We have investigated the regulation of mRNA levels of alpha- and beta-subunits of guanine nucleotide-binding regulatory proteins (G proteins) by peptide hormones in prolactin producing rat pituitary adenoma cells (GH3 cells) in culture. The cells were treated with thyroliberin (1 microM), vasoactive intestinal peptide (1 microM) or somatostatin (10 microM) for 6 to 48 hours. Thyroliberin and vasoactive intestinal peptide increased the levels of Gs alpha Go alpha, Gi-2 alpha, Gi-3 alpha, Gx alpha, G beta 36 and mRNAs. The effect of vasoactive intestinal peptide was however earlier and more pronounced. Gi-2 alpha mRNA levels showed the quantitatively largest alterations. Somatostatin upregulated Gs alpha and downregulated Go alpha and Gi-2 mRNAs. G protein mRNAs for Gi-2 alpha and Go alpha were increased by exposure of the cells to a medium devoid of serum. We conclude that G protein mRNA levels are subjected to alterations by hormones that act through the corresponding G proteins in the regulation of prolactin synthesis and secretion.