[Abstract unavailable

Respiring mitochondria maintain a membrane potential (⌬⌿) 1 of Ϫ150 to Ϫ180 mV (⌬⌿, inside negative). This high ⌬⌿ constitutes a large driving force for the electrophoretic influx of cations, either through specific channels or by diffusion through the membrane. Several cation channels have been characterized physiologically (reviewed in Refs. 1 and 2), and recently, a single one has been identified molecularly (3). These transport systems seem to have intrinsic control mechanisms which ensure that the matrix cation concentrations stay within physiological ranges, far below chemical equilibrium.Diffusive permeability of the inner mitochondrial membrane to ions is generally low but physiologically significant, as it lowers the pH gradient and membrane potential. Moreover, if not counteracted by extrusion, steadily increasing concentrations of matrix cations (and of compensating anions) will lead to an imbalance of osmotic pressure across the inner mitochondrial membrane. As a consequence, water will pass through the membrane, causing excessive swelling and eventual rupture of the organelle (1, 2, 4).As first proposed by P. Mitchell (5), mitochondria have carrier systems allowing the electroneutral exchange of cations against H ϩ (and anions against OH Ϫ ). These exchangers counteract the ⌬⌿-driven cation leakage of the membrane and also cation imbalances due to changes in mitochondrial physiology. Mitochondrial cation distribution is, therefore, a steady state, in which the accumulation ratio is modulated by the relative rates of cation influx and efflux by means of separate pathways.Many physiological studies have been devoted to cation/H ϩ exchange systems (reviewed in Ref.

Loss of the MDM38 gene product in yeast mitochondria results in a variety of phenotypic effects including reduced content of respiratory chain complexes, altered mitochondrial morphology and loss of mitochondrial K þ /H þ exchange activity resulting in osmotic swelling. By use of doxycycline-regulated shut-off of MDM38 gene expression, we show here that loss of K þ /H þ exchange activity and mitochondrial swelling are early events, associated with a reduction in membrane potential and fragmentation of the mitochondrial reticulum. Changes in the pattern of mitochondrially encoded proteins are likely to be secondary to the loss of K þ /H þ exchange activity. The use of a novel fluorescent biosensor directed to the mitochondrial matrix revealed that the loss of K þ /H þ exchange activity was immediately followed by morphological changes of mitochondria and vacuoles, the close association of these organelles and finally uptake of mitochondrial material by vacuoles. Nigericin, a K þ /H þ ionophore, fully prevented these effects of Mdm38p depletion. We conclude that osmotic swelling of mitochondria triggers selective mitochondrial autophagy or mitophagy.

Standard animal behavior paradigms incompletely mimic nature and thus limit our understanding of behavior and brain function. Virtual reality (VR) can help, but it poses challenges. Typical VR systems require movement restrictions but disrupt sensorimotor experience, causing neuronal and behavioral alterations. We report the development of FreemoVR, a VR system for freely moving animals. We validate immersive VR for mice, flies, and zebrafish. FreemoVR allows instant, disruption-free environmental reconfigurations and interactions between real organisms and computer-controlled agents. Using the FreemoVR platform, we established a height-aversion assay in mice and studied visuomotor effects in Drosophila and zebrafish. Furthermore, by photorealistically mimicking zebrafish we discovered that effective social influence depends on a prospective leader balancing its internally preferred directional choice with social interaction. FreemoVR technology facilitates detailed investigations into neural function and behavior through the precise manipulation of sensorimotor feedback loops in unrestrained animals.

Cardiac and skeletal muscle critically depend on mitochondrial energy metabolism for their normal function. Recently, we showed that apoptosis-inducing factor (AIF), a mitochondrial protein implicated in programmed cell death, plays a role in mitochondrial respiration. However, the in vivo consequences of AIFregulated mitochondrial respiration resulting from a loss-of-function mutation in Aif are not known. Here, we report tissue-specific deletion of Aif in the mouse. Mice in which Aif has been inactivated specifically in cardiac and skeletal muscle exhibit impaired activity and protein expression of respiratory chain complex I. Mutant animals develop severe dilated cardiomyopathy, heart failure, and skeletal muscle atrophy accompanied by lactic acidemia consistent with defects in the mitochondrial respiratory chain. Isolated hearts from mutant animals exhibit poor contractile performance in response to a respiratory chain-dependent energy substrate, but not in response to glucose, supporting the notion that impaired heart function in mutant animals results from defective mitochondrial energy metabolism. These data provide genetic proof that the previously defined cell death promoter AIF has a second essential function in mitochondrial respiration and aerobic energy metabolism required for normal heart function and skeletal muscle homeostasis.In animals, the growth and maintenance of tissues absolutely depend on energy metabolism, which is met principally through mitochondrial oxidative phosphorylation (OXPHOS). OXPHOS is a complex biochemical process in which electrons generated from the catabolism of energy substrates flow through a series of catalysts known as the mitochondrial respiratory chain; the free energy liberated from this oxidative process drives formation of an electrochemical potential difference across the inner mitochondrial membrane which is utilized to generate energy for the cell in the form of ATP.

Human Wolf-Hirschhorn syndrome (WHS) is a multigenic disorder resulting from a hemizygous deletion on chromosome 4. LETM1 is the best candidate gene for seizures, the strongest haploinsufficiency phenotype of WHS patients. Here, we identify the Drosophila gene CG4589 as the ortholog of LETM1 and name the gene DmLETM1. Using RNA interference approaches in both Drosophila melanogaster cultured cells and the adult fly, we have assayed the effects of down-regulating the LETM1 gene on mitochondrial function. We also show that DmLETM1 complements growth and mitochondrial K 1 /H 1 exchange (KHE) activity in yeast deficient for LETM1. Genetic studies allowing the conditional inactivation of LETM1 function in specific tissues demonstrate that the depletion of DmLETM1 results in roughening of the adult eye, mitochondrial swelling and developmental lethality in third-instar larvae, possibly the result of deregulated mitophagy. Neuronal specific down-regulation of DmLETM1 results in impairment of locomotor behavior in the fly and reduced synaptic neurotransmitter release. Taken together our results demonstrate the function of DmLETM1 as a mitochondrial osmoregulator through its KHE activity and uncover a pathophysiological WHS phenotype in the model organism D. melanogaster.

YOL027c in yeast and LETM1 in humans encode integral proteins of the inner mitochondrial membrane. They have been implicated in mitochondrial K+ homeostasis and volume control. To further characterize their role, we made use of submitochondrial particles (SMPs) with entrapped K+- and H+-sensitive fluorescent dyes PBFI and BCECF, respectively, to study the kinetics of K+ and H+ transport across the yeast inner mitochondrial membrane. Wild-type SMPs exhibited rapid, reciprocal translocations of K+ and H+ driven by concentration gradients of either of them. K+ and H+ translocations have stoichiometries similar to those mediated by the exogenous K+/H+ exchanger nigericin, and they are shown to be essentially electroneutral and obligatorily coupled. Moreover, [K+] gradients move H+ against its concentration gradient, and vice-versa. These features, as well as the sensitivity of K+ and H+ fluxes to quinine and Mg2+, qualify these activities as K+/H+ exchange reactions. Both activities are abolished when the yeast Yol027p protein is absent (yol027Delta mutant SMPs), indicating that it has an essential role in this reaction. The replacement of the yeast Yol027p by the human Letm1 protein restores K+/H+ exchange activity confirming functional homology of the yeast and human proteins. Considering their newly identified function, we propose to refer to the yeast YOL027c gene and the human LETM1 gene as yMKH1 and hMKH1, respectively.

Ca2+ transport across the inner membrane of mitochondria (IMM) is of major importance for their functions in bioenergetics, cell death and signaling. It is therefore tightly regulated. It has been recently proposed that LETM1—an IMM protein with a crucial role in mitochondrial K+/H+ exchange and volume homeostasis—also acts as a Ca2+/H+ exchanger. Here we show for the first time that lowering LETM1 gene expression by shRNA hampers mitochondrial K+/H+ and Na+/H+ exchange. Decreased exchange activity resulted in matrix K+ accumulation in these mitochondria. Furthermore, LETM1 depletion selectively decreased Na+/Ca2+ exchange mediated by NCLX, as observed in the presence of ruthenium red, a blocker of the Mitochondrial Ca2+ Uniporter (MCU). These data confirm a key role of LETM1 in monovalent cation homeostasis, and suggest that the effects of its modulation on mitochondrial transmembrane Ca2+ fluxes may reflect those on Na+/H+ exchange activity.