Figure 1.

Phosphorylation of ERK_1∕2, a known target of 5-HT_2B receptor stimulation in astrocytes. After treatment with phosphate-buffered saline, PBS (Control), transfection solution without siRNA (siRNA (−)) or siRNA specific to 5-HT_2B receptor [siRNA (+)], culturing of well differentiated astrocytes (at least 3 weeks in culture and treated with dibutyryl cAMP [dBcAMP]), was continued for another week. Since 5-HT_2B receptor expression in the cultures had been normal until the exposure to siRNA the result indicates an effect on mature astrocytes. This contrasts the non-conditional gene knockouts of intact mice used by Diaz et al. (2012), where effects during development cannot be ruled out. For the phosphorylation experiments, the cells were incubated for 20 min in serum-free medium in the absence of any drug (Control) or in the presence of 10 μM fluoxetine. (1) Immunoblot showing gene expression of phosphorylated and non-phosphorylated ERK 1 and 2 (p-ERK₁ and ERK₁ and p-ERK₂ and ERK₂) from representative experiments. Similar results were obtained in three independent experiments. Average ERK phosphorylation was quantitated as ratios between p-ERK₁ and ERK₁ (2) and between p-ERK₂ and ERK₂ (3). SEM values are indicated by vertical bars. * Indicates statistically significant (P < 0.05) difference from all other groups for ERK₁ and ERK₂. (From Li et al., 2008).