Raman micro-spectroscopy as a viable tool to monitor and estimate the ionic transport in epithelial cells

Leonardo Puppulin,Yasuaki Kumamoto,Yoshinori Marunaka,Hideo Tanaka,Takashi Nakahari,Hongxin Sun,Giuseppe Pezzotti,Toshio Inui,Shigekuni Hosogi

doi:10.1038/s41598-017-03595-y

Leonardo Puppulin, Yasuaki Kumamoto + Show 7 more

Open Access

https://doi.org/10.1038/s41598-017-03595-y

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Abstract

The typical response to the lowering of plasma Na+ concentration and blood pressure in our body involves the release of aldosterone from the adrenal glands, which triggers the reabsorption of sodium in the kidney. Although the effects of aldosterone on this physiological mechanism were extensively studied in the past decades, there are still some aspects to be fully elucidated. In the present study, we propose for the first time a new approach based on Raman spectroscopy to monitor the ionic activity in aldosterone-treated A6 renal epithelial cells. This spectroscopic technique is capable of probing the cells through their thickness in a non-destructive and nimble way. The spectroscopic variations of the Raman bands associated to the O-H stretching of water were correlated to the variations of ionic concentration in the intracellular and extracellular fluids. The increase of Na+ concentration gradients was clearly visualized in the cytosol of aldosterone-treated cells. The enhancement of the Na+ current density induced by aldosterone was estimated from the variation of the ionic chemical potential across the intracellular space. In addition, the variation of the O-H Raman bands of water was used to quantify the cell thickness, which was not affected by aldosterone.

Highlights

The daily excretion/reabsorption of electrolytes in the kidney represents a pivotal physiological mechanism to maintain the appropriate fluid balance in our body, which in turn regulates blood pressure and enables acid-base homeostasis
We propose for the first time a new approach based on Raman micro-spectroscopy to monitor ionic activity in epithelial cells treated with aldosterone
In previous studies it was clearly demonstrated that the intensities of the O-H stretching bands are affected by the presence of ions in solutions, which alters the populations of the water clusters[30, 35, 36]

Summary

Introduction

The daily excretion/reabsorption of electrolytes in the kidney represents a pivotal physiological mechanism to maintain the appropriate fluid balance in our body, which in turn regulates blood pressure and enables acid-base homeostasis. Short-circuit current measurements supported by mathematical simulation enabled to calculate ENaC translocation in the intracellular space[19, 20] The outcomes of these studies emphasized the necessity of combining different experimental and theoretical approaches to elaborate a synergetic method of analysis aimed at obtaining the most complete picture of the phenomenon under investigation. We propose for the first time a new approach based on Raman micro-spectroscopy to monitor ionic activity in epithelial cells treated with aldosterone. This spectroscopic technique is non-destructive and it offers the advantage of micrometric spatial resolution, which is suitable to probe the cells through their thickness in an accurate and nimble way. The spectroscopic variations of the Raman bands associated to the O-H stretching of water were deeply investigated and correlated to the variations of concentration in physiologically relevant solutions, namely fluids in the intracellular and extracellular compartments

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