Muscarinic Research Articles

Neurochemical organization of cortical proteinopathy and neurophysiology along the Alzheimer's disease continuum.

Despite parallel research indicating amyloid-β accumulation, alterations in cortical neurophysiological signaling, and multi-system neurotransmitter disruptions in Alzheimer's disease (AD), the relationships between these phenomena remains unclear. Using magnetoencephalography, positron emission tomography, and an atlas of 19 neurotransmitters, we studied the alignment between neurophysiological alterations, amyloid-β deposition, and the neurochemical gradients of the cortex. In patients with mild cognitive impairment and AD, changes in cortical rhythms were topographically aligned with cholinergic, serotonergic, and dopaminergic systems. These alignments correlated with the severity of clinical impairments. Additionally, cortical amyloid-β plaques were preferentially deposited along neurochemical boundaries, influencing how neurophysiological alterations align with muscarinic acetylcholine receptors. Most of the amyloid-β-neurochemical and alpha-band neuro-physio-chemical alignments replicated in an independent dataset of individuals with asymptomatic amyloid-β accumulation. Our findings demonstrate that AD pathology aligns topographically with the cortical distribution of chemical neuromodulator systems and scales with clinical severity, with implications for potential pharmacotherapeutic pathways. Changes in cortical rhythms in Alzheimer's are organized along neurochemical boundaries. The strength of these alignments is related to clinical symptom severity. Deposition of amyloid-β (Aβ) is aligned with similar neurotransmitter systems. Aβ deposition mediates the alignment of beta rhythms with cholinergic systems. Most alignments replicate in participants with pre-clinical Alzheimer's pathology.

Voltage dependence of M2 muscarinic receptor antagonists and allosteric modulators

Muscarinic receptors are G protein-coupled receptors (GPCRs) that play a role in various physiological functions. Previous studies have shown that these receptors, along with other GPCRs, are voltage-sensitive; both their affinity toward agonists and their activation are regulated by membrane potential. To our knowledge, whether the effect of antagonists on these receptors is voltage-dependent has not yet been studied. In this study, we used Xenopus oocytes expressing the M2 muscarinic receptor (M2R) to investigate this question. Our results indicate that the potencies of two M2R antagonists, atropine and scopolamine, are voltage-dependent; they are more effective at resting potential than under depolarization. In contrast, the M2R antagonist AF-DX 386 did not exhibit voltage-dependent potency.Furthermore, we discovered that the voltage dependence of M2R activation by acetylcholine remains unchanged in the presence of two allosteric modulators, the negative modulator gallamine and the positive modulator LY2119620. These findings enhance our understanding of GPCRs’ voltage dependence and may have pharmacological implications.

The Role of Muscarinic Acetylcholine Receptor M3 in Cardiovascular Diseases.

The muscarinic acetylcholine receptor M3 (M3-mAChR) is involved in various physiological and pathological processes. Owing to specific cardioprotective effects, M3-mAChR is an ideal diagnostic and therapeutic biomarker for cardiovascular diseases (CVDs). Growing evidence has linked M3-mAChR to the development of multiple CVDs, in which it plays a role in cardiac protection such as anti-arrhythmia, anti-hypertrophy, and anti-fibrosis. This review summarizes M3-mAChR's expression patterns, functions, and underlying mechanisms of action in CVDs, especially in ischemia/reperfusion injury, cardiac hypertrophy, and heart failure, opening up a new research direction for the treatment of CVDs.

Identification of Psychoplastogenic Tropanes Lacking Muscarinic Activity.

Tropane-containing small molecules like scopolamine are a promising class of psychoplastogens. However, their potent antagonism of all muscarinic receptor subtypes presents the potential for undesirable anticholinergic side effects. In an effort to decouple their neuroplasticity-promoting effects from their muscarinic activity, we performed phenotypic structure-activity relationship studies across a variety of structurally distinct subclasses of tropanes. We discovered several novel tropanes capable of significantly increasing cortical neuronal growth while exhibiting drastically reduced activity at all muscarinic receptor subtypes compared to scopolamine.

Assessing the interplay between off-target promiscuity, cytotoxicity, and tolerability in rodents to improve the safety profile of novel anti-malarial plasmepsin X inhibitors.

Within drug development, high off-target promiscuity as well as potent cytotoxicity, are associated with a high attrition rate. We investigated the safety profile of novel plasmepsin X (PMX) inhibitors for the treatment of malaria. In our screening cascade, a total of 249 PMX compounds were profiled in a panel of in vitro secondary pharmacology assays containing 44 targets (SafetyScreen44 panel) and in a cytotoxicity assay in HepG2 cells using ATP as an endpoint. Six of the lead compounds were subsequently tested in a 7-d rat toxicology study, and/or in a cardiovascular study in guinea pigs. Overall, compounds with high cytotoxicity in HepG2 cells correlated with high promiscuity (off-target hit rate >20%) in the SafetyScreen44 panel and were associated with poor tolerability in vivo (decedents, morbidity, adverse clinical signs, or severe cardiovascular effects). Some side effects observed in rats or guinea pigs could putatively be linked with hits in the secondary pharmacological profiling, such as the M1 or M2 muscarinic acetylcholine receptor, opioid µ and/or κ receptors or hERG/CaV1.2/Na+ channels, which were common to >50% the compounds tested in vivo. In summary, compounds showing high cytotoxicity and high promiscuity are likely to be poorly tolerated in vivo. However, such associations do not necessarily imply a causal relationship. Identifying the targets that cause these undesirable effects is key for early safety risk assessment. A tiered approach, based on a set of in vitro assays, helps selecting the compounds with highest likelihood of success to proceed to in vivo toxicology studies.

Preclinical evaluation of [18F]SYN1 and [18F]SYN2, novel radiotracers for PET myocardial perfusion imaging

BackgroundPositron emission tomography (PET) is now an established diagnostic method for myocardial perfusion imaging (MPI) in coronary artery disease, which is the main cause of death globally. The available tracers show several limitations, therefore, the 18F-labelled tracer is in high demand nowadays. The preclinical studies on normal Wistar rats aimed to characterise two potential, novel radiotracers, [18F]SYN1 and [18F]SYN2, to evaluate which is a better candidate for PET MPI cardiotracer.ResultsThe dynamic microPET images showed rapid myocardial uptake for both tracers. However, the uptake was higher and also stable for [18F]SYN2, with an average standardized uptake value of 3.8. The biodistribution studies confirmed that [18F]SYN2 uptake in the cardiac muscle was high and stable (3.02%ID/g at 15 min and 2.79%ID/g at 6 h) compared to [18F]SYN1 (1.84%ID/g at 15 min and 0.32%ID/g at 6 h). The critical organs determined in dosimetry studies were the small intestine and the kidneys. The estimated effective dose for humans was 0.00714 mSv/MBq for [18F]SYN1 and 0.0109 mSv/MBq for [18F]SYN2. The tested dose level of 2 mg/kg was considered to be the No Observed Adverse Effect Level (NOAEL) for both candidates. The better results were achieved for [18F]SYN2, therefore, further preclinical studies were conducted only for this tracer. Radioligand binding assays showed significant responses in 3 from 68 assays: muscarinic acetylcholine M1 and M2 receptors and potassium channel hERG. The compound was mostly metabolised via an oxidative N-dealkylation, while the fluor substituent was not separated from the molecule.Conclusion[18F]SYN2 showed a favourable pharmacodynamic and pharmacokinetic profile, which enabled a clear visualization of the heart in microPET. The compound was well-tolerated in studies in normal rats with moderate radiation exposure. The results encourage further exploration of [18F]SYN2 in clinical studies.

Muscarinic and Nitric oxide Pathway Involvement in the Intestinal Transit and Gastric Emptying delay of Salvia barrelieri Methanol Extract in Mice

This study investigated the influence of Salvia barrelieri (SBA) methanol and decocted extracts (ME and DE) on intestinal transit (IT) and gastric emptying (GE) in mice. Only the doses of ME SBA induced a strong inhibition of GE at 46.82 ± 4.34, 54.71 ± 3.29 and 48.45 ± 1.33% (P≤0.0001) for the dosages 100, 200 or 400 mg·kg-1 respectively. The extracts by themselves had no effects on intestinal movement (only a slight, non–significant increase at 400 mg·kg-1). However, blocking muscarinic receptors resulted in a decrease in IT by 10.34 and 17.53% with ME and DE extracts, respectively, compared to control. Conversely, co–administration with L–arginine (Nitric Oxide donor) significantly decreased transit (47.31 and 50.80% for ME and DE, respectively), while inhibiting Nitric Oxide Synthase (NOS) with L–Nω–Nitro–Arginine (L–NNA) had a smaller effect (12.24 and 17.24% for ME and ED, respectively). Only ME SBA extracts significantly inhibited GE (46.82–54.71% decrease across doses), mimicking atropine’s effect. DE extracts and combining ME with atropine showed no significant impact. Interestingly, L–arginine only affected emptying with DE SBA (27.8% decrease), not ME SBA. Inhibiting NOS partially blocked the effect of ME SBA. These findings suggest that ME SBA extracts primarily target GE through mechanisms involving both muscarinic and NO pathways, while DE extracts have minimal effects. This study highlights the intricate interplay of pathways in gut function and the potential influence of extract type and formulation on their effectiveness.

Cholinergic modulation of interhemispheric inhibition in the mouse motor cortex.

Interhemispheric inhibition of the homotopic motor cortex is believed to be effective for accurate unilateral motor function. However, the cellular mechanisms underlying interhemispheric inhibition during unilateral motor behavior remain unclear. Furthermore, the impact of the neuromodulator acetylcholine on interhemispheric inhibition and the associated cellular mechanisms are not well understood. To address this knowledge gap, we conducted recordings of neuronal activity from the bilateral motor cortex of mice during the paw-reaching task. Subsequently, we analyzed interhemispheric spike correlation at the cell-pair level, classifying putative cell types to explore the underlying cellular circuitry mechanisms of interhemispheric inhibition. We found a cell-type pair-specific enhancement of the interhemispheric spike correlation when the mice were engaged in the reaching task. We also found that the interhemispheric spike correlation was modulated by pharmacological acetylcholine manipulation. The local field responses to contralateral excitation differed along the cortical depths, and muscarinic receptor antagonism enhanced the inhibitory component of the field response in deep layers. The muscarinic subtype M2 receptor is predominantly expressed in deep cortical neurons, including GABAergic interneurons. These results suggest that GABAergic interneurons expressing muscarinic receptors in deep layers mediate the neuromodulation of interhemispheric inhibition in the homotopic motor cortex.

Discovery of VU6008677: A Structurally Distinct Tricyclic M4 Positive Allosteric Modulator with Improved CYP450 Profile.

This Letter details our efforts to develop novel tricyclic muscarinic acetylcholine receptor subtype 4 (M4) 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 M4 and greatly reduced cytochrome P450 inhibition when compared with parent compound ML253.

Efficacy of Topical Anticholinergic in Control of Symptoms in Patients with Vasomotor Rhinitis and non-Vasomotor Rhinitis Patients and Normal Individuals. A Controlled Comparative Study

Abstract Background The use of a topical anticholinergic medication, ipratropium bromide, and its ability to inhibit methacholine and rhinitis-induced hypersecretion is emphasized. Ipratropium bromide appears to be both safe and effective in reducing this troublesome symptom. This study is designed to show the therapeutic effect of anticholinergic local treatment on nasal mucosa in patients with vasomotor rhinitis. Objective To assess the therapeutic effect of local anticholinergic treatment on nasal mucosa in patients with vasomotor rhinitis compared to non-vasomotor (allergic) patients and normal individuals, and if there any down regulation of the muscarinic receptors or not. Patients and Methods This prospective interventional study was conducted in Otorhinolaryngology Department, Faculty of Medicine, Ain Shams University. This study was conducted on 60 cases. All patients were divided into 3 groups: Study group (1): includes 20 patients diagnosed clinically non allergic rhinitis suggestive to be vasomotor rhinitis. Study group (2): includes 20 patients diagnosed as allergic rhinitis. Study group (3): includes 20 patients performing surgery for non-vasomotor rhinitis non allergic causes (as septoplasty or rhinoplasty). Results Comparison between the pre and post SNOT questionnaire of symptoms among vasomotor group showed that there was a significant difference between the pre and post treatment SNOT questionnaire symptoms; Nasal obstruction, runny nose, post nasal drip, thick nasal discharge and lack of good night sleep. Among allergic cases group, there was a significant difference between the pre and post treatment immunohistochemistry findings considering epithelium, glands, arteries and veins. 20% of cases had grade 3 epithelium before treatment while after treatment, this was dropped to 0%. Comparison between the pre and post SNOT questionnaire of symptoms among allergic group showed that there was a significant difference between the pre and post treatment SNOT questionnaire symptoms; runny nose, post nasal drip, and thick nasal discharge. Among non- vasomotor rhinitis non allergic causes (as septoplasty or rhinoplasty) group, there was a significant difference between the pre and post treatment IHC findings considering epithelium, glands, and arteries, however no significant difference between the pre and post IHC veins grade. Also, there was no significant difference between the pre and post treatment SNOT questionnaire symptoms. Conclusion Topical anticholinergic such as ipratropium bromide nasal spray are effective in treating rhinorrhea symptoms in allergic and non-allergic rhinitis. Further controlled studies with larger sample size and longer follow-up are needed to confirm our results and to identify risk factors of adverse events.

Muscarinic Receptors and Alzheimer's Disease: New Perspectives and Mechanisms.

Alzheimer's disease (AD) is one of the most prevalent neurodegenerative diseases on a global scale. Historically, this pathology has been linked to cholinergic transmission, and despite the scarcity of effective therapies, numerous alternative processes and targets have been proposed as potential avenues for comprehending this complex illness. Nevertheless, the fundamental pathophysiological mechanisms underpinning AD remain largely enigmatic, with a growing body of evidence advocating for the significance of muscarinic receptors in modulating the brain's capacity to adapt and generate new memories. This review summarizes the current state of the art in the field of muscarinic receptors' involvement in AD. A specific key factor was the relationship between comorbidity and the emergence of new mechanisms.

Antihistamines H1 as Potential Anthelmintic Agents against the Zoonotic Parasite Angiostrongylus cantonensis.

Infections caused by parasitic helminths pose significant health concerns for both humans and animals. The limited efficacy of existing drugs underscores the urgent need for novel anthelmintic agents. Given the reported potential of antihistamines against various parasites, including worms, this study conducted a screening of clinically available antihistamines against Angiostrongylus cantonensis-a nematode with widespread implications for vertebrate hosts, including humans. Twenty-one anti-H1 antihistamines were screened against first-stage larvae (L1) of A. cantonensis obtained from the feces of infected rats. Standard anthelmintic drugs ivermectin and albendazole were employed for comparative analysis. The findings revealed four active compounds (promethazine, cinnarizine, desloratadine, and rupatadine), with promethazine demonstrating the highest potency (EC50 = 31.6 μM). Additionally, morphological analysis showed that antihistamines induced significant changes in larvae. To understand the mechanism of action, antimuscarinic activities were reported based on average pK i values for human muscarinic receptor (mAChR) subtypes of the evaluated compounds. Furthermore, an analysis of the physicochemical and pharmacodynamic properties of antihistamines revealed that their anthelmintic activity does not correlate with their activity at H1 receptors. This study marks the first documentation of antihistamines' activity against A. cantonensis, offering a valuable contribution to the quest for novel agents effective against zoonotic helminths.

Postural Orthostatic Tachycardia Syndrome (POTS) of Autoimmune Origin - Case and Review

Introduction: Postural Orthostatic Tachycardia Syndrome (POTS) belongs to a group of orthostatic intolerance syndromes. It is defined by the increase in heart rate ≥30 bpm or heart rate higher than 120/min during first 10 minutes after standing, without significant decrease in blood pressure. It is a rare disease, typically affecting younger females. The dysregulation of autonomic nervous system plays a key role in the pathogenesis. The autonomic dysbalance can be caused by various neurohumoral mechanisms, receptor hypersensitivity, hypovolemia, mast cell dysregulation and last but not least, autoimmunity. The presence of autoantibodies targeted against various adrenergic and muscarinic receptors, as well, as angiotensin or endothelin receptors, has been identified in some but not all patients with POTS. Case Description: We present a case of a young patient with an overlap syndrome of POTS and vasovagal vasodepressoric syncope diagnosed with the head up tilt test, without any identified trigger. The laboratory investigation documented the presence of anti-alpha-1, beta-1 and beta-2 adrenergic receptor antibodies, antimuscarinic receptor 3 antibodies and antibodies against angiotensin II type 1 receptor. MIBG (metaiodobenzylguanidine) scan has not revealed any changes in the sympathetic inervation of the heart. The initial treatment by betablockers, as well as by ivabradine failed. On midodrine medication, the symptoms of postural tachycardia and the reflex syncope improved. Conclusion: The role of autoimmunity in POTS is currently studied, though the role of autoantibodies against autonomic nervous system in the pathogenesis of vasovagal syncope remains unclear.

Intracellular signaling pathways involved in the regulation of gene expression by pilocarpine

ObjectivesPilocarpine is commonly used clinically to treat dry mouth. The long-term administration of pilocarpine reportedly improves salivary secretion more effectively than short-term administration. Therefore, we hypothesized that pilocarpine alters gene expression in salivary glands via muscarinic receptor stimulation. This study aimed to investigate the effects of pilocarpine use on gene expression mediated by mitogen-activated protein kinase (MAPK) activity. MethodsThe effects of pilocarpine on gene expression were investigated in rats and human salivary gland (HSY) cells using several inhibitors of intracellular signaling pathways. Gene expression in the rat submandibular gland and HSY cells was determined using reverse transcription–quantitative polymerase chain reaction analysis of total RNA. ResultsIn animal experiments, at 7 days after pilocarpine stimulation, Ctgf and Sgk1 expressions were increased in the submandibular gland. In cell culture experiments, pilocarpine increased Ctgf expression in HSY cells. The mitogen-activated protein kinase kinase inhibitor trametinib, the Src inhibitor PP2, and the muscarinic acetylcholine receptor antagonist atropine suppressed the effect of pilocarpine on gene expression. ConclusionsPilocarpine enhances Ctgf and Sgk1 expressions by activating Src-mediated MAPK activity. Although further studies are required to fully understand the roles of Ctgf and Sgk1, changes in gene expression may play an important role in improving salivary secretions.

Assessment of antinociceptive property of Cynara scolymus L. and possible mechanism of action in the formalin and writhing models of nociception in mice.

Cynara scolymus has bioactive constituents and has been used for therapeutic actions. The present study was undertaken to investigate the mechanisms underlying pain-relieving effects of the hydroethanolic extract of C. scolymus (HECS). The antinociceptive activity of HECS was assessed through formalin and acetic acid-induced writhing tests at doses of 50, 100 and 200 mg/kg intraperitoneally. Additionally, naloxone (non-selective opioid receptors antagonist, 2 mg/kg), atropine (non-selective muscarinic receptors antagonist, 1 mg/kg), chlorpheniramine (histamine HH1-receptor antagonist, 20 mg/kg), cimetidine (histamine H2-receptor antagonist, 12.5 mg/kg), flumazenil (GABAA/BDZ receptor antagonist, 5 mg/kg) and cyproheptadine (serotonin receptor antagonist, 4 mg/kg) were used to determine the systis implicated in HECS-induced analgesia. Impact of HECS on locomotor activity was executed by open-field test. Determination of total phenolic content (TPC) and total flavonoid content (TFC) was done. Evaluation of antioxidant activity was conducted iploying 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging assay. HECS (50, 100 and 200 mg/kg) significantly indicated dose dependent antinociceptive activity against pain-related behavior induced by formalin and acetic acid (P < 0.001). Pretreatment with naloxone, atropine and flumazenil significantly reversed HECS-induced analgesia. Antinociceptive effect of HECS riained unaffected by chlorpheniramine, cimetidine and cyproheptadine. Locomotor activity was not affected by HECS. TPC and TFC of HECS were 59.49 ± 5.57 mgGAE/g dry extract and 93.39 ± 17.16 mgRE/g dry extract, respectively. DPPH free radical scavenging activity (IC50) of HECS was 161.32 ± 0.03 μg/mL. HECS possesses antinociceptive activity which is mediated via opioidergic, cholinergic and GABAergic pathways.

Correlation between the expression of the iNOS, caspase-3 and α-SMA genes in the parotid glands of albino rats following the administration of two antihistamines from two different generations.

Several medications, including antihistamines, can alter salivary gland function, causing dry mouth or xerostomia. Antihistamines are commonly used for treating allergic rhinitis. The aim of the present study was to compare and correlate the effects of first-generation vs. second-generation H1-antihistamines on the parotid glands of rats. Twelve adult male albino rats were used; 4 rats served as a control group (group I) and the remaining rats were divided into 2 groups: group II received promethazine hydrochloride; and group III received cetirizine dihydrochloride for 3 weeks. The parotid salivary glands were dissected, and examined histologically and analyzed histomorphometrically for the acinar area percentage. In addition, mRNA gene expression of iNOS, caspase-3 and α-SMA was assessed using quantitative realtime polymerase chain reaction (qRT-PCR). Finally, all the obtained data was statistically analyzed. Histologically, group I showed the typical architecture of the gland. In group II, degenerative changes were noticed, including acinar degeneration and shrinkage with widened connective tissue septa, intracellular vacuolization, and increased inflammatory cell infiltration. In group III, similar histological features were detected as in group II, but to a lesser extent. Histomorphometric results revealed significant differences in the acinar area percentage between various groups. In addition, qRT-PCR results showed a significant increase in iNOS expression in both groups II and III as compared to group I, caspase-3 gene expression was significantly increased in group II, while in group III, it increased non-significantly. Finally, α-SMA gene expression non-significantly decreased in both groups II and III. A significant positive correlation was observed between caspase-3 and iNOS gene expression, while an inverse correlation was noticed between caspase-3 and α-SMA gene expression. The administration of antihistamines resulted in changes in the rat salivary glands, which could be due to the induction of oxidative stress and the resultant apoptotic effect. These changes were suggested to occur mainly through action on muscarinic receptors; yet, action on histamine receptors could not be excluded. However; these effects were less marked with the second-generation antihistamine.

Co-stimulation of muscarinic M1 and M4 acetylcholine receptors prevents later cocaine reinforcement in male and female mice, but not place-conditioning

Acute stimulation of M1 or M4 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. M1 and M1 + M4 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 M1 receptor partial agonist VU0364572, M4 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 M1 + M4 receptor stimulation. Thus, in mice, simultaneous stimulation of both receptor subtypes seems to produce potential neuroplastic changes that yield lasting effects.

Allosteric modulators of M1 muscarinic receptors enhance acetylcholine efficacy and decrease locomotor activity and turning behaviors in zebrafish

Allosteric modulation of muscarinic acetylcholine receptors (mAChR) has been identified as a potential strategy for regulating cholinergic signaling in the treatment of various neurological disorders. Most positive allosteric modulators (PAMs) of mAChR enhance agonist affinity and potency, while very few PAMs (e.g., amiodarone) selectively enhance G protein coupling efficacy. The key structural features of amiodarone responsible for enhancement of mAChR efficacy were examined in CHO cells expressing M1 receptors. Subsequent incorporation of these structural features into previously identified allosteric modulators of potency (i.e., n-benzyl isatins) generated ligands that demonstrated similar or better enhancement of mAChR efficacy, lower in vivo toxicity, and higher allosteric binding affinity relative to amiodarone. Notable ligands include 8a, c which respectively demonstrated the strongest binding affinity and the most robust enhancement of mAChR efficacy as calculated from an allosteric operational model. Amiodarone derivatives and hybrid ligands were additionally screened in wildtype zebrafish (Danio rerio) to provide preliminary in vivo toxicity data as well as to observe effects on locomotor and turning behaviors relative to other mAChR PAMs. Several compounds, including 8a, c, reduced locomotor activity and increased measures of turning behaviors in zebrafish, suggesting that allosteric modulation of muscarinic receptor efficacy might be useful in the treatment of repetitive behaviors associated with autism spectrum disorder (ASD) and other neuropsychiatric disorders.

Binding of interleukin-1 receptor antagonist to cholinergic receptor muscarinic 4 promotes immunosuppression and neuroendocrine differentiation in prostate cancer

The tumor microenvironment (TME) of prostate cancer (PCa) is characterized by high levels of immunosuppressive molecules, including cytokines and chemokines. This creates a hostile immune landscape that impedes effective immune responses. The interleukin-1 (IL-1) receptor antagonist (IL1RN), a key anti-inflammatory molecule, plays a significant role in suppressing IL-1-related immune and inflammatory responses. Our research investigates the oncogenic role of IL1RN in PCa, particularly its interactions with muscarinic acetylcholine receptor 4 (CHRM4), and its involvement in driving immunosuppressive pathways and M2-like macrophage polarization within the PCa TME. We demonstrate that following androgen deprivation therapy (ADT), the IL1RN-CHRM4 interaction in PCa activates the MAPK/AKT signaling pathway. This activation upregulates the transcription factors E2F1 and MYCN, stimulating IL1RN production and creating a positive feedback loop that increases CHRM4 abundance in both PCa cells and M2-like macrophages. This ADT-driven IL1RN/CHRM4 axis significantly enhances immune checkpoint markers associated with neuroendocrine differentiation and treatment-resistant outcomes. Higher serum IL1RN levels are associated with increased disease aggressiveness and M2-like macrophage markers in advanced PCa patients. Additionally, elevated IL1RN levels correlate with better clinical outcomes following immunotherapy. Clinical correlations between IL1RN and CHRM4 expression in advanced PCa patients and neuroendocrine PCa organoid models highlight their potential as therapeutic targets. Our data suggest that targeting the IL1RN/CHRM4 signaling could be a promising strategy for managing PCa progression and enhancing treatment responses.

Ameliorative effect of vanillic acid against scopolamine-induced learning and memory impairment in rat via attenuation of oxidative stress and dysfunctional synaptic plasticity

Alzheimer’s disease (AD) is characterized by cognitive impairment, loss of learning and memory, and abnormal behaviors. Scopolamine (SCOP) is a non-selective antagonist of muscarinic acetylcholine receptors that exhibits the behavioral and molecular hallmarks of AD. Vanillic acid (VA), a phenolic compound, is obtained from the roots of a traditional plant called Angelica sinensis, and has several pharmacologic effects, including antimicrobial, anti-inflammatory, anti-angiogenic, anti-metastatic, and antioxidant properties. Nevertheless, VA’s neuroprotective potential associated with the memory has not been thoroughly investigated. Therefore, this study investigated whether VA treatment has an ameliorative effect on the learning and memory impairment induced by SCOP in rats. Behavioral experiments were utilized to assess the learning and memory performance associated with the hippocampus. Using western blotting analysis and assay kits, the neuronal damage, oxidative stress, and acetylcholinesterase activity responses of hippocampus were evaluated. Additionally, the measurement of long-term potentiation was used to determine the function of synaptic plasticity in organotypic hippocampal slice cultures. In addition, the synaptic vesicles’ density and the length and width of the postsynaptic density were evaluated using electron microscopy. Consequently, the behavioral, biochemical, electrophysiological, and ultrastructural analyses revealed that VA treatment prevents learning and memory impairments caused by SCOP in rats. The study’s findings suggest that VA has a neuroprotective effect on SCOP-induced learning and memory impairment linked to the hippocampal cholinergic system, oxidative damage, and synaptic plasticity. Therefore, VA may be a prospective therapeutic agent for treating AD.