Insulin and insulin-like growth factor (somatomedin) receptors on cloned rat pituitary tumor cells

Specific receptors for insulin and the somatomedin peptides insulin-like growth factors I and II (IGF-I and IGF-II) have been characterized on three separate cloned strains of rat pituitary tumor cells (GH₃, GH₁, and GC). Binding of ¹²⁵I-labeled peptides was time, temperature, and pH dependent for all three cell lines. Specific binding of [¹²⁵I]insulin, which was extremely low in normal rat adenohypophyseal cells, was demonstrable for all three lines, with the K_d for the high affinity receptor ranging from 10^-10 to 4 × 10^-10 M/liter. A specific high affinity IGF-I receptor was also identified, with a K_d of approximately 10^-9 M/liter. IGF-II and insulin were, respectively, 10% and 1% as potent as IGF-I in competing for this receptor. When [¹²⁵I]insulin and [¹²⁵I]IGF-I were cross-linked to GH₃ cells with disuccinimidyl suberate, followed by sodium dodecyl sulfatepolyacrylamide gel electrophoresis, both receptors were found to have an apparent mol wt greater than 300, 000 in the unreduced state, with subunits of apparent mol wt 125, 000 after reduction. A third discrete receptor, which bound [¹²⁵I]IGF-II, was also identified on all three cell lines. IGF-I was only 10% as potent as IGF-II at displacing [¹²⁵I]IGF-II, and insulin was virtually unreactive. When [¹²⁵I]IGF-II was cross-linked to GH3 cells and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, two receptors were identified. One had an apparent mol wt of 205, 000 unreduced and 250, 000 upon reduction, and presumably represents the type II receptor. Additionally, a band was observed at an apparent mol wt greater than 300, 000 unreduced and 125, 000 upon reduction, probably representing IGFII binding to the IGF-I or insulin receptor. The presence of specific high affinity receptors for insulin, IGF-I, and IGF-II in these transformed cell lines is consistent with previous observations in normal rat and human pituitary cells and suggests a role for these peptides in the modulation of pituitary function.