IGF-I causes an ultrasensitive reduction in GH mRNA levels via an extracellular mechanism involving IGF binding proteins

IGF-I-dependent decreases in endogenous GH mRNA expression were studied in individual rat MtT/S somatotroph cells using in situ hybridization. It was first shown that increasing IGF-I concentrations (0-90 nm) decreased GH mRN A levels in a ultrasensitive manner when averaged over the entire populatio n, such that the decrease occurred over a narrow range of IGF-I concentrati on with an EC50 of 7.1 nm. The degree of ultrasensitivity of the population average was expressed by calculating the Hill coefficient (nA), which had a value of -2.0. GH mRNA levels in individual dispersed cells from these cu ltures were then measured. These results were first summed for all cells to show that the average response of the population remained ultrasensitive ( nA = -2.6, EC50 = 8.1 nm). Then, parameters for individual cells of the pop ulation were calculated using mathematical modeling of the distribution of individual cell GH mRNA levels after treatment with 0-90 nm IGF-I. Solution of the data from the individual cells yielded a Hill coefficient (nl = -0. 65) and a heterogeneity coefficient (ml = -1.2) indicative of individual ce ll responsiveness to IGF-I that was not ultrasensitive and very heterogeneo us. These results suggested that ultrasensitivity in the population may lik ely be caused by an extracellular mechanism regulating IGF-I concentrations , such as IGF binding proteins. Increasing concentrations of long (Arg)(3)I GF-1, an analog that binds the IGF type-1 receptor but not IGF binding prot eins, showed a linear inhibition of GH mRNA levels. Treatment with IGF bind ing protein ligand inhibitor, an IGF-I analog that binds to IGF binding pro teins but not the IGF type-1 receptor, decreased GH mRNA levels in the abse nce of exogenous IGF-I. Thus, IGF binding proteins provide the extracellula r sequestration of IGF-I necessary for the precise and ultrasensitive regul ation of GH mRNA levels in the entire cell population, although expression within individual cells is regulated in a graded fashion.