Matos J E, Robaye B, Boeynaems J M, Beauwens R, Leipziger J
Institute of Physiology and Biophysics, The Water and Salt Research Center, University of Aarhus, Denmark.
J Physiol. 2005 Apr 1;564(Pt 1):269-79. doi: 10.1113/jphysiol.2004.080002. Epub 2005 Feb 17.
Extracellular nucleotides are important regulators of epithelial ion transport, frequently exerting their action from the luminal side. Luminal P2Y receptors have previously been identified in rat distal colonic mucosa. Their activation by UTP and ATP stimulates K+ secretion. The aim of this study was to clarify which of the P2Y receptor subtypes are responsible for the stimulated K+ secretion. To this end P2Y2 and P2Y4 knock-out mice were used to measure distal colonic ion transport in an Ussing chamber. In mouse (NMRI) distal colonic mucosa, luminal UTP and ATP with similar potency induced a rapid and transient increase of the transepithelial voltage (V(te)) (UTP: from -0.81 +/- 0.23 to 3.11 +/- 0.61 mV, n = 24), an increase of equivalent short circuit current (I(sc)) by 166.9 +/- 22.8 microA cm(-2) and a decrease of transepithelial resistance (R(te)) from 29.4 +/- 2.4 to 23.5 +/- 2.0 Omega cm2. This effect was completely inhibited by luminal Ba2+ (5 mm, n = 5) and iberiotoxin (240 nm, n = 6), indicating UTP/ATP-stimulated K+ secretion. RT-PCR analysis of isolated colonic crypts revealed P2Y2, P2Y4 and P2Y6 specific transcripts. The luminal UTP-stimulated K+ secretion was still present in P2Y2 receptor knock-out mice, but significantly reduced (DeltaV(te): 0.83 +/- 0.26 mV) compared to wild-type littermates (DeltaV(te): 2.08 +/- 0.52 mV, n = 9). In P2Y4 receptor knock-out mice the UTP-induced K+ secretion was similarly reduced. Luminal UTP-stimulated K+ secretion was completely absent in P2Y2/P2Y4 double receptor KO mice. Basolateral UTP showed no effect. In summary, these results indicate that both the P2Y2 and P2Y4 receptors are present in the luminal membrane of mouse distal colonic mucosa, and stimulation of these receptors leads to K+ secretion.
细胞外核苷酸是上皮离子转运的重要调节因子，常从管腔侧发挥作用。此前已在大鼠远端结肠黏膜中鉴定出管腔P2Y受体。UTP和ATP对其激活可刺激钾离子分泌。本研究的目的是阐明哪种P2Y受体亚型负责刺激钾离子分泌。为此，使用P2Y2和P2Y4基因敲除小鼠在尤斯灌流小室中测量远端结肠离子转运。在小鼠（NMRI）远端结肠黏膜中，管腔UTP和ATP以相似效力诱导跨上皮电压（V(te)）迅速短暂升高（UTP：从-0.81±0.23 mV升至3.11±0.61 mV，n = 24），等效短路电流（I(sc)）增加166.9±22.8 μA cm(-2)，跨上皮电阻（R(te)）从29.4±2.4降至23.5±2.0 Ω cm2。这种效应被管腔Ba2+（5 mM，n = 5）和埃博毒素（240 nM，n = 6）完全抑制，表明UTP/ATP刺激钾离子分泌。对分离的结肠隐窝进行RT-PCR分析显示有P2Y2、P2Y4和P2Y6特异性转录本。在P2Y2受体基因敲除小鼠中，管腔UTP刺激的钾离子分泌仍然存在，但与野生型同窝小鼠相比显著降低（ΔV(te)：0.83±0.26 mV）（野生型：ΔV(te)：2.08±0.52 mV，n = 9）。在P2Y4受体基因敲除小鼠中，UTP诱导的钾离子分泌同样降低。在P2Y2/P2Y4双受体基因敲除小鼠中，管腔UTP刺激的钾离子分泌完全不存在。基底侧UTP无作用。总之，这些结果表明P2Y2和P2Y4受体均存在于小鼠远端结肠黏膜的管腔膜中，刺激这些受体可导致钾离子分泌。
