Figure 2.

E₂ induces Akt and ERK1/2 phosphorylation in a time-dependent manner. Rat cortical neuron cells were exposed to E₂ (10 nm) at specific time intervals (0, 5, 10, 20, 30, 45, and 60 min), and cell lysates were immunoblotted for detection of pAkt or pERK1/2. A, Western Blot analysis showed that E₂-induced Akt phosphorylation was first apparent within 5 min of exposure, was maximally induced at 10 min, diminished at 20 min, and reached maximal induction again at 30 min in a temporal pattern, consistent with ER–p85 coupling with significant and maximal efficacy at 10 and 30 min. B, pERK1/2 was linearly and significantly induced at 5 min, continued to rise 10 and 20 min, reached maximal activation at 30 min, and returned to baseline at 45 min. As a loading control, membranes were reblotted for total Akt and ERK1/2. Densitometry of phosphorylated forms were normalized to either total Akt or total ERK1/2, respectively. Expression of total proteins did not change across conditions or time. Data shown are from a single experiment and are representative of three independent experiments. *p < 0.05 versus control neurons; **p < 0.01 versus control neurons. Ctrl, Control.