Purinoceptors (ARs and P2Rs) are expressed on the cellular components of the glomerular filtration barrier and their activation may affect glomerular permeability to albumin, which may ultimately lead to albuminuria, a well-established risk factor for progression of chronic kidney disease and the development of cardiovascular diseases. We have investigated mechanisms underlying the in vitro and in vivo purinergic action on glomerular filter permeability to albumin, measuring convectional albumin permeability (P_alb) in a single isolated rat glomerulus based on the video-microscopy method. Primary cultured rat podocytes were used for analysis of albumin permeability, cGMP accumulation, PKG Iα dimerization and immunofluorescence. In vitro, natural nucleotides (ATP, ADP, UTP, UDP) and non-metabolized ATP analogues (2-meSATP, ATP--S) increased P_alb in a time- and concentration-dependent manner. The effects were dependent on P2Rs activation, nitric oxide synthase and cytoplasmic guanylate cyclase. ATP analogues significantly increased the permeability to albumin, cGMP accumulation and sub-cortical actin reorganization in a PKG-dependent but non dimmer-mediated route in cultured podocytes. In vivo, 2-meSATP and ATP--S increased Palb but did not significantly affect urinary albumin excretion. Both agonists enhanced the clathrin-mediated endocytosis of albumin in podocytes. A product of adenine nucleotides hydrolysis, adenosine, increased the permeability of the glomerular barrier via ARs in a dependent and independent manner. Our results suggest that the extracellular nucleotides that stimulate an increase of glomerular permeability to albumin involve nitric oxide synthase and cytoplasmic guanylate cyclase with actin reorganization in podocytes.