Abstract
The culture of Nil hamster fibroblasts in MEM lacking nicotinamide (NAm^β^ MEM) leads to: (1) the rapid loss of intracellular total nicotinamide adenine dinucleotide (NAD(H)) content in these cells from a level of 150β200 pmoles/10^5^ cells to less than 20 pmoles/10^5^ cells; (2) the cessation of cell division and inhibition of DNA synthesis; and (3) a reduction of glucose consumption and lactic acid production. In most situations, following nicotinamide starvation, the restoration of intracellular NAD(H) follows rapidly the readdition of NAD^+^ (oxidized), nicotinamide mononucleotide (NMN), nicotinamide, or nicotinic acid. Resumption of cell division occurs after only a lag of about 24 hours. Nil cells subcultured for three consecutive times in the absence of nicotinamide (3Β° NAmβ cells) exhibit different behavior. These severely starved cells are incapable of quickly restoring their intracellular NAD(H) content to normal levels when provided with any pyridine ring compound except NAD^+^. Oneβhour exposure of such cells to NAD^+^ allows utilization of nicotinamide to rapidly restore intracellular NAD(H). This short incubation with NAD^+^ does not result in any significant restoration of intracellular NAD(H) or lead to the accumulation of an intracellular pool of some precursor. This function of NAD^+^ as a stimulatory signal to the NAD(H)βbiosynthetic pathway in severely starved Nil cells is a previously unreported role of NAD^+^, and does not require protein synthesis.