Localization of muscarinic receptors in human heart biopsies using rabbit anti-peptide antibodies

Abstract

Immunocytochemistry of muscarinic receptors on human heart biopsies from patients with heart disease was studied using rabbit antibodies against a synthetic peptide corresponding to amino acids 168-192 of the second extracellular loop of the human M2 muscarinic receptor. By using both light and electron microscopic immunocytochemistry techniques, muscarinic receptors were visualized on sarcolemma of human myocytes from patients with different heart diseases such as coronary heart disease and dilated cardiomyopathy in adults and congenital heart disease in children. The patchy distribution of immunoreactivity suggests a muscarinergic activity in vivo. These reactivities were abolished by preincubation of antibodies with antigenic peptide and were not shown in the absence of antibodies. Moreover, these antibodies were able to interfere with muscarinic ligand binding in myocardium from human dilated cardiomyopathy as shown by decreases in binding sites and antagonist affinity. These results demonstrate that the antibodies against the second extracellular loop of the human M2 muscarinic receptor can specifically recognize muscarinic receptors in human tissue and display pharmacological activity in human diseased myocardium, confirming their usefulness for the study of localization and function of muscarinergic activity in the human heart.