Supplementary Material for: Coincidence Signaling of Dopamine D<sub>1</sub>-Like and M<sub>1</sub> Muscarinic Receptors in the Regulation of Cyclic AMP Formation and CREB Phosphorylation in Mouse Prefrontal Cortex

2012-03-28T00:00:00Z (GMT) by Olianas M.C. Dedoni S. Onali P.
In the prefrontal cortex, dopamine D<sub>1</sub>-like and M<sub>1</sub> muscarinic receptors are both involved in the regulation of attentional, cognitive and emotional processes but so far no information has been provided on their functional interaction. In the present study we show that in mouse medial prefrontal cortex, concomitant activation of M<sub>1</sub> muscarinic receptors potentiated D<sub>1</sub>-like receptor-induced cyclic AMP formation through a mechanism involving activation of G<sub>q/11</sub> and the release of G protein βγ subunits. Immunohistochemical studies indicated that the adenylyl cyclase isoforms AC2 and AC4 are expressed in mouse prefrontal cortex and that they colocalize with D<sub>1</sub>-like receptors with a greater association for AC4. In primary cultures of frontal cortex neurons, D<sub>1</sub>-like receptor-induced Ser133 phosphorylation of the transcription factor cyclic AMP-responsive element binding protein (CREB) was potentiated by concurrent stimulation of M<sub>1</sub> receptors. Suppression of AC4 expression with small interfering RNA transfection reduced D<sub>1</sub> stimulation of cyclic AMP formation and CREB phosphorylation and abolished the M<sub>1</sub> potentiation, whereas knockdown of AC2 had no significant effects. These data indicate that in mouse prefrontal cortex G<sub>q/11</sub>-coupled M<sub>1</sub> receptor and G<sub>s</sub>-coupled D<sub>1</sub>-like receptor inputs converge on AC4 with a consequent enhancement of cyclic AMP formation and signaling to the nucleus.