The corticosteroid receptor was investigated in isolated rat adipocytes with a new technique which characterizes the corticosteroid receptors that can be activated and tightly bound to the nucleus. The binding reaction with [3H]triamcinolone was performed with intact isolated adipocytes and the radioactivity associated with the nucleus was subsequently determined after cell lysis. Scatchard analysis revealed a homogeneous class of nuclear corticosteroid receptors in rat epididymal adipocytes with an apparent Kd of 4.93 ± 1.5 mM and a Bmax of 21.8 ± 6.6 fmol/106 cells corresponding to about 13 000 receptors per nucleus. The corticosteroid binding exhibited regional variations in isolated adipocytes. The highest receptor number was found in epididymal adipocytes (Bmax 25.8 ± 3.9 fmol/106 cells) whereas there were significantly lower nuclear binding sites in perirenal adipocytes (16.5 ± 5.5 fmol/106 cells) (P < 0.05) and subscutaneous adipocytes (4.8 ± 1.5 fmol/106 cells) (P <0.01). The apparent affinity in the three fat depots were similar with Kd values about 4 nM. The nuclear corticosteroid receptor in adipocytes was steroid specific, as neither unlabelled estradiol nor testosterone were able to displace the [3H]triamcinolone binding at concentrations up to 100 μM. However, unlabelled progesterone and promegestrone (R5020) were able to compete with triamcinolone-binding (by 50-80%). In order to investigate whether the nuclear corticosteroid binding in adipocytes were under influence of other hormones we examined the effects of lipocytic and antilipolytic compounds on the binding. Preincubation with isoproterenol and dibutryl-cAMP for 1 h was able to decrease the corticosteroid binding by 30-50%. However, the antilipolytic hormone insulin had no effect in preincubations performed for up to 2 h. In conclusion, high affinity nuclear corticosteroid receptors were found in rat adipocytes. These receptors exhibited regional variations and were modulated by lipolytic hormones.