Regulation of prolactin mRNA by 1,25-dihydroxyvitamin D3 in GH4C1 cells

GH4C1 cells, which possess specific cytosolic binding for 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3), have previously been found to respond to this hormone with a highly selective increase in prolactin synthesis. Here we extend our previous observations by measuring the effect of 1,25-(OH)2D3 on the accumulation of specific mRNA in GH4C1 cells, using cytoplasmic dot hybridization. In a hormone-free, chemically defined medium containing 0.4 mM Ca2+, stimulation of prolactin mRNA accumulation was 200-215% of control after 96 h of treatment with 1 X 10(-9) M 1,25-(OH)2D3. Thyrotropin-releasing hormone (1 X 10(-7) M) caused a similar increase in prolactin mRNA (206% of control). Neither hormone increased growth hormone mRNA. Stimulation of prolactin mRNA by 1,25-(OH)2D3 was 163% of control after 48 h and 200% after 96 h. No consistent effect was observed in the first 24 h of treatment. The same response to 48 h treatment with 1,25-(OH)2D3 was observed whether the cells were withdrawn from serum-containing medium for 24 h (as above) or 72 h. In cells withdrawn for 96 h, 1,25-(OH)2D3 raised prolactin mRNA within 24 h. The effect of 1,25-(OH)2D3 was significant at a concentration of 10(-11) M, and half-maximal at approximately 10(-10) M. In low-Ca2+ medium, the response to greater than 10(-10) M 1,25-(OH)2D3 was eliminated. Extracellular Ca2+ (0.4 mM) increased prolactin mRNA in the presence of, but not in the absence of, 1,25-(OH)2D3. These findings demonstrate that 1,25-(OH)2D3 enhanced prolactin gene expression with a distinctive time course and interactively with calcium.