Abstract Studies have established the critical roles of a superfamily of phosphodiesterases (PDEs) in hydrolyzing cAMP and its subcellular distribution. We aim to explore the role of PDEs in differential regulation of the cAMP signals at the plasma membrane and in the nucleus. Specifically, we will uncover the regulation of the nuclear cAMP signals under the CNS β2-adrenergic receptor (b2AR) stimulation in hippocampal (HC) neurons during learning and memory. Interestingly, PDE4 inhibitors benefit learning and memory in rodents and humans, indicating that PDE4 may control the βAR-induced cAMP signal in the nucleus. PDE4D isoforms are associated with β2AR to fine-tune subcellular cAMP-PKA signals in fibroblasts and myocytes, and PDE4D5 is associated with an AKAP95/PKA complex in the nucleus. Our preliminary data show that βAR stimulation promotes the nuclear export of PDE4D5 in HC neurons. This relocation of PDE4D5 depends on endosome GRK-phosphorylated b2AR and is critically necessary for delivering cAMP signals into the nucleus. Besides the GRK-phosphorylated b2AR, we have recently characterized another distinct subpopulation of b2AR that are PKA-phosphorylated and located at the PM after agonist stimulation in HC neurons. We hypothesize that two b2AR subpopulations synergistically promote nuclear cAMP signal by arrestin3-dependent export of PDE4D5 from the nucleus. We will characterize the mechanisms underlying the PDE4D5-dependent regulation of nuclear cAMP signaling and gene expression and how the regulation may affect the b2AR signaling in learning and memory (Aim 3). This proposed study will define the role of PDE4D5 in nuclear cAMP signaling, gene expression, and learning and memory, which not only offer new strategies to treat disorders associated with the CNS adrenergic system but also offer an example to study many other Gs-coupled receptors, such as dopamine receptors in the regulation of gene expression.