The co-transporter activity of Na⁺-K⁺-2Cl⁻ 1 (NKCC1) is dependent on phosphorylation. In this study we show the energy-sensing kinase AMPK inhibits NKCC1 activity. Three separate AMPK activators (AICAR, Phenformin and A-769662) inhibited NKCC1 flux in a variety of nucleated cells. Treatment with A-769662 resulted in a reduction of NKCC1^T212/T217 phosphorylation, and this was reversed by treatment with the non-selective AMPK inhibitor Compound C. AMPK dependence was confirmed by treatment of AMPK null mouse embryonic fibroblasts, where A-769662 had no effect on NKCC1 mediated transport. AMPK was found to directly phosphorylate a recombinant human-NKCC1 N-terminal fragment (1–293) with the phosphorylated site identified as S77. Mutation of Serine 77 to Alanine partially prevented the inhibitory effect of A-769662 on NKCC1 activity. In conclusion, AMPK can act to reduce NKCC1-mediated transport. While the exact mechanism is still unclear there is evidence for both a direct effect on phosphorylation of S77 and reduced phosphorylation of T212/217.

钠的共转运蛋白活性⁺ -K ⁺ -2Cl ^- 1（NKCC1）是依赖于磷酸化。在这项研究中，我们显示了能量感应激酶AMPK抑制NKCC1活性。三种单独的AMPK激活剂（AICAR，苯乙双胍和A-769662）抑制了各种有核细胞中的NKCC1通量。用A-769662处理导致NKCC1 ^{T212 / T217}减少磷酸化，并通过使用非选择性AMPK抑制剂化合物C的治疗逆转。通过对AMPK空小鼠胚胎成纤维细胞进行处理，证实了AMPK依赖性，其中A-769662对NKCC1介导的转运没有影响。发现AMPK可以直接磷酸化重组人NKCC1 N端片段（1-293），其磷酸化位点为S77。丝氨酸77突变为丙氨酸部分阻止了A-769662对NKCC1活性的抑制作用。总之，AMPK可以减少NKCC1介导的转运。虽然确切的机制尚不清楚，但有证据表明对S77的磷酸化有直接作用，而对T212 / 217的磷酸化作用降低。