Barbiturate inhibition of growth and of the synthesis of deoxyribonucleic acid, ribonucleic acid and protein in cultured mammalian cells

Abstract

Pentobarbital (PB) inhibited growth and the synthesis of nucleic acids and protein in murine, mastocytoma cells (P815Y) grown in culture. The inhibition increased with an increase in the concentration of drug and was also time-dependent with a high level of drug. For example, 0.5 mM PB ( id 50) reduced both the rate of division of the cells and the synthesis of DNA, RNA and protein by about one-half, compared with the control, over a 12-hr period. In contrast, treatment with a 2-fold higher concentration of PB (1 mM; id 100) blocked both cell division and the synthesis of protein promptly. It also reduced the synthesis of DNA and RNA by about one-half, compared with the control, during the first 4 hr of treatment. After this time, however, the synthesis of both DNA and RNA stopped abruptly. It is concluded that the inhibition of DNA synthesis caused by barbiturate in the latter case may have been secondary to the inhibition of protein synthesis. Transfer of the inhibited (1 mM PB; 12 hr) cells to drugfree medium caused the synthesis of protein and RNA to begin without apparent delay. In contrast, the synthesis of DNA proceeded slowly for about 8 hr; then the amount of DNA increased parasynchronously. Cell division, which also proceeded slowly during the first 10 hr, occurred as a parasynchronous wave some 2 hr after DNA synthesis began. The inhibitory effects of PB were also studied in murine, lymphoblastic cells (L5178Y) synchronized by sequential treatment with thymidine (5 hr) and deoxycytidine plus Colcemid (5 hr). When the mitotically inhibited cells were transferred to normal medium containing PB (1.5 mM; id 100), slightly more than one-half of the cells failed to complete mitosis and the synthesis of DNA, RNA and protein was blocked by the drug. The synthesis of DNA, RNA and protein was also blocked by PB addition in the middle of the S-phase. Its addition at the onset of the second wave of mitosis prevented mitosis as before and blocked the initiation of DNA synthesis.