Muscarinic Receptors in Developing Rat Colon

Muscarinic receptor subtypes were characterized in fetal (21 day), newborn (3 day), and adult (3 month) rat colon smooth muscle. Saturation binding of the nonselective muscarinic antagonist radioligand [3H]quinuclidinyl benzilate revealed a single class of binding sites in all three age groups. The binding affinities of [3H]quinuclidinyl benzilate were not significantly different among three age groups (KD: 0.19-->0.27 nM). In contrast, the receptor densities (Bmax, fmol/mg protein) showed a significant age-related decrease with fetus (518.9 +/- 7.4) > newborn (480.3 +/- 45.6) >> adult (192.4 +/- 32.8). In both newborn and adult tissues, the muscarinic agonist carbachol bound to two sites with high and low affinities. Although the agonist binding affinities in the newborn tissue were not significantly different from those in the adult tissue, the high-affinity binding sites for carbachol were significantly increased in the later (41%-->61%). Addition of guanosine-5'-O-(3-thio)triphosphate (100 microM) abolished apparent high-affinity binding sites in both newborn and adult tissues. Antagonist competition binding in the newborn tissue indicated a homogeneous population of muscarinic M2 receptors. Unlike in newborn tissues, the heterogeneous binding of pirenzepine and 4-diphenylacetoxy-N-methylpiperidine methobromide in adult tissues revealed coexistence of muscarinic M3 (45%) and M2 (55%) receptors. In accordance, activation of muscarinic receptors in the adult tissue stimulated synthesis of inositol 1,4,5-trisphosphate. These results suggest maturational changes of muscarinic receptor subtypes and their coupling to G proteins in rat colonic smooth muscle. These changes may account, at least in part, for developmental alterations of functional responses in colonic smooth muscle.