Desensitization and Sequestration of Human m2 Muscarinic Acetylcholine Receptors by Autoantibodies from Patients with Chagas' Disease

Abstract

Chronic Chagas' disease is associated with pathologic changes of the cardiovascular, digestive, and autonomic nervous system, culminating in autonomic denervation and congestive heart failure. Previously, circulating autoantibodies that activate signaling by cardiac muscarinic acetylcholine receptors (mAChRs) have been described. However, it remains unclear whether the chagasic IgGs directly interact with the m2 mAChRs (predominant cardiac subtype), and, if so, whether chronic exposure of the mAChRs to such activating IgGs would result in receptor desensitization. Here we performed studies with purified and reconstituted hm2 mAChRs and demonstrate that IgGs from chagasic serum immunoprecipitated the mAChRs in a manner similar to an anti-m2 mAChR monoclonal antibody tested in parallel. The chagasic antibodies did not directly interact with the ligand binding site, because the binding of radiolabeled antagonist was unchanged by the addition of the chagasic IgG. In intact cells stably expressing the hm2 mAChR, the chagasic IgGs, but not normal IgGs, mimicked the ability of the agonist acetylcholine to induce two effects associated with agonist-induced receptor desensitization: a decrease in affinity for agonist binding to m2 mAChR and sequestration of the hm2 mAChRs from the cell surface. The results demonstrate that the chagasic IgGs can directly interact with and desensitize m2 mAChRs and provide support for the hypothesis of autoimmune mechanisms having a role in the pathogenesis of Chagas' cardioneuromyopathy. Chronic Chagas' disease is associated with pathologic changes of the cardiovascular, digestive, and autonomic nervous system, culminating in autonomic denervation and congestive heart failure. Previously, circulating autoantibodies that activate signaling by cardiac muscarinic acetylcholine receptors (mAChRs) have been described. However, it remains unclear whether the chagasic IgGs directly interact with the m2 mAChRs (predominant cardiac subtype), and, if so, whether chronic exposure of the mAChRs to such activating IgGs would result in receptor desensitization. Here we performed studies with purified and reconstituted hm2 mAChRs and demonstrate that IgGs from chagasic serum immunoprecipitated the mAChRs in a manner similar to an anti-m2 mAChR monoclonal antibody tested in parallel. The chagasic antibodies did not directly interact with the ligand binding site, because the binding of radiolabeled antagonist was unchanged by the addition of the chagasic IgG. In intact cells stably expressing the hm2 mAChR, the chagasic IgGs, but not normal IgGs, mimicked the ability of the agonist acetylcholine to induce two effects associated with agonist-induced receptor desensitization: a decrease in affinity for agonist binding to m2 mAChR and sequestration of the hm2 mAChRs from the cell surface. The results demonstrate that the chagasic IgGs can directly interact with and desensitize m2 mAChRs and provide support for the hypothesis of autoimmune mechanisms having a role in the pathogenesis of Chagas' cardioneuromyopathy. Chagas' disease, one of the most common determinants of congestive heart failure and sudden death in the world, is transmitted by the parasite Trypanosoma cruzi (1WHO Expert Committee, Technical Report Series. 1991; 811: 1-10Google Scholar, 2WORLD BANK WHO, Special Programme for Research and Training in Tropical Diseases: VI. Programme Reports. 1982; (, 7, 31, 32): 1Google Scholar). Most patients survive the acute phase of the disease asymptomatically; the chronic stage of the disease develops over 20–30 years and is manifested by cardiovascular, digestive, and autonomic nervous system disorders (3Koberle F. Bol. Org. Panam. Salud. 1961; 51: 404-428PubMed Google Scholar, 4Laranja F.S. Dias E. Nobrega G. Miranda A. Circulation. 1956; 14: 1035-1059Crossref PubMed Google Scholar, 5Mott K.D. Hagstrom J.W.C. Circulation. 1965; 31: 273-286Crossref PubMed Scopus (60) Google Scholar, 6Tafuri W.L. Virchows Arch A. 1971; 354: 136-149Crossref Scopus (27) Google Scholar). Several studies have focused on the disease-related loss of function of the autonomic receptors, the β-adrenergic and muscarinic cholinergic receptors (mAChR) 1The abbreviations used are: mAChR, muscarinic cholinergic receptor; CHO, Chinese hamster ovary; PBS, phosphate-buffered saline; hm2, human m2; DMEM:F-12, Dulbecco's modified Eagle's medium:Ham's F-12; Gpp(NH)p, 5′-guanylylimidodiphosphate; NMS, N-methylscopolamine; QNB, quinuclidinyl-benzilate. 1The abbreviations used are: mAChR, muscarinic cholinergic receptor; CHO, Chinese hamster ovary; PBS, phosphate-buffered saline; hm2, human m2; DMEM:F-12, Dulbecco's modified Eagle's medium:Ham's F-12; Gpp(NH)p, 5′-guanylylimidodiphosphate; NMS, N-methylscopolamine; QNB, quinuclidinyl-benzilate. (5Mott K.D. Hagstrom J.W.C. Circulation. 1965; 31: 273-286Crossref PubMed Scopus (60) Google Scholar, 6Tafuri W.L. Virchows Arch A. 1971; 354: 136-149Crossref Scopus (27) Google Scholar, 7Iosa D. DeQuattro V. De Ping Lee D. Elkayan U. Palmero H. Am. Heart J. 1989; 117: 882-887Crossref PubMed Scopus (57) Google Scholar, 8Machado A.B.M. Machado C.R.S. Gomez M.V. Exp. Parasitol. 1978; 47: 105-115Google Scholar, 9Tanowitz H.B. Davies P. Wittner M. J. Infect. Dis. 1983; 147: 460-466Crossref PubMed Scopus (13) Google Scholar, 10Morris S.A. Rowin K.A. Tanowitz H.B. Wittner M. Bilezikian J.P. Mol. Biochem. Parasitol. 1984; 13: 227-234Crossref PubMed Scopus (9) Google Scholar, 11Morris S.A. Tanowitz H.B. Factor S.M. Bilezikian J.P. Wittner M. Circ. Res. 1988; 62: 800-810Crossref PubMed Scopus (25) Google Scholar). The paradoxical severe involvement of the heart in the absence of any form of the parasite there has prompted proposals that autoimmune mechanisms participate in the pathogenesis of this cardiac neuromyopathy (12Acosta A.M. Santos-Buch C.A. Circulation. 1985; 71: 1255-1261Crossref PubMed Scopus (73) Google Scholar, 13Morris S.A. Tanowitz H.B. Wittner M. Bilezikian J.P. Circulation. 1990; 82: 1900-1909Crossref PubMed Scopus (145) Google Scholar, 14Sterin-Borda L. Borda E.S. Acta Physiol. Pharmacol. Ther. Latinoam. 1994; 44: 109-123PubMed Google Scholar). Chagasic patients have been found to possess circulating antibodies that are able to interact with and modulate the activity of cardiac β-adrenergic and mAChRs (15Borda E.S. Pascual J. Cossio P.M. Vega M. Arana R.M. Sterin-Borda L. Clin. Exp. Immunol. 1984; 57: 679-686PubMed Google Scholar, 16Sterin-Borda L. Perez Leiros C. Wald M. Cremaschi G. Borda E.S. Clin. Exp. Immunol. 1988; 74: 349-354PubMed Google Scholar, 17Goin J.C. Borda E.S. Segovia A. Sterin-Borda L. Proc. Soc. Exp. Biol. Med. 1991; 197: 186-192Crossref PubMed Scopus (28) Google Scholar, 18Goin J.C. Borda E.S. Perez Leiros C. Storino R. Sterin-Borda L. J. Auton. Nerv. Sys. 1994; 47: 45-52Abstract Full Text PDF PubMed Scopus (89) Google Scholar, 19Goin J.C. Perez Leiros C. Borda E.S. Sterin-Borda L. Neuroimmunomodulation. 1994; 1: 284-291Crossref PubMed Scopus (26) Google Scholar). This report focuses on the molecular events initiated by the interaction of the chagasic antibodies with the mAChRs to begin to understand how the antibodies could ultimately account for the abnormalities of cardiac autonomic function characteristic of the late stages of Chagas' disease. Previously, chagasic antibodies were shown to mimic agonists of mAChR to cause inhibition of cardiac contractility, stimulation of cGMP levels, and attenuation of cAMP synthesis (18Goin J.C. Borda E.S. Perez Leiros C. Storino R. Sterin-Borda L. J. Auton. Nerv. Sys. 1994; 47: 45-52Abstract Full Text PDF PubMed Scopus (89) Google Scholar, 19Goin J.C. Perez Leiros C. Borda E.S. Sterin-Borda L. Neuroimmunomodulation. 1994; 1: 284-291Crossref PubMed Scopus (26) Google Scholar). Thus, the in vivo situation during the course of Chagas' disease could be similar to that of a persistent stimulus, i.e. circulating antibodies that activate cardiac mAChRs could result in receptor desensitization. Such a scenario could cause receptor/G-protein uncoupling and loss of receptors from the cell surface and explain the progressive blockade of receptors that is observed in the disease (20Iosa D. DeQuattro V. De Ping Lee D. Elkayan U. Caeiro T. Palmero H. J. Aut. Nerv. Sys. 1990; 30: S83-S88Abstract Full Text PDF PubMed Scopus (37) Google Scholar,21Iosa D. Casadei Massari M. Dorsey F.C. Am. Heart J. 1991; 122: 775-785Crossref PubMed Scopus (27) Google Scholar). To test this hypothesis, we used both purified human m2 mAChRs and mammalian CHO cells transfected with human m2 mAChRs to assess the ability of chagasic IgG to directly interact with and desensitize the mAChR. The results support a role for the chagasic antibodies in causing loss of mAChR function. Trypanosoma cruzi-infected patients from metropolitan Buenos Aires were studied. These patients, classified as Group I based on WHO Expert Committee on Chagas' disease criteria (1WHO Expert Committee, Technical Report Series. 1991; 811: 1-10Google Scholar), had positive T. cruzi serology and the absence of clinical symptoms with normal electrocardiogram and chest x-ray. Among this group, sera were chosen for their previously demonstrated ability to activate rat atrial mAChR in an atropine-sensitive manner (18Goin J.C. Borda E.S. Perez Leiros C. Storino R. Sterin-Borda L. J. Auton. Nerv. Sys. 1994; 47: 45-52Abstract Full Text PDF PubMed Scopus (89) Google Scholar, 19Goin J.C. Perez Leiros C. Borda E.S. Sterin-Borda L. Neuroimmunomodulation. 1994; 1: 284-291Crossref PubMed Scopus (26) Google Scholar). Normal sera taken as controls were from volunteers who had negative serology for Chagas' disease, presented no evidence of cardiovascular or chronic systemic disease or acute viral or febrile disease, and had normal electrocardiogram and chest x-ray. The age of patients and controls ranged between 25 and 50 years. IgG was isolated from chagasic or normal sera by means of DEAE-cellulose chromatography (Bio-Rad) as shown (18Goin J.C. Borda E.S. Perez Leiros C. Storino R. Sterin-Borda L. J. Auton. Nerv. Sys. 1994; 47: 45-52Abstract Full Text PDF PubMed Scopus (89) Google Scholar) and tested for purity by immunoelectrophoresis with goat anti-whole serum or anti-IgG antiserum (Sigma). The IgG concentration was determined by radioimmunodifussion assay. F(ab′)2 fragments were obtained from normal or chagasic patients by pepsin digestion of IgG fractions in 0.1 m sodium acetate buffer (pH 4.5) for 22 h at 37 °C, dialysis against phosphate-buffered saline (PBS), and chromatography on Sephadex G-200. Purification of IgG was assessed by SDS-polyacrylamide gel electrophoresis and immunoelectrophoresis. Human mAChRs (hm2 mAChR) from Spodoptera frugiperda (Sf9) insect cells infected with recombinant baculovirus encoding the human m2 mAChR were purified and reconstituted as described (22Haga K. Haga T. J. Biol. Chem. 1983; 258: 13575-13579Abstract Full Text PDF PubMed Google Scholar, 23Richardson R.M. Kim C. Benovic J.L. Hosey M.M. J. Biol. Chem. 1993; 268: 13650-13656Abstract Full Text PDF PubMed Google Scholar, 24Parker E.M. Kameyama K. Higashijima T. Ross E.M. J. Biol. Chem. 1991; 266: 519-527Abstract Full Text PDF PubMed Google Scholar). The hm2 mAChR purified from Sf9 cells is expressed as a 55-kDa protein, which is smaller than that observed in mammalian cells because it lacks the extensive glycosylation associated with the mammalian expressed receptor (24Parker E.M. Kameyama K. Higashijima T. Ross E.M. J. Biol. Chem. 1991; 266: 519-527Abstract Full Text PDF PubMed Google Scholar). Recombinant virus was kindly provided by Drs. Eric M. Parker and Elliot Ross (University of Texas, Dallas, TX). Chinese hamster ovary cells stably transfected with hm2 mAChRs (m2 CHO cells; Ref. 25Ashkenazi A. Peralta E.G. Winslow J.W. Ramachandran J. Capon D.J. Cell. 1989; 56: 487-493Abstract Full Text PDF PubMed Scopus (174) Google Scholar) were kindly provided by Dr. Ernest Peralta (Harvard University, Cambridge, MA). Cells were grown in Dulbecco's modified Eagle's medium:Ham's F-12 (DMEM:F-12) supplemented with 10% dialyzed fetal bovine serum, 100 units/ml of penicillin and streptomycin, and 2 μmglutamine in the presence of 250 nm methotrexate. Cells were plated in 60- or 100-mm dishes and used at about 80% confluency. The density of mAChRs per cell was determined by saturation binding of [3H]QNB to intact cells (26Luetje C. Brunwell C. Norman M.G. Peterson G.L. Schimerlik M.I. Nathanson N.M. Biochemistry. 1987; 26: 6892-6896Crossref PubMed Scopus (29) Google Scholar). Purified reconstituted hm2 mAChRs were immunoprecipitated with chagasic antibodies or a monoclonal anti-m2 antibody (27Pals-Rylaarsdam R. Xu Y. Witt-Enderby P. Benovic J.L. Hosey M.M. J. Biol. Chem. 1995; 270: 29004-29011Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar) generously provided by Dr. M. Schimerlik from Oregon State University (Corvallis, OR). Chagasic or normal sera or anti-m2 hybridoma supernatant were precoupled to protein A-agarose beads (50 μl, Pierce) overnight at 4 °C. The beads were then incubated with ∼0.7 pmol of reconstituted mAChRs at 4 °C for 5 h and washed thoroughly. The immunoprecipitates were subjected to SDS-polyacrylamide gel electrophoresis using 8.5% gels and then transferred to were by means of the anti-m2 monoclonal antibody and a goat IgG to by the as described (26Luetje C. Brunwell C. Norman M.G. Peterson G.L. Schimerlik M.I. Nathanson N.M. Biochemistry. 1987; 26: 6892-6896Crossref PubMed Scopus (29) Google Scholar). The m2 CHO cells were grown in 100-mm washed with PBS, and then to for at 37 °C with the addition of to induce or To assess the effects of chagasic sera to cause decrease in affinity agonist cells were with a no IgG the agonist or chagasic or normal Cells were incubated for h at 37 °C in a with one of the and extensive were in buffer (pH m were performed at 4 °C. Cells were at for and were as described previously (27Pals-Rylaarsdam R. Xu Y. Witt-Enderby P. Benovic J.L. Hosey M.M. J. Biol. Chem. 1995; 270: 29004-29011Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar). To test for effects of with agonist or IgGs on the affinity of the receptors for the agonist was used to for [3H]QNB binding to mAChR in the were incubated at 37 °C for in buffer (pH 2 [3H]QNB and of in the presence or the absence of the at 100 The were with the D. Biochem. PubMed Scopus Google Scholar). To assess for sequestration of hm2 were performed on intact m2 CHO cells using the ligand to changes in cell surface m2 mAChR the ligand [3H]QNB was used to assess with or IgGs as described the dishes were on and the cells were washed with were at in and assessed for by and with than cells were used for binding of or [3H]QNB were incubated with cells in in the presence or the absence of to binding for 2 h at 4 °C for and for at 37 °C for Cells were and was in a of surface and receptors were as the in cells the as but with the of of or IgG. These were as of assay. The of to or IgG was as the of loss of surface receptors with in with was as a decrease in the receptor as assessed by [3H]QNB binding in cell density per were by on the of To test the ability of chagasic antibodies to directly interact with purified hm2 were performed with chagasic or normal sera and purified hm2 mAChRs reconstituted in a an of the hm2 mAChR were immunoprecipitated with a monoclonal anti-m2 mAChR antibody (26Luetje C. Brunwell C. Norman M.G. Peterson G.L. Schimerlik M.I. Nathanson N.M. Biochemistry. 1987; 26: 6892-6896Crossref PubMed Scopus (29) Google Scholar). The immunoprecipitates were by SDS-polyacrylamide gel and immunoprecipitated mAChR was by with the anti-m2 mAChR monoclonal antibody The results demonstrated that the chagasic sera immunoprecipitated the hm2 mAChR and were by the anti-m2 mAChR one to a protein of the for the hm2 mAChR purified from Sf9 and a at that the receptor (24Parker E.M. Kameyama K. Higashijima T. Ross E.M. J. Biol. Chem. 1991; 266: 519-527Abstract Full Text PDF PubMed Google Scholar). The that were in the immunoprecipitates from the chagasic sera to immunoprecipitated by the m2 The of mAChR immunoprecipitated by the chagasic was than the of receptors immunoprecipitated by the m2 in the of mAChR immunoprecipitated by the chagasic sera was and than the immunoprecipitated by the normal sera used as and These results provide a of the of the hm2 mAChR by antibodies in chagasic To test whether the chagasic sera directly with the ligand binding of the m2 mAChR or with we tested whether the chagasic sera could with the antagonist [3H]QNB in ligand binding studies we from CHO cells expressing the hm2 mAChR and performed saturation binding in and with normal or chagasic IgG. in the or were observed the of normal and chagasic IgG on the binding of the antagonist were from CHO cells expressing the hm2 mAChR as described in the and incubated in the presence or the absence of normal or chagasic IgG for at 37 °C. The were then washed with binding buffer and used in saturation binding with [3H]QNB The results are the means of The were with the D. Biochem. PubMed Scopus Google Scholar). in a were from CHO cells expressing the hm2 mAChR as described in the and incubated in the presence or the absence of normal or chagasic IgG for at 37 °C. The were then washed with binding buffer and used in saturation binding with [3H]QNB The results are the means of The were with the D. Biochem. PubMed Scopus Google Scholar). The that the persistent of m2 mAChRs by chagasic antibodies could to receptor was of receptors is associated with in receptor/G-protein uncoupling and sequestration of the receptors from their normal of receptor uncoupling from is the of receptors from a to affinity The affinity for agonists is to be a receptor/G-protein the affinity is to the In agonist binding studies in a loss of affinity agonist binding can be the addition of or to the of the and of the from the of the β-adrenergic receptor has been associated with the loss of affinity agonist binding and of receptors to a affinity J.P. J. Biol. Chem. 1984; Full Text PDF PubMed Google Scholar). To test for effects of chagasic antibodies on agonist binding we the effects of of intact cells with the agonist acetylcholine or normal or chagasic IgG on agonist were and used in binding in the presence or the absence of a In of from cells with agonist were obtained in the absence of Gpp(NH)p, both and affinity agonist binding 2 The addition of and the as for the of affinity affinity 2 The and changes In of the with the D. Biochem. PubMed Scopus Google Scholar) demonstrated the in that a and affinity were observed and a of receptors from the to affinity of cells with the agonist acetylcholine that to (27Pals-Rylaarsdam R. Xu Y. Witt-Enderby P. Benovic J.L. Hosey M.M. J. Biol. Chem. 1995; 270: 29004-29011Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar) or with chagasic IgG of the of in the absence of with cells to 2 The effects by were not as extensive as by the addition of to the in The digestion of chagasic had similar effects to the chagasic IgG not The by chagasic IgG was not observed were performed with normal IgG 2 of the with the that the decrease in affinity by the chagasic IgG was to an in the of there was no in the of receptors in the affinity of the cells with agonist a similar The that the of the cells by agonist or chagasic IgG effects from because the of receptors in the affinity was by the addition of to the in The obtained with the normal IgG were similar to from the cells and for the agonist from from shown are and for the in The results are shown from the obtained in the absence and the presence of and are presented as the means the to 2 for from from in a obtained from of of m2 CHO of from no shown in 2 were with the D. Biochem. PubMed Scopus Google Scholar). and for binding to and affinity of the and to the of receptors in affinity are the means from similar to the one shown in not from no in a The shown are and for the in The results are shown from the obtained in the absence and the presence of and are presented as the means the to 2 for The shown in 2 were with the D. Biochem. PubMed Scopus Google Scholar). and for binding to and affinity of the and to the of receptors in affinity are the means from similar to the one shown in not is not the effects of the with agonist or chagasic IgG in the decrease in H. is that the chagasic IgG or agonist the in studies of the of the β-adrenergic similar results were obtained J.P. J. Biol. Chem. 1984; Full Text PDF PubMed Google Scholar). a decrease in of affinity β-adrenergic receptors was and the to be a in J.P. J. Biol. Chem. 1984; Full Text PDF PubMed Google Scholar) as was observed The results were similar to we previously obtained in heart with of for agonist binding to the mAChR was the of affinity receptors was M.M. Hosey M.M. J. Biol. Chem. Full Text PDF PubMed Google Scholar). In we observed similar in studies with expressed hm2 mAChR in Sf9 cells R.M. Hosey M.M. J. Biol. Chem. Full Text PDF PubMed Google Scholar). that the effects of on the affinity of agonist binding to the m2 mAChR are extensive than can be in the addition of to ligand binding studies be to of that are by desensitization. the results are with the that acute exposure of the m2 CHO cells to the chagasic IgG in a affinity of the receptor for These effects of the chagasic IgG were similar to by exposure to The results are with the that the activity of the chagasic sera can induce of the m2 mAChR in intact To test if chagasic IgG sequestration of receptors, cells expressing m2 mAChRs were incubated for with of chagasic or normal IgGs or F(ab′)2 binding were with acetylcholine were in parallel. The ligand and the ligand [3H]QNB were used in cell binding to assess cell surface and receptors, the chagasic IgG and acetylcholine a sequestration of the mAChRs to a similar normal IgG had no The effects of both acetylcholine and the chagasic IgG were the observed with the chagasic IgG at and was similar to that by 100 nm acetylcholine The chagasic F(ab′)2 but with a Normal IgG was at any concentration In the no of receptors by chagasic IgGs, normal IgGs, or acetylcholine was observed based on of mAChR with [3H]QNB not The report evidence of a interaction between antibodies from the serum of chagasic patients and hm2 mAChRs and of the persistent of the receptors by the The results obtained support the hypothesis that antibodies having an activity can participate in the of the receptors their normal The decrease in agonist affinity h of of m2 CHO cells with chagasic IgGs or was extensive by similar to the of The demonstrate that the of the antibodies is to induce the of associated with of the The results are with on the ability of chagasic IgG to agonist-induced inhibition of of rat with IgGs (18Goin J.C. Borda E.S. Perez Leiros C. Storino R. Sterin-Borda L. J. Auton. Nerv. Sys. 1994; 47: 45-52Abstract Full Text PDF PubMed Scopus (89) Google Scholar). is that the results could have from because the in demonstrated an ability of the chagasic antibodies to directly interact with the m2 mAChR. In studies have effects of chagasic serum on signaling (15Borda E.S. Pascual J. Cossio P.M. Vega M. Arana R.M. Sterin-Borda L. Clin. Exp. Immunol. 1984; 57: 679-686PubMed Google Scholar, 16Sterin-Borda L. Perez Leiros C. Wald M. Cremaschi G. Borda E.S. Clin. Exp. Immunol. 1988; 74: 349-354PubMed Google Scholar, 17Goin J.C. Borda E.S. Segovia A. Sterin-Borda L. Proc. Soc. Exp. Biol. Med. 1991; 197: 186-192Crossref PubMed Scopus (28) Google Scholar, 18Goin J.C. Borda E.S. Perez Leiros C. Storino R. Sterin-Borda L. J. Auton. Nerv. Sys. 1994; 47: 45-52Abstract Full Text PDF PubMed Scopus (89) Google Scholar, 19Goin J.C. Perez Leiros C. Borda E.S. Sterin-Borda L. Neuroimmunomodulation. 1994; 1: 284-291Crossref PubMed Scopus (26) Google Scholar), the CHO cells used in this receptors, it that the observed effects of the chagasic antibodies on the mAChR were to from effects on signaling of the Chagasic patients of function L. Borda E.S. Acta Physiol. Pharmacol. Ther. Latinoam. 1994; 44: 109-123PubMed Google the molecular mechanisms Thus, the of report is that events of of m2 mAChR can be initiated by autoantibodies that to and activate the studies be to the of whether molecular in the function of the cardiac autonomic nervous system during Chagas' disease are to the presence of autoantibodies receptor desensitization. the of autoantibodies against autonomic receptors has been in such as human C. Circ. Res. 1989; PubMed Scopus Google Scholar, Y. F. A. J. J. Clin. 1993; PubMed Scopus Google Scholar, M. A. J. J. 1994; PubMed Scopus Google Scholar) and heart Sterin-Borda L. Arana R.M. Borda E.S. J. 1994; PubMed Scopus (27) Google Scholar), but evidence is about the mechanisms their or not autoantibodies a role in remains to be for and of the m2 mAChR.