Muscarinic acetylcholine receptors in cat iris

The binding of tritium labeled N-methyl-4-piperidylbenzilate([³H]-4-NMPB) to fractions obtained from cat irides was investigated. The binding of this highly potent muscarinic antagonist to the cat iris consisted of two components: (a) high affinity binding sites with low capacity and (b) low affinity binding sites with high capacity. By a simple fractionation procedure those two components were reasonably separated, and a relatively pure fraction with high affinity binding sites was obtained. The high affinity binding sites are attributed to the muscarinic receptor sites, since they showed characteristic behaviour: saturability, high affinity, reduction of the binding by muscarinic ligands, and no effect on binding by non-muscarinic ligands (at a pharmacological range of concentration). Hill coefficients for the antagonists were approximately 1, and for the agonists less than 1. The dissociation constant of [³H]-4-NMPB (0.4-0.6 nM) is similar to that found in albino rabbit irides and in mouse brain. Furthermore, the capacity of the high affinity binding sites (468 fmole/iris) was similar to that found in albino rabbit irides (447 fmole/iris) and in pigmented rabbits irides (512 fmole/iris). According to these findings, the low affinity binding sites cannot be related to the muscarinic receptor and are probably related to the presence of abundant melanin pigment. The interactions of [³H]-4-NMPB with the components of the cat irides and their possible physiological significance are discussed.