Cyclic nucleotides and growth regulation in suspension cultures of mammalian cells

Abstract

Cyclic(c)AMP and cyclic(c)GMP were measured by radioimmunoassay in suspension cultures of WRL-10A cells, a subline of L-929 mouse cells of fibroblastic origin, in which prolonged arrest in the early G1 or G0 phase and reactivation of growth is determined by the availability of small MW nutrients in relation to population density. In high-density growth-inhibited cell population cAMP (.apprx. 3.4 pmol/mg protein) was 50% lower than in low-density exponentially growing cultures, whereas cGMP (.apprx. 0.3 pmol/mg protein) was the same in both populations. With initiation of growth by dilution of high-density cultures, cAMP and cGMP rose promptly with a cAMP peak (3.1-fold increase) and a cGMP peak (1.8-fold increase) before initiation of DNA synthesis. Growth inhibition during medium depletion was accompanied by a slightly lower cAMP and by a transient 2-fold rise in cGMP. After initiation of growth by medium renewal in the depleted cultures, cAMP rose and cGMP fell. In both cases the cAMP/cGMP ratios fell with growth inhibition and rose with initiation of growth. This behavior appears to be a reflection of cell synchrony and of changes in ATP levels characteristic of growth inhibition and reactivation in this cell culture system. These results, in conjunction with related findings by others, lead to the conclusion that the most consistent relationship between growth regulation and cyclic nucleotides is the elevation of cAMP observed in growth inhibition caused by serum restriction.