Characterization of cholinergic muscarinic receptors in the rabbit iris.

The sphincter smooth muscle of the iris is innervated by excitatory parasympathetic nerve fibers, and the activation of these fibers results in the breakdown of phosphatidylinositol 4,5-bisphosphate into its derived second messengers, myosin light chain phosphorylation and muscle contraction. The present study characterizes the muscarinic acetylcholine receptors (mAChRs) of the rabbit iris employing [3H]N-methylscopolamine ([3H]NMS) and L-[3H]quinuclidinyl benzilate ([3H]QNB) as probes. Binding studies indicated that [3H]NMS and [3H]QNB bound to homogeneous populations of mAChRs with apparent Bmax values of 0.67 and 1.09 pmol/mg protein respectively. Binding of radioligands was rapid, saturable, stereospecific, reversible, and inhibited by specific muscarinic agonists and antagonists in a competitive manner. [3H]NMS displayed a lower amount of nonspecific binding and a faster association and dissociation rate than [3H]QNB. The relative potencies for displacement of both radioligands, based on their Ki values, were (-)QNB greater than atropine greater than (+)QNB greater than pirenzepine greater than pilocarpine. Antagonist displacement of the radioligands appeared to obey the law of mass action, indicating interaction with a single binding site. However, displacement of the radioligands by the agonists carbamylcholine and methacholine indicated interaction with both high and low affinity binding sites. Comparison of the displacement of [3H]NMS and [3H]QNB by pirenzepine in microsomal fractions from rabbit iris, ileal muscle and cerebral cortex revealed the presence of a single subtype of mAChR in the iris which had an affinity for PZ that was slightly higher than that of ileal M2 receptors, but lower than that of brain M1 receptors. This suggests that the mAChRs in the iris may represent a subclass of receptors within the M2 subtype, or they may constitute an entirely different subtype of mAChRs.