Distribution of Like-muscarinic Acetylcholine Receptor M2 in the Brain of Three Castes of Polyrhachis vicina

The cholinergic system plays an important role in the central nervous system of insects and is closely related to the complex behavior of insects. The immunohistochemical technique was performed to detect the expression of like-muscarinic acetylcholine receptor M2 in the brain of three castes of Polyrhachis vicina. A positive expression of like-muscarinic acetylcholine receptor M2 was observed in the mushroom body, central body and antennal lobes of the ant brain; but there is great diversity in their location and intensity among worker, queen and male ants. It is speculated that like-muscafinic acetylcholine receptor M2 plays a critical role in the central nervous system, in terms of projecting visual information and olfactory information into the protocerebrum and integrating many inputs.

Research on Autoantibodies Against Myocardialβ_1-adrenergic and M_2 Cholinergic Receptors in Patients With Chronic Keshan Disease

Objectives To explore the relationship between serum autoantibodies against myocardial β1-adrenergic, M2-cholinergic receptors and chronic Keshan disease （CKD）. Methods The second extracellular loops of β1 and M2 receptors on human cardiomyocytes were used as the antigens. Enzyme linked immunosorbent assay （ELISA） was applied to determine serum autoantibodies against myocardial β1 and ME receptors in 32 CKD patients. 31 healthy subjects from endemic area were selected as the control. Results Positive rate of autoantibodies against myocardial β1 adrenergic （51.3%, 17/32） and M2 cholinergic （56.3% , 18/32） receptors were significantly higher than those in the control （9.7%, 3/ 31; 12.9%, 4/31） （both P〈 0.01）. Both positive rate and titers of above autoantibodies in NYHA Ⅱ - Ⅲ CKD patients were significantly higher than those in NYHA Ⅳ, demonstrating an apparently positive correlation between serum antibodies against myocardial β1 and M2 receptors （r=0.95）. Conclusions Autoantibodies against myocardial β1 and M2 receptors were found in sera of CKD patients; distribution of positive rate and titers of the autoantibodies in CKD patients in various NYHA are significantly different. classes of cardiac function

Mechanisms mediating cholinergic antral circular smooth muscle contraction in rats

AIM:To investigate the pathway(s)mediating rat antral circular smooth muscle contractile responses to the cholinomimetic agent,bethanechol and the subtypes of muscarinic receptors mediating the cholinergic contraction. METHODS:Circular smooth muscle strips from the antrum of Sprague-Dawley rats were mounted in muscle baths in Krebs buffer.Isometric tension was recorded.Cumulative concentration-response curves were obtained for(+)-cis- dioxolane(cD),a nonspecific muscarinic agonist,at 10^(-8)- 10^(-4)mol/L,in the presence of tetrodotoxin(TTX,10^(-7)mol/L). Results were normalized to cross sectional area.A repeat concentration-response curve was obtained after incubation of the muscle for 90 min with antagonists for M1(pirenzepine), M2(methoctramine)and M3(darifenadn)muscarinic receptor subtypes.The sensitivity to PTX was tested by the ip injection of 100 mg/kg of PTX 5 d before the experiment.The antral circular smooth muscles were removed from PTX-treated and non-treated rats as strips and dispersed smooth muscle cells to identify whether PTX-linked pathway mediated the contractility to bethanechol. RESULTS:A dose-dependent contractile response observed with bethanechol,was not affected by TTx.The pretreatment of rats with pertussis toxin decreased the contraction induced by bethanechol.Lack of calcium as well as the presence of the L-type calcium channel blocker,nifedipine,also inhibited the cholinergic contraction,with a reduction in response from 2.5±0.4 g/mm^2 to 1.2±0.4 g/mm^2(P<0.05).The dose- response curves were shifted to the right by muscarinic antagonists in the following order of affinity:darifenacin (M_3)>methocramine(M_2)>pirenzepine(M_1). CONCLUSION:The muscarinic receptors-dependent contraction of rat antral circular smooth muscles was linked to the signal transduction pathway(s)involving pertussis-toxin sensitive GTP-binding proteins and to extracellular calcium via L-type voltage gated calcium channels.The presence of the residual contractile response after the treatment with nifedipine,suggests that an additional pathway could mediate the cholinergic contraction.The involvement of more than one muscarinic receptor(functionally predominant type 3 over type 2)also suggests more than one pathway mediating the cholinergic contraction in rat antrum.

Effect of organophosphorus insecticides on phosphorylation of the M_2 muscarinic acetylcholine receptor

BACKGROUND： Organophosphorus insecticides may promote the accumulation of acetylcholine at synapses and the neuromuscular junction by inhibiting acetylcholinesterase activity to cause disturbance of neural signal conduction and induce a toxic reaction. Organophosphorus insecticides may act on M2 muscarinic acetylcholine receptors, whose combination with G proteins is regulated by phosphorylation of G protein-coupled receptor kinase 2. OBJECTIVE： To investigate the effects of organophosphorus insecticides on the phosphorylation of G protein-coupled receptor kinase 2-mediated M2 muscarinic acetylcholine receptors and to reveal other possible actions of organophosphorus insecticides. DESIGN, TIME AND SETTING： An observational study, which was performed in the Central Laboratory of Shenyang Medical College, and Department of Physiological Sciences, College of Veterinary Medicine, Oklahoma State University from June 2002 to December 2004. MATERIALS： Paraoxon, parathion, chlorpyrifos, and chlorpyrifos oxon were provided by Chem Service Company, USA, [γ -p^32] ATP and [^35S]GTP γ S by New England Nuclear Life Science Products, and recombinant β 2-adrenergic receptor membrane protein by Sigma Company, USA. METHODS： The M2 muscarinic acetylcholine receptor was extracted and purified from pig brain using affinity chromatography. Subsequently, the purified M2 muscarinic acetylcholine receptor, G protein-coupled receptor kinase 2, and [γ -p^32] ATP were incubated with different concentrations of paraoxon and chlorpyrifos oxon together. The mixture then underwent polyacrylamide gel electrophoresis, and the gel film was dried and radioactively autographed to detect phosphorylation of the M2 muscarinic acetylcholine receptor. Finally, the radio-labeled phosphorylated M2 receptor protein band was excised for counting with an isotope liquid scintillation counter. MAIN OUTCOME MEASURES： Effects of chlorpyrifos oxon, paraoxon, chlorpyrifos, and parathion in different concentrations on the phosphorylation of the M2 muscarinic acetylcholine receptor; effects of chlorpyrifos oxon on the phosphorylation of the β -adrenergic receptor. RESULTS： Chlorpyrifos oxon could completely inhibit the phosphorylation of the M2 muscarinic acetylcholine receptor, and its IC50 was 70 μ mol/L. Chlorpyrifos could also inhibit the phosphorylation of the M2 muscarinic acetylcholine receptor. However, paraoxon and parathion could not inhibit the phosphorylation of the M2 muscarinic acetylcholine receptor. Chlorpyrifos oxon in different concentrations could also not inhibit the phosphorylation of the β 2-adrenergic receptor catalyzed by G protein-coupled receptor kinase 2. CONCLUSION： Different kinds of organophosphorus insecticides have different effects on the phosphorylation of the G protein-coupled receptor kinase 2-mediated M2 muscarinic acetylcholine receptor. Organophosphorus insecticides possibly have different toxic effects.