Pericardial Delivery of SDF-1α Puerarin Hydrogel Promotes Heart Repair and Electrical Coupling.

The stromal-derived factor 1α/chemokine receptor 4 (SDF-1α/CXCR4) axis contributes to myocardial protection after myocardial infarction (MI) by recruiting endogenous stem cells into the ischemic tissue. However, excessive inflammatory macrophages are also recruited simultaneously, aggravating myocardial damage. More seriously, the increased inflammation contributes to abnormal cardiomyocyte electrical coupling, leading to inhomogeneities in ventricular conduction and retarded conduction velocity. It is highly desirable to selectively recruit the stem cells but block the inflammation. In this work, SDF-1α-encapsulated Puerarin (PUE) hydrogel (SDF-1α@PUE) is capable of enhancing endogenous stem cell homing and simultaneously polarizing the recruited monocyte/macrophages into a repairing phenotype. Flow cytometry analysis of the treated heart tissue shows that endogenous bone marrow mesenchymal stem cells, hemopoietic stem cells, and immune cells are recruited while SDF-1α@PUE efficiently polarizes the recruited monocytes/macrophages into the M2 type. These macrophages influence the preservation of connexin 43 (Cx43) expression which modulates intercellular coupling and improves electrical conduction. Furthermore, by taking advantage of the improved "soil", the recruited stem cells mediate an improved cardiac function by preventing deterioration, promoting neovascular architecture, and reducing infarct size. These findings demonstrate a promising therapeutic platform for MI that not only facilitates heart regeneration but also reduces the risk of cardiac arrhythmias.

Oral sazetidine-A, a selective α4ß2* nicotinic receptor desensitizing agent, reduces nicotine self-administration in rats.

Sazetidine-A selectively desensitizes α4ß2 nicotinic receptors and also has partial agonist effects. We have shown that subcutaneous acute and repeated injections as well as chronic infusions of sazetidine-A significantly reduce intravenous (IV) nicotine self-administration in rats. To further investigate the promise of sazetidine-A as a smoking cessation aid, it is important to determine sazetidine-A effects with oral administration and the time-effect function for its action on nicotine self-administration. Young adult female Sprague-Dawley rats were trained to self-administer IV nicotine at the benchmark dose of 0.03 mg/kg/infusion dose in an operant FR1 schedule in 45-min sessions. After five sessions of training, they were tested for the effects of acute oral doses of sazetidine-A (0, 0.3, 1 and 3 mg/kg) given 30 min before testing. To determine the time-effect function, these rats were administered 0 or 3 mg/kg of sazetidine-A 1, 2, 4 or 23 h before the onset of testing. Our previous study showed that with subcutaneous injections, only 3 mg/kg of sazetidine-A significantly reduced nicotine self-administration, however, with oral administration of sazetidine-A lower dose of 1 mg/kg was also effective in reducing nicotine intake. A similar effect was seen in the time-effect study with 3 mg/kg of oral sazetidine-A causing a significant reduction in nicotine self-administration across all the time points of 1, 2, 4 or 23 h after oral administration. These results advance the development of sazetidine-A as a possible aid for smoking cessation by showing effectiveness with oral administration and persistence of the effect over the course of a day.

α4ß2 Nicotinic receptor desensitizing compounds can decrease self-administration of cocaine and methamphetamine in rats.

Sazetidine-A [6-(5(((S)-azetidine-2-yl)methoxy)pyridine-3-yl)hex-5-yn-1-ol] is a selective α4ß2 nicotinic receptor desensitizing agent and partial agonist. Sazetidine-A has been shown in our previous studies to significantly reduce nicotine and alcohol self-administration in rats. The question arises whether sazetidine-A would reduce self-administration of other addictive drugs as well. Nicotinic receptors on the dopaminergic neurons in the ventral tegmental area play an important role in controlling the activity of these neurons and release of dopamine in the nucleus accumbens, which is critical mechanism for reinforcing value of drugs of abuse. Previously, we showed that the nonspecific nicotinic antagonist mecamylamine significantly reduces cocaine self-administration in rats. In this study, we acutely administered systemically sazetidine-A and two other selective α4ß2 nicotinic receptor-desensitizing agents, VMY-2-95 and YL-2-203, to young adult female Sprague-Dawley rats and determined their effects on IV self-administration of cocaine and methamphetamine. Cocaine self-administration was significantly reduced by 0.3 mg/kg of sazetidine-A. In another set of rats, sazetidine-A (3 mg/kg) significantly reduced methamphetamine self-administration. VMY-2-95 significantly reduced both cocaine and methamphetamine self-administration with threshold effective doses of 3 and 0.3 mg/kg, respectively. In contrast, YL-2-203 did not significantly reduce cocaine self-administration at the same dose range and actually significantly increased cocaine self-administration at the 1 mg/kg dose. YL-2-203 (3 mg/kg) did significantly decrease methamphetamine self-administration. Sazetidine-A and VMY-2-95 are promising candidates to develop as new treatments to help addicts successfully overcome a variety of addictions including tobacco, alcohol as well as the stimulant drugs cocaine and methamphetamine.

Absolute Configuration and Pharmacology of the Poison Frog Alkaloid Phantasmidine.

Phantasmidine, a rigid congener of the well-known nicotinic acetylcholine receptor agonist epibatidine, is found in the same species of poison frog ( Epipedobates anthonyi). Natural phantasmidine was found to be a 4:1 scalemic mixture, enriched in the (2a R,4a S,9a S) enantiomer by chiral-phase LC-MS comparison to the synthetic enantiomers whose absolute configurations were previously established by Mosher's amide analysis. The major enantiomer has the opposite S configuration at the benzylic carbon to natural epibatidine, whose benzylic carbon is R. Pharmacological characterization of the synthetic racemate and separated enantiomers established that phantasmidine is ∼10-fold less potent than epibatidine, but ∼100-fold more potent than nicotine in most receptors tested. Unlike epibatidine, phantasmidine is sharply enantioselective in its activity and the major natural enantiomer whose benzylic carbon has the 4a S configuration is more active. The stereoselective pharmacology of phantasmidine is ascribed to its rigid and asymmetric shape as compared to the nearly symmetric conformations previously suggested for epibatidine enantiomers. While phantasmidine itself is too toxic for direct therapeutic use, we believe it is a useful platform for the development of potent and selective nicotinic agonists, which may have value as pharmacological tools.

The (α4)3(ß2)2 Stoichiometry of the Nicotinic Acetylcholine Receptor Predominates in the Rat Motor Cortex.

The α4ß2 nicotinic acetylcholine receptor (nAChR) is important in central nervous system physiology and in mediating several of the pharmacological effects of nicotine on cognition, attention, and affective states. It is also the likely receptor that mediates nicotine addiction. This receptor assembles in two distinct stoichiometries: (α4)2(ß2)3 and (α4)3(ß2)2, which are referred to as high-sensitivity (HS) and low-sensitivity (LS) nAChRs, respectively, based on a difference in the potency of acetylcholine to activate them. The physiologic and pharmacological differences between these two receptor subtypes have been described in heterologous expression systems. However, the presence of each stoichiometry in native tissue currently remains unknown. In this study, different ratios of rat α4 and ß2 subunit cDNA were transfected into human embryonic kidney 293 cells to create a novel model system of HS and LS α4ß2 nAChRs expressed in a mammalian cell line. The HS and LS nAChRs were characterized through pharmacological and biochemical methods. Isolation of surface proteins revealed higher amounts of α4 or ß2 subunits in the LS or HS nAChR populations, respectively. In addition, sazetidine-A displayed different efficacies in activating these two receptor stoichiometries. Using this model system, a neurophysiological "two-concentration" acetylcholine or carbachol paradigm was developed and validated to determine α4/ß2 subunit stoichiometry. This paradigm was then used in layers I-IV of slices of the rat motor cortex to determine the percent contribution of HS and LS α4ß2 receptors in this brain region. We report that the majority of α4ß2 nAChRs in this brain region possess a stoichiometry of the (α4)3(ß2)2 LS subtype.

Differential efficacies of the nicotinic α4ß2 desensitizing agents in reducing nicotine self-administration in female rats.

RATIONALE AND OBJECTIVES: Desensitization of neuronal nicotinic acetylcholine receptors holds promise as an effective treatment of tobacco addiction. Previously, we found that sazetidine-A (Saz-A), which selectively desensitizes α4ß2 nicotinic receptors, significantly decreased intravenous (IV) nicotine self-administration (SA) in rats with an effective dose of 3 mg/kg in acute and repeated injection studies. We also found that chronic infusions of Saz-A at doses of 2 and 6 mg/kg/day significantly reduced nicotine SA in rats. In continuing studies, we have characterized other Saz-A analogs, YL-2-203 and VMY-2-95, to determine their efficacies in reducing nicotine SA in rats. METHODS: Young adult female Sprague-Dawley rats were fitted with IV catheters and were trained for nicotine SA (0.03 mg/kg/infusion) on a fixed ratio 1 schedule for ten sessions. The same rats were also implanted subcutaneously with osmotic minipumps to continually deliver 2 or 6 mg/kg body weight YL-2-203, VMY-2-95, or saline for four consecutive weeks. RESULTS: Chronic administration of VMY-2-95 at doses of 2 and 6 mg/kg/day caused significant (p < 0.01) decreases in nicotine SA over the 2 weeks of continued nicotine SA and for the 1-week period of resumed access after a week of enforced abstinence, whereas chronic administration of YL-2-203 at the same doses was not found to be effective. CONCLUSIONS: These studies, together with our previous studies of Saz-A, revealed a spectrum of efficacies for these α4ß2 nicotinic receptor desensitizing agents and provide a path forward for the most effective compounds to be further developed as possible aids to smoking cessation.

Anxiolytic- and antidepressant-like effects of the methadone metabolite 2-ethyl-5-methyl-3,3-diphenyl-1-pyrroline (EMDP).

The enhancement of GABAergic and monoaminergic neurotransmission has been the mainstay of pharmacotherapy and the focus of drug-discovery for anxiety and depressive disorders for several decades. However, the significant limitations of drugs used for these disorders underscores the need for novel therapeutic targets. Neuronal nicotinic acetylcholine receptors (nAChRs) may represent one such target. For example, mecamylamine, a non-competitive antagonist of nAChRs, displays positive effects in preclinical tests for anxiolytic and antidepressant activity in rodents. In addition, nicotine elicits similar effects in rodent models, possibly by receptor desensitization. Previous studies (Xiao et al., 2001) have identified two metabolites of methadone, EMDP (2-ethyl-5-methyl-3,3-diphenyl-1-pyrroline) and EDDP (2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine), which are considered to be inactive at opiate receptors, as relatively potent noncompetitive channel blockers of rat α3ß4 nAChRs. Here, we show that these compounds are likewise highly effective blockers of human α3ß4 and α4ß2 nAChRs. Moreover, we show that they display relatively low affinity for opiate binding sites labeled by [(3)H]-naloxone. We then evaluated these compounds in rats and mice in preclinical behavioral models predictive of potential anxiolytic and antidepressant efficacy. We found that EMDP, but not EDDP, displayed robust effects predictive of anxiolytic and antidepressant efficacy without significant effects on locomotor activity. Moreover, EMDP at behaviorally active doses, unlike mecamylamine, did not produce eyelid ptosis, suggesting it may produce fewer autonomic side effects than mecamylamine. Thus, the methadone metabolite EMDP may represent a novel therapeutic avenue for the treatment of some affective disorders.

Competition, Selectivity and Efficacy of Analogs of A-84543 for Nicotinic Acetylcholine Receptors with Repositioning of Pyridine Nitrogen.

Nicotinic acetylcholine receptors (nAChRs) play a crucial role in a number of clinically relevant mental and neurological pathways, as well as autonomic and immune functions. The development of subtype-selective ligands for nAChRs therefore is potentially useful for targeted therapeutic management of conditions where nAChRs are involved. We tested if selectivity for a particular nAChR subtype can be achieved through small structural modifications of a lead compound containing the nicotinic pharmacophore by changing the distance between the electronegative elements. For this purpose, analogs of A-84543 were designed, synthesized and characterized as potentially new nAChR subtype-selective ligands. Compounds were tested for their binding properties in rat cerebral cortical tissue homogenates, and subtype-selectivity was determined using stably transfected HEK cells expressing different nAChR subtypes. All compounds synthesized were found to competitively displace [(3)H]-epibatidine ([(3)H]EB) from the nAChR binding site. Of all the analogues, H-11MNH showed highest affinity for nAChRs compared to a ~ fivefold to tenfold lower affinity of A-84543. All other compounds had affinities >10,000 nM. Both A-84543 and H-11MNH have highest affinity for α2ß2 and α4ß2 nAChRs and show moderate affinity for ß4- and α7-containing receptors. H-11MNH was found to be a full agonist with high potency at α3ß4, while A-84543 is a partial agonist with low potency. Based on their unique pharmacological binding properties we suggest that A-84543 and its desmethylpyrrolidine analog can be useful as pharmacological ligands for studying nAChRs if selective pharmacological and/or genetic tools are used to mask the function of other receptors subtypes.

AT-1001 Is a Partial Agonist with High Affinity and Selectivity at Human and Rat α3ß4 Nicotinic Cholinergic Receptors.

AT-1001 [N-(2-bromophenyl)-9-methyl-9-azabicyclo[3.3.1] nonan-3-amine] is a high-affinity and highly selective ligand at α3ß4 nicotinic cholinergic receptors (nAChRs) that was reported to decrease nicotine self-administration in rats. It was initially reported to be an antagonist at rat α3ß4 nAChRs heterologously expressed in HEK293 cells. Here we compared AT-1001 actions at rat and human α3ß4 and α4ß2 nAChRs similarly expressed in HEK 293 cells. We found that, as originally reported, AT-1001 is highly selective for α3ß4 receptors over α4ß2 receptors, but its binding selectivity is much greater at human than at rat receptors, because of a higher affinity at human than at rat α3ß4 nAChRs. Binding studies in human and rat brain and pineal gland confirmed the selectivity of AT-1001 for α3ß4 nAChRs and its higher affinity for human compared with rat receptors. In patch-clamp electrophysiology studies, AT-1001 was a potent partial agonist with 65-70% efficacy at both human and rat α3ß4 nAChRs. It was also a less potent and weaker (18%) partial agonist at α4ß2 nAChRs. Both α3ß4 and α4ß2 nAChRs are upregulated by exposure of cells to AT-1001 for 3 days. Similarly, AT-1001 desensitized both receptor subtypes in a concentration-dependent manner, but it was 10 and 30 times more potent to desensitize human α3ß4 receptors than rat α3ß4 and human α4ß2 receptors, respectively. After exposure to AT-1001, the time to recovery from desensitization was longest for the human α3ß4 nAChR and shortest for the human α4ß2 receptor, suggesting that recovery from desensitization is primarily related to the dissociation of the ligand from the receptor.

Structure-activity relationship studies of SEN12333 analogues: determination of the optimal requirements for binding affinities at α7 nAChRs through incorporation of known structural motifs.

Alpha7 nicotinic acetylcholine receptors (nAChRs) have implications in the regulation of cognitive processes such as memory and attention and have been identified as a promising therapeutic target for the treatment of the cognitive deficits associated with schizophrenia and Alzheimer's disease (AD). Structure affinity relationship studies of the previously described α7 agonist SEN12333 (8), have resulted in the identification of compound 45, a potent and selective agonist of the α7 nAChR with enhanced affinity and improved physicochemical properties over the parent compound (SEN12333, 8).

Investigations of amide bond variation and biaryl modification in analogues of α7 nAChR agonist SEN12333.

Several lines of experimental evidence support the involvement of the α7 nAChR in schizophrenia and Alzheimer's disease. Modulators of the α7 nAChR have been extensively reviewed for the treatment of the cognitive deficits associated with these pathologies. SEN12333 represents a novel α7 nAChR agonist chemotype with potential for reduced side effects but requiring further SAR exploration. The present work investigates the amide bond of SEN12333, specifically its connectivity and replacement with the tetrazole functionality, a known cis amide isostere. The results reveal the original amide bond connectivity of SEN12333 to be favorable for binding affinity and agonist activity at α7 nAChRs. The use of a tetrazole isostere completely abolishes affinity and functional activity and suggests that SEN12333 binds in a linear conformation. Results reported herein also suggest the pyridine nitrogen within the terminal aromatic ring of SEN12333 is not essential for binding affinity or functional activity. Further SAR investigations involving manipulation of other moieties contained within SEN12333 are warranted.

Synthesis and pharmacological characterization of new neuronal nicotinic acetylcholine receptor ligands derived from Sazetidine-A.

The enantiomers of two analogs of Sazetidine-A as well as several other novel biosteric analogues were synthesized. Their binding affinities at three major nAChRs subtypes and selectivity profiles were determined. Though many (S)-enantiomers of Sazetidine-A analogs have high binding affinities and good subtype selectivities, it is not a general rule that (S)-enantiomers are better than their (R) counterparts. Compound 11, of which the ethynyl group was replaced by its' bioisostere-the triazole via click chemistry, showed a high binding affinity to α4ß2 subtype (Ki=1.3 nM) and better selectivity to the α4ß2 subtype over α3ß4 subtype with that of Sazetidine-A. The azide compound 15, a potential photoaffinity label, showed improved high selectivity and similar binding property profile with that of Sazetidine-A. The biaryl analog 17 exhibited a much lower affinity as compared to Sazetidine-A indicating the importance of a 'long tail' side chain for α4ß2 nAChR binding.

18F-ASEM, a radiolabeled antagonist for imaging the α7-nicotinic acetylcholine receptor with PET.

UNLABELLED: The α7-nicotinic cholinergic receptor (α7-nAChR) is a key mediator of brain communication and has been implicated in a wide variety of central nervous system disorders. None of the currently available PET radioligands for α7-nAChR are suitable for quantitative PET imaging, mostly because of insufficient specific binding. The goal of this study was to evaluate the potential of (18)F-ASEM ((18)F-JHU82132) as an α7-nAChR radioligand for PET. METHODS: The inhibition binding assay and receptor functional properties of ASEM were assessed in vitro. The brain regional distribution of (18)F-ASEM in baseline and blockade were evaluated in DISC1 mice (dissection) and baboons (PET). RESULTS: ASEM is an antagonist for the α7-nAChR with high binding affinity (Ki = 0.3 nM). (18)F-ASEM readily entered the baboon brain and specifically labeled α7-nAChR. The in vivo specific binding of (18)F-ASEM in the brain regions enriched with α7-nAChRs was 80%-90%. SSR180711, an α7-nAChR-selective partial agonist, blocked (18)F-ASEM binding in the baboon brain in a dose-dependent manner, suggesting that the binding of (18)F-ASEM was mediated by α7-nAChRs and the radioligand was suitable for drug evaluation studies. In the baboon baseline studies, the brain regional volume of distribution (VT) values for (18)F-ASEM were 23 (thalamus), 22 (insula), 18 (hippocampus), and 14 (cerebellum), whereas in the binding selectivity (blockade) scan, all regional VT values were reduced to less than 4. The range of regional binding potential values in the baboon brain was from 3.9 to 6.6. In vivo cerebral binding of (18)F-ASEM and α7-nAChR expression in mutant DISC1 mice, a rodent model of schizophrenia, was significantly lower than in control animals, which is in agreement with previous postmortem human data. CONCLUSION: (18)F-ASEM holds promise as a radiotracer with suitable imaging properties for quantification of α7-nAChR in the human brain.

Chronic sazetidine-A maintains anxiolytic effects and slower weight gain following chronic nicotine without maintaining increased density of nicotinic receptors in rodent brain.

Chronic nicotine administration increases the density of brain α4ß2* nicotinic acetylcholine receptors (nAChRs), which may contribute to nicotine addiction by exacerbating withdrawal symptoms associated with smoking cessation. Varenicline, a smoking cessation drug, also increases these receptors in rodent brain. The maintenance of this increase by varenicline as well as nicotine replacement may contribute to the high rate of relapse during the first year after smoking cessation. Recently, we found that sazetidine-A (saz-A), a potent partial agonist that desensitizes α4ß2* nAChRs, does not increase the density of these receptors in brain at doses that decrease nicotine self-administration, increase attention in rats, and produce anxiolytic effects in mice. Here, we investigated whether chronic saz-A and varenicline maintain the density of nAChRs after their up-regulation by nicotine. In addition, we examined the effects of these drugs on a measure of anxiety in mice and weight gain in rats. After increasing nAChRs in the rodent brain with chronic nicotine, replacing nicotine with chronic varenicline maintained the increased nAChR binding, as well as the α4ß2 subunit proteins measured by western blots. In contrast, replacing nicotine treatments with chronic saz-A resulted in the return of the density of nAChRs to the levels seen in saline controls. Nicotine, saz-A and varenicline each demonstrated anxiolytic effects in mice, but only saz-A and nicotine attenuated the gain of weight over a 6-week period in rats. These findings suggest that apart from its modest anxiolytic and weight control effects, saz-A, or drugs like it, may be useful in achieving long-term abstinence from smoking.

Design, synthesis and discovery of picomolar selective α4ß2 nicotinic acetylcholine receptor ligands.

Developing novel and selective compounds that desensitize α4ß2 nicotinic acetylcholine receptors (nAChRs) could provide new effective treatments for nicotine addiction, as well as other disorders. Here we report a new class of nAChR ligands that display high selectivity and picomolar binding affinity for α4ß2 nicotinic receptors. The novel compounds have Ki values in the range of 0.031-0.26 nM and properties that should make them good candidates as drugs acting in the CNS. The selected lead compound 1 (VMY-2-95) binds with high affinity and potently desensitizes α4ß2 nAChRs. At a dose of 3 mg/kg, compound 1 significantly reduced rat nicotine self-administration. The overall results support further characterizations of compound 1 and its analogues in preclinical models of nicotine addiction and perhaps other disorders involving nAChRs.

Effects of the sazetidine-a family of compounds on the body temperature in wildtype, nicotinic receptor ß2-/- and α7-/- mice.

Nicotine elicits hypothermic responses in rodents. This effect appears to be related to nicotinic receptor desensitization because sazetidine-A, an α4ß2 nicotinic receptor desensitizing agent, produces marked hypothermia and potentiates nicotine-induced hypothermia in mice. To determine the specificity of sazetidine-A induced hypothermia to ß2 subunit-containing nicotinic receptors, we tested its efficacy in ß2 knockout (ß2(-/-)) mice. These effects were compared with wildtype (WT) and α7 knockout (α7(-/-)) mice. Confirming our earlier results, sazetidine-A elicited a pronounced and long-lasting hypothermia in WT mice. In comparison, sazetidine-A induced a much attenuated and shorter hypothermic response in ß2(-/-) mice. This indicates that the greater proportion of sazetidine-A induced hypothermia is mediated via actions on ß2-containing nicotinic receptors, while a smaller component of hypothermia induced by sazetidine-A is mediated by non-ß2 receptors. Similar to WT mice, α7(-/-) mice showed the full extent of the sazetidine-A effect, suggesting that the hypothermia produced by sazetidine-A did not depend on actions on α7 nicotinic receptor subtype. Three other novel nicotinic receptor desensitizing agents derived from sazetidine-A, triazetidine-O, VMY-2-95 and YL-1-127 also produced hypothermia in WT and α7(-/-) mice. Furthermore, unlike sazetidine-A, triazetidine-O and YL-1-127 did not show any hint of a hypothermic effect in ß2(-/-) mice. VMY-2-95 like sazetidine-A did show a residual hypothermic effect in the ß2(-/-) mice. These studies show that the hypothermic effects of sazetidine-A and the related compound VMY-2-95 are mainly mediated by nicotinic receptors containing ß2 subunit, but that a small component of the effect is apparently mediated by non-ß2 containing receptors.

Derivatives of dibenzothiophene for positron emission tomography imaging of α7-nicotinic acetylcholine receptors.

A new series of derivatives of 3-(1,4-diazabicyclo[3.2.2]nonan-4-yl)dibenzo[b,d]thiophene 5,5-dioxide with high binding affinities and selectivity for α7-nicotinic acetylcholine receptors (α7-nAChRs) (Ki = 0.4-20 nM) has been synthesized for positron emission tomography (PET) imaging of α7-nAChRs. Two radiolabeled members of the series [(18)F]7a (Ki = 0.4 nM) and [(18)F]7c (Ki = 1.3 nM) were synthesized. [(18)F]7a and [(18)F]7c readily entered the mouse brain and specifically labeled α7-nAChRs. The α7-nAChR selective ligand 1 (SSR180711) blocked the binding of [(18)F]7a in the mouse brain in a dose-dependent manner. The mouse blocking studies with non-α7-nAChR central nervous system drugs demonstrated that [(18)F]7a is highly α7-nAChR selective. In agreement with its binding affinity the binding potential of [(18)F]7a (BPND = 5.3-8.0) in control mice is superior to previous α7-nAChR PET radioligands. Thus, [(18)F]7a displays excellent imaging properties in mice and has been chosen for further evaluation as a potential PET radioligand for imaging of α7-nAChR in non-human primates.

Chemistry and pharmacological studies of 3-alkoxy-2,5-disubstituted-pyridinyl compounds as novel selective α4ß2 nicotinic acetylcholine receptor ligands that reduce alcohol intake in rats.

Neuronal acetylcholine receptors mediate the addictive effects of nicotine and may also be involved in alcohol addiction. Varenicline, an approved smoking cessation medication, showed clear efficacy in reducing alcohol consumption in heavy-drinking smokers. More recently, sazetidine-A, which selectively desensitizes α4ß2 nicotinic receptors, was shown to significantly reduce alcohol intake in a rat model. To develop novel therapeutics for treating alcohol use disorder, we designed and synthesized novel sazetidine-A analogues containing a methyl group at the 2-position of the pyridine ring. In vitro pharmacological studies revealed that some of the novel compounds showed overall pharmacological property profiles similar to that of sazetidine-A but exhibited reduced agonist activity across all nicotinic receptor subtypes tested. In rat studies, compound (S)-9 significantly reduced alcohol uptake. More importantly, preliminary results from studies in a ferret model indicate that these novel nAChR ligands have an improved adverse side-effect profile in comparison with that of varenicline.

Effects of chronic sazetidine-A, a selective α4ß2 neuronal nicotinic acetylcholine receptors desensitizing agent on pharmacologically-induced impaired attention in rats.

RATIONALE: Nicotine and nicotinic agonists have been shown to improve attentional function. Nicotinic receptors are easily desensitized, and all nicotinic agonists are also desensitizing agents. Although both receptor activation and desensitization are components of the mechanism that mediates the overall effects of nicotinic agonists, it is not clear how each of the two opposed actions contributes to attentional improvements. Sazetidine-A has high binding affinity at α4ß2 nicotinic receptors and causes a relatively brief activation followed by a long-lasting desensitization of the receptors. Acute administration of sazetidine-A has been shown to significantly improve attention by reversing impairments caused by the muscarinic cholinergic antagonist scopolamine and the NMDA glutamate antagonist dizocilpine. METHODS: In the current study, we tested the effects of chronic subcutaneous infusion of sazetidine-A (0, 2, or 6 mg/kg/day) on attention in Sprague-Dawley rats. Furthermore, we investigated the effects of chronic sazetidine-A treatment on attentional impairment induced by an acute administration of 0.02 mg/kg scopolamine. RESULTS: During the first week period, the 6-mg/kg/day sazetidine-A dose significantly reversed the attentional impairment induced by scopolamine. During weeks 3 and 4, the scopolamine-induced impairment was no longer seen, but sazetidine-A (6 mg/kg/day) significantly improved attentional performance on its own. Chronic sazetidine-A also reduced response latency and response omissions. CONCLUSIONS: This study demonstrated that similar to its acute effects, chronic infusions of sazetidine-A improve attentional performance. The results indicate that the desensitization of α4ß2 nicotinic receptors with some activation of these receptors may play an important role in improving effects of sazetidine-A on attention.

Sub-anesthetic concentrations of (R,S)-ketamine metabolites inhibit acetylcholine-evoked currents in α7 nicotinic acetylcholine receptors.

The effect of the (R,S)-ketamine metabolites (R,S)-norketamine, (R,S)-dehydronorketamine, (2S,6S)-hydroxynorketamine and (2R,6R)-hydroxynorketamine on the activity of α7 and α3ß4 neuronal nicotinic acetylcholine receptors was investigated using patch-clamp techniques. The data indicated that (R,S)-dehydronorketamine inhibited acetylcholine-evoked currents in α7-nicotinic acetylcholine receptor, IC(50) = 55 ± 6 nM, and that (2S,6S)-hydroxynorketamine, (2R,6R)-hydroxynorketamine and (R,S)-norketamine also inhibited α7-nicotinic acetylcholine receptor function at concentrations ≤ 1 µM, while (R,S)-ketamine was inactive at these concentrations. The inhibitory effect of (R,S)-dehydronorketamine was voltage-independent and the compound did not competitively displace selective α7-nicotinic acetylcholine receptor ligands [(125)I]-α-bungarotoxin and [(3)H]-epibatidine indicating that (R,S)-dehydronorketamine is a negative allosteric modulator of the α7-nicotinic acetylcholine receptor. (R,S)-Ketamine and (R,S)-norketamine inhibited (S)-nicotine-induced whole-cell currents in cells expressing α3ß4-nicotinic acetylcholine receptor, IC(50) 3.1 and 9.1 µM, respectively, while (R,S)-dehydronorketamine, (2S,6S)-hydroxynorketamine and (2R,6R)-hydroxynorketamine were weak inhibitors, IC(50) >100 µM. The binding affinities of (R,S)-dehydronorketamine, (2S,6S)-hydroxynorketamine and (2R,6R)-hydroxynorketamine at the NMDA receptor were also determined using rat brain membranes and the selective NMDA receptor antagonist [(3)H]-MK-801. The calculated K(i) values were 38.95 µM for (S)-dehydronorketamine, 21.19 µM for (2S,6S)-hydroxynorketamine and>100 µM for (2R,6R)-hydroxynorketamine. The results suggest that the inhibitory activity of ketamine metabolites at the α7-nicotinic acetylcholine receptor may contribute to the clinical effect of the drug.