Search Results (325)

Brain-derived neurotrophic factor (BDNF) impacts parasympathetic nervous system function by increasing the excitability of cardioinhibitory parasympathetic neurons in the brainstem, ultimately lowering heart rate (HR) and heightening resting parasympathetic tone. Yet, whether BDNF and its high-affinity receptor—tropomyosin receptor kinase B (TrkB)—also act more distally, i.e., at the level of cholinergic-sensitive intrinsic cardiac ganglia (ICGs), remains unclear. Hence, we conducted morphological and functional studies in neural crest-specific BDNF knockout mice (ncBDNF KO), a model that selectively ablates BDNF signaling in neural crest-derived autonomic structures, including the intrinsic cardiac nervous system. ncBDNF mice exhibited a significant rise in resting heart rate with unchanged baseline contractile performance, thus supporting the role of endogenous BDNF in maintaining physiological parasympathetic restraint. When examining the ICGs, immunofluorescence analysis revealed a highly compartmentalized organization, with BDNF being predominantly confined to cholinergic neuronal somata and TrkB mainly clustered instead in S100-positive satellite glial cells, thus unveiling a previously unrecognized neuron–glia BDNF/TrkB ICG pattern. Next, we directly infused BDNF in Langendorff-perfused isolated WT mouse hearts and observed a rapid and reproducible bradycardic response that was abrogated by atropine but potentiated by neostigmine, hence attesting to the cholinergic nature of such bradycardia. Of note, BDNF maintained its positive inotropic effects under muscarinic blockade, as witnessed by the enhanced left ventricular developed pressure, maximal dP/dt, and rate-pressure product, congruent with direct BDNF-evoked myocardial TrkB agonism. Thus, ICGs are additional relevant relay stations interposed between BDNF/TrkB signaling and parasympathetic modulation of heart function. Although through different molecular paths, BDNF-mediated modulation of ICG firing can coordinate with the previously reported BDNF positive inotropy/lusitropy to adapt cardiac performance to increased workload and/or stress conditions. Full article

Background and Aims: Postoperative ileus (POI) is a motility dysfunction associated with inflammation in the gastrointestinal (GI) tract after abdominal surgery. Management relies on supportive care, as there is no effective medical treatment. Clinical trials found that coffee consumption improves bowel movement and shortens hospital stays in POI. We aimed to investigate the mechanisms underlying the beneficial effect of coffee in an animal model of POI. Methods: Mouse POI was established by manipulation of the small intestine with wet swab applicators for 5 min. Mice were then treated with water, regular or decaffeinated coffee (10 mg per day) by oral gavage and euthanized 24 h after the operation. Key Results: Intestinal manipulation slowed GI transit rate from 5.64 ± 0.47 to 3.77 ± 0.16 (N = 6, measured by geometric center), reduced intestinal muscle contractility, and induced an acute inflammatory response with increased expression of proinflammatory mediators such as IL-6, IL-1, CCL2, and CXCL-1 in the POI intestine. Coffee treatment (regular or decaffeinated) did not reduce inflammation or the expression of inflammatory mediators but significantly improved muscle contractility and increased GI transit to 4.59 ± 0.31 (N = 5) and 4.80 ± 0.29 (N = 5) in POI mice (regular and decaffeinated coffee, respectively). Regular or decaffeinated coffee dose-dependently (0.1–10 mg/mL) increased contractility of intestinal muscle strips. The contractile effect was not affected by neural toxin tetrodotoxin (10⁻⁶ M) or cholinergic nicotinic antagonist hexamethonium (10⁻⁴ M) but was completely abolished by muscarinic receptor antagonist atropine (10⁻⁶ M). Conclusions: Coffee consumption does not attenuate inflammatory response but improves GI motor function and stimulates intestinal smooth muscle contractions in the mouse model of POI. Coffee stimulates contractions in a caffeine-independent manner through a cholinergic muscarinic receptor-dependent mechanism. Full article

Background/Objectives: Understanding the unique properties of human neurons is crucial for advancing knowledge of brain function and facilitating clinical translation. This study explores the use of photopharmacology, specifically a photoswitchable ligand, Phthalimide-Azo-Iperoxo (PAI), to modulate cortical activity in human brain slices. Methods: Human cortical tissue was obtained from patients undergoing resective neurosurgery for pharmacoresistant epilepsy. The inactive cis isomer of PAI (200 nM) was bath-applied to brain slices exhibiting slow-oscillatory activity, producing no effect on the ongoing network activity. Subsequent illumination of the slices with white light induced photoconversion to trans-PAI, the active form of the compound. Results: Photoactivation of PAI selectively activated M₂ muscarinic acetylcholine receptors, resulting in a significant increase in oscillatory frequency accompanied by reductions in both Up-state and Down-state durations. Conclusions: In this proof-of-concept study, our findings provide preliminary evidence that photopharmacology can selectively modulate slow oscillations in human cortical circuits, highlighting its potential as a tool for investigating human cortical dynamics. By extending observations previously made in animal models to human tissue, this work establishes the feasibility of photopharmacological modulation in human cortical tissue and provides a foundation for future translational research. Full article

Background/Objectives: Opioids inhibit breathing that can lead to fatal overdose, highlighting the need for testing effective countermeasure agents and potential mechanisms of action. Here we examined the role muscarinic cholinergic receptors play in the ability of L-cysteine ethyl ester (L-CYSee) to overcome the deleterious effects of morphine on ventilatory parameters in male Sprague Dawley rats and the ventilatory responses during a subsequent hypoxic-hypercapnic (HH) challenge. Methods: Ventilatory parameters were measured by whole body plethysmography. Results: The injection of the muscarinic receptor antagonist, atropine (1.0 mg/kg, IV), elicited an array of ventilatory responses (e.g., an increase in frequency of breathing coupled with a fall in tidal volume). Injection of morphine (10 mg/kg, IV) to vehicle-treated rats elicited a depression of breathing, including sustained decreases in tidal volume, minute ventilation, peak inspiratory flow, and peak inspiratory and expiratory drives, which were associated with marked increases in end inspiratory pause (EIP) and end expiratory pause (EEP), expiratory flow at 50% expired tidal volume (EF₅₀), and rate of achieving peak expiratory flow (Rpef). Most effects of morphine (10 mg/kg, IV) were not altered in atropine-treated rats, except that increases in EIP, EEP and Rpef were reduced. Subsequent injections of L-CYSee (2 × 500 μmol/kg, IV given 15 min apart) overcame the adverse actions of morphine on ventilatory parameters in vehicle-treated rats. The effects of L-CYSee, such as reversal of the effects of morphine on frequency of breathing, tidal volume and minute ventilation, were markedly reduced in atropine-treated rats. The ability of L-CYSee to reverse the adverse effects of morphine to a HH gas challenge was markedly diminished in atropine-treated rats. Conclusions: These findings demonstrate that muscarinic receptors play a vital role in the ability of L-CYSee to overcome the deleterious effects of morphine. Full article

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy that requires novel therapeutic targets and selective repurposed drugs with low systemic toxicity. Here, we evaluated the clinically approved M3 muscarinic receptor antagonist, Darifenacin (DF), against Jurkat and MOLT-4 T-ALL cells, as well as healthy human T lymphocytes, under identical conditions. DF induced selective, concentration-dependent cytotoxicity with IC₅₀ values of 26.7 ± 1.07 µM (Jurkat) and 30.5 ± 1.54 µM (MOLT-4), whereas healthy T lymphocytes exhibited an apparent IC₅₀ of 197.9 ± 1.29 µM, corresponding to a 6.5–7.4-fold therapeutic window. Mechanistically, DF decreased M3 receptor and choline acetyltransferase expression and increased acetylcholinesterase activity. Convergent multi-assay validation confirmed that this cytotoxicity was driven by activation of the intrinsic mitochondrial apoptotic pathway, as evidenced by increased Bax, decreased Bcl-2, and cleavage of the executioner caspase-3. Furthermore, DF induced glycative stress through the accumulation of methylglyoxal (MG) and advanced glycation end products (AGEs), and selectively inhibited lysosomal Cathepsins B and C. Molecular docking predicted highly favorable molecular binding within the catalytic cavities of these proteases. These findings suggest that DF exhibits selective antileukemic activity by simultaneously disrupting cholinergic signaling, inducing glycative stress, inhibiting cathepsins, and triggering apoptosis. Thus, DF emerges as a promising candidate for multi-target drug repurposing in T-ALL therapy. Full article

The structural hybridization of bioactive quinones is a promising strategy for generating pharmacologically active compounds through sustainable synthetic approaches. This study aimed to synthesize and evaluate a dapsone-derived phenylaminojuglone as a potential intestinal smooth muscle relaxant. Juglone (AJ) was functionalized with dapsone (D) via a solvent-free mechanochemical aza-Michael reaction using silica gel as a mild acid catalyst. The resulting compound (AJ-D) was characterized and evaluated in isolated rat ileum preparations. Pharmacological studies were complemented by molecular docking, density functional theory (DFT) calculations, and in silico ADMET predictions. AJ-D was obtained with complete regioselectivity at the C-3 position and required shorter reaction times than conventional solution-based methods. The compound exhibited significant spasmolytic and antispasmodic effects under basal conditions and against acetylcholine- and KCl-induced contractions. Its relaxant activity was not significantly affected by muscarinic receptor blockade or K⁺ channel inhibition, whereas verapamil reduced its potency. Calcium reintroduction experiments suggested the involvement of extracellular Ca²⁺ influx pathways. Docking studies suggested favorable interactions with the Ca_V1.2 L-type calcium channel, whereas DFT and ADMET analyses indicated suitable electronic and drug-like properties. AJ-D is a promising juglone-derived scaffold with antispasmodic activity, likely associated with the modulation of extracellular calcium influx pathways involved in intestinal smooth muscle contraction. Full article

Background/Objectives: We examined the role of muscarinic cholinergic receptors in the ventilatory depressant effects of morphine in male Sprague Dawley rats, and in responses elicited by a hypoxic-hypercapnic gas (HH) challenge and upon return to room air. Methods: Ventilatory parameters were measured in freely moving rats by whole body plethysmography that continuously recorded these parameters. Results: The injection of morphine (10 mg/kg, IV) elicited a relatively pronounced depression of breathing, including sustained decreases in tidal volume, minute ventilation, peak inspiratory flow, and inspiratory and expiratory drives that were associated with marked increases in end inspiratory pause (EIP), expiratory flow at 50% expired tidal volume (EF₅₀), and the rate of achieving peak expiratory flow (Rpef). Subsequent injection of the muscarinic receptor antagonist, atropine (1 mg/kg, IV), in morphine-treated rats did not affect the frequency of breathing, tidal volume, minute ventilation, inspiratory and expiratory times, end expiratory pause, peak inspiratory and expiratory flows, relaxation time, expiratory delay, apneic pause, inspiratory and expiratory drives, and the non-eupneic breathing index (p > 0.05 for all comparisons). In contrast, atropine dramatically reduced the morphine-induced increases in EIP, Rpef and EF₅₀ (p < 0.05, for all comparisons). In addition, the changes in many ventilatory parameters (e.g., frequency of breathing, tidal volume and minute ventilation) that occurred during a subsequent HH-challenge and upon the return to room air were markedly reduced in atropine-treated rats (p < 0.05, for all comparisons). Conclusions: These findings suggest that muscarinic receptors play a limited, but important role, in the expression of the ventilatory effects of morphine in male rats and a substantial role in the expression of responses elicited during and following HH-challenge. Full article

Cantharidin (CANT) and sodium fluoride inhibit serine/threonine phosphatase (PP) 1 and PP2A in the heart. We hypothesized that Cant and sodium fluoride could weaken the reduction in force by carbachol in the presence of serotonin in human right atrial preparations (HAP). We measured contractility in HAP, and for comparison, in left atria from wild type mice (CD1, WT), mice with human 5-HT₄-serotonin receptor cardiac overexpression (5-HT₄-TG), or in mice with human H₂-histamine receptor cardiac overexpression (H₂-TG). Isoprenaline (1 µM), histamine (1 µM), and serotonin (1 µM) raised contraction in HAP. Likewise, isoprenaline (1 µM), histamine (1 µM), and serotonin (1 µM) raised contractility in the left atria of WT, H₂-TG, and 5-HT₄-TG. Carbachol (1 µM), a muscarinic cholinoceptor agonist, diminished the force of contraction in a time-dependent manner after prior stimulation of force by serotonin, isoprenaline, or histamine in HAP but also in left atria from 5-HT₄-TG, WT, and H₂-TG. These anti-β-adrenergic, anti-serotoninergic and anti-histaminergic effects of carbachol on force of contraction were attenuated by 100 µM or 30 µM cantharidin or 3 mM sodium fluoride in HAP and in left atrial preparations from WT, H₂-TG, and 5-HT₄-TG, respectively. We conclude that muscarinic cholinoceptor activation may exert a negative inotropic effect in the HAP by stimulating PP1 and/or PP2A, which had previously been inhibited by isoprenaline or histamine or serotonin. Full article

Background: Bladder dysfunction is a complicated condition that substantially impairs quality of life for both men and women. Due to the adverse effects and limited efficacy of current therapies, new strategies must be rapidly developed. Female bladder dysfunction arises from multifaceted etiologies distinct from the predominantly male benign prostatic hyperplasia (BPH) that is the focus of existing drug development. In this study, we investigated the therapeutic potential of Quisqualis indica extract (QIE), a traditional medicinal herb that attenuates BPH-induced lower urinary symptoms (LUTS), to elucidate its underlying mechanisms in a female bladder dysfunction model. Methods and Results: A bladder dysfunction model was established by inducing partial bladder outlet obstruction (pBOO) in female Sprague Dawley rats, followed by the oral administration of QIE for 7 weeks. Voiding pattern analysis and cystometry were conducted to evaluate indicators such as voiding frequency, voiding volume, and intravesical pressure. Histological analysis of excised bladder tissue quantified smooth muscle hypertrophy and collagen deposition. Gene expression profiling of inflammatory cytokines and fibrosis-related markers within the bladder tissue was performed to assess tissue remodeling. Furthermore, pharmacological contraction studies examined the direct effects of QIE on detrusor muscle responsiveness to muscarinic and purinergic agonists. QIE administration significantly improved the elevated voiding pressure and abnormal inter-contraction intervals observed in the pBOO rats, restoring normal voiding patterns. Histological examination revealed a marked decrease in muscle hypertrophy and collagen deposition. Expression levels of pro-inflammatory cytokines (TNFα, IL-1β) and fibrosis-associated genes (TGF-β, α-SMA) were downregulated. Pharmacological contraction assays demonstrated that QIE attenuated the hypercontractile response of bladder smooth muscle to a muscarinic agonist, with concurrent reduced expression of muscarinic receptors (M₂, M₃) at the mRNA level. Conclusions:QIE ameliorates key aspects of bladder dysfunction, voiding abnormalities, inflammation, fibrosis, and hypercontractility by modulating muscarinic receptor signaling and fibrotic pathways. This study suggests that QIE warrants further investigation as a natural product-based therapeutic candidate for female bladder dysfunction. Full article

Little is known about how nicotinic signalling in human astrocytes may contribute to the functional neurotoxicity of compounds related to tobacco alkaloids and neonicotinoid pesticides. We generated a single-cell Ca²⁺-imaging assay in induced pluripotent stem cell (iPSC)-derived astrocytes, and profiled functional expressions of some neurotoxicologically relevant receptors. Responses to pharmacological tool compounds indicated the expression of nicotinic, muscarinic, purinergic, glutamatergic receptors and voltage-gated Na⁺/Ca²⁺ channels. Closer investigation of the nicotinic system, e.g., using the alpha7 nicotinic acetylcholine receptor (nAChR)-selective positive allosteric modulator PNU-120596 and alpha7-preferring agonist (AR-R17779) demonstrated that Ca²⁺ signals elicited by nicotine and neonicotinoids are dominated by alpha7 nAChRs and depend on the downstream activation of L-type Ca²⁺ channels and tetrodotoxin-sensitive Na⁺ channels. Crosstalk of nAChR activation/desensitization was not observed for the inflammatory response elicited by TNF or for activation of glutamatergic or purinergic signalling. However, pre-stimulation of nAChR by neonicotinoids significantly blunted the response to the neurotransmitter acetylcholine. Comparative experiments in the human neuronal cultures (LUHMES cells) revealed similar potency ranges and pharmacological fingerprints for several neonicotinoids and their human-relevant metabolites descyanothiacloprid and desnitroimidacloprid. The pesticide metabolites showed a high potency, compared with their respective parent compounds. After this basic system characterization, the hitherto data-poor pesticides cycloxaprid and flupyradifurone were comparatively profiled in astrocytic and neuronal test systems. They showed the typical features of alpha7 nAChR agonists. The disruption of cholinergic signalling in astrocytes suggests that neonicotinoids affect not only neurons in human brains. Therefore, future neurotoxicity screening approaches may need to consider astrocyte toxicity. Full article

Schizophrenia remains a debilitating global health challenge where pharmacologic treatment has been stagnant for over seventy years, relying almost exclusively on the blockade of dopamine receptors. While this mechanism controls positive psychosis, it frequently fails to address negative symptoms or cognitive impairment and carries a significant burden of metabolic and motor adverse effects. This review evaluates the scientific and clinical validation of xanomeline–trospium (Cobenfy^®; investigational name KarXT), the first approved antipsychotic with a completely non-dopaminergic mechanism of action. We synthesize data ranging from the unique structural biology of the bitopic M1/M4 muscarinic receptor agonist xanomeline to the pharmacokinetic innovation of using the peripheral antagonist trospium chloride to mitigate systemic toxicity. Comprehensive analyses of the EMERGENT clinical trial program demonstrate that this combination significantly reduces heterogeneous schizophrenia symptoms with a safety profile distinct from current standards of care, specifically avoiding weight gain and extrapyramidal movement disorders. Furthermore, we contrast this success with the recent failures of other novel mechanisms and explore the potential for precision medicine through the identification of muscarinic receptor deficit biotypes. We conclude that M1/M4 muscarinic receptor agonism represents an important advance toward circuit-based therapeutics that may help overcome some of the limitations inherent to dopamine-centered pharmacotherapy. Full article

We analyze data indicating that a set of currently marketed FDA/EMA-approved drugs used to treat parkinsonism, extrapyramidal side effects of antipsychotic drugs, or overactive bladder may have the potential to slow the growth of glioblastoma; diffuse midline glioma, H3K27-altered (DMG); and a particular form of DMG growing in the pons of children, diffuse intrinsic pontine glioma (DIPG). These gliomas are typically associated with poor prognosis. Clinical trials evaluating conventional chemotherapeutic drugs have failed to improve DIPG survival. Our analysis of the biochemistry and physiology of DMG and DIPG concludes that neuronal acetylcholinergic agonisms at muscarinic receptors M1 and M3 on primitive oligodendrocyte precursor cells (OPCs) are trophic, growth-stimulating factors in DMG/DIPG growth. A set of muscarinic receptor inhibitors—benztropine, biperiden, and trihexyphenidyl—is used clinically to treat Parkinson’s disease or the parkinsonian side effects from antipsychotic medicines. Another muscarinic inhibitor, oxybutynin, is used to treat overactive bladder. All four drugs may impose dose-related side effects inherent to muscarinic receptor inhibition, such as xerostomia, asthenia, and mild cognitive impairment. We recount the evidence for the inhibition of OPC proliferation and migration mediated by these four M1/M3 inhibitors and report details on the rationale for selecting oxybutynin as the primary candidate for adjuvant therapy in DMG/DIPG. We chose oxybutynin as the first choice to study in DMG and DIPG compared to other antimuscarinic drugs based on its (i) high brain-tissue concentration, (ii) relatively stronger M3 inhibition, (iii) lower side-effect propensity than scopolamine, (iv) wide availability, and (v) the absence of H1 antihistamine or dopaminergic effects. Given the rapidly fatal nature of DMG and DIPG, the potential of oxybutynin for growth slowing may outweigh the associated risks and mild side-effect burdens. Full article

Positron emission tomography (PET) of muscarinic acetylcholine receptors has evolved from a receptor-mapping exercise into a potential translational tool for probing cholinergic dysfunction in neurodegenerative disease. However, the field still lacks a clear hierarchy of clinical value across receptor subtypes, tracers, and quantitative analysis strategies, including acquisition protocols and kinetic modeling approaches. In the M₂ arena, [¹⁸F]FP-TZTP has been associated with higher distribution volumes in older APOE-ε4 carriers, but the biological meaning of this signal remains uncertain in the absence of longitudinal conversion data and with quantification approaches that are difficult to implement routinely. Among M₄ ligands, [¹¹C]MK-6884 is the most advanced and reproducible tracer to date, yet its clinical evidence still rests on proof-of-concept studies, mainly in moderate-to-severe Alzheimer’s disease, with important methodological limitations. In our view, muscarinic PET should now be reframed less as a stand-alone diagnostic biomarker and more as a platform for mechanistic and pharmacodynamic studies, especially for the development and monitoring of muscarinic-positive allosteric modulators. Future progress will depend on longitudinal multicenter validation, simplified quantification pipelines, and next-generation PET systems capable of capturing tracer kinetics more efficiently. Full article

PVP-stabilized silver nanoparticles (Ag NPs) were functionalized with sildenafil (Sil), leading to spherical NPs (Ag@Sil NPs) with a size of about 30 nm as observed through transmission electron microscopy and dynamic light scattering. Fourier-transformed IR spectroscopy confirmed the covering of the particles with Sil. The Ag@Sil NPs were incorporated into a 0.1 wt% ointment and tested for the treatment of acute anal fissures in a preliminary medical study involving 50 patients. Typical symptoms such as pain, bleeding, itching, and mass sensation were improved in the intervention group with no adverse effects. Molecular docking showed strong interactions with docking scores slightly above −10 kcal/mol between sildenafil and two different model complexes [Ag–Sil]⁺ for the Ag-bound sildenafil with either piperazine-N- or pyrazole-N-bound Ag⁺ ions and the muscarinic M2 and the nicotinic acetylcholine α3β4 receptor, which are both involved in anal sphincter regulation. All three showed superior binding compared with nitroglycerin and L-arginine. The residue analysis revealed a higher number of relevant interactions for the sildenafil and the two Ag⁺ complexes, compared to nitroglycerin and L-arginine, fully in line with the differences in the docking scores. Full article

Asthma is a chronic airway disease characterized by inflammation, bronchial hyperresponsiveness, and airflow limitation, highlighting the need for novel bronchodilator agents. Cryptotanshinone (CT), a bioactive diterpenoid derived from Salvia miltiorrhiza, exhibits anti-inflammatory and vasodilatory properties; however, its direct effects on airway smooth muscle remain poorly characterized. This study investigated the bronchodilatory activity of CT and its pharmacological mechanisms. Molecular docking was performed to evaluate potential interactions with M₃ muscarinic receptors and L-type calcium channels. Functional experiments were conducted using isolated guinea pig tracheal smooth muscle preparations. The relaxant effects of CT (10⁻⁷–3 × 10⁻⁴ M) were evaluated against carbachol (1 µM)- and high-K⁺ (80 mM)-induced contractions. Docking predicted favorable binding of CT to the M₃ receptor and L-type Ca²⁺ channel, with binding energies of −9.854 and −9.951 kcal/mol, respectively. In vitro, CT produced concentration-dependent relaxation of CCh-induced contractions, reaching a maximal effect of 41.9 ± 2.58% at 3 × 10⁻⁴ M (pEC₅₀ = 4.60). CT produced minimal relaxation in high-K⁺-induced contractions, suggesting receptor-mediated rather than non-selective smooth muscle inhibition. CT also produced a parallel rightward shift of the CCh concentration–response curve at 10⁻⁵ M, whereas a higher concentration (10⁻⁴ M) altered the maximal contractile response, suggesting concentration-dependent pharmacological effects. Pharmacological inhibition studies indicated the involvement of muscarinic receptor-mediated mechanisms, with additional contributions from calcium channel-related mechanisms and partial involvement of the NO/cGMP pathway, while β₂-adrenergic signaling and potassium channels were not significantly involved. These findings suggest that CT exerts bronchodilatory effects through the involvement of multiple pharmacological pathways relevant to airway smooth muscle regulation and provide preliminary mechanistic evidence supporting further investigation. Full article