The influence of cortisol on intestinal DNA-dependent RNA polymerase activity was studied in purified nuclei of vitamin D-deficient or 1,25-dihydroxyvitamin D3-treated chicks. Six- to 7-week-old vitamin D-deficient cockerels were given 5 mg of cortisol or vehicle intraperitoneally 24 and 48 hours before sacrifice. Three hours before sacrifice, 200 ng of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) was administered intracardially. Cortisol did not alter the uptake or metabolism of 1,25(OH)2D3 in the intestinal mucosa. After a 200 ng dose of 1,25(OH)2D3 the in situ intestinal ligated loop technique revealed a 39% increase in calcium absorption compared to control birds (P less than 0.001). The administration of cortisol (5 mg) to chickens given 1,25(OH)2D3, however, resulted in a significant decrease in intestinal calcium transport in vivo (P less than 0.0025). When intestinal nuclei were prepared from birds treated in a manner identical with that described above, 1,25(OH)2D3-treated and 1,25(OH)2D3 plus cortisol-treated chicks had intestinal RNA polymerase II transcriptional activities that were significantly greater than those of vitamin D-deficient controls (P less than or equal to 0.02, P less than or equal to 0.005). There was no difference between RNA polymerase II and I + III activities of the 1,25(OH)2D3-treated birds and that of the cortisol plus 1,25(OH)2D3-treated birds. Vitamin D-deficient chicks treated with cortisol alone showed RNA polymerase I + III activity that was significantly higher (P less than or equal to 0.01) than that of birds treated with vehicle alone.(ABSTRACT TRUNCATED AT 250 WORDS)