Fatty acids do not activate UCP2 in pancreatic beta cells: comparison with UCP1

UCP2 is expressed in pancreatic β cells where its postulated uncoupling activity will modulate glucose-induced changes in ATP/ADP ratio and insulin secretion. The consequences of UCP2 over/underexpression on β-cell function has mainly been studied in the basal state; however, a UCP has no uncoupling activity unless stimulated by fatty acids and/or reactive oxygen species. Here, UCP2 was overexpressed in INS-1 cells and parameters reflecting mitochondrial coupling measured in the basal state and after stimulation by fatty acids. For comparison, UCP1 was expressed to similar levels and the same parameters measured. Neither UCP1 expression nor UCP2 overexpression modified basal or glucose-stimulated metabolic changes. Upon addition of fatty acids, UCP1-expressing cells displayed the expected mitochondrial uncoupling effect, while UCP2 did not elicit any measurable change in mitochondrial function. Taken together, our data demonstrate that, in pancreatic β-cells, UCP2 has no uncoupling activity in the basal state or after fatty acid stimulatio

Control of mitochondrial pH by uncoupling protein 4 in astrocytes promotes neuronal survival.

Brain activity is energetically costly and requires a steady and highly regulated flow of energy equivalents between neural cells. It is believed that a substantial share of cerebral glucose, the major source of energy of the brain, will preferentially be metabolized in astrocytes via aerobic glycolysis. The aim of this study was to evaluate whether uncoupling proteins (UCPs), located in the inner membrane of mitochondria, play a role in setting up the metabolic response pattern of astrocytes. UCPs are believed to mediate the transmembrane transfer of protons, resulting in the uncoupling of oxidative phosphorylation from ATP production. UCPs are therefore potentially important regulators of energy fluxes. The main UCP isoforms expressed in the brain are UCP2, UCP4, and UCP5. We examined in particular the role of UCP4 in neuron-astrocyte metabolic coupling and measured a range of functional metabolic parameters including mitochondrial electrical potential and pH, reactive oxygen species production, NAD/NADH ratio, ATP/ADP ratio, CO2 and lactate production, and oxygen consumption rate. In brief, we found that UCP4 regulates the intramitochondrial pH of astrocytes, which acidifies as a consequence of glutamate uptake, with the main consequence of reducing efficiency of mitochondrial ATP production. The diminished ATP production is effectively compensated by enhancement of glycolysis. This nonoxidative production of energy is not associated with deleterious H2O2 production. We show that astrocytes expressing more UCP4 produced more lactate, which is used as an energy source by neurons, and had the ability to enhance neuronal survival

Lentiviral Transgenesis as a Tool to Study the Role of Uncoupling Protein Isoforms in the Nervous System

A tight coupling exists between synaptic activity and glucose utilization by astrocytes. Metabolic cooperation between neurons and astrocytes mediates this coupling. During synaptic activation, glutamate that is released in the synaptic cleft as a neurotransmitter by neurons is rapidly cleared by an active uptake into astrocytes. One glutamate is co-transported with three Na+. Intracellular astrocytic Na+ homeostasis is re-established by the Na+/K+-ATPase which requires ATP provided mainly by glycolysis. The resulting lactate is released in extracellular space and contributes to the energetic balance of activated neurons. This coupling between astrocytes and neurons is known as the astrocyte-neuron lactate shuttle hypothesis (ANLSH). The role of mitochondria in this coupling remains to be determined and more specifically the role of uncoupling proteins (UCP) which have been recently identified in neural tissues. UCPs belong to a family of mitochondrial carriers which are present in the inner mitochondrial membrane. There are five isoforms named UCP1 to UCP5. The well-known characterized is UCP1, also named thermogenin, which is present in the brown adipose tissue (BAT) of the small rodents and is responsible of non-shivering thermogenesis. It dissipates the proton gradient across the inner mitochondrial membrane, thereby producing heat instead of ATP. UCP2 is ubiquitous and is implicated in protection against excessive reactive oxygen (ROS) production. The expression of UCP3 is restricted to skeletal muscle where it is linked to fatty acid metabolism. The role of the brain isoforms UCP4 and UCP5 is still poorly understood. Initially, we set out to evaluate their function in energy metabolism of astrocytes and neurons. We used the lentiviral strategy to overexpress or silence the UCPs isoforms in primary cultures of astrocytes and neurons. We found that only UCP4 and UCP5 had an uncoupling activity in brain cells. Indeed, we observed a reduced mitochondrial potential and a reduced ATP/ADP ratio in astrocytes as well as in neurons overexpressing UCP4 or UCP5. UCP4 reduced the oxidative pathway of astrocytes without modifying the basal glucose metabolism and without having a lethal effect on the cell. Oxygen consumption rate (OCR) of brain cells ovexpressing UCPs was reduced. Moreover, UCP4 increased lactate release by astrocytes, suggesting an implication in the astrocyte-neuron coupling. We also brought evidence that both UCP4 and UCP5 partially protect brain cells from oxidative damage. All these results were confirmed by experiments on UCPs silencing. In a second step, we used a co-culture model to study the impact of astrocytic uncoupling protein on neuronal survival and we demonstrated that the presence of uncoupling protein in astrocytes prevents neuronal death after glutamate excitotoxicity. Taken together, all these results support an important role of uncoupling proteins in the regulation of astrocytic energy production, thus promoting local export of lactate which can be used by neurons

Fatty acids do not activate UCP2 in pancreatic beta cells: comparison with UCP1

UCP2 is expressed in pancreatic beta cells where its postulated uncoupling activity will modulate glucose-induced changes in ATP/ADP ratio and insulin secretion. The consequences of UCP2 over/underexpression on beta-cell function has mainly been studied in the basal state; however, a UCP has no uncoupling activity unless stimulated by fatty acids and/or reactive oxygen species. Here, UCP2 was overexpressed in INS-1 cells and parameters reflecting mitochondrial coupling measured in the basal state and after stimulation by fatty acids. For comparison, UCP1 was expressed to similar levels and the same parameters measured. Neither UCP1 expression nor UCP2 overexpression modified basal or glucose-stimulated metabolic changes. Upon addition of fatty acids, UCP1-expressing cells displayed the expected mitochondrial uncoupling effect, while UCP2 did not elicit any measurable change in mitochondrial function. Taken together, our data demonstrate that, in pancreatic beta-cells, UCP2 has no uncoupling activity in the basal state or after fatty acid stimulation

Distinct fission signatures predict mitochondrial degradation or biogenesis

Comment in Revolutionary view of two ways to split a mitochondrion. Chakrabarti R, Higgs HN. Nature. 2021 May;593(7859):346-347. doi: 10.1038/d41586-021-01173-x. PMID: 33953387International audienceMitochondrial fission is a highly regulated process that, when disrupted, can alter metabolism, proliferation and apoptosis1-3. Dysregulation has been linked to neurodegeneration3,4, cardiovascular disease3 and cancer5. Key components of the fission machinery include the endoplasmic reticulum6 and actin7, which initiate constriction before dynamin-related protein 1 (DRP1)8 binds to the outer mitochondrial membrane via adaptor proteins9-11, to drive scission12. In the mitochondrial life cycle, fission enables both biogenesis of new mitochondria and clearance of dysfunctional mitochondria through mitophagy1,13. Current models of fission regulation cannot explain how those dual fates are decided. However, uncovering fate determinants is challenging, as fission is unpredictable, and mitochondrial morphology is heterogeneous, with ultrastructural features that are below the diffraction limit. Here, we used live-cell structured illumination microscopy to capture mitochondrial dynamics. By analysing hundreds of fissions in African green monkey Cos-7 cells and mouse cardiomyocytes, we discovered two functionally and mechanistically distinct types of fission. Division at the periphery enables damaged material to be shed into smaller mitochondria destined for mitophagy, whereas division at the midzone leads to the proliferation of mitochondria. Both types are mediated by DRP1, but endoplasmic reticulum- and actin-mediated pre-constriction and the adaptor MFF govern only midzone fission. Peripheral fission is preceded by lysosomal contact and is regulated by the mitochondrial outer membrane protein FIS1. These distinct molecular mechanisms explain how cells independently regulate fission, leading to distinct mitochondrial fates