Muscarinic Research Articles

Effects of Different Photoperiods on Growth Performance, Glucose Metabolism, Acetylcholine, and Its Relative Acetylcholine Receptor Modulation in Broiler Chickens.

Photoperiods are crucial environmental factors in the growth and health of modern intensive broiler chicken production. To date, the effects of different photoperiods on glucose metabolism, acetylcholine (ACh), and its relative acetylcholine receptor modulation in broilers remain elusive. Herein, we aimed to identify the effects of different photoperiods on regulating glucose metabolism, ACh, nicotinic acetylcholine receptor alpha 4 (α4 nAChR) mRNA, and M3 muscarinic acetylcholine receptor (M3 mAChR) modulation in broilers. A total of 216 healthy 5-day-old Arbor Acres (AA) male broilers was randomly assigned to 12L:12D, 18L:6D, and 24L:0D photoperiods for 4 weeks. The results show that, compared with the 12L:12D photoperiod, the 18L:6D and 24L:0D photoperiods significantly increase the average daily gain (ADG) and average daily feed intake (ADFI) of broilers (p < 0.05). However, the feed efficiency (FE) of broilers significantly decreased in the 18L:6D and 24L:0D photoperiods (p < 0.05). Moreover, compared with the 12L:12D photoperiod, the ACh concentrations and α4 nAChR mRNA expression levels in the hypothalamus and medulla oblongata of broilers significantly increased (p < 0.05); M3 mAChR mRNA expression levels in cecum significantly reduced in the 18L:6D photoperiod and the 24L:0D photoperiod (p < 0.05). Compared with the 12L:12D photoperiod, the serum glucose (GLU), serum insulin (INS), serum triglyceride (TG) levels, and homeostasis model assessment of insulin resistance (HOMA-IR) of broilers significantly enhanced in the 18L:6D and 24L:0D photoperiods (p < 0.05). Our results indicate that extending the photoperiod can promote the growth rate, ACh expression, and α4 nAChR mRNA expression of broilers while reducing the feed efficiency, inhibiting M3 mAChR mRNA expression, and inducing glucose metabolism disorders in broilers.

Involvement of Cholinergic and Cyclooxygenase Pathways in the Diuretic Effects of Rosmarinic Acid.

Rosmarinic acid (RA) is a natural antioxidant known for its diverse biological activities. Although its diuretic activity has been previously established, the mode of action remained unclear. To investigate this, we examined the diuretic activity of RA alone and in combination with hydrochlorothiazide (HCTZ), amiloride (AML), atropine (ATR), and indomethacin (INDO) to see if any of these drugs could interfere with RA's activity in an 8-hour acute diuresis animal model. We observed that RA increases urine excretion and may have a synergistic effect in the HCTZ+RA group, with a potassium-sparing capacity. In the AML+RA group, we also noted increased urine excretion while sodium and potassium excretion decreased. ATR and INDO prevented RA from increasing urine excretion, suggesting a potential interaction with muscarinic receptors or a role in promoting prostaglandin production. Additionally, molecular docking analysis indicated possible interactions with key receptors involved in increased diuresis or free-electrolyte water excretion.

Long-Term Neurotoxic Effects and Alzheimer's Disease Risk of Early EHDPP Exposure in Zebrafish: Insights from Molecular Mechanisms to Adult Pathology.

2-Ethylhexyl diphenyl phosphate (EHDPP), ubiquitously monitored in environmental media, is highly bioaccumulative and may pose long-term risks, even after short-term exposure. In this investigation, larval zebrafish were exposed to 0.05, 0.5, and 5.0 μg/L EHDPP from 4 to 120 h postfertilization (hpf) to examine the long-term neurotoxicity effects of early exposure. Exposure to 5.0 μg/L EHDPP yielded hyperactive locomotor behavior, which was characterized by increased swimming speed, larger turning angles, and heightened sensitivity to light-dark stimulation. The predicted targets of EHDPP (top 100 potential macromolecules) were primarily associated with brain diseases like Alzheimer's disease (AD). Comparisons of differentially expressed genes (DEGs) from AD patients (GSE48350) and RNA-seq data from EHDPP-exposed zebrafish confirmed consistently abnormal regulatory pathways. EHDPP's interaction with M1 and M5 muscarinic acetylcholine receptors likely disrupted calcium homeostasis, leading to mitochondrial dysfunction and neurotransmitter imbalance as well as abnormal locomotor behavior. Especially, 5.0 μg/L EHDPP exposure during early development (4-120 hpf) triggered early- and midstage AD-like symptoms in adulthood (180 dpf), characterized by cognitive confusion, aggression, blood-brain barrier disruption, and mitochondrial damage in brains. These findings provide deep insights into the long-term neurotoxicity effects and Alzheimer's disease risks of early EHDPP exposure at extremely low dosages.

Berberine and Berberine Nanoparticles Mitigate Scopolamine-Induced Amnesia in Rats

Abstract Introduction/Objective Berberine (BER) has garnered attention in research for its multifaceted biochemical and pharmacological properties, positioning it as a promising natural remedy. Conversely, scopolamine (SCO) functions as a non-selective muscarinic receptor antagonist, disrupting central cholinergic neurotransmission and impairing learning and short-term memory. Methods/Case Report Forty-two male Wistar rats were procured from the National Research Center in Cairo, Egypt, and allocated randomly into seven groups, each comprising six rats. The groups were as follows: Sham Control (no treatment), BSA Control (received oral administration of 1 ml Bovine Serum Albumin-NP solution daily for 28 days), BRB-NP Control (received oral administration of BRB-NP nanoparticles at 5 mg/kg/day for 28 days), Induced Group (intraperitoneal injection of SCO at 2 mg/kg dissolved in saline once daily for 28 days), BER Treated Group (received BER at 50 mg/kg suspended in saline followed by intraperitoneal injection of SCO at 2 mg/kg after 40 minutes daily for 28 days), BSA Treated Group (received oral administration of 1 ml BSA-NP solution followed by intraperitoneal injection of SCO at 2 mg/kg after 40 minutes daily for 28 days), and BER-NP Treated Group (received oral administration of BER-NP at 5 mg/kg followed by intraperitoneal injection of SCO at 2 mg/kg after 40 minutes daily for 28 days). Whole blood was collected via heart puncture, and brain tissue was promptly extracted for brain homogenate preparation. Routine biochemical assays were conducted on serum and brain samples. Results (if a Case Study enter NA) Statistically significant differences were observed among the seven experimental rat groups concerning plasma AST, albumin, total protein, and triglyceride levels, while differences in ALT and total cholesterol were non-significant. Regarding brain parameters, significant differences were noted among groups in total protein, cholesterol, phospholipids, and uric acid levels. Conclusion Treatment with BRB NPs or free berberine led to a progressive increase in brain protein levels and a decrease in cholesterol content (p&amp;lt;0.05). Additionally, SCO administration resulted in a gradual reduction in brain phospholipid content, while treatment with BER NPs, regardless of the rat’s condition, exhibited the highest phospholipid levels (p&amp;lt;0.05). Furthermore, administration of BSA or free BRB improved brain uric acid levels (p&amp;lt;0.05).

Spasmolytic Activity and Anti-Inflammatory Effect of Novel Mebeverine Derivatives.

Background: Irritable bowel syndrome (IBS) has a major negative influence on quality of life, causing cramps, stomach pain, bloating, constipation, etc. Antispasmodics have varying degrees of efficacy. Mebeverine, for example, works by controlling bowel movements and relaxing the muscles of the intestines but has side effects. Therefore, more efficient medication is required. Methods: In the current study, we investigated the synthesis of novel mebeverine analogs and determined ex vivo their spasmolytic and in vitro and ex vivo anti-inflammatory properties. The ability to influence both contractility and inflammation provides a dual-action approach, offering a comprehensive solution for the prevention and treatment of both conditions. Results: The results showed that all the compounds have better spasmolytic activity than mebeverine and good anti-inflammatory potential. Among the tested compounds, 3, 4a, and 4b have been pointed out as the most active in all the studies conducted. To understand their mechanism of activity, molecular docking simulation was investigated. The docking analysis explained the biological activities with their calculated Gibbs energies and possibilities for binding both centers of albumin. Moreover, the calculations showed that molecules can bind also the two muscarinic receptors and interleukin-β, hence these structures would exert a positive therapeutic effect owed to interaction with these specific receptors/cytokine. Conclusions: Three of the tested compounds have emerged as the most active and effective in all the studies conducted. Future in vivo and preclinical experiments will contribute to the establishment of these novel mebeverine derivatives as potential drug candidates against inflammatory diseases in the gastrointestinal tract.

Exploring the Benzazoles Derivatives as Pharmacophores for AChE, BACE1, and as Anti-Aβ Aggregation to Find Multitarget Compounds against Alzheimer’s Disease

Despite the great effort that has gone into developing new molecules as multitarget compounds to treat Alzheimer’s disease (AD), none of these have been approved to treat this disease. Therefore, it will be interesting to determine whether benzazoles such as benzimidazole, benzoxazole, and benzothiazole, employed as pharmacophores, could act as multitarget drugs. AD is a multifactorial disease in which several pharmacological targets have been identified—some are involved with amyloid beta (Aβ) production, such as beta secretase (BACE1) and beta amyloid aggregation, while others are involved with the cholinergic system as acetylcholinesterase (AChE) and butirylcholinesterase (BChE) and nicotinic and muscarinic receptors, as well as the hyperphosphorylation of microtubule-associated protein (tau). In this review, we describe the in silico and in vitro evaluation of benzazoles on three important targets in AD: AChE, BACE1, and Aβ. Benzothiazoles and benzimidazoles could be the best benzazoles to act as multitarget drugs for AD because they have been widely evaluated as AChE inhibitors, forming π–π interactions with W286, W86, Y72, and F338, as well as in the AChE gorge and catalytic site. In addition, the sulfur atom from benzothiazol interacts with S286 and the aromatic ring from W84, with these compounds having an IC50 value in the μM range. Also, benzimidazoles and benzothiazoles can inhibit Aβ aggregation. However, even though benzazoles have not been widely evaluated on BACE1, benzimidazoles evaluated in vitro showed an IC50 value in the nM range. Therefore, important chemical modifications could be considered to improve multitarget benzazoles’ activity, such as substitutions in the aromatic ring with electron withdrawal at position five, or a linker 3 or 4 carbons in length, which would allow for better interaction with targets.

Healing and Hallucinations: A Comprehensive Look at Datura

The Solanaceae plant family is a large clade that includes Datura, a genus of plants with many species recognized for their severe pharmacological effects via tropane alkaloids like atropine, hyoscyamine and scopolamine. These alkaloids act as antagonists of muscarinic acetylcholine receptors with leading to a truly diverse pharmacological action, ranging from anticholinergic effects all the way through hallucinogenic properties. Datura plants have been used historically in traditional medicine and rituals worldwide but are infamous for poisoning due to the risk of severe anticholinergic toxicity, including death. Nevertheless, their pharmacology has become of interest for research due to potential therapeutic applications: specifically in controlling nauseousness and perhaps neurological disorders. In this review, we provide a summary of the botanical description and cultivation methods in addition to those on traditional use practices concerning phytochemistry, pharmacology and toxicology as compared with evidence-based information together emphasizing cautions against availability without stringent regulatory consideration for its modern medical uses or ritualistic purposes.

Involvement of muscarinic acetylcholine receptor-mediated cholinergic neurotransmission in TMS–EEG responses

The combination of transcranial magnetic stimulation and electroencephalography (TMS–EEG) is emerging as a valuable tool for investigating brain functions in health and disease. However, the detailed neural mechanisms underlying TMS–EEG responses, including TMS-evoked EEG potentials (TEPs) and TMS-induced EEG oscillations (TIOs), remain largely unknown. Combining TMS–EEG with pharmacological interventions provides a unique opportunity to elucidate the roles of specific receptor-mediated neurotransmissions in these responses. Here, we investigated the involvement of muscarinic acetylcholine receptor (mAChR)-mediated cholinergic neurotransmission in TMS–EEG responses by evaluating the effects of mAChR antagonists on TEPs and TIOs in twenty-four healthy participants using a randomized, placebo-controlled crossover design. TEPs and TIOs were measured before and after administering a single oral dose of scopolamine (a non-selective mAChR antagonist), biperiden (an M1 mAChR antagonist), or placebo, with TMS targeting the left medial prefrontal cortex (mPFC), angular gyrus (AG), and supplementary motor area (SMA). The results indicated that mAChR-mediated cholinergic neurotransmission played a role in TEPs, but not TIOs, in a target-specific manner. Specifically, scopolamine significantly increased the amplitude of a local TEP component between approximately 40 and 63 ms post-stimulus when TMS was applied to the SMA, but not the mPFC or AG. Biperiden produced a similar but less pronounced effect. Importantly, the effects of these mAChR antagonists on TEPs were independent of those on sensory-evoked EEG potentials caused by TMS-associated sensory stimulation. These findings expand our understanding of TMS–EEG physiology, providing insights for its application in physiological and clinical research.

Acute Hypobaric Hypoxia Causes Alterations in Acetylcholine-Mediated Signaling Through Varying Expression of Muscarinic Receptors in the Prefrontal Cortex and Cerebellum of Rats' Brain.

Sharma, Narendra Kumar, Mansi Srivastava, Tikam Chand Dakal, Vipin Ranga, and Pawan Kumar Maurya. Acute hypobaric hypoxia (HH) causes alterations in acetylcholine-mediated signaling through varying expression of muscarinic receptors in the PFC and cerebellum of rats' brain. High Alt Med Biol. 00:00-00, 2024. Background: Muscarinic receptor (CHRM) proteins are G-protein-associated acetylcholine receptors found in neuronal membranes. Five major subtypes, CHRM1-CHRM5, modulate acetylcholine in central nervous system signaling cascades. CHRM1, CHRM3, and CHRM5 are linked to Gαq/Gα11 proteins, whereas CHRM2 and CHRM4 are linked to Gαi/Gαo proteins. Objective: Limited research has been conducted to explore the impact of HH on CHRM gene expressions. It is caused by low oxygen availability at high altitudes, which impairs neurotransmission, cognitive performance, and physiological functions. Previous studies have shown that exposure to hypoxia leads to a reduction in CHRM receptors, which in turn causes alteration in signal transduction, physiological responses, cognitive deficits, and mood alterations. Method: In the present study, we have used semiquantitative PCR to measure muscarinic receptor gene expression after 6, 12, and 24 hours of HH exposure at 25,000 feet using a decompression chamber in rat brain's PFC and cerebellum. Result: We have found that CHRM1-CHRM5 downregulated after acute exposure to hypoxia until 12 hours, and then, the expression level of these receptors increased to 24 hours when compared with 12 hours in PFC. All subtypes have shown a similar pattern in PFC regions under hypoxia exposure. On the other hand, these receptors have shown altered expression at different time points in the cerebellum. CHRM1 and CHRM4 acutely downregulated, CHRM2 and CHRM5 downregulated, while CHRM3 upregulated after hypoxia exposure. Conclusion: Our study, for the first time, has shown the altered expressions of muscarinic receptors under temporal hypoxia exposure. The altered expression pattern has shown an association with acclimatization and protection against necrosis due to hypoxia. This study may pave further investigations for understanding and addressing the cognitive, behavioral, and physiological impacts of hypoxia and therapeutic development.

Photo-BQCA: Positive Allosteric Modulators Enabling Optical Control of the M1 Receptor.

The field of G protein-coupled receptor (GPCR) research has greatly benefited from the spatiotemporal resolution provided by light controllable, i.e., photoswitchable ligands. Most of the developed tools have targeted the Rhodopsin-like family (Class A), the largest family of GPCRs. However, to date, all such Class A photoswitchable ligands were designed to act at the orthosteric binding site of these receptors. Herein, we report the development of the first photoswitchable allosteric modulators of Class A GPCRs, designed to target the M1 muscarinic acetylcholine receptor. The presented benzyl quinolone carboxylic acid (BQCA) derivatives, Photo-BQCisA and Photo-BQCtrAns, exhibit complementary photopharmacological behavior and allow reversible control of the receptor using light as an external stimulus. This makes them valuable tools to further investigate M1 receptor signaling and a proof of concept for photoswitchable allosteric modulators at Class A receptors.

TRPC5 controls the adrenaline-mediated counter regulation of hypoglycemia.

Hypoglycemia triggers autonomic and endocrine counter-regulatory responses to restore glucose homeostasis, a response that is impaired in patients with diabetes and its long-term complication hypoglycemia-associated autonomic failure (HAAF). We show that insulin-evoked hypoglycemia is severely aggravated in mice lacking the cation channel proteins TRPC1, TRPC4, TRPC5, and TRPC6, which cannot be explained by alterations in glucagon or glucocorticoid action. By using various TRPC compound knockout mouse lines, we pinpointed the failure in sympathetic counter-regulation to the lack of the TRPC5 channel subtype in adrenal chromaffin cells, which prevents proper adrenaline rise in blood plasma. Using electrophysiological analyses, we delineate a previously unknown signaling pathway in which stimulation of PAC1 or muscarinic receptors activates TRPC5 channels in a phospholipase-C-dependent manner to induce sustained adrenaline secretion as a crucial step in the sympathetic counter response to insulin-induced hypoglycemia. By comparing metabolites in the plasma, we identified reduced taurine levels after hypoglycemia induction as a commonality in TRPC5-deficient mice and HAAF patients.

Sublingual administration of atropine eye drops for treating sialorrhea after stroke: A randomized controlled trial

BackgroundSialorrhea is a common concern in patients with swallowing disorders after stroke. Atropine sulfate blocks the muscarinic receptors in the salivary glands and leads to reduced saliva production. ObjectiveThe present study aimed to assess the safety, efficacy, and tolerability of sublingual administration of atropine eye drops for treating sialorrhea after stroke. DesignThis was a prospective cohort study. SettingThis study was conducted at Xiangyang No. 1 People's Hospital, Hubei University of Medicine, Hubei Province, China. PopulationStroke patients with sialorrhea were analyzed. MethodsOne hundred stroke patients with sialorrhea were randomly assigned to the control group and the test group (n = 50 per group). The control group received routine swallowing rehabilitation training and neuromuscular electrical stimulation. The test group received therapy with 1% atropine eye drops, wherein one drop was administered sublingually 3 times per day. The Sialorrhea Scoring Scale and the incidence of adverse events were used to compare the severity of sialorrhea in the two groups. ResultsThe mean (standard deviation) sialorrhea score improved from 5.12 for the control group with routine rehabilitation training to 3.94 for the test group with atropine eye drop administration (P < 0.01). No significant differences in the incidence of adverse events were observed between the two groups. ConclusionsThe sublingual administration of 1% atropine eye drops three times per day can reduce the degree of sialorrhea to an extent more than that achieved with routine rehabilitation training; thus, this approach is effective, safe, and minimally invasive for treating sialorrhea after stroke.

New potent muscarinic receptor ligands bearing the 1,4-dioxane nucleus: Investigation on the nature of the substituent in position 2.

A new series of muscarinic acetylcholine receptor (mAChR) ligands obtained by inserting different substituents in position 2 of the potent 6,6-diphenyl-1,4-dioxane antagonists 4 and 5 was designed and synthesized to investigate the influence of steric bulk on the mAChR affinity. Specifically, the insertion of a 2-methyl group, affording compounds 6 and 9, resulted as the most favorable modification in terms of affinity for all muscarinic subtypes. As supported by computational studies performed on the hM1 receptor, this substituent may contribute to stabilize the ligand within the binding site by favoring the formation of stable interactions between the cationic head of the ligand and the residue D105. The increase of steric bulk, obtained by replacing the methyl group with an ethyl (7 and 10) and especially a phenyl substituent (8 and 11), caused a marked decrease of mAChR affinity, demonstrating the crucial role played by the steric bulk of the 2-substituent in the mAChR interaction. The most intriguing result was obtained with the tertiary amine 9, which, surprisingly, showed two different pKi values for all mAChRs, with preferential subpicomolar affinities for the M1, M3, and M4 subtypes. Interestingly, biphasic curves were also observed with both the eutomer (S)-(-)-9 and the distomer (R)-( + )-9.

M1 muscarinic receptor activation reverses age-related memory updating impairment in mice

Previously consolidated memories can become temporarily labile upon reactivation.Reactivation-based memory updating is chiefly studied in young subjects, so we aimed to assess this process across the lifespan. To do this, we developed a behavioural paradigm wherein a reactivated object memory is updated with contextual information; 3-month-old and 6-month-old male C57BL/6 mice displayed object memory updating, but 12-month-old mice did not. We found that M1 muscarinic acetylcholine receptor signaling during reactivation was necessary for object memory updating in the young mice. Next, we targeted this mechanism in an attempt to facilitate object memory updating in aging mice. Remarkably, systemic pharmacological M1 receptor activation reversed the age-related deficit. Quantification of cholinergic system markers within perirhinal cortex revealed subtle cellular changes that may contribute to differential performance across age groups. These findings suggest that natural cholinergic change across the lifespan contributes to inflexible memory in the aging brain.

Implications of the non-neuronal cholinergic system for therapeutic interventions of inflammatory skin diseases.

The pivotal roles of acetylcholine (ACh) in physiological processes encompass both the nervous and non-neuronal cholinergic systems (NNCS). This review delineates the synthesis, release, receptor interactions, and degradation of ACh within the nervous system, and explores the NNCS in depth within skin cells including keratinocytes, endothelial cells, fibroblasts, macrophages, and other immune cells. We highlight the NNCS's essential functions in maintaining epidermal barrier integrity, promoting wound healing, regulating microcirculation, and modulating inflammatory responses. The potential of the NNCS as a therapeutic target for localized ACh regulation in the skin is discussed, though the translation of these findings into clinical practice remains uncertain due to the complexity of cholinergic signalling and the lack of comprehensive human studies. The review progresses to therapeutic modulation strategies of the NNCS, including AChE inhibitors, nicotinic and muscarinic receptor agonists and antagonists, choline uptake enhancers, and botulinum toxin, highlighting their relevance in dermatology. We highlight the impact of the NNCS on prevalent skin diseases such as psoriasis, atopic dermatitis, rosacea, acne, bullous diseases, hyperhidrosis and hypohidrosis, illustrating its significance in disease pathogenesis and therapy. This comprehensive overview aims to enhance understanding of the NNCS's role in skin health and disease, offering a foundation for future research and therapeutic innovation.

Herbal Amara extract induces gastric fundus relaxation via inhibition of the M2 muscarinic receptor.

Impaired gastric accommodation is one of the most frequent symptoms of functional dyspepsia. The safety and efficacy of conventional treatments remain to be proven and alternative herbal therapies have been proposed to alleviate gastrointestinal symptoms. This preclinical study examined the role of herbal Amara extract (containing Artemisia absinthium, Centaurium erythraea, Cichorium intybus, Gentiana lutea, Juniperus communis, Achillea millefolium, Peucedanum ostruthium, Salvia officinalis, and Taraxacum extracts) on gastric (fundus) accommodation and the possible implication of muscarinic receptors in its regulation. The effect of Amara extract on fundus motility was investigated in organ baths of smooth muscle strips isolated from the fundus of guinea pigs, and the role of the muscarinic receptor pathway was evaluated using functional and radioligand binding assays in cell lines expressing the M2 or M3 muscarinic receptor. Amara extract inhibited carbachol-induced contraction of guinea pig smooth muscle strips in a dose-dependent manner. This relaxant effect was not affected by the M3 antagonist J-104129. Amara extract also inhibited M2, but not M3, receptor activity in CHO-K1 cells (IC50 219 μg mL-1), and specifically bound the M2 receptor (IC50 294 μg mL-1). Of the nine herbal components of Amara extract, Juniperus communis, P. ostruthium, and Salvia officinalis inhibited M2 receptor activity (IC50 32.0, 20.8, and 20.1 μg mL-1, respectively), and P. ostruthium was sufficient to reverse carbachol-induced exvivo contraction of guinea pig fundic smooth muscles. Amara extract relaxes gastric smooth muscles by inhibiting the M2 muscarinic receptor. This study suggests the potential benefit of Amara extract for patients with impaired gastric accommodation.

Cholinergic Signaling Mediated by Muscarinic Receptors Triggers the Ultraviolet-Induced Release of Melanosome in Cultured Melanoma Cells.

In skin, melanin is synthesized and stored in melanosomes. In epidermal melanocytes, melanosomes are transported to and internalized by the neighboring keratinocytes, subsequently leading to skin pigmentation. Ultraviolet (UV) radiation induces the release of acetylcholine (ACh) from keratinocytes, which in turn activates ACh receptors (AChRs) on nearby melanocytes, forming a proposed "skin synapse". Here, we illustrated that the UV-induced melanosome release from cultured B16F10 melanoma cells could be mediated by co-actions of ACh. In the cell cultures, UV exposure robustly elicited melanosome release. Applied bethanechol (BeCh), an agonist of muscarinic AChR (mAChR), could significantly enhance the release. In parallel, the intracellular Ca2+ mobilization was regulated. The applied antagonists of M1 and/or M3 mAChRs could block the UV-induced melanosome release and the mobilization of intracellular Ca2+. The phosphorylation of PKC, triggered by UV and BeCh treatments, could be suppressed by the applied mAChR antagonists. The expressions of tethering complex for exocytosis, for example, Sec8, Exo70, and Rab11b, as well as synaptotagmin, were increased under UV exposure together with mAChR agonist: The inductions were fully abolished by M1 or M3 antagonist. Here, we hypothesize that the cholinergic signaling is playing roles in UV-induced exocytosis of melanosomes.

The Activation of Muscarinic Acetylcholine Receptors Protects against Neuroinflammation in a Mouse Model through Attenuating Microglial Inflammation.

Neuroinflammation is a critical factor that contributes to neurological impairment and is closely associated with the onset and progression of neurodegenerative diseases. In the central nervous system (CNS), microglia play a pivotal role in the regulation of inflammation through various signaling pathways. Therefore, mitigating microglial inflammation is considered a promising strategy for restraining neuroinflammation. Muscarinic acetylcholine receptors (mAChRs) are widely expressed in the CNS and exhibit clear neuroprotective effects in various disease models. However, whether the activation of mAChRs can harness benefits in neuroinflammation remains largely unexplored. In this study, the anti-inflammatory effects of mAChRs were found in a neuroinflammation mouse model. The expression of various cytokines and chemokines was regulated in the brains and spinal cords after the administration of mAChR agonists. Microglia were the primary target cells through which mAChRs exerted their anti-inflammatory effects. The results showed that the activation of mAChRs decreased the pro-inflammatory phenotypes of microglia, including the expression of inflammatory cytokines, morphological characteristics, and distribution density. Such anti-inflammatory modulation further exerted neuroprotection, which was found to be even more significant by the direct activation of neuronal mAChRs. This study elucidates the dual mechanisms through which mAChRs exert neuroprotective effects in central inflammatory responses, providing evidence for their application in inflammation-related neurological disorders.

Discovery of VU6016235: A Highly Selective, Orally Bioavailable, and Structurally Distinct Tricyclic M4 Muscarinic Acetylcholine Receptor Positive Allosteric Modulator (PAM).

Herein, we report structure-activity relationship (SAR) studies to develop novel tricyclic M4 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 M4 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.

A cholinergic signaling pathway underlying cortical circuit activation of quiescent neural stem cells in the lateral ventricle.

Neural stem cells (NSCs) in the subventricular zone (SVZ) located along the lateral ventricles (LVs) of the mammalian brain continue to self-renew to produce new neurons after birth and into adulthood. Quiescent LV cells, which are situated close to the ependymal cells lining the LVs, are activated by choline acetyltransferase-positive (ChAT+) neurons within the subependymal (subep) region of the SVZ when these neurons are stimulated by projections from the anterior cingulate cortex (ACC). Here, we uncovered a signaling pathway activated by the ACC-subep-ChAT+ circuit responsible for the activation and proliferation of quiescent LV NSCs specifically in the ventral area of the SVZ. This circuit activated muscarinic M3 receptors on quiescent LV NSCs, which subsequently induced signaling mediated by the inositol 1,4,5-trisphosphate receptor type 1 (IP3R1). Downstream of IP3R1 activation, which would be expected to increase intracellular Ca2+, Ca2+-/calmodulin-dependent protein kinase II δ and the MAPK10 signaling pathway were stimulated and required for the proliferation of quiescent LV NSCs in the SVZ. These findings reveal the mechanisms that regulate quiescent LV NSCs and underscore the critical role of projections from the ACC in promoting their proliferative activity within the ventral SVZ.