Guidelines on experimental methods to assess mitochondrial dysfunction in cellular models of neurodegenerative diseases

Neurodegenerative diseases are a spectrum of chronic, debilitating disorders characterised by the progressive degeneration and death of neurons. Mitochondrial dysfunction has been implicated in most neurodegenerative diseases, but in many instances it is unclear whether such dysfunction is a cause or an effect of the underlying pathology, and whether it represents a viable therapeutic target. It is therefore imperative to utilise and optimise cellular models and experimental techniques appropriate to determine the contribution of mitochondrial dysfunction to neurodegenerative disease phenotypes. In this consensus article, we collate details on and discuss pitfalls of existing experimental approaches to assess mitochondrial function in in vitro cellular models of neurodegenerative diseases, including specific protocols for the measurement of oxygen consumption rate in primary neuron cultures, and single-neuron, time-lapse fluorescence imaging of the mitochondrial membrane potential and mitochondrial NAD(P)H. As part of the Cellular Bioenergetics of Neurodegenerative Diseases (CeBioND) consortium ( www.cebiond.org ), we are performing cross-disease analyses to identify common and distinct molecular mechanisms involved in mitochondrial bioenergetic dysfunction in cellular models of Alzheimer's, Parkinson's, and Huntington's diseases. Here we provide detailed guidelines and protocols as standardised across the five collaborating laboratories of the CeBioND consortium, with additional contributions from other experts in the field

Thermal and hydraulic design of water-based cooling systems for electrical machines.

Thermal management of electrical machines is gaining more and more attention from the research and industrial community, due to the demanding power density and reliability conditions of present and future applications. The aim of this research work is to draw some conclusions about which is the best way of designing and optimizing two types of cooling systems for electrical machines. More precisely, the cooling systems studied use water as a secondary coolant, and are classified by IEC60034 as IC71W and IC81W. These systems are very popular in traction and marine applications. For this purpose, both algebraic and CFD based models have been developed in order to precisely calculate the behavior of the main components of these cooling systems. Once these models have been validated experimentally, they have been employed to derive design and optimization criteria for the systems. Thus, as a first step, the main considerations for the development of an algebraic thermal model for the IC71W cooling system are examined. A detailed classification of the most critical convective correlations and a review of the modelling of the water jacket is given. Once the model is described, the solving procedure and its experimental validation is exposed. Secondly, using the previously validated model a complete design methodology for the water jacket of an IC71W cooling system is proposed. Different elements such as the shaft or the housing are analyzed in order to obtain some criteria for their design. The inclusion of wafters in this kind of cooling systems is examined, proposing an optimum configuration and evaluating the effect of this element on the temperatures of the stator end-windings. Regarding the IC81W cooling system, the same process has been followed. As a first step, both a hydraulic and a thermal model have been developed. The hydraulic model is defined in more detail, because it is crucial to obtain a realistic behavior of airflow inside the machine. After that, a method for the computation and solving of these models and its experimental validation process is presented. Finally, with the developed models, the main elements defining an IC81W cooling system (which is very extended for marine applications), the fan and the primary circuit, are studied. A design methodology for the fan fixed to the shaft is proposed, including the effect of the main parameters defining this element on the airflow and the pressure rise. Furthermore, different elements in the primary circuit are studied, obtaining the thermal behavior of different configurations and determining the best choice in order to reduce temperatures in the active parts of the machine

Pyruvate kinase and aspartate-glutamate carrier distributions reveal key metabolic links between neurons and glia in retina.

Symbiotic relationships between neurons and glia must adapt to structures, functions, and metabolic roles of the tissues they are in. We show here that Müller glia in retinas have specific enzyme deficiencies that can enhance their ability to synthesize Gln. The metabolic cost of these deficiencies is that they impair the Müller cell’s ability to metabolize Glc. We show here that the cells can compensate for this deficiency by using metabolites produced by neurons. Müller glia are deficient for pyruvate kinase (PK) and for aspartate/glutamate carrier 1 (AGC1), a key component of the malate-aspartate shuttle. In contrast, photoreceptor neurons express AGC1 and the M2 isoform of pyruvate kinase, which is commonly associated with aerobic glycolysis in tumors, proliferating cells, and some other cell types. Our findings reveal a previously unidentified type of metabolic relationship between neurons and glia. Müller glia compensate for their unique metabolic adaptations by using lactate and aspartate from neurons as surrogates for their missing PK and AGC1.This work was supported by National Eye Institute Grants EY06641 and EY023346 (to J.B.H.), by Vision Core Grant EY1730 and Grants BFU2011-30456-C02-01/BMC and S2010/BMD-2402 (to J.S.), and by the Fundación Ramón Areces to the Centro de Biología Molecular Severo OchoaPeer Reviewe