Immunocytochemical localization of M2 muscarinic receptors in rat ventricles with anti-peptide antibodies.
We produced antibodies against a synthetic peptide corresponding to amino acids 168-192 of the second extracellular loop of the M2 human muscarinic receptor in rabbits. In immunoblot, affinity-purified antibodies specifically recognized a major band of rat ventricular muscarinic receptor protein with a molecular weight of about 80 KD. This recognition could be blocked by pre-incubation with peptide. Moreover, with both light (LM) and electron microscopic (EM) immunocytochemistry techniques, muscarinic receptors were detected on sarcolemma and T-tubules of rat cardiomyocytes. In addition, immunoreactions were localized in membranes of capillaries. Likewise, these reactivities were abolished by pre-incubation with peptide. These results suggest that the antibodies against the second extracellular loop of human M2 muscarinic receptor could specifically recognize rat ventricular muscarinic receptor protein and could be a powerful tool to study the fate of this receptor under different pathological or physiological conditions.
Fu
M
M
Wallenberg Laboratory, Sahlgren's Hospital, University of Göteborg, Sweden.
Schulze
W
W
Wolf
W P
WP
Hjalmarson
A
A
Hoebeke
J
J
D000109:Acetylcholine / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D005455:Fluorescent Antibody Technique; D006352:Heart Ventricles / Q000737:chemistry* / Q000648:ultrastructure; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D016253:Microscopy, Immunoelectron / Q000379:methods; D008969:Molecular Sequence Data; D010455:Peptides; D011817:Rabbits; D051381:Rats; D011976:Receptors, Muscarinic / Q000032:analysis* / Q000737:chemistry / Q000276:immunology
D000906:Antibodies; D010455:Peptides; D011976:Receptors, Muscarinic; D000109:Acetylcholine