Proceedings of The Physiological Society
University College London (2011) Proc Physiol Soc 24, C10 and PC10
P2X4 receptors and the epithelial sodium channel in mouse collecting ducts in vivo
E. Craigie1, R. J. Unwin1, S. S. Wildman2, D. G. Shirley1
1. Centre for Nephrology, University College London, London, United Kingdom. 2. Department of Veterinary Basic Sciences, Royal Veterinary College, London, United Kingdom.
Extracellular nucleotides, acting via P2 (purine) receptors, can modify epithelial sodium channel (ENaC)-mediated sodium reabsorption in collecting ducts (Bailey & Shirley 2009), but the P2 receptor subtype(s) involved have not been clearly defined. Pharmacological profiling and the use of genetically engineered mice point to an important role for P2Y2 receptors (Pochynyuk et al 2010), but a patch-clamp investigation of rat cortical collecting duct principal cells has provided evidence that apical P2X4 and/or P2X4/6 receptors may also modulate ENaC activity (Wildman et al 2008). The present study was designed to assess, in vivo, the possible role of the P2X4 receptor subunit in the regulation of ENaC activity. P2X4 null mice (P2X4-/-), and their wild-type littermate controls (P2X4+/+), were anaesthetised (thiobutabarbital sodium, 100 mg/kg, I.P.; Sigma) and surgically prepared for renal clearance studies. Baseline clearance measurements were made for 40 minutes; mice were then given the ENaC inhibitor benzamil (1 mg/kg bolus, IV), and, after a 20 minute equilibration period, clearance collections were repeated for a further 40 minutes. Data are presented as means ± SEM; statistical comparisons were made using ANOVA. In animals used as drug-vehicle controls, glomerular filtration rate (GFR) and fractional sodium excretion (FENa) remained stable. Benzamil treatment did not affect GFR significantly in either P2X4+/+ or P2X4-/- mice. In P2X4+/+ mice, benzamil caused an increase in FENa from 0.6 ± 0.2 % to 1.9 ± 0.3% (n = 8, P<0.001; ΔFENa = 1.3 ± 0.2 %). In P2X4-/- mice, benzamil increased FENa from 0.8 ± 0.2 % to 1.8 ± 0.2 % (n = 8, P<0.001; ΔFENa = 1.0 ± 0.3 %). The benzamil-induced increase in FENa was not significantly different between the two groups. These data provide no solid evidence for a role for P2X4 receptor subunits in the regulation of ENaC-mediated sodium reabsorption in mice on a normal sodium intake. Experiments are now under way to assess whether this remains the case when ENaC expression and activity is up regulated under sodium-restricted conditions.
Where applicable, experiments conform with Society ethical requirements