Rat thymocytes incubated in RPMI medium for 120-300 min release a soluble factor of molecular weight below 10,000 Da which inhibits the binding of 3H-labeled muscarinic antagonists in an uncompetitive manner, i.e. it reduces maximal specific binding (Bmax) without changing the affinity of the ligand (KD). This factor inhibited muscarinic antagonist binding on thymocytes and rat cerebral cortex cellular membranes. Thymocytes from hydrocortisone-treated rats produced more factor per mg cell protein than thymocytes from untreated rats. The activity of the factor was unaffected by incubation with Cd2+ (1 mM) or Zn2+ (1 mM) or EDTA (1 mM), and its protein nature is supported by the following findings: it was trypsin sensitive, heat denaturated at 56 degrees C and precipitated by (NH4)2SO4 (40%, w/v). Inhibition by the factor was apparently irreversible after 1 h of incubation.