Using the in vitro autoradiographic technique with tritium-sensitive LKB sheet film and the liquid scintillation counting method, the distribution and the binding parameters of the muscarinic cholinergic receptors (MChR) were determined in various discrete regions of the rat brain. The results obtained in the present study were as follows: (1) Specific binding of [3H]QNB to the slide-mounted tissue sections increased slowly when incubated at room temperature; saturation occurred 2 h after incubation. Only 23% of [3H]QNB bound to the tissue section was dissociated 5 h after the addition of 20 microM atropine to the medium. These findings were very different from those obtained in the study using the tissue homogenates. (2) The regional distribution of MChR was determined using both autoradiographic and liquid scintillation counting methods. The distribution of MChR was heterogeneous, with highest densities in the striatum and nucleus accumbens and lowest in the globus pallidus, nucleus interpeduncularis and nucleus septi. Moreover, MChR were unevenly distributed within the subfields of each region. (3) In saturation binding studies using the slide-mounted tissue sections of 20 micron thickness the (Kd)app-values were similar but not exactly identical in 5 discrete regions, i.e. the striatum, somatosensory cortex, hippocampus (the subiculum + CA1 field), nucleus accumbens and gyrus dentatus, determined in the present study. The (Kd)app-value of each region was about 700 pM which was about 20 times higher than that obtained in the study using the tissue homogenates. However (Kd)app-values obtained with 5 and 10 micron tissue sections were approximately 3-fold lower.