Real‐time electrochemical detection of endogenous substance release in freshly isolated organs

Abstract

We established a novel approach to measure acute changes in endogenous substances in the freshly isolated kidney. This method is based on enzymatic microelectrode biosensors and utilizes amperometry technique. In addition to quantitation of basal levels this technique allows real‐time detection of variety of substances, reflecting changes and fluctuations in their concentrations in freshly isolated organs during vehicle or drug perfusion. Basal levels of H2O2 in the kidney cortex and medulla of Sprague Dawley (SD) rats were measured by biosensors and conventional interstitial fluid collection method. Real‐time changes in ATP and H2O2 concentrations in response to angiotensin II (Ang II) were determined. Freshly isolated kidneys of SD and Dahl salt‐sensitive (SS) rats were perfused with 1 μM Ang II under constant laminar flow. This produced a rapid simultaneous release of both ATP and H2O2 in the kidney cortex area with a strong enhancement in SS rats, especially when animals were fed a high‐salt diet. Biosensors specificity revealed by kidney perfusion with AT1 receptor antagonist losartan and decomposition of H2O2 with catalase enzyme. Established method represents a unique powerful approach for the real‐time monitoring of the endogenous substance level in organs or tissue and could be used for ex vivo and in vivo measurements during normal or pathological conditions. This research was supported by NHLBI.