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Kidney Week

Abstract: TH-PO021

A Novel Kidney Slice Culture System Visualizing Intrarenal ATP and Segment-Dependent Energy Metabolism

Session Information

Category: Acute Kidney Injury

  • 103 AKI: Mechanisms

Authors

  • Yamamoto, Shigenori, Department of Nephrology, Kyoto University Graduate School of Medicine, Kyoto, Japan
  • Yamamoto, Shinya, Department of Nephrology, Kyoto University Graduate School of Medicine, Kyoto, Japan
  • Yamamoto, Masamichi, Department of Nephrology, Kyoto University Graduate School of Medicine, Kyoto, Japan
  • Yanagita, Motoko, Department of Nephrology, Kyoto University Graduate School of Medicine, Kyoto, Japan
Background

The kidney constantly utilizes adenosine 5’ triphosphate (ATP), and ATP depletion plays a crucial role in the progression of kidney diseases. Recently, we generated a mouse line, which expresses the FRET-based biosensor systemically, and reported spatiotemporal ATP dynamics in the kidney during AKI model using two-photon microscope. In our previous observation from the kidney surface, however, deeper nephron segments such as S3 segment of proximal tubules (PTs), glomeruli, and thick ascending limbs of Henle (TALs) cannot be observed. Additionally, we cannot analyze ATP dynamics in the presence of the reagents with systemic effects in vivo.

Methods

We established ATP imaging system using the kidney slice culture of ATP visualizing mice and evaluated ATP dynamics in the presence of pharmacological inhibitors of oxidative phosphorylation (OXPHOS) and glycolysis. We also evaluated ATP dynamics after cisplatin administration in this system.

Results

While the ATP levels of PTs, TALs and DTs rapidly and significantly decreased by the administration of 4mM NaN3, an OXPHOS inhibitor, those of podocytes were well maintained as long as 60 min after the administration. On the other hand, the administration of 0.2mM phloretin, a glucose transporters inhibitor, decreased ATP levels in podocytes, but not apparently in PTs, TALs, and DTs. When the kidney slice was incubated in the buffer containing 1mM cisplatin, the ATP levels of PTs and DTs decreased after 60 min, while those of podocytes and principle cells showed no apparent changes even after 120 min. Interestingly, mitochondrial cristae deformation were observed by electron microscopy in the slice incubated with cisplatin for 120min.

Conclusion

Utilizing this novel slice culture system, we, for the first time, directly demonstrated the segment-specific changes in ATP metabolism. While PTs, TALs and DTs are more or less dependent on OXPHOS for ATP production, suggesting their possible vulnerability to ischemia, podocytes rely more on glycolysis for ATP production than on OXPHOS. In addition, we succeeded in demonstrating the different sensitivity to cisplatin among nephron segments. This method could be useful for the elucidation of the metabolic changes in the pathophysiological conditions and for screening of renal toxic drugs.