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Schizophrenia is a complex disease involving the dysregulation of numerous brain circuits and patients exhibit positive symptoms (hallucinations, delusions), negative symptoms (anhedonia), and cognitive impairments. We have shown that the antipsychotic efficacy of positive allosteric modulators (PAMs) of both the M 4 muscarinic receptor and metabotropic glutamate receptor 1 (mGlu 1 ) involve the retrograde activation of the presynaptic cannabinoid type-2 (CB 2 ) receptor, indicating that CB 2 activation or potentiation could result in a novel therapeutic strategy for schizophrenia. We used two complementary assays, receptor-mediated phosphoinositide hydrolysis and GIRK channel activation, to characterize a CB 2 PAM scaffold, represented by the compound EC21a, to explore its potential as a starting point to optimize therapeutics for schizophrenia. These studies revealed that EC21a acts as an allosteric inverse agonist at CB 2 in both assays and exhibits a mixed allosteric agonist/negative allosteric modulator profile at CB 1 depending upon the assay used for profiling. A series of compounds related to EC21a also functioned as CB 2 inverse agonists. Overall, these results suggest that EC21a exhibits complicated and potentially assay-dependent pharmacology, which may impact interpretation of in vivo studies.

Medetomidine is an FDA-approved α 2 -adrenoreceptor (α 2 -AR) agonist used as a veterinary sedative due to its analgesic, sedative, and anxiolytic properties. While it is marketed for veterinary use as a racemic mixture under the brand name Domitor, the pharmacologically active enantiomer, dexmedetomidine, is approved for sedation and analgesia in the hospital setting. Medetomidine has recently been detected in the illicit drug supply alongside fentanyl, xylazine, cocaine, and heroin, producing pronounced sedative effects that are not reversed by naloxone. The pharmacological effects along with the low cost of supply and lack of regulation for medetomidine has made it a target for misuse. Since 2022, medetomidine has been found as an adulterant in samples of seized drugs, as well as in toxicological analyses of patients admitted to the emergency department after suspected overdoses across several U.S. states and Canada. This Review will discuss the history, chemistry, structure-activity relationships, drug metabolism and pharmacokinetics (DMPK), pharmacology, and emergence of medetomidine as an adulterant in drug mixtures in the context of the current opioid drug crisis.

Tianeptine ( 1 ) is an unusual antidepressant in that its mechanism of action appears to be independent from any activity at serotonin receptors or monoamine transporters. In fact, tianeptine has been shown to be a moderately potent agonist for the mu opioid receptor (MOR) and to a lesser extent the delta opioid receptor (DOR). Additionally, tianeptine's efficacy may be related to its action on glutamate-mediated pathways of neuroplasticity. Regardless of which neurotransmitter system is primarily responsible for the observed efficacy, the MOR agonist activity is problematic with respect to abuse liability. Increasing numbers of case reports have demonstrated that tianeptine is indeed being used recreationally at doses far beyond what are considered therapeutically relevant or safe, and scheduling reclassifications or outright bans on tianeptine products are ongoing around the world. It is the aim of this review to discuss the medicinal chemistry and pharmacology of tianeptine and to summarize this intriguing discrepancy between tianeptine's historical use as a safe and effective antidepressant and its emerging potential for abuse.

Herein, we report structure-activity relationship (SAR) studies to develop novel tricyclic M 4 PAM scaffolds with improved pharmacological properties. This endeavor involved a "tie-back" strategy to replace a 5-amino-2,4-dimethylthieno[2,3- d ]pyrimidine-6-carboxamide core, which led to the discovery of two novel tricyclic cores. While both tricyclic cores displayed low nanomolar potency against both human and rat M 4 and were highly brain-penetrant, the 2,4-dimethylpyrido[4',3':4,5]thieno[2,3- d ]pyrimidine tricycle core provided lead compound, VU6016235 , with an overall superior pharmacological and drug metabolism and pharmacokinetics (DMPK) profile, as well as efficacy in a preclinical antipsychotic animal model.

Herein we report progress toward a backup clinical candidate to the M 1 positive allosteric modulator (PAM) VU319/ACP-319. Scaffold-hopping from the pyrrolo[2,3- b ]pyridine-based M 1 PAM VU6007477 to isomeric pyrrolo[3,2- b ]pyridine and thieno[3,2- b ]pyridine congeners identified several backup contenders. Ultimately, VU6007496, a pyrrolo[3,2- b ]pyridine, advanced into late stage profiling, only to be plagued with unanticipated, species-specific metabolism and active/toxic metabolites which were identified in our phenotypic seizure liability in vivo screen, preventing further development. However, VU6007496 proved to be a highly selective and CNS penetrant M 1 PAM, with minimal agonism, that displayed excellent multispecies IV/PO pharmacokinetics (PK), CNS penetration, no induction of long-term depression (or cholinergic toxicity) and robust efficacy in novel object recognition (minimum effective dose = 3 mg/kg p.o.). Thus, VU6007496 can serve as another valuable in vivo tool compound in rats and nonhuman primates, but not mouse, to study selective M 1 activation.

Acute stimulation of M 1 or M 4 muscarinic cholinergic receptors reduces cocaine abuse-related effects in mice and rats. The combined activation of these receptor subtypes produces synergistic effects on some behavioural endpoints in mice. M 1 and M 1  + M 4 receptor stimulation in a cocaine vs. food choice assay in rats and microdialysis in rats showed delayed and lasting "anticocaine effects". Here, we tested whether these putative lasting neuroplastic changes are sufficient to occlude the reinforcing effects of cocaine at the behavioural level in mice. Mice were pre-treated with the M 1 receptor partial agonist VU0364572, M 4 receptor positive allosteric modulator VU0152100, or VU0364572 + VU0152100 two weeks prior to acquisition of cocaine intravenous self-administration (IVSA). Male C57BL/6JRj mice received vehicle, VU0364572, VU0152100, or VU0364572 + VU0152100. Female mice were tested with two VU0364572 + VU0152100 dose combinations or vehicle. To attribute potential effects to either reduced rewarding effects or increased aversion to cocaine, we tested VU0364572 alone and VU0364572 + VU0152100 in acquisition of cocaine-conditioned place preference (CPP) in male mice using an unbiased design. The acquisition of cocaine IVSA was drastically reduced and/or slowed in male and female mice receiving VU0364572 + VU0152100, but not either drug alone. Food-maintained operant behaviour was unaffected, indicating that the treatment effects were cocaine-specific. No treatment altered the acquisition of cocaine-CPP, neither in the post-test, nor in a challenge 14 days later. The cocaine IVSA findings confirm unusual long-lasting "anticocaine" effects of muscarinic M 1  + M 4 receptor stimulation. Thus, in mice, simultaneous stimulation of both receptor subtypes seems to produce potential neuroplastic changes that yield lasting effects., Competing Interests: Declaration of competing interest Craig W. Lindsley and P. Jeffrey Conn are inventors on multiple pending and issued patents that protect composition of matter for M(1) and M(4) receptor ligands. All other authors declare no conflict of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

While the muscarinic acetylcholine receptor mAChR subtype 5 (M 5 ) has been studied over decades, recent findings suggest that more in-depth research is required to elucidate a thorough understanding of its physiological function related to neurological and psychiatric disorders. Our efforts to identify potent, selective, and pharmaceutically favorable next-generation M 5 antagonist tool compounds have led to the discovery of a novel triazolopyridine-based series. In particular, VU6036864 ( 45 ) showed exquisite potency (human M 5 IC 50 = 20 nM), good subtype selectivity (>500 fold selectivity against human M 1-4 ), desirable brain exposure ( K p = 0.68, K p,uu = 0.65), and high oral bioavailability (% F > 100%). VU6036864 ( 45 ) and its close analogues will support further studies of M 5 as advanced antagonist tool compounds and play an important role in the emerging biology of M 5 .

Background: Polymorphisms in the gene encoding for metabotropic glutamate receptor 3 (mGlu 3 ) are associated with an increased likelihood of schizophrenia diagnosis and can predict improvements in negative symptoms following treatment with antipsychotics. However, the mechanisms by which mGlu 3 can regulate brain circuits involved in schizophrenia pathophysiology are not clear., Methods: We employed selective pharmacological tools and a variety of approaches including whole-cell patch-clamp electrophysiology, slice optogenetics, and fiber photometry to investigate the effects of mGlu 3 activation on phencyclidine (PCP)-induced impairments in thalamo-accumbal transmission and sociability deficits. A chemogenetic approach was used to evaluate the role of thalamo-accumbal transmission in PCP-induced sociability deficits., Results: We first established that PCP treatment augmented excitatory transmission onto dopamine D 1 receptor-expressing medium spiny neurons (D1-MSNs) in the nucleus accumbens (NAc) and induced sociability deficits. Our studies revealed a selective increase in glutamatergic synaptic transmission from thalamic afferents to D1-MSNs in the NAc shell. Chemogenetic silencing of thalamo-accumbal inputs rescued PCP-induced sociability deficits. Pharmacological activation of mGlu 3 normalized PCP-induced impairments in thalamo-accumbal transmission and sociability deficits. Mechanistic studies revealed that mGlu 3 activation induced robust long-term depression at synapses from the thalamic projections onto D1-MSNs in the NAc shell., Conclusions: These data demonstrate that activation of mGlu 3 decreases thalamo-accumbal transmission and thereby rescues sociability deficits in mouse modeling schizophrenia-like symptoms. These findings provide novel insights into the NAc-specific mechanisms and suggest that agents modulating glutamatergic signaling in the NAc may provide a promising approach for treating negative symptoms in schizophrenia., (Copyright © 2023 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.)

This Letter details our efforts to develop novel tricyclic muscarinic acetylcholine receptor subtype 4 (M 4 ) positive allosteric modulator (PAM) scaffolds with improved pharmacological properties. This endeavor involved a "tie-back" strategy to replace the 3-amino-5-chloro-4,6-dimethylthieno[2,3- b ]pyridine-2-carboxamide core, which led to the discovery of two novel tricyclic cores: an 8-chloro-9-methylpyrido[3',2':4,5]thieno[3,2- d ]pyrimidin-4-amine core and 8-chloro-7,9-dimethylpyrido[3',2':4,5]furo[3,2- d ]pyrimidin-4-amine core. Both tricyclic cores displayed low nanomolar potency against human M 4 and greatly reduced cytochrome P450 inhibition when compared with parent compound ML253 ., Competing Interests: The authors declare the following competing financial interest(s): R.A.C., S.R.B., J.L.E., M.F.L., C.W.L., P.J.C., T.M.B., D.W.E., and K.J.T. are inventors on applications for composition of matter patents that protect several series of M4 positive allosteric modulators., (© 2024 The Authors. Published by American Chemical Society.)

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the progressive loss of dopaminergic neurons in the substantia nigra, resulting in motor dysfunction. Current treatments are primarily centered around enhancing dopamine signaling or providing dopamine replacement therapy and face limitations such as reduced efficacy over time and adverse side effects. To address these challenges, we identified selective dopamine receptor subtype 4 (D 4 R) antagonists not previously reported as potential adjuvants for PD management. In this study, a library screening and artificial neural network quantitative structure-activity relationship (QSAR) modeling with experimentally driven library design resulted in a class of spirocyclic compounds to identify candidate D 4 R antagonists. However, developing selective D 4 R antagonists suitable for clinical translation remains a challenge.

Here, we demonstrate a structure-based small molecule virtual screening and lead optimization pipeline using a homology model of a difficult-to-drug G-protein-coupled receptor (GPCR) target. Protease-activated receptor 4 (PAR4) is activated by thrombin cleavage, revealing a tethered ligand that activates the receptor, making PAR4 a challenging target. A virtual screen of a make-on-demand chemical library yielded a one-hit compound. From the single-hit compound, we developed a novel series of PAR4 antagonists. Subsequent lead optimization via simultaneous virtual library searches and structure-based rational design efforts led to potent antagonists of thrombin-induced activation. Interestingly, this series of antagonists was active against PAR4 activation by the native protease thrombin cleavage but not the synthetic PAR4 agonist peptide AYPGKF., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)

Sp 3 -enriched small molecules play a critical role in developing drug candidates. While designing analogues with greater sp 3 character, a methodology utilizing a less explored cyclic-aziridine amide ring-opening reaction to generate sp 3 -enriched scaffolds has been developed and reported. This methodology enables rapid access to substructures with higher fsp 3 values, attracting greater attention within the past few decades. The reaction exhibits a wide reaction scope, featuring a highly sterically hindered phenolic ether, thiophenolic ethers, protected aniline formations, and aliphatic/heteroaromatic ring-containing aziridine amides as substrates. Additionally, this reaction provides access to congested tertiary ether formations through regioselective transformation, applicable to an extensive range of drug discovery targets, construction of complex small molecules, and natural product syntheses. The scaffolds developed show improved physicochemical properties.

Vascular smooth muscle K ATP channels critically regulate blood flow and blood pressure by modulating vascular tone and therefore represent attractive drug targets for treating several cardiovascular disorders. However, the lack of potent inhibitors that can selectively inhibit Kir6.1/SUR2B (vascular K ATP ) over Kir6.2/SUR1 (pancreatic K ATP ) has eluded discovery despite decades of intensive research. We therefore screened 47,872 chemically diverse compounds for novel inhibitors of heterologously expressed Kir6.1/SUR2B channels. The most potent inhibitor identified in the screen was an N -aryl- N '-benzyl urea compound termed VU0542270. VU0542270 inhibits Kir6.1/SUR2B with an IC 50 of approximately 100 nM but has no apparent activity toward Kir6.2/SUR1 or several other members of the Kir channel family at doses up to 30 µM (>300-fold selectivity). By expressing different combinations of Kir6.1 or Kir6.2 with SUR1, SUR2A, or SUR2B, the VU0542270 binding site was localized to SUR2. Initial structure-activity relationship exploration around VU0542270 revealed basic texture related to structural elements that are required for Kir6.1/SUR2B inhibition. Analysis of the pharmacokinetic properties of VU0542270 showed that it has a short in vivo half-life due to extensive metabolism. In pressure myography experiments on isolated mouse ductus arteriosus vessels, VU0542270 induced ductus arteriosus constriction in a dose-dependent manner similar to that of the nonspecific K ATP channel inhibitor glibenclamide. The discovery of VU0542270 provides conceptual proof that SUR2-specific K ATP channel inhibitors can be developed using a molecular target-based approach and offers hope for developing cardiovascular therapeutics targeting Kir6.1/SUR2B. SIGNIFICANCE STATEMENT: Small-molecule inhibitors of vascular smooth muscle K ATP channels might represent novel therapeutics for patent ductus arteriosus, migraine headache, and sepsis; however, the lack of selective channel inhibitors has slowed progress in these therapeutic areas. Here, this study describes the discovery and characterization of the first vascular-specific K ATP channel inhibitor, VU0542270., (Copyright © 2024 by The Author(s).)

Herein, we report the synthesis and characterization of a novel set of substituted indazole-ethanamines and indazole-tetrahydropyridines as potent serotonin receptor subtype 2 (5-HT 2 ) agonists. Specifically, we examine the 5-HT 2 pharmacology of the direct indazole analogs of 5-methoxy- N , N -dimethyltryptamine (5-MeO-DMT) and related serotonergic tryptamines, and highlight the need for rigorous characterization of 5-HT 2 subtype selectivity for these analogs, particularly for the 5-HT 2B receptor subtype. Within this series, the potent analog VU6067416 ( 19d ) was optimized to have suitable preclinical pharmacokinetic properties for in vivo dosing, although potent 5-HT 2B agonist activity precluded further characterization for this series. Additionally, in silico docking studies suggest that the high potency of 19d may be a consequence of a halogen-bonding interaction with Phe234 5.38 in the 5-HT 2A orthosteric pocket., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)

M 4 muscarinic receptors are highly expressed in the striatum and cortex, brain regions that are involved in diseases such as Parkinson's disease, schizophrenia, and dystonia. Despite potential therapeutic advantages of specifically targeting the M 4 receptor, it has been historically challenging to develop highly selective ligands, resulting in undesired off-target activity at other members of the muscarinic receptor family. Recently, we have reported first-in-class, potent, and selective M 4 receptor antagonists. As an extension of that work, we now report the development and characterization of a radiolabeled M 4 receptor antagonist, [ 3 H]VU6013720, with high affinity (pK d of 9.5 ± 0.2 at rat M 4 , 9.7 at mouse M 4 , and 10 ± 0.1 at human M 4 with atropine to define nonspecific binding) and no significant binding at the other muscarinic subtypes. Binding assays using this radioligand in rodent brain tissues demonstrate loss of specific binding in Chrm4 knockout animals. Dissociation kinetics experiments with various muscarinic ligands show differential effects on the dissociation of [ 3 H]VU6013720 from M 4 receptors, suggesting a binding site that is overlapping but may be distinct from the orthosteric site. Overall, these results demonstrate that [ 3 H]VU6013720 is the first highly selective antagonist radioligand for the M 4 receptor, representing a useful tool for studying the basic biology of M 4 as well for the support of M 4 receptor-based drug discovery. SIGNIFICANCE STATEMENT: This manuscript describes the development and characterization of a novel muscarinic (M) acetylcholine subtype 4 receptor antagonist radioligand, [ 3 H]VU6013720. This ligand binds to or overlaps with the acetylcholine binding site, providing a highly selective radioligand for the M 4 receptor that can be used to quantify M 4 protein expression in vivo and probe the selective interactions of acetylcholine with M 4 versus the other members of the muscarinic receptor family., (Copyright © 2023 by The American Society for Pharmacology and Experimental Therapeutics.)

Ligands for the serotonin 2B receptor (5-HT 2B ) have shown potential to treat pulmonary arterial hypertension in preclinical models but cannot be used in humans because of predicted off-target neurological effects. The aim of this study was to develop novel systemically restricted compounds targeting 5-HT 2B . Here, we show that mice treated with VU6047534 had decreased RVSP compared with control treatment in both the prevention and intervention studies using Sugen-hypoxia. VU6047534 is a novel 5-HT 2B partial agonist that is peripherally restricted and able to both prevent and treat Sugen-hypoxia-induced pulmonary arterial hypertension. We have synthesized and characterized a structurally novel series of 5-HT 2B ligands with high potency and selectivity for the 5-HT 2B receptor subtype. Next-generation 5-HT 2B ligands with similar characteristics, and predicted to be systemically restricted in humans, are currently advancing to investigational new drug-enabling studies., Competing Interests: This work was supported by the National Institutes of Health (HL135790, HL095797, and HL087738) and Vanderbilt University Discovery Grant. Research undertaken in the Vanderbilt Mouse Neurobehavior Core was supported by the EKS NICHD of the National Institutes of Health under Award P50HD103537. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. The authors have reported that they have no relationships relevant to the contents of this paper to disclose., (© 2023 The Authors.)

The cardiotoxicity associated with des -ethyl-dexfenfluramine (norDF) and related agonists of the serotonin receptor 2B (5-HT 2B ) has solidified the receptor's place as an "antitarget" in drug discovery. Conversely, a growing body of evidence has highlighted the utility of 5-HT 2B antagonists for the treatment of pulmonary arterial hypertension (PAH), valvular heart disease (VHD), and related cardiopathies. In this Perspective, we summarize the link between the clinical failure of fenfluramine-phentermine (fen-phen) and the subsequent research on the role of 5-HT 2B in disease progression, as well as the development of drug-like and receptor subtype-selective 5-HT 2B antagonists. Such agents represent a promising class for the treatment of PAH and VHD, but their utility has been historically understudied due to the clinical disasters associated with 5-HT 2B . Herein, it is our aim to examine the current state of 5-HT 2B drug discovery, with an emphasis on the receptor's role in the central nervous system (CNS) versus the periphery.

Previously described approaches for the alkylation of N H-sulfoximines typically rely either on transition metal catalysis, or the use of traditional alkylation reagents and strong bases. Herein, we report a straightforward alkylation of diverse N H-sulfoximines under simple Mitsunobu-type conditions, despite the unusually high p K a of the N H center.

Allosteric modulation of G protein-coupled receptors (GPCRs) is a major paradigm in drug discovery. Despite decades of research, a molecular-level understanding of the general principles that govern the myriad pharmacological effects exerted by GPCR allosteric modulators remains limited. The M 4 muscarinic acetylcholine receptor (M 4 mAChR) is a validated and clinically relevant allosteric drug target for several major psychiatric and cognitive disorders. In this study, we rigorously quantified the affinity, efficacy, and magnitude of modulation of two different positive allosteric modulators, LY2033298 (LY298) and VU0467154 (VU154), combined with the endogenous agonist acetylcholine (ACh) or the high-affinity agonist iperoxo (Ipx), at the human M 4 mAChR. By determining the cryo-electron microscopy structures of the M 4 mAChR, bound to a cognate G i1 protein and in complex with ACh, Ipx, LY298-Ipx, and VU154-Ipx, and applying molecular dynamics simulations, we determine key molecular mechanisms underlying allosteric pharmacology. In addition to delineating the contribution of spatially distinct binding sites on observed pharmacology, our findings also revealed a vital role for orthosteric and allosteric ligand-receptor-transducer complex stability, mediated by conformational dynamics between these sites, in the ultimate determination of affinity, efficacy, cooperativity, probe dependence, and species variability. There results provide a holistic framework for further GPCR mechanistic studies and can aid in the discovery and design of future allosteric drugs., Competing Interests: ZV, JW, VP, JM, MB, AB, WB, GT, MY, VN, KL, Ev, EK, YL, AG, CL, AT, RD, CV, YM, DT No competing interests declared, DW, PS, AC P.M.S, D.W., and A.C. are shareholders of Septerna Inc, (© 2023, Vuckovic, Wang, Pham et al.)

Kir6.2/SUR1 is an ATP-regulated potassium channel that acts as an intracellular metabolic sensor, controlling insulin and appetite-stimulatory neuropeptides secretion. In this Letter, we present the SAR around a novel Kir6.2/SUR1 channel opener scaffold derived from an HTS screening campaign. New series of compounds with tractable SAR trends and favorable potencies are reported., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Optimization of effort-related choices is impaired in depressive disorders. Acetylcholine (ACh) and dopamine (DA) are linked to depressive disorders, and modulation of ACh tone in the ventral tegmental area (VTA) affects mood-related behavioral responses in rats. However, it is unknown if VTA ACh mediates effort-choice behaviors. Using a task of effort-choice, rats can choose to lever press on a fixed-ratio 5 (FR5) schedule for a more-preferred food or consume freely available, less-preferred food. VTA administration of physostigmine (1 μg and 2 μg/side), a cholinesterase inhibitor, reduced FR5 responding for the more-preferred food while leaving consumption of the less-preferred food intact. VTA infusion of the M5 muscarinic receptor negative allosteric modulator VU6000181 (3 μM, 10 μM, 30 μM/side) did not affect lever pressing or chow consumption. However, VU6000181 (30 μM/side) coadministration with physostigmine (2 μg/side) attenuated physostigmine-induced decrease in lever pressing in female and male rats and significantly elevated lever pressing above vehicle baseline levels in male rats. In in vivo voltammetry experiments, VTA infusion of combined physostigmine and VU6000181 did not significantly alter evoked phasic DA release in the nucleus accumbens core (NAc) in female rats. In male rats, combined VTA infusion of physostigmine and VU6000181 increased phasic evoked DA release in the NAc compared with vehicle, physostigmine, or VU6000181 infusion alone. These data indicate a critical role and potential sex differences of VTA M5 receptors in mediating VTA cholinergic effects on effort choice behavior and regulation of DA release. SIGNIFICANCE STATEMENT: Effort-choice impairments are observed in depressive disorders, which are often treatment resistant to currently available thymoleptics. The role of ventral tegmental area (VTA) acetylcholine muscarinic M5 receptors, in a preclinical model of effort-choice behavior, is examined. Using the selective negative allosteric modulator of the M5 receptor VU6000181, we show the role of VTA M5 receptors on effort-choice and regulation of dopamine release in the nucleus accumbens core. This study supports M5 receptors as therapeutic targets for depression., (Copyright © 2023 by The American Society for Pharmacology and Experimental Therapeutics.)

Antagonists of the serotonin receptor 2B (5-HT 2B ) have shown great promise as therapeutics for the treatment of pulmonary arterial hypertension, valvular heart disease, and related cardiopathies. Herein, we describe a high-throughput screen campaign that led to the identification of highly potent and selective 5-HT 2B antagonists. Furthermore, selected compounds were profiled for their predicted ability to cross the blood-brain barrier. Two exemplary compounds, VU0530244 and VU0631019, were predicted to have very limited potential for brain penetration in human subjects, a critical profile for the development of 5-HT 2B antagonists devoid of centrally-mediated adverse effects.

Recent evidence suggests that inhibition of the M 5 muscarinic acetylcholine receptor (mAChR) may provide a novel non-opioid mechanism for the treatment of opioid use disorder (OUD). Previous studies from our group and others have demonstrated that acute administration of the long-acting M 5 negative allosteric modulator (NAM) ML375 attenuates established self-administration of cocaine, ethanol, oxycodone, and remifentanil in rats. In the present study, we characterized the effects of acute and repeated administration of the novel, short-acting M 5 NAM VU6008667 on the reinforcing effects of oxycodone and reinstatement of oxycodone-seeking behaviors in male Sprague-Dawley rats, as well as on physiological withdrawal from oxycodone. Acute VU6008667 decreased oxycodone self-administration under both fixed ratio 3 (FR3) and progressive ratio (PR) schedules of reinforcement and attenuated cue-induced reinstatement of lever pressing following extinction from oxycodone self-administration, a commonly used relapse model. When administered daily to opioid-naïve rats, VU6008667 prevented acquisition of oxycodone self-administration behavior. VU6008667 had minimal effects on naloxone-precipitated withdrawal. After acute administration, VU6008667 did not inhibit sucrose self-administration and, when given chronically, delayed but did not prevent acquisition of sucrose maintained self-administration. VU6008667 also did not impact oxycodone induced anti-nociception or motor coordination, but mildly decreased novelty exploration. Finally, acute or daily VU6008667 administration did not impair cued fear conditioning. Overall, these results suggest that inhibition of the M 5 mAChR may provide a novel, non-opioid based treatment for distinct aspects of OUD by inhibiting opioid intake in established OUD, reducing relapse during abstinence, and by reducing the risk of developing OUD., Competing Interests: Declaration of competing interest C. K. J. and C. W. L. declare the following conflict of interest: Inventors on composition of matter patents that protect several series of M(5) antagonists., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Commonly known as "Quaaludes," methaqualone ( 1 ) is a sedative-hypnotic medication, with effects resembling barbiturates and other downers, that exerts its effects through modulation of γ-aminobutyric acid type A receptors (GABA A R). Following the discovery of the sedative and euphoric effects of methaqualone ( 1 ), it was quickly adopted by pharmaceutical companies and promoted by clinicians around the world as a "safe" sleeping pill option, and for a period it was available over the counter. The popularity of methaqualone ( 1 ) soared worldwide, and many people began to use it recreationally for its sedative-hypnotic-like psychoactive effects. Not long after its introduction, many individuals began to misuse the drug leading to overdoses and drug dependence which brought to light methaqualone's ( 1 ) addictive nature. In this review, the background, synthesis, pharmacology, metabolism, and pharmacokinetics of methaqualone ( 1 ) will be covered along with its discovery, history, and the derivatives that are currently available around the world through manufacture in clandestine laboratories.

Degeneration of the cholinergic basal forebrain is implicated in the development of cognitive deficits and sleep/wake architecture disturbances in mild cognitive impairment (MCI) and Alzheimer's disease (AD). Indirect-acting muscarinic cholinergic receptor agonists, such as acetylcholinesterase inhibitors (AChEIs), remain the only FDA-approved treatments for the cognitive impairments observed in AD that target the cholinergic system. Novel direct-acting muscarinic cholinergic receptor agonists also improve cognitive performance in young and aged preclinical species and are currently under clinical development for AD. However, little is known about the effects of direct-acting muscarinic cholinergic receptor agonists on disruptions of sleep/wake architecture and arousal observed in nonpathologically aged rodents, nonhuman primates, and clinical populations. The purpose of the present study was to provide the first assessment of the effects of the direct-acting M 1 /M 4 -preferring muscarinic cholinergic receptor agonist xanomeline on sleep/wake architecture and arousal in young and nonpathologically aged mice, in comparison with the AChEI donepezil, when dosed in either the active or inactive phase of the circadian cycle. Xanomeline produced a robust reversal of both wake fragmentation and disruptions in arousal when dosed in the active phase of nonpathologically aged mice. In contrast, donepezil had no effect on either age-related wake fragmentation or arousal deficits when dosed during the active phase. When dosed in the inactive phase, both xanomeline and donepezil produced increases in wake and arousal and decreases in nonrapid eye movement sleep quality and quantity in nonpathologically aged mice. Collectively, these novel findings suggest that direct-acting muscarinic cholinergic agonists such as xanomeline may provide enhanced wakefulness and arousal in nonpathological aging, MCI, and AD patient populations.

To fertilize an oocyte, the membrane potential of both mouse and human sperm must hyperpolarize (become more negative inside). Determining the molecular mechanisms underlying this hyperpolarization is vital for developing new contraceptive methods and detecting causes of idiopathic male infertility. In mouse sperm, hyperpolarization is caused by activation of the sperm-specific potassium (K + ) channel SLO3 [C. M. Santi  et al. ,  FEBS Lett. 584 , 1041-1046 (2010)]. In human sperm, it has long been unclear whether hyperpolarization depends on SLO3 or the ubiquitous K + channel SLO1 [N. Mannowetz, N. M. Naidoo, S. A. S. Choo, J. F. Smith, P. V. Lishko, Elife   2 , e01009 (2013), C. Brenker  et al. ,  Elife 3 , e01438 (2014), and S. A. Mansell, S. J. Publicover, C. L. R. Barratt, S. M. Wilson,  Mol. Hum. Reprod. 20 , 392-408 (2014)]. In this work, we identified the first selective inhibitor for human SLO3-VU0546110-and showed that it completely blocked heterologous SLO3 currents and endogenous K + currents in human sperm. This compound also prevented sperm from hyperpolarizing and undergoing hyperactivated motility and induced acrosome reaction, which are necessary to fertilize an egg. We conclude that SLO3 is the sole K + channel responsible for hyperpolarization and significantly contributes to the fertilizing ability of human sperm. Moreover, SLO3 is a good candidate for contraceptive development, and mutation of this gene is a possible cause of idiopathic male infertility.

Herein, we report on the further chemical optimization of the first reported mGlu 7 positive allosteric modulator (PAM), VU6027459. Replacement of the quinoline core by a cinnoline scaffold increased mGlu 7 PAM potency by ∼ 10-fold, and concomitant introduction of a chiral tricyclic motif led to potent mGlu 7 PAMs with enantioselective mGlu receptor selectivity profiles. Of these, VU6046980 emerged as a putative in vivo tool compound with excellent CNS penetration (K p  = 4.1; K p,uu  = 0.7) and efficacy in preclinical models. However, either off-target activity at the sigma-1 receptor or activity at a target not elucidated by large ancillary pharmacology panels led to sedation not driven by activation of mGlu 7 (validated in Grm7 knockout mice). Thus, despite a significant advance, a viable mGlu 7 PAM in vivo tool remains elusive., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

This Letter describes our ongoing effort to improve the clearance of selective M 5 antagonists. Herein, we report the replacement of the previously disclosed piperidine amide (4, disclosed in Part 1) with a pyrrolidine amide core. Several compounds within this series provided good potency, subtype selectivity, and low to moderate clearance profiles. Interestingly, the left-hand side SAR for this series diverged from our earlier efforts., Competing Interests: Declaration of Competing Interest Some authors are inventors on application for composition of matter patents that protect several series of M(5) antagonists., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The lack of potent and selective tool compounds with pharmaceutically favorable properties limits the in vivo understanding of muscarinic acetylcholine receptor subtype 5 (M 5 ) biology. Previously, we presented a highly potent and selective M 5 antagonist VU6019650 with a suboptimal clearance profile as our second-generation tool compound. Herein, we disclose our ongoing efforts to generate next-generation M 5 antagonists with improved clearance profiles. A mix and match approach between VU6019650 (lead) and VU0500325 (HTS hit) generated a piperidine amide-based novel M 5 antagonist series. Several analogs within this series, including 29f, provided good on-target potency with improved clearance profiles, though room for improvement remains., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Some authors are inventors on the application for the composition of matter patents that protect several series of M(5) antagonists., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Many dementias are propagated through the spread of "prion-like" misfolded proteins. This includes prion diseases themselves (such as Creutzfeldt-Jakob disease) and Alzheimer's disease (AD), for which no treatments are available to slow or stop progression. The M 1 acetylcholine muscarinic receptor (M 1 receptor) is abundant in the brain, and its activity promotes cognitive function in preclinical models and in patients with AD. Here, we investigated whether activation of the M 1 receptor might slow the progression of neurodegeneration associated with prion-like misfolded protein in a mouse model of prion disease. Proteomic and transcriptomic analysis of the hippocampus revealed that this model had a molecular profile that was similar to that of human neurodegenerative diseases, including AD. Chronic enhancement of the activity of the M 1 receptor with the positive allosteric modulator (PAM) VU0486846 reduced the abundance of prion-induced molecular markers of neuroinflammation and mitochondrial dysregulation in the hippocampus and normalized the abundance of those associated with neurotransmission, including synaptic and postsynaptic signaling components. PAM treatment of prion-infected mice prolonged survival and maintained cognitive function. Thus, allosteric activation of M 1 receptors may reduce the severity of neurodegenerative diseases caused by the prion-like propagation of misfolded protein.

We describe here a series of metabotropic glutamate receptor 7 (mGlu 7 ) negative allosteric modulators (NAMs) with a saturable range of activity in inhibiting responses to an orthosteric agonist in two distinct in vitro pharmacological assays. The range of inhibition among compounds in this scaffold provides highly structurally related ligands with differential degrees of receptor blockade that can be used to understand inhibitory efficacy profiles in native tissue or in vivo., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Glutamate acts at eight metabotropic glutamate (mGlu) receptor subtypes expressed in a partially overlapping fashion in distinct brain circuits. Recent evidence indicates that specific mGlu receptor protomers can heterodimerize and that these heterodimers can exhibit different pharmacology when compared to their homodimeric counterparts. Group III mGlu agonist-induced suppression of evoked excitatory potentials and induction of long-term potentiation at Schaffer collateral-CA1 (SC-CA1) synapses in the rodent hippocampus can be blocked by the selective mGlu 7 negative allosteric modulator (NAM), ADX71743. Curiously, a different mGlu 7 NAM, 6-(4-methoxyphenyl)-5-methyl-3-pyridin-4-ylisoxazonolo[4,5-c]pyridin-4(5H)-one, failed to block these responses in brain slices despite its robust activity at mGlu 7 homodimers in vitro. We hypothesized that this might result from heterodimerization of mGlu 7 with another mGlu receptor protomer and focused on mGlu 8 as a candidate given the reported effects of mGlu 8 -targeted compounds in the hippocampus. Here, we used complemented donor acceptor-resonance energy transfer to study mGlu 7/8 heterodimer activation in vitro and observed that ADX71743 blocked responses of both mGlu 7/7 homodimers and mGlu 7/8 heterodimers, whereas 6-(4-methoxyphenyl)-5-methyl-3-pyridin-4-ylisoxazonolo[4,5-c]pyridin-4(5H)-one only antagonized responses of mGlu 7/7 homodimers. Taken together with our electrophysiology observations, these results suggest that a receptor with pharmacology consistent with an mGlu 7/8 heterodimer modulates the activity of SC-CA1 synapses. Building on this hypothesis, we identified two additional structurally related mGlu 7 NAMs that also differ in their activity at mGlu 7/8 heterodimers, in a manner consistent with their ability to inhibit synaptic transmission and plasticity at SC-CA1. Thus, we propose that mGlu 7/8 heterodimers are a key molecular target for modulating the activity of hippocampal SC-CA1 synapses., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Extensive evidence supports the hypothesis that deficits in inhibitory GABA transmission in the prefrontal cortex (PFC) may drive pathophysiological changes underlying symptoms of schizophrenia that are not currently treated by available medications, including cognitive and social impairments. Recently, the mGlu 1 subtype of metabotropic glutamate (mGlu) receptor has been implicated as a novel target to restore GABAergic transmission in the PFC. A recent study reported that activation of mGlu 1 increases inhibitory transmission in the PFC through excitation of somatostatin-expressing GABAergic interneurons, implicating mGlu 1 PAMs as a potential treatment strategy for schizophrenia. Here, we leveraged positive allosteric modulators (PAMs) of mGlu 1 to examine whether mGlu 1 activation might reverse physiological effects and behavioral deficits induced by MK-801, an NMDA receptor antagonist commonly used to model cortical deficits observed in schizophrenia patients. Using ex vivo whole-cell patch-clamp electrophysiology, we found that MK-801 decreased the frequency of spontaneous inhibitory postsynaptic currents onto layer V pyramidal cells of the PFC and this cortical disinhibition was reversed by mGlu 1 activation. Furthermore, acute MK-801 treatment selectively induced inhibitory deficits onto layer V pyramidal cells that project to the basolateral amygdala, but not to the nucleus accumbens, and these deficits were restored by selective mGlu 1 activation. Importantly, the mGlu 1 PAM VU6004909 effectively reversed deficits in sociability and social novelty preference in a three-chamber assay and improved novel objection recognition following MK-801 treatment. Together, these findings provide compelling evidence that mGlu 1 PAMs could serve as a novel approach to reduce social and cognitive deficits associated with schizophrenia by enhancing inhibitory transmission in the PFC, thus providing an exciting improvement over current antipsychotic medication., (© 2022. The Author(s), under exclusive licence to American College of Neuropsychopharmacology.)