Figure 5

CREB phosphorylation is dependent on purinergic receptor signaling. (A) Representative 12 μm cryosection stained with antibodies to pCREB (Ser-133) after pre-treatment with PPADS/Suramin and treatment with 2 h odorant, CREB phosphorylation is markedly reduced in the sustentacular cell layer. Control sections show epithelium without odorant treatment and epithelium without odorant treatment, but with PPADS/Suramin incubation. Scale bar 20 μm. (B) Fluorescence intensities were quantified as before. Bar chart shows mean ± SD. Sustentacular cells 2 h odorant (7.10 ± 0.59 n = 79), +PPADS/Suramin (1.55 ± 0.48 n = 80), OSNs 2 h odorant (4.16 ± 3.11 n = 75), +PPADS/Suramin (4.21 ± 2.76 n = 71, p = 0.48), immature cells 2 h odorant (4.03 ± 2.04 n = 79), +PPADS/Suramin (3.51 ± 1.02 n = 80, p = 0.11), p*** < 0.001. (C) Western blot of whole olfactory epithelia lysates of OMP-GFP mice (2 mice pooled per condition) treated for 2 h with a mixture of 100 odorants or pre-treated with PPADS/Suramin. Blot representative of 3 independent experiments. (D) Bar chart shows average densitometry values, mean ± SEM of phospho-CREB (Ser133), shown as percentage of total CREB. Control (3.70 ± 1.29), 2 hours odorant treatment (68.04 ± 4.98), PPADS/suramin pre-treatment (45.93 ± 6.06), p** < 0.01.