Retinoic acid-mediated enhancement of the cholinergic/neuronal nitric oxide synthase phenotype of the medial septal SN56 clone: Establishment of a nitric oxide-sensitive proapoptotic state

It is unclear what mechanisms lead to the degeneration of basal forebrain c holinergic neurons in Alzheimer's or other human brain diseases. Some brain cholinergic neurons express neuronal nitric oxide (NO) synthase (nNOS), wh ich produces a free radical that has been implicated in some forms of neuro degeneration. We investigated nNOS expression and NO toxicity in SN56 cells , a clonal cholinergic model derived from the medial septum of the mouse ba sal forebrain. We show here that, in addition to expressing choline acetylt ransferase (ChAT), SN56 cells express nNOS. Treatment of SN56 cells with re tinoic acid (RA; 1 mu M) for 48 h increased ChAT mRNA (+126%), protein (+88 %), and activity (+215%) and increased nNOS mRNA (+98%), protein (+400%), a nd activity (+15%). After RA treatment, SN56 cells became vulnerable to NO excess generated with S-nitro-N-acetyl-DL-penicillamine (SNAP) and exhibite d increased nuclear DNA fragmentation that was blocked with a caspase-3 inh ibitor. Treatment with dexamethasane, which largely blocked the RA-mediated increase in nNOS expression. or inhibition of nNOS activity with methylthi ocitrulline strongly potentiated the apoptotic response to SNAP in RA-treat ed SN56 cells. Caspase-3 activity was reduced when SNAP was incubated with cells or cell lysates, suggesting that NO can directly inhibit the protease . Thus, whereas RA treatment converts SN56 cells to a proapoptotic state se nsitive to NO excess, endogenously produced NO appears to be anti-apoptotic , possibly by tonically inhibiting caspase-3.