FIGURE 6.

Antisense inhibition of zebrafish Mg²⁺-dependent neutral SMase 1 blocks heat shock-mediated apoptosis and ceramide generation. A, effect of antisense oligo treatment on Mg²⁺-dependent neutral SMase activity. ZE cells were pretreated with an antisense (0–10 μm) or sense (10 μm) oligonucleotide targeting neutral SMase 1 mRNA for 48 h; heat-shocked at 38 °C for 1 h; allowed to recover at 28.5 °C; and harvested after 0, 1, 2, 3, 6, or 12 h. Some cells were also treated with 20 μm Z-VAD-fmk for 1 h before being heat-shocked. The neutral SMase activity in the cells was measured as described under “Experimental Procedures.” B and C, Mg²⁺-dependent neutral SMase activity. D and E, ceramide content. F, sphingomyelin content. G, caspase-3 activity. H, DAPI assay. I, cell viability. Values and bars indicate the means and standard deviations, respectively, of three independent experiments. ^*, p < .05 versus sense oligonucleotide-treated cells.