APOPTOSIS IS INDUCED IN POSTMITOTIC RAT SYMPATHETIC NEURONS BY ARABINOSIDES AND TOPOISOMERASE-II INHIBITORS IN THE PRESENCE OF NGF

Sympathetic neurons depend on nerve growth factor (NGF) for their surv ival and die by apoptosis when NGF is withdrawn, despite their post-mi totic state. Martin et al. (1990, J. Neurosci. 10, 184-193) showed tha t cytosine arabinoside, but no other arabinofuranosyl nucleoside, coul d induce cell death in the presence of NGF and they suggested that it may block a critical step in the NGF-signalling pathway. We show that cytosine arabinoside is not the only nucleoside capable of inducing ap optosis in sympathetic neurons in the presence of NGF. In newly isolat ed neurons from P0 rat pups cultured in the presence of NGF, all the a rabinose nucleosides (adenine, cytosine, guanine. and thymine) induce apoptosis at 10 mu M when combined with 5-fluorodeoxyuridine treatment . Because 1-beta-arabinofuranosylcytosine is associated with double-st rand breaks and chromosomal abberrations, we examined whether topoisom erase II inhibitors, which also cause double-strand breaks by stabilis ing the enzyme-DNA 'cleavable complex', were capable of promoting apop tosis in these neurons. Although P0 rat neurons are strictly postmitot ic, topoisomerase II inhibitors teniposide and mitoxantrone induced th em to die by apoptosis in the presence of NGF with the same apparent t ime-course as arabinose treatment or NGF withdrawal. By contrast, ICRF 193, a catalytic inhibitor of topoisomerase II, reduced the extent of apoptosis induced by mitoxantrone or teniposide by 80% if added simul taneously with the latter but by 2 hours it had no rescue effect, sugg esting that topoisomerase II is highly active in these neurons. ICRF 1 93 also partially reduced the induction of fluorodeoxyuridine-dependen t apoptosis by the arabinose nucleosides. These data suggest that indu ction of double-strand breaks in the DNA or NGF withdrawal activate si milar apoptotic programmes in sympathetic neurons. We propose that tre atment with FdUr results in dUTP incorporation in the neurons, that th e presence of arabinose nucleosides causes disruption of the repair sy stem that involves topoisomerase II and this causes double-strand brea ks and thus apoptosis.