicotinic Agonists; Pyridines; Quinolizines; Receptors, Nicotinic

The biodistribution of the nicotinic acetylcholine receptor (nAChR) radioligand 2-[18F]fluoro-3-[2(S)-2-azetidinylmethoxy]pyridine ([18F]fluoro-A-85380, half-life of fluorine-18 = 110 min) in selected rat brain areas was assessed in vivo. The radiotracer showed a good penetration in the brain. The regional distribution of the radioligand was consistent with the density of nAChRs determined from previous studies in vitro. Sixty minutes post-injection, the highest uptake was observed in the thalamus, (1% I.D./g tissue), an intermediate one in the frontal cortex (0.78% I.D./g tissue), and the lowest in the cerebellum (0.5% I.D./g tissue). Pretreatment with several nAChR ligands (nicotine, cytisine, epibatidine, unlabeled fluoro-A-85380) substantially reduced uptake of the radioligand in the three cerebral areas. Pretreatment with the nAChR channel blocker mecamylamine or with the muscarinic receptor antagonist dexetimide had no appreciable effect on the uptake of fluoro-A-85380. These results support the high in vivo selectivity and specificity of fluoro-A-85380. Therefore, [18F]fluoro-A-85380 may be useful for positron emission tomography study of nAChRs in humans.

Topics: Alkaloids; Animals; Azetidines; Azocines; Binding, Competitive; Brain; Bridged Bicyclo Compounds, Heterocyclic; Cerebellum; Fluorine Radioisotopes; Frontal Lobe; Kinetics; Ligands; Male; Mecamylamine; Nicotine; Nicotinic Agonists; Pyridines; Quinolizines; Radioligand Assay; Rats; Rats, Sprague-Dawley; Receptors, Nicotinic; Thalamus; Tissue Distribution

Analogs of A-98593 (1) and its enantiomer ABT-594 (2) with diverse substituents on the pyridine ring were prepared and tested for affinity to nicotinic acetylcholine receptor binding sites in rat brain and for analgesic activity in the mouse hot plate assay. Numerous types of modifications were consistent with high affinity for [3H]cytisine binding sites. By contrast, only selected modifications resulted in retention of analgesic potency in the same range as 1 and 2. Analogs of 2 with one or two methyl substituents at the 3-position of the azetidine ring also were prepared and found to be substantially less active in both assays.

Topics: Alkaloids; Analgesics, Non-Narcotic; Animals; Azetidines; Azocines; Binding Sites; Brain; Mice; Nicotinic Agonists; Pain Measurement; Pyridines; Quinolizines; Rats; Receptors, Nicotinic; Stereoisomerism; Structure-Activity Relationship; Tritium

The in vivo biodistribution profile of the novel nicotinic acetylcholine receptor (nAChR) radioligand 5-[I-125/123]Iodo-3(2(S)-azetidinylmethoxy)pyridine, [I-125/123]-5-IA, in mouse brain was examined. This radiotracer displayed good brain penetration (3.1% of the injected dose (ID) in whole brain at 15 min post-radioligand injection). Radioligand distribution was consistent with the density of high affinity nAChRs with highest uptake observed in the nAChR-rich thalamus (14.9 %ID/g at 60 min), moderate uptake in cortex (8.5 %ID/g at 60 min), and lowest uptake in the cerebellum (2.4 %ID/g at 60 min). Pretreatment with several different nAChR agonists (A-85380, (-)-nicotine, cytisine) significantly inhibited [I-125]-5-IA binding in all brain regions studied (P < 0.01) demonstrating the high specificity of the radioligand for nAChRs. Blocking doses of the muscarinic antagonist scopolamine and the non-competitive nAChR channel blocker mecamylamine had no significant effect on radioactive uptake supporting the in vitro selectivity of [I-125]-5-IA for the nAChR component of the cholinergic system. [I-125]-5-IA binding sites were shown to be saturable with unlabeled 5-IA. With a relatively low acute toxicity (LD50 > 3 mg/kg via intravenous injection in mice) and high in vivo specificity and selectivity, 5-IA labeled with the imaging radionuclide I-123 may prove useful for single photon emission computed tomography (SPECT) studies of nAChRs in human subjects.

Topics: Alkaloids; Animals; Azetidines; Azocines; Binding Sites; Brain; Dose-Response Relationship, Drug; Iodine Radioisotopes; Male; Mecamylamine; Mice; Muscarinic Antagonists; Nicotine; Nicotinic Antagonists; Quinolizines; Radionuclide Imaging; Receptors, Nicotinic; Scopolamine; Tissue Distribution

Recent evidence indicating the therapeutic potential of cholinergic channel modulators for the treatment of central nervous system (CNS) disorders as well as the diversity of brain neuronal nicotine acetylcholine receptors (nAChRs) have suggested an opportunity to develop subtype-selective nAChR ligands for the treatment of specific CNS disorders with reduced side effect liabilities. We report a novel series of 3-pyridyl ether compounds which possess subnanomolar affinity for brain nAChRs and differentially activate subtypes of neuronal nAChRs. The synthesis and structure-activity relationships for the leading members of the series are described, including A-85380 (4a), which possesses ca.50 pM affinity for rat brain [(3)H]-(-)-cytisine binding sites and 163% efficacy compared to nicotine to stimulate ion flux at human alpha4beta2 nAChR subtype, and A-84543 (2a), which exhibits 84-fold selectivity to stimulate ion flux at human alpha4beta2 nAchR subtype compared to human ganglionic type nAChRs. Computational studies indicate that a reasonable superposition of a low energy conformer of 4A with (S)-nicotine and (-)-epibatidine can be achieved.

Topics: Alkaloids; Animals; Azocines; Binding, Competitive; Brain; Cell Line; Cell Membrane; Ethers; Ganglia; Humans; Molecular Structure; Neurons; Nicotinic Agonists; Pyridines; Quinolizines; Radioligand Assay; Rats; Receptors, Nicotinic; Structure-Activity Relationship; Tritium