Background and purpose Diabetic nephropathy (DN) is a leading cause of chronic kidney diseases (CKD) characterized by mesangial matrix expansion that involves dysfunctional mesangial cells (MCs). However, the underlying mechanisms remain unclear. This study aims to delineate the spatiotemporal contribution of adrenergic signaling in diabetic kidney fibrosis to reveal potential therapeutic target. Experimental Approach Db/db mice were used to study DN. RNA sequence analyses, western blot and immunostaining were conducted to profile gene expression in kidneys. Subcellular-localized fluorescence resonance energy transfer (FRET) biosensors determined adrenergic signaling microdomains in MCs. Rolipram, a PDE4 inhibitor, was orally administrated in db/db mice to test its impact on the kidney. Key Results Db/db mice exhibited impaired kidney function with elevated adrenergic and fibrotic gene expression, including ADRB1, PDEs, ACTA2, and TGF-β. MCs with dysregulated YAP pathway played a pivotal role in extracellular matrix secretion in DN. TGF-β up-regulated β1-adrenergic receptors (β1ARs) and α-SMA in MCs. Diminished nuclear-specific cAMP signaling in MCs by TGF-β triggered profibrotic gene transcription via reducing PKA-dependent phosphorylation of YAP. The profibrotic formation was partially alleviated by norepinephrine administration and prevented by combined treatment of norepinephrine and rolipram. In db/db mice, rolipram treatment alleviated kidney fibrosis and restored kidney filtration function. Conclusion and Implications DN impairs nuclear-localized β1AR-cAMP signaling microdomain through upregulating PDE4 expression, promoting fibrosis in MCs via PKA dephosphorylation-dependent YAP activation. Results suggest PDE4 inhibition as a promising strategy for alleviating kidney fibrosis in DN.