The detection of the non-M2 muscarinic receptor subtype in the rat heart atria and ventricles

Mammal heart tissue has long been assumed to be the exclusive domain of the M-2 subtype of muscarinic receptor, but data supporting the presence of other subtypes also exist. We have tested the hypothesis that muscarinic receptors other than the M-2 subtype are present in the heart as minor populations. We used several approaches: a set of competition binding experiments with pirenzepine, AFDX-116, 4-DAMP, PD 102807, p-F-HHSiD, AQ-RA 741, DAU 5884, methoctramine and tripinamide, blockage of M-1 muscarinic receptors using MT7 toxin, subtype-specific immunoprecipitation experiments and determination of phospholipase C activity. We also attempted to block M-1-M-4 receptors using co-treatment with MT7 and AQ-RA 741. Our results show that only the M-2 subtype is present in the atria. In the ventricles, however, we were able to determine that 20% (on average) of the muscarinic receptors were subtypes other than M-2, with the majority of these belonging to the M-1 subtype. We were also able to detect a marginal fraction (6 +/- 2%) of receptors that, based on other findings, belong mainly to the M-5 muscarinic receptors. Co-treatment with MT7 and AQ-RA 741 was not a suitable tool for blocking of M-1-M-4 receptors and can not therefore be used as a method for M-5 muscarinic receptor detection in substitution to crude venom. These results provide further evidence of the expression of the M-1 muscarinic receptor subtype in the rat heart and also show that the heart contains at least one other, albeit minor, muscarinic receptor population, which most likely belongs to the M-5 muscarinic receptors but not to that of the M-3 receptors.