Modulation of thromboxane receptor activation in rat glomerular mesangial cells

R. F. Spurney, J. P. Middleton, J. R. Raymond and T. M. Coffman Department of Medicine, Duke University, Durham, North Carolina. Rat glomerular mesangial cells were used to investigate mechanisms of thromboxane A2 (TxA2) receptor regulation in the kidney. Exposure of mesangial cells to the TxA2 agonist U-46619 for 10 min reduced subsequent TxA2-induced increases in inositol phosphates and intracellular Ca2+ levels by approximately 70%. This loss of receptor responsiveness could be blocked by the TxA2 receptor antagonist SQ-29548 and was reversible after removal of agonist from the incubation medium. Radioligand binding studies using the TxA2 agonist [125I]BOP suggested that exposure of mesangial cells to U-46619 for 10 min reduced TxA2 receptor responsiveness without a loss of receptor sites from plasma membrane fractions of the cell, although the density of mesangial cell TxA2 receptors was decreased by approximately 60% after more prolonged exposure of mesangial cells to thromboxane agonists. Both desensitization to U-46619 and loss of TxA2 binding sites could be attenuated by the protein kinase C (PKC) inhibitors staurosporine, sphingosine, or H-7, and TxA2 receptor responsiveness was reduced in cells incubated with phorbol esters before stimulation with thromboxane agonists. We conclude that 1) agonist-specific decreases in TxA2 receptor responsiveness may involve initial uncoupling of the receptor from its effector systems, followed by a loss of TxA2 receptor sites from plasma membrane fractions of the cell, and 2) PKC may be involved in these processes.