Phosphorylation of CREB (CRE binding protein) at its serine 133 residue has been implicated in plasticity in the hippocampus and neocortex. CREB binds to a DNA region known as CRE (cAMP response element) to initiate gene expression. CRE-mediated gene expression increases in the barrel cortex following single whisker experience (Barth et al. 2000). In this study we tested the hypothesis that the MEK and PKA pathways were responsible for activating experience-dependent gene expression in barrel cortex by using specific inhibitors in transgenic mice containing a CRE reporter gene (CRE-LacZ).

CRE-Lac Z mice were anaesthetized with avertin (I.P. injection 0.1 ml, 5 g), and implanted with osmotic mini-pumps (Azlet), which had a canula attached that was placed several millimetres anterior to the barrel cortex in the left cerebral hemisphere. We used three groups, a control group (n = 4) which had pumps filled with saline (0.9 %), a MEK inhibitor (UO126) group (50 mM) (Promega) (n = 4), and a cAMP inhibitor (Rp-8-ClcAMPS) group (5 mM) (Biomol) (n = 2). All but the D1 whiskers on both sides of the face were removed, and the mice were placed in an environmentally enriched cage for 16 h, after which they were anaesthetized with isoflurane (5 %), and killed by decapitation. The brains were then removed and reacted to quantify β-galactosidase activity using X-gal. Quantification was carried out by sectioning horizontally, mounting and counterstaining with propidium iodide. Numerical analysis was carried out by counting the number of X-Gal positive cells within the D1 barrel, and was expressed as a percentage of the total number of cells within the barrel. At least two observers counted the cells, and the results were averaged. An ANOVA statistical test was carried out between the MEK inhibited and the control group of hemispheres. The same was carried out for the deprived C1 and E1 barrels.

There was significant CRE-Lac Z expression in the D1 barrel of both hemispheres in the control animals. In animals receiving inhibitors, there was a significant reduction in CRE-LacZ expression in the left hemisphere treated with UO126 though not complete abolition (P < 0.005). This implies that MEK is an important factor for experience-dependent CRE-mediated gene expression in the barrel cortex. Similarly in brains treated with the cAMP inhibitor, CRE-mediated gene expression was reduced but not abolished. These preliminary observations also implicate adenylate cyclase and PKA in CRE-mediated gene expression.