Single-nucleotide polymorphisms in mitochondrial DNA (mtDNA) encoded genes of respiratory chain complexes are known to be associated with severe diseases and life-threatening syndromes. In the assembly of the ATP synthase, the enzyme that in the final steps of oxidative phosphorylation generates ATP from ADP and inorganic phosphate, two subunits (ATP6 and ATP8) are mtDNA-encoded. In our study, we investigated the impact of a single-nucleotide polymorphism in MT-ATP8 with respect to memory function in a preclinical model. Here, we have employed two conplastic mouse strains. The mouse strain C57BL/6 J-mt ^AKR/J served as a control with wild-type sequence in MT-ATP8 , while C57BL/6 J-mt ^FVB/NJ exhibited an m.7778G > T transversion. Using two age groups (3 months and 24 months), levels of reactive oxygen species (ROS), spatial learning in the Morris-Water-Maze, and long-term potentiation were assessed. Immunohistologically, the expressions of NeuN and GFAP were quantified. Additionally, the lifespan of both strains was registered. In comparison to young C57BL/6 J-mt ^FVB/NJ mice, aged animals had higher ROS levels in the hippocampus. A decreased NeuN/GFAP level was found in C57BL/6 J-mt ^FVB/NJ mice as well as in old animals of the control strain. Aged animals performed worse in the swimming trials, but no significant differences between both strains were detected. The long-term potentiation recordings revealed reduced synaptic plasticity in young C57BL/6 J-mt ^FVB/NJ mice. Interestingly, C57BL/6 J-mt ^FVB/NJ mice presented an extended lifespan compared to animals of the control strain. Together, our data suggest a minor impact of a single-nucleotide polymorphism in MT-ATP8 on spatial learning and oxidative stress depending on the neuronal tissue. In line with the concept of mitohormesis, our findings may be linked to the longevity of mice harbouring single-nucleotide polymorphisms.

Galvanotaxis, the migration along direct current electrical fields, may contribute to the invasion of brain cancer cells in the tumor-surrounding tissue. We hypothesized that pharmacological perturbation of the epidermal growth factor (EGF) receptor and downstream phosphatidylinositol 3-kinase (PI3K)/AKT pathway prevent galvanotactic migration. In our study, patient-derived glioblastoma and brain metastases cells were exposed to direct current electrical field conditions. Velocity and direction of migration were estimated. To determine the effects of EGF receptor antagonist afatinib and AKT inhibitor capivasertib, assays of cell proliferation, apoptosis and immunoblot analyses were performed. Both inhibitors attenuated cell proliferation in a dose-dependent manner and induced apoptosis. We found that most of the glioblastoma cells migrated preferentially in an anodal direction, while brain metastases cells were unaffected by direct current stimulations. Afatinib presented only a mild attenuation of galvanotaxis. In contrast, capivasertib abolished the migration of glioblastoma cells without genetic alterations in the PI3K/AKT pathway, but not in cells harboring PTEN mutation. In these cells, an increase in the activation of ERK1/2 may in part substitute the inhibition of the AKT pathway. Overall, our data demonstrate that glioblastoma cells migrate in the electrical field and the PI3K/AKT pathway was found to be highly involved in galvanotaxis.

Glioblastoma is the most common and aggressive primary brain tumor. Multiple genetic and epigenetic alterations in several major signaling pathways—including the phosphoinositide 3-kinases (PI3K)/AKT/mTOR and the Raf/MEK/ERK pathway—could be found. We therefore aimed to investigate the biological and molecular effects of small-molecule kinase inhibitors that may interfere with those pathways. For this purpose, patient-derived glioblastoma cells were challenged with dactolisib, ipatasertib, MK-2206, regorafenib, or trametinib. To determine the effects of the small-molecule kinase inhibitors, assays of cell proliferation and apoptosis and immunoblot analyses were performed. To further investigate the effects of ipatasertib on organotypic brain slices harboring glioblastoma cells, the tumor growth was estimated. In addition, the network activity in brain slices was assessed by electrophysiological field potential recordings. Multi-kinase inhibitor regorafenib and both MK-2206 and dactolisib were very effective in all preclinical tumor models, while with respect to trametinib, two cell lines were found to be highly resistant. Only in HROG05 cells, ipatasertib showed anti-tumoral effects in vitro and in organotypic brain slices. Additionally, ipatasertib diminished synchronous network activity in organotypic brain slices. Overall, our data suggest that ipatasertib was only effective in selected tumor models, while especially regorafenib and MK-2206 presented a uniform response pattern.

Mutations in the mitochondrial DNA (mtDNA) are widely known to impact on lifespan and tissue integrity. For example, more than 250 pathogenic mtDNA mutations are known, many of which lead to neurological symptoms. In addition, major neurodegenerative diseases share key components of their etiopathogenesis with regard to mtDNA mutations, mitochondrial dysfunction and oxidative stress. In our study we used a set of conplastic mouse models carrying stable point mutations in mitochondrial genes of transfer RNA (tRNA) and oxidative phosphorylation (OXPHOS)-proteins. We analyzed the impact of these mutations on complex traits like lifespan, learning and memory in the ageing process. The combination of both point mutations in the OXPHOS complex IV gene and adenine insertions in the mitochondrially encoded tRNA arginine (tRNA-Arg) gene ( mt-Tr ) leads to an age-dependent phenotype with elevated mitochondrial superoxide production in the neocortex. Mice with this combination of tRNA and OXPHOS mutations show significantly reduced lifespan and poor physical constitution at the age of 24 months, whereas single point mutations in OXPHOS or mt-tRNA(Arg) do not have this impact. Therefore, we suggest a synergistic effect of these mutations.

Cerebral ischemia–reperfusion injury is a key determinant of a poor outcome after stroke. The mitochondrial uncoupling protein 2 (UCP2) has been implicated in cerebral ischemia-reperfusion injury and in the outcome of ischemic stroke, although its role remains controversial. In C57BL/6J and B6.129S4-Ucp2^tm1Lowl/J (UCP2^−/−) mice, we analyzed cognitive function and lifespan. In an MCAO model induced for one hour, infarct volumes, neurological deficits, and gene expression patterns were determined after 24 h. The UCP2 inhibitor genipin was used in an oxygen-glucose deprivation (OGD) model to investigate synaptic transmission in the hippocampus. Compared to controls, UCP2^−/− mice exhibited a reduced lifespan and displayed impaired cognition. However, in 6-month-old UCP2^−/− mice, the infarct volume was reduced, primarily due to a smaller core size, but not in 18-month-old animals. In both strains, ischemia induced upregulation of antioxidant defense genes, including catalase and SOD1. In the ex vivo ODG model, synaptic transmission was depressed, but pretreatment with genipin prevented the tissue from this impairment. Our findings indicate an infarct-reducing effect of UCP2 deficiency, especially in young-adult mice, and, mechanistically, a neuroprotective effect by genipin in hippocampal slices.