Department of Cellular and Integrative Physiology, University of Texas Health Science Center at San Antonio , San Antonio, Texas.
Division of Nephrology, Department of Internal Medicine, University of Michigan , Ann Arbor, Michigan.
Am J Physiol Renal Physiol. 2019 Apr 1;316(4):F758-F767. doi: 10.1152/ajprenal.00612.2018. Epub 2019 Feb 6.
Stimulation of metabotropic G-coupled purinergic P2Y receptors decreases activity of the epithelial Na channel (ENaC) in renal principal cells of the distal nephron. The physiological consequences of P2Y receptor signaling disruption in the P2Y receptor knockout mouse are decreased Na excretion and increased arterial blood pressure. However, because of the global nature of this knockout model, the quantitative contribution of ENaC and distal nephron compared with that of upstream renal vascular and tubular elements to changes in urinary excretion and arterial blood pressure is obscure. Moreover, it is uncertain whether stimulation of P2Y receptor inhibition of ENaC is sufficient to drive renal (urinary) Na excretion (UV). Here, using a pharmacogenetic approach and selective agonism of the P2Y receptor, we test the sufficiency of targeted stimulation of G signaling in principal cells of the distal nephron and P2Y receptors to increase UV. Selective stimulation of the P2Y receptor with the ligand MRS2768 decreased ENaC activity in freshly isolated tubules (as assessed by patch-clamp electrophysiology) and increased UV (as assessed in metabolic cages). Similarly, selective agonism of hM3Dq-designer receptors exclusively activated by designer drugs (DREADD) restrictively expressed in principal cells of the distal nephron with clozapine- N-oxide decreased ENaC activity and, consequently, increased UV. Clozapine- N-oxide, when applied to control littermates, failed to affect ENaC and UV. This study represents the first use of pharmacogenetic (DREADD) technology in the renal tubule and demonstrated that selective activation of the P2Y receptor and G signaling in principal cells is sufficient to promote renal salt excretion.
代谢型 G 蛋白偶联嘌呤能 P2Y 受体的刺激可降低远曲小管上皮钠通道 (ENaC) 的活性。在 P2Y 受体敲除小鼠中，P2Y 受体信号中断的生理后果是钠排泄减少和动脉血压升高。然而，由于这种敲除模型的全局性，ENaC 和远曲小管与上游肾血管和管状元件相比，对尿排泄和动脉血压变化的定量贡献尚不清楚。此外，刺激 P2Y 受体抑制 ENaC 是否足以驱动肾脏（尿）钠排泄 (UV) 尚不确定。在这里，我们使用药理学方法和 P2Y 受体的选择性激动剂，测试了远曲小管主细胞中 G 信号的靶向刺激和 P2Y 受体刺激的充分性，以增加 UV。用配体 MRS2768 选择性刺激 P2Y 受体可降低新鲜分离的小管 (通过膜片钳电生理学评估) 中的 ENaC 活性，并增加 UV (通过代谢笼评估)。同样，选择性激动 hM3Dq-designer 受体，该受体仅被 designer drugs (DREADD) 激活，并且在远曲小管主细胞中受到 clozapine-N-oxide 的限制表达，可降低 ENaC 活性，从而增加 UV。当 clozapine-N-oxide 应用于对照同窝仔时，未能影响 ENaC 和 UV。本研究代表了药理学 (DREADD) 技术在肾小管中的首次应用，并证明了 P2Y 受体和 G 信号在主细胞中的选择性激活足以促进肾脏盐排泄。
