Pharmacological characterization of the P2 receptors profile in the podocytes of the freshly isolated rat glomeruli.
Calcium flux in the podocytes is critical for normal and pathophysiological regulation of these types of cells, and excessive calcium signaling results in podocytes damage and improper glomeruli function. Purinergic activation of P2 receptors is a powerful and rapid signaling process; however, the exact physiological identity of P2 receptors subtypes in podocytes remains essentially unknown. The goal of this study was to determine the P2 receptor profile in podocytes of the intact Sprague-Dawley rat glomeruli using available pharmacological tools. Glomeruli were isolated by differential sieving and loaded with Fluo-4/Fura Red cell permeable calcium indicators, and the purinergic response in the podocytes was analyzed with ratiometric confocal fluorescence measurements. Various P2 receptors activators were tested and compared with the effect of ATP, specifically, UDP, MRS 2365, bzATP, αβ-methylene, 2-meSADP, MRS 4062, and MRS 2768, were analyzed. Antagonists (MRS 2500, 5-BDBD, A438079, and NF 449) were tested when 10 μM ATP was applied as the EC50 for ATP activation of the calcium influx in the podocytes was determined to be 10.7 ± 1.5 μM. Several agonists including MRS 2365 and 2-meSADP caused calcium flux. Importantly, only the P2Y1-specific antagonist MRS 2500 (1 nM) precluded the effects of ATP concentrations of the physiological range. Immunohistochemical analysis confirmed that P2Y1 receptors are highly expressed in the podocytes. We conclude that P2Y1 receptor signaling is the predominant P2Y purinergic pathway in the glomeruli podocytes and P2Y1 might be involved in the pathogenesis of glomerular injury and could be a target for treatment of kidney diseases.
Department of Physiology, Medical College of Wisconsin, Milwaukee, Wisconsin.
Daria V
DV
Ilatovskaya
Oleg
O
Palygin
Vladislav
V
Levchenko
Alexander
A
Staruschenko
D000818:Animals; D002118:Calcium / Q000378:metabolism; D005786:Gene Expression Regulation / Q000502:physiology; D007678:Kidney Glomerulus / Q000166:cytology*; D008297:Male; D050199:Podocytes / Q000378:metabolism*; D058910:Purinergic P2 Receptor Agonists / Q000494:pharmacology*; D058919:Purinergic P2 Receptor Antagonists / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D058469:Receptors, Purinergic P2X / Q000235:genetics / Q000378:metabolism; D058472:Receptors, Purinergic P2Y1 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D046509:Tissue Culture Techniques
D058910:Purinergic P2 Receptor Agonists; D058919:Purinergic P2 Receptor Antagonists; D058469:Receptors, Purinergic P2X; D058472:Receptors, Purinergic P2Y1; D002118:Calcium
ATP; P2Y1; calcium signaling; glomeruli; kidney; podocytes