Acetylcholine Receptor Protein Research Articles

Recombinant Acetylcholine Receptor Immunization Induces a Robust Model of Experimental Autoimmune Myasthenia Gravis in Mice.

Myasthenia gravis (MG) is a prototypical autoimmune disease of the neuromuscular junction (NMJ). The study of the underlying pathophysiology has provided novel insights into the interplay of autoantibodies and complement-mediated tissue damage. Experimental autoimmune myasthenia gravis (EAMG) emerged as a valuable animal model, designed to gain further insight and to test novel therapeutic approaches for MG. However, the availability of native acetylcholine receptor (AChR) protein is limited favouring the use of recombinant proteins. To provide a simplified platform for the study of MG, we established a model of EAMG using a recombinant protein containing the immunogenic sequence of AChR in mice. This model recapitulates key features of EAMG, including fatigable muscle weakness, the presence of anti-AChR-antibodies, and engagement of the NMJ by complement and a reduced NMJ density. Further characterization of this model demonstrated a prominent B cell immunopathology supported by T follicular helper cells. Taken together, the herein-presented EAMG model may be a valuable tool for the study of MG pathophysiology and the pre-clinical testing of therapeutic applications.

PATOFISIOLOGI MIASTENIA GRAVIS

Myasthenia Gravis (MG) is an autoimmune disease that disrupts transmission at the neuromuscular junction (NMJ). In myasthenia gravis, the immune system will attack the acetylcholine receptor (AChR) or other proteins involved in neuromuscular transmission or due to abnormalities in the thymus, which plays a role in immunity. This causes characteristic manifestations in the form of muscle weakness, which will improve after rest. So, in its management, the use of immunosuppressive therapy and removal of the thymus can be a therapeutic option depending on the type and severity of the disease.

Monosodium Glutamate Effect on The Expression of ɑ7nACHR and ɑ4nACHR Subunits in The Testicular Tissue.

Monosodium glutamate (MSG) is a popular food flavor enhancer, and a glutamate subset that induces different toxicities such as hepatotoxicity, neurotoxicity, reproductive toxicity, and nephrotoxicity. This study was conducted to assess the effects of MSG on the α7 and α4 nicotinic acetylcholine receptor (nACHR) protein subunits expression of adult rat testis and the safety role of vitamin C. For this experimental research, 24 rats were haphazardly grouped into four equal groups (n=6) and orally gavaged for 30 days as follows: control group (distilled water gavage), MSG group (3 g/kg/b.w/ day), vitamin C group (150 mg/kg/b.w/day), and MSG+vitamin C group (3 g/kg/b.w/day+150 mg/kg/b.w/day, respectively) that rats of all groups on the 30th day were anesthetized, and the left testes were used for of α4 and α7 nACHR protein subunit evaluation by immunohistochemistry (IHC). Statistical computations were performed using Graph Pad Prism software. The present study revealed a significant reduction in the expression and optical density (OD) of α7 nACHR and α4 nACHR in the seminiferous tubules and intertubular connective tissue in the MSG group compared to the control group. In the MSG+vitamin C group, the expression and OD of α7 nACHR and α4 nACHR increased in the seminiferous tubules and intertubular connective tissue but this improvement was not significant compared to the MSG group. MSG decreased the expression level of nACHR protein subunits, α7 and α4, in the seminiferous tubules and interstitial testicular tissue. Vitamin C in the MSG+vitamin C group could not significantly improve the expression of α7 and α4 nACHR subunits in testicular tissue. Probably, MSG toxicity can be compensated with higher doses of vitamin C.

Swimming exercise and clove oil can improve memory by molecular responses modification and reduce dark cells in rat model of Alzheimer's disease

Alzheimer's disease (AD) is marked by reduced acetylcholine receptor (AChR) density and an increase in nucleotide oligomerization domain (NOD)-like receptors NLR family, pyrin domain containing 1 (NLRP1). We examined the effect of swimming and consumption of clove supplements on memory, dark cells, and α7nAChR and NLRP1 mRNA and protein expression in the hippocampus of the rat model of AD. Forty-eight rats were divided into six groups: sham (sh), healthy-control (HC), Alzheimer (-control (AC), -training (AT), -training-supplement (ATS), and -supplement (AS)). Alzheimer was induced by injection of amyloid β1–42 (Aβ1–42). Swimming exercise protocol (30 min) and gavaging clove supplement (0.1 mg/kg) were administered daily for three weeks. The results indicated that in response to AD, α7 nicotinic acetylcholine receptor (α7nAChR) mRNA and protein rate (p = 0.001) and memory (p = 0.003) were significantly decreased. In contrast, NLRP1 mRNA and protein rate (p = 0.001) and dark cells (p = 0.001) were significantly increased. This is while exercise and clove supplementation improved Alzheimer-induced changes in α7nAChR, NLRP1, memory, and dark cells (p < 0/05). The present study indicated that exercising and consuming clove supplementation could improve memory by increasing α7nAChR and decreasing NLRP1 and dark cells.

Potential antiproliferative and apoptotic effects of pilocarpine combined with TNF alpha in chronic myeloid leukemia cells.

Pilocarpine is a selective M1/M3 agonist of muscarinic acetylcholine receptor subtypes. Muscarinic acetylcholine receptors are G protein-coupled receptors. These receptors are different drug targets. The aim of the present work was to investigate the effect of pilocarpine on the expression of M3 muscarinic acetylcholine receptor, the AChE activity, IL-8 release response, and proliferation in K562 cells, via muscarinic receptor activation.Human chronic myeloid leukemic cell cultures were incubated with drugs. Proliferation assays were performed by BrdU assay. Expression of M3 muscarinic acetylcholine receptor and apoptosis proteins such as bcl, bax, cyt C, and caspases was assessed with the semiquantitative Western blotting method.Pilocarpine inhibits chronic myeloid cell proliferation and M3 muscarinic acetylcholine receptor protein expression. Pilocarpine increases caspase-8 and -9 expression levels, upregulating the proapoptotic protein Bax and downregulating the expression levels of the antiapoptotic protein Bcl-2. The apoptotic activity of pilocarpine is associated with an increase in AChE activity. M3 muscarinic acetylcholine receptors can activate multiple signal transduction systems and mediate inhibitory effects on chronic myeloid K562 cell proliferation depending on the presence of 1% FBS conditions. This apoptotic effect of pilocarpine may be due to the concentration of pilocarpine and the increase in AChE level.Our results suggest that inhibition of cell proliferation by inducing apoptosis of pilocarpine in K562 cells may be one of the targets. M3 selective agonist may have therapeutic potential in chronic myeloid leukemia.

MiRNAs as the important regulators of myasthenia gravis: involvement of major cytokines and immune cells.

Myasthenia gravis (MG) is a type of muscle paralysis created by immune responses against acetylcholine receptor proteins in neuromuscular synapses. This disease is characterized by muscle weakness, especially ocular weakness symptoms that could be ptosis (fall of the upper eyelid) or diplopia (double vision of a single object). Some patients also identified with speech and swallowing problems. The main goals of MG therapeutic approaches are to achieve remission, reduce symptoms, and improve life quality. Recently, other studies have revealed the potential role of various microRNAs (miRNAs) in the development of MG through different mechanisms and have proposed these molecules as effective biomarkers for the treatment of MG. This review was aimed at providing an overview of the critical regulatory roles of various miRNAs in the pathogenesis of this autoimmune disease focusing on human MG studies and the interaction between different miRNAs with important cytokines and immune cells during the development of this autoimmune disease.

Neuregulin-1/ErbB4 upregulates acetylcholine receptors via Akt/mTOR/p70S6K: a study in a rat model of obstetric brachial plexus palsy and in vitro.

In obstetric brachial plexus palsy (OBPP), the operative time window for nerve reconstruction of the intrinsic muscles of the hand (IMH) is much shorter than that of biceps. The reason is that the atrophy of IMH becomes irreversible more quickly than that of biceps. A previous study confirmed that the motor endplates of denervated intrinsic muscles of the forepaw (IMF) were destabilized, while those of denervated biceps remained intact. However, the specific molecular mechanism of regulating the self-repair of motor endplates is still unknown. In this study, we use a rat model of OBPP with right C5-C6 rupture plus C7-C8-T1 avulsion and left side as a control. Bilateral IMF and biceps are harvested at 5 weeks postinjury to assess relative protein and mRNA expression. We also use L6 skeletal myoblasts to verify the effects of signaling pathways regulating acetylcholine receptor (AChR) protein synthesis in vitro. The results show that in the OBPP rat model, the protein and mRNA expression levels of NRG-1/ErbB4 and phosphorylation of Akt/mTOR/p70S6K are lower in denervated IMF than in denervated biceps. In L6 myoblasts stimulated with NRG-1, overexpression and knockdown of ErbB4 lead to upregulation and downregulation of AChR subunit protein synthesis and Akt/mTOR/p70S6K phosphorylation, respectively. Inhibition of mTOR abolishes protein synthesis of AChR subunits elevated by NRG-1/ErbB4. Our findings suggest that in the OBPP rat model, lower expression of AChR subunits in the motor endplates of denervated IMF is associated with downregulation of NRG-1/ErbB4 and phosphorylation of Akt/mTOR/p70S6K. NRG-1/ErbB4 can promote protein synthesis of the AChR subunits in L6 myoblasts via phosphorylation of Akt/mTOR/p70S6K.

Motoneurons innervation determines the distinct gene expressions in multinucleated myofibers

BackgroundNeuromuscular junctions (NMJs) are peripheral synapses connecting motoneurons and skeletal myofibers. At the postsynaptic side in myofibers, acetylcholine receptor (AChR) proteins are clustered by the neuronal agrin signal. Meanwhile, several nuclei in each myofiber are specially enriched around the NMJ for postsynaptic gene transcription. It remains mysterious that how gene expressions in these synaptic nuclei are systematically regulated, especially by motoneurons.ResultsWe found that synaptic nuclei have a distinctive chromatin structure and gene expression profiling. Synaptic nuclei are formed during NMJ development and maintained by motoneuron innervation. Transcriptome analysis revealed that motoneuron innervation determines the distinct expression patterns in the synaptic region and non-synaptic region in each multinucleated myofiber, probably through epigenetic regulation. Myonuclei in synaptic and non-synaptic regions have different responses to denervation. Weighted gene co-expression network analysis revealed that the histone lysine demethylases Kdm1a is a negative regulator of synaptic gene expression. Inhibition of Kdm1a promotes AChR expression but impairs motor functions.ConclusionThese results demonstrate that motoneurons innervation determines the distinct gene expressions in multinucleated myofibers. Thus, dysregulation of nerve-controlled chromatin structure and muscle gene expression might cause muscle weakness and atrophy in motoneuron degenerative disorders.

Clinical value of cell-based assays in the characterisation of seronegative myasthenia gravis

ObjectivePatients with myasthenia gravis without acetylcholine receptor (AChR) or muscle-specific kinase (MuSK) antibodies detected by radioimmunoprecipitation assays (RIAs) are classified as seronegative myasthenia gravis (SNMG). Live cell-based assays (l-CBAs) can...

Scrutinizing pathways of nicotine effect on renal Alpha-7 nicotinic acetylcholine receptor and Mitogen-activated protein kinase (MAPK) expression in Ehrlich ascites carcinoma-bearing mice: Role of Chlorella vulgaris

Nicotine is one of several physiologically stable and active chemicals found in tobacco. The mechanism through which nicotine causes kidney damage is still obscure. As a result, the goal of this research was to investigate how oral nicotine intake can lead to kidney damage. Naturaly occurring superfood green algae are immense supplements help us using extra chemicals during cancer prevalence if the patient is exposed to nicotine. Hence, the mitigating role of Chlorella vulgaris extract (CVE) against nicotine-nephrotoxic impact in Ehrlich ascites carcinoma (EAC)-bearing mice was studied. For this purpose, four groups of Swiss female mice were assigned, nicotine group (NIC) (100 µg/ml/kg), CVE group (100 mg/kg), CVE + Nicotine, and a control group. Renal dysfunction was evaluated by estimating serum biomarkers ofrenal damage. The expression pattern of Nf-KB, MAPK, P53, and α7-nAchR, lipid peroxidation biomarker, and antioxidant enzyme activities were evaluated in kidney tissue. Also, micro-morphometric examination and apoptosis immunohistochemical reactivity of kidney tissue were applied. The obtained results indicated up-regulation of all estimated genes and oxidative stress. Moreover, a significant (P < 0.05) increment in the apoptotic marker Caspase-3 and declined BCL-2 proteins were recorded. In serum, a significant (P < 0.05) elevation of urea, creatinine, TNF-α, IL-1β, and Kim-1 were evident. Histological investigation reinforced the aforementioned data, revealing structural changes involving the tubules, glomeruli, and interstitium of mice kidneys. CVE may be a strong contender for protecting renal tissue damage since it reduces renal tissue injury and oxidative stress. Cancer patients who regularly use nicotine through direct smoking or second-hand exposure can benefit from CVE usage as a dietary supplement.

Fluorescence Studies of Nicotinic Acetylcholine Receptor and Its Associated Lipid Milieu: The Influence of Erwin London's Methodological Approaches.

Erwin London dedicated considerable effort to understanding lipid interactions with membrane-resident proteins and how these interactions shaped the formation and maintenance of lipid phases and domains. In this endeavor, he developed ad hoc techniques that greatly contributed to advancements in the field. We have employed and/or modified/extended some of his methodological approaches and applied them to investigate lipid interaction with the nicotinic acetylcholine receptor (nAChR) protein, the paradigm member of the superfamily of rapid pentameric ligand-gated ion channels (pLGIC). Our experimental systems ranged from purified receptor protein reconstituted into synthetic lipid membranes having known effects on receptor function, to cellular systems subjected to modification of their lipid content, e.g., varying cholesterol levels. We have often employed fluorescence techniques, including fluorescence quenching of diphenylhexatriene (DPH) extrinsic fluorescence and of nAChR intrinsic fluorescence by nitroxide spin-labeled phospholipids, DPH anisotropy, excimer formation of pyrene-phosphatidylcholine, and Förster resonance energy transfer (FRET) from the protein moiety to the extrinsic probes Laurdan, DPH, or pyrene-phospholipid to characterize various biophysical properties of lipid-receptor interactions. Some of these strategies are revisited in this review. Special attention is devoted to the anionic phospholipid phosphatidic acid (PA), which stabilizes the functional resting form of the nAChR. The receptor protein was shown to organize its PA-containing immediate microenvironment into microdomains with high lateral packing density and rigidity. PA and cholesterol appear to compete for the same binding sites on the nAChR protein.

Skeletal Muscle-Restricted Expression of Human SOD1 in Transgenic Mice Causes a Fatal ALS-Like Syndrome.

Amyotrophic lateral sclerosis (ALS) is a fatal heterogeneous neurodegenerative disease that causes motor neuron (MN) loss and skeletal muscle paralysis. It is uncertain whether this degeneration of MNs is triggered intrinsically and is autonomous, or if the disease initiating mechanisms are extrinsic to MNs. We hypothesized that skeletal muscle is a primary site of pathogenesis in ALS that triggers MN degeneration. Some inherited forms of ALS are caused by mutations in the superoxide dismutase-1 (SOD1) gene, that encodes an antioxidant protein, so we created transgenic (tg) mice expressing wild-type-, G37R-, and G93A-human SOD1 gene variants only in skeletal muscle. Presence of human SOD1 (hSOD1) protein in skeletal muscle was verified by western blotting, enzyme activity gels, and immunofluorescence in myofibers and satellite cells. These tg mice developed limb weakness and paresis with motor deficits, limb and chest muscle wasting, diaphragm atrophy, and age-related fatal disease with a lifespan shortening of 10–16%. Brown and white adipose tissue also became wasted. Myofibers of tg mice developed crystalline-like inclusions, individualized sarcomere destruction, mitochondriopathy with vesiculation, DNA damage, and activated p53. Satellite cells became apoptotic. The diaphragm developed severe loss of neuromuscular junction presynaptic and postsynaptic integrity, including decreased innervation, loss of synaptophysin, nitration of synaptophysin, and loss of nicotinic acetylcholine receptor and scaffold protein rapsyn. Co-immunoprecipitation identified hSOD1 interaction with rapsyn. Spinal cords of tg mice developed gross atrophy. Spinal MNs formed cytoplasmic and nuclear inclusions, axonopathy, mitochondriopathy, accumulated DNA damage, activated p53 and cleaved caspase-3, and died. Tg mice had a 40–50% loss of MNs. This work shows that hSOD1 in skeletal muscle is a driver of pathogenesis in ALS, that involves myofiber and satellite cell toxicity, and apparent muscle-adipose tissue disease relationships. It also identifies a non-autonomous mechanism for MN degeneration explaining their selective vulnerability as likely a form of target-deprivation retrograde neurodegeneration.

AChR β-Subunit mRNAs Are Stabilized by HuR in a Mouse Model of Congenital Myasthenic Syndrome With Acetylcholinesterase Deficiency.

Collagen Q (COLQ) is a specific collagen that anchors acetylcholinesterase (AChE) in the synaptic cleft of the neuromuscular junction. So far, no mutation has been identified in the ACHE human gene but over 50 different mutations in the COLQ gene are causative for a congenital myasthenic syndrome (CMS) with AChE deficiency. Mice deficient for COLQ mimic most of the functional deficit observed in CMS patients. At the molecular level, a striking consequence of the absence of COLQ is an increase in the levels of acetylcholine receptor (AChR) mRNAs and proteins in vivo and in vitro in murine skeletal muscle cells. Here, we decipher the mechanisms that drive AChR mRNA upregulation in cultured muscle cells deficient for COLQ. We show that the levels of AChR β-subunit mRNAs are post-transcriptionally regulated by an increase in their stability. We demonstrate that this process results from an activation of p38 MAPK and the cytoplasmic translocation of the nuclear RNA-binding protein human antigen R (HuR) that interacts with the AU-rich element located within AChR β-subunit transcripts. This HuR/AChR transcript interaction induces AChR β-subunit mRNA stabilization and occurs at a specific stage of myogenic differentiation. In addition, pharmacological drugs that modulate p38 activity cause parallel modifications of HuR protein and AChR β-subunit levels. Thus, our study provides new insights into the signaling pathways that are regulated by ColQ-deficiency and highlights for the first time a role for HuR and p38 in mRNA stability in a model of congenital myasthenic syndrome.

Efficient screening of ligand-receptor complex formation using fluorescence labeling and size-exclusion chromatography

Evidence of a complex formation is a crucial step in the structural studies of ligand-receptor interactions. Here we presented a simple and fast approach for qualitative screening of the complex formation between the chimeric extracellular domain of the nicotinic acetylcholine receptor (α7-ECD) and three-finger proteins. Complex formation of snake toxins α-Bgtx and WTX, as well as of recombinant analogs of human proteins Lynx1 and SLURP-1, with α7-ECD was confirmed using fluorescently labeled ligands and size-exclusion chromatography with simultaneous absorbance and fluorescence detection. WTX/α7-ECD complex formation also was confirmed by cryo-EM. The proposed approach could easily be adopted to study the interaction of other receptors with their ligands.

Muscarinic-Dependent miR-182 and QR2 Expression Regulation in the Anterior Insula Enables Novel Taste Learning.

In a similar manner to other learning paradigms, intact muscarinic acetylcholine receptor (mAChR) neurotransmission or protein synthesis regulation in the anterior insular cortex (aIC) is necessary for appetitive taste learning. Here we describe a parallel local molecular pathway, where GABAA receptor control of mAChR activation causes upregulation of miRNA-182 and quinone reductase 2 (QR2) mRNA destabilization in the rodent aIC. Damage to long-term memory by prevention of this process, with the use of mAChR antagonist scopolamine before novel taste learning, can be rescued by local QR2 inhibition, demonstrating that QR2 acts downstream of local muscarinic activation. Furthermore, we prove for the first time the presence of endogenous QR2 cofactors in the brain, establishing QR2 as a functional reductase there. In turn, we show that QR2 activity causes the generation of reactive oxygen species, leading to modulation in Kv2.1 redox state. QR2 expression reduction therefore is a previously unaccounted mode of mAChR-mediated inflammation reduction, and thus adds QR2 to the cadre of redox modulators in the brain. The concomitant reduction in QR2 activity during memory consolidation suggests a complementary mechanism to the well established molecular processes of this phase, by which the cortex gleans important information from general sensory stimuli. This places QR2 as a promising new target to tackle neurodegenerative inflammation and the associated impediment of novel memory formation in diseases such as Alzheimer’s disease.

Expression Levels of Inflammasome Complexes in Experimental Autoimmune Myasthenia Gravis Mouse Model (EAMG)

Objective: Despite the clues that myasthenia gravis (MG) disease may be associated with inflammasomes, there are no studies in the literature on MG disease and inflammasome complexes. Hence, to address this question, we investigated the possible participation of inflammasomes in experimental autoimmune myasthenia gravis mouse model (EAMG). Material and Method: EAMG was induced in mouse using acetylcholine receptor (AChR) protein, and Anti-AChR IgG antibody levels detected by ELISA in the experimental group confirmed our model. Levels of CASP1, IL-1β, NLRP3, P2X7R, and AKT1 of the experimental and control (complete Freund’s adjuvant -CFA immunized) groups were measured by qRT-PCR. Results: After immunization, the AChR IgG antibody levels were significantly higher in the AChR-immunized group than in the control group (p=0.042). IL-1β levels in the experimental group were significantly higher, compared to the control group (p=0.01). CASP1, NLRP3, and P2X7R levels were also higher compared to the control group. However, these differences did not attain statistical significance (p>0.05). AKT1 levels were lower compared to the control group. There was no correlation between serum antibody concentration and gene expression levels. Conclusion: Our results suggest that there might be inflammasome involvement in the pathology of MG disease. Increase in IL-1β levels indicates the importance of the inflammatory response; however, further studies are necessary to confirm this.

Stress degradation, structural optimization, molecular docking, ADMET analysis of tiemonium methylsulphate and its degradation products

Attempts have been taken to study stress degradation of tiemonium methylsulphate (TMS). Quantum mechanical approach was applied to investigate the structural information, protein binding affinity and pharmacokinetic properties of the TMS and degradation products. Forced degradation study revealed that TMS degraded significantly under acid hydrolysis and photolytic degradation conditions than basic, thermal or oxidative conditions. Density functional theory (DFT) with B3LYP/6-31G+ (d, p) has been employed to optimize the structures. Frontier molecular orbital features (HOMO–LUMO) gap, hardness, softness), dipole moment, atomic partial charge, electrostatic potential and thermodynamic properties (electronic energy, enthalpy, Gibb's free energy) are investigated for optimization of degradation products. Molecular docking has been performed against muscarinic acetylcholine receptor protein 5DSG to search the binding affinity and mode(s). ADMET calculation predicts that all the products are non-carcinogenic and safe for oral administration.

Myasthenia Gravis: Pathogenic Effects of Autoantibodies on Neuromuscular Architecture.

Myasthenia gravis (MG) is an autoimmune disease of the neuromuscular junction (NMJ). Autoantibodies target key molecules at the NMJ, such as the nicotinic acetylcholine receptor (AChR), muscle-specific kinase (MuSK), and low-density lipoprotein receptor-related protein 4 (Lrp4), that lead by a range of different pathogenic mechanisms to altered tissue architecture and reduced densities or functionality of AChRs, reduced neuromuscular transmission, and therefore a severe fatigable skeletal muscle weakness. In this review, we give an overview of the history and clinical aspects of MG, with a focus on the structure and function of myasthenic autoantigens at the NMJ and how they are affected by the autoantibodies’ pathogenic mechanisms. Furthermore, we give a short overview of the cells that are implicated in the production of the autoantibodies and briefly discuss diagnostic challenges and treatment strategies.

Myasthenia gravis.

Myasthenia gravis (MG) is an autoimmune disease caused by antibodies against the acetylcholine receptor (AChR), muscle-specific kinase (MuSK) or other AChR-related proteins in the postsynaptic muscle membrane. Localized or general muscle weakness is the predominant symptom and is induced by the antibodies. Patients are grouped according to the presence of antibodies, symptoms, age at onset and thymus pathology. Diagnosis is straightforward in most patients with typical symptoms and a positive antibody test, although a detailed clinical and neurophysiological examination is important in antibody-negative patients. MG therapy should be ambitious and aim for clinical remission or only mild symptoms with near-normal function and quality of life. Treatment should be based on MG subgroup and includes symptomatic treatment using acetylcholinesterase inhibitors, thymectomy and immunotherapy. Intravenous immunoglobulin and plasma exchange are fast-acting treatments used for disease exacerbations, and intensive care is necessary during exacerbations with respiratory failure. Comorbidity is frequent, particularly in elderly patients. Active physical training should be encouraged.

Effect of Pei Yuan Tong Nao capsules on neuronal function and metabolism in cerebral ischemic rats

Ethnopharmacological significancePei Yuan Tong Nao (PYTN) capsules have been used in traditional Chinese medicine (TCM) for many centuries for the treatment of renal and vascular diseases, including cerebral ischemia. Although they are used in clinical practice, the evidence of their efficacy is lacking. Materials and methodsThe cerebral ischemic rat model was established by bilateral carotid artery ligation and shear surgery. The laser speckle blood flow imaging apparatus was used to observe the changes in cerebral blood flow before and after surgery. The working memory of rats were analyzed using the Morris water maze (MWM) test. The 2-deoxy-2-[18F]fluoro-D-glucose (18-FDG) standard uptake values (SUVs) of rats with cerebral ischemia were evaluated by small-living animal positron emission tomography (PET) imaging. The quantity of acetylcholine in the hippocampus and the protein quantity of α7 nicotinic acetylcholine receptor in the hippocampus were detected with high-resolution fluid phase chromatography and Western blot analysis. ResultsThere was no significant difference in the preoperative mean cerebral blood perfusion between the groups. There was a significant decrease (P < 0.01) in cerebral perfusion in the model-operation, nimodipine, and PYTN groups after bilateral common carotid artery occlusion (BCCAo) compared with presurgery. The escape latency is shortened in the PYTN group (P < 0.05) compared with the model-operation group. SUVs of the nimodipine and PYTN groups increased (P < 0.05) compared with the model-operation group. After treatment with nimodipine and PYTN, acetylcholine levels in the hippocampus of the rats in the respective groups were lower than that of the sham-operation group (P < 0.01), and acetylcholine levels in the hippocampus of the nimodipine and PYTN groups were higher than that of the model-operation group (P < 0.05). α7 nicotinic acetylcholine receptor analysis showed that the protein levels of the nimodipine and PYTN groups increased after treatment compared with the model-operation group (P < 0.05). ConclusionPYTN capsules improved the performance of rats with cerebral ischemia (analyzed using the MWM test), which may be due the to improvement of glucose metabolism in the hippocampus (evaluated by estimating acetylcholine and α7 nicotinic acetylcholine receptor protein).