Electrically wired mitochondrial electrodes for measuring mitochondrial function for drug screening† Robert L. Arechederra, a Abdul Waheed, b William S. Sly b and Shelley D. Minteer * ac Received 1st May 2011, Accepted 5th July 2011 DOI: 10.1039/c1an15370f In the continual search of new therapeutics, many possible drug candidates are excluded, because they are found to negatively affect mitochondrial function. We have developed an approach for directly, electrochemically assaying mitochondrial metabolic activity as a function of metabolic substrate to determine drug toxicity. By wiring mouse mitochondria to a carbon electrode surface, electrons can be intercepted before they reach Complex IV, the terminal step of electron transport chain. The electrons are rerouted, to a separate electrode of the electrochemical cell, the cathode. This allows for the direct measurement of electrical current and potential of the mitochondria during their oxidation of substrates such as pyruvate and fatty acids when there are different concentrations of drug present. This analytical technique has been shown to reliably assay several classical mitochondrial toxins and exhibits potential for the further development of a drug candidate screening technique, as well as other applications where the quantitative study of mitochondrial dysfunction is important. Introduction Screening of mitochondria function in the presence of drug compounds is becoming of increased interest. As more research accumulates for fundamental drug mechanisms and fundamental causes of disease, more is being discovered that points directly to mitochondria function. 1–11 One classical example is the treatment of bipolar disorder with lithium salts. Until recently, the funda- mental mechanism of action has been unknown. The use of lithium salts can be traced back over 100 years ago for the treatment of mania. In 1970, the FDA approved its use as a treatment for bipolar disorder, and it has been heavily used ever since. Clinically, the mechanism has been traced to the cell/tissue level, but the molecular biology was still uncertain. In the last few years, research has shown that the lithium ion stimulates oxidative phosphorylation in the mitochondria by acting as an activator for the electron transport chain. 12 In another example several potent analgesics including Demerol and barbiturates, reversibly inhibit complex I of the electron transport chain causing the nerve cells to not be as active thereby reducing the pain an individual is experiencing. 13 In cancer cells, the mitochondrial activity is suppressed because most of the metabolism is routed through glycolysis to produce lactate. This suppression also causes programmed cell death, which is caused by cytochrome release from mitochondria, allowing the cancer cell to proliferate. If compounds are targeted to specific complexes and proteins of the mitochondria, then the therapeutic could effectively gain control of the mitochondria and stabilize the cell function or allow the cell to achieve programmed cell death, as in the case of cancer. 14 Mitochondria are important to cellular function through their important role in energy conversion, but they are also important in calcium homeostasis, heme synthesis, steroid synthesis, and programmed cell death. 15 Mitochondrial dysfunction is increas- ingly being found to be the cause of drug-induced toxicities. Due to this issue, pharmaceutical companies are realizing the importance of early identification of drug effects of mitochon- drial function, in order to avoid late-stage attrition during drug development. 16,17 Screening strategies to screen chemical drug libraries early in the drug development for mitochondrial dysfunction would be very valuable. This technique of electri- cally wiring mitochondria to carbon electrodes allows for the ability to monitor the rate of metabolite oxidation by the mito- chondria, making it a unique way to assay a compounds effect on the mitochondrial metabolic rate. Screening has not been part of the preclinical drug develop- ment process until recently due to four main issues: 1) it is difficult to accurately assess mitochondrial function; 2) there is no high throughput method for screening drug candidates; 3) drug companies did not fully understand how common mito- chondrial dysfunction was a cause of drug toxicity; and, 4) the lack of evidence as to whether in vitro responses correspond to a Department of Chemistry, Saint Louis University, St. Louis, MO 63103, USA. E-mail: minteers@slu.edu; Fax: +1 314-977-2521; Tel: +1 314- 977-3624 b Edward Doisy Department of Biochemistry & Molecular Biology, Saint Louis University, St. Louis, MO 63104, USA c Departments of Chemistry and Materials Science and Engineering, University of Utah, Salt Lake City, UT 84112, USA. E-mail: minteer@ chem.utah.edu; Fax: +1 801-581-8181; Tel: +1 801-587-8325 † Electronic supplementary information (ESI) available. See DOI: 10.1039/c1an15370f This journal is ª The Royal Society of Chemistry 2011 Analyst, 2011, 136, 3747–3752 | 3747 Dynamic Article Links C < Analyst Cite this: Analyst, 2011, 136, 3747 www.rsc.org/analyst PAPER