Endocrinology, Vol 112, 1964-1972, Copyright © 1983 by Endocrine Society

ARTICLES

Effect of environmental hydrogen ion concentration on regulation of insulin receptors in cultured R3230AC mammary carcinoma cells

LK Sorge and R Hilf

Insulin binding and responsiveness in primary cultures of R3230AC rat mammary tumor cells were studied as a function of the hydrogen ion concentration of the culture medium. When insulin binding was assayed at pH 7.4, cultures that were maintained for a sufficient length of time in acidic medium demonstrated a significant increase in insulin receptor concentration compared to control cultures. The increase in insulin binding, which occurred as cultures approached confluency, was attributed to the increased acidity rather than nutrient depletion of the culture medium. In contrast, maximum binding was observed when the pH of the assay buffer was above 8.0, independent of the culture conditions. Binding of Concanavalin A, reflecting more generalized cell surface glycoproteins, decreased as cultures approached confluency, but was unaffected by the pH of either the culture medium or the assay buffer. The effect of pH on insulin responsiveness was studied. Insulin receptors generated by acidic culture conditions demonstrated insulin- induced down-regulation. Regardless of the pH environment, all cells demonstrated the same amount of insulin binding after exposure to 10(- 6) M insulin. Under the in vitro conditions employed, cultured cells did not demonstrate a significant response to added insulin by alteration in growth, substrate transport, or incorporation of precursors into macromolecules, although the basal rates of these parameters were lower in cells maintained in acidic pH environments. The data presented indicate the necessity of considering the pH of the culture medium in studies of receptor regulation. It is possible that tumor cells, due to increased lactic acid production, may be especially prone to these changes.