Fig. 3.

N-myc promotes sympathetic neuron survival after NGF withdrawal. A, AdN-myc does not decrease the survival of sympathetic neurons in the presence of NGF. MTT survival assays of sympathetic neurons selected in NGF, infected with 100–400 MOI of N-myc, GFP, or hypophosphorylated pRb adenovirus, maintained in NGF, and assayed 4 d after infection are shown. All conditions were performed in triplicate, and error bars represent the SD of the mean. Values derive from one representative experiment of three and are normalized to those obtained for uninfected neurons maintained in 10 ng/ml NGF, which are considered to be 100% survival. No significant alterations are seen in any of the conditions.B–D, Adenovirus-mediated expression of N-myc rescues sympathetic neurons from apoptosis induced by NGF withdrawal, as monitored by MTT assays (B), nuclear morphology (C), and Trypan blue exclusion (D). In all experiments, neurons were infected with adenovirus, withdrawn from NGF 2 d later, and analyzed after an additional 2 d. B, MTT assays of neurons infected with 80–320 MOI N-myc, GFP, or hypophosphorylated pRb adenovirus. Values were normalized to those obtained for uninfected neurons either maintained in NGF (100%) or withdrawn from NGF (1%). Values represent the average of three independent experiments, each of which was performed in triplicate. *p < 0.05; Student's t test comparing results obtained with the N-myc versus control GFP adenovirus. C,D, Percentage of living cells or nonapoptotic cells counted by Hoechst-labeled intact nuclei (C) or Trypan blue exclusion (D) after virus infection and NGF withdrawal. For both methods, values were obtained by counting three randomly selected microscope fields in each condition, and the error bars represent the SD of the mean. Values are representative results of one of two independent experiments. **p< 0.01; Student's t test comparing results obtained using the N-myc versus control β-galactosidase (LacZ) adenovirus.