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Lactate, a metabolic end product of glycolysis in mammals, has emerged as an important energy substrate for the brain. In addition to its energetic role, lactate was shown to modulate the excitability of neurons, to have a neuroprotective role and to participate in long-term memory formation. One previous investigation from our group reported that lactate modulates 4 synaptic plasticity-associated genes and potentiates the activity of the N-Methyl-D-aspartic acid (NMDA) receptor, a major receptor type involved in glutamatergic neurotransmission. The current thesis aimed at first to extend these findings by examining genome-wide transcriptional responses to this metabolite in cortical neurons. Using ribonucleic acid(RNA) sequencing to evaluate expression changes in protein-coding genes, we found that lactate modulates robustly after 1h, 20 genes involved in the mitogen-activated protein kinase (MAPK) signaling pathway and in synaptic plasticity in a NMDA receptor activitydependent manner and that nicotinamide adenine dinucleotide, reduced (NADH), but not pyruvate, reproduces the modulatory effects of lactate on 70% of all differentially expressed genes. In a time course experiment, genes modulated after lactate treatment for 6h and 24h were also identified; these are involved in 9 signaling pathways including circadian rhythm, drug addiction, and retrograde endocannabinoid signaling. Bioinformatics analyses indicated CREB1 and CREM as candidate master regulators of gene expression and the modulatory effect of lactate was prevented by inhibitors of Ca2+/calmodulin-dependent protein kinase II (CaMKII) activity, indicating a role for this kinase in mediating lactate signaling. An examination of changes in dendritic spines’ morphology and density - a morphologicalcorrelate of synaptic plasticity – has shown that lactate modulated spine density changes induced by potassium chloride (KCl) and carbachol. An additional investigation described in this thesis indicated that different gut microbiota manipulations (germ-free, prebiotics, high-fat diet) regulated mRNA expression of genes involved in the Astrocyte-Neuron Lactate Shuttle (ANLS) - a metabolic cooperation mechanism between astrocytes and glutamatergic neurons. Overall, the results of this thesis help to establish a role for lactate as a signaling molecule in the brain, highlight mechanisms implicated in its signaling, and open new avenues for investigation of links between the gut microbiota and brain energy metabolism.

Nanostructures fabricated by different methods have become increasingly important for various applications at the cellular level. In order to understand how these nanostructures “behave” and for studying their internalization kinetics, several attempts have been made at tagging and investigating their interaction with living cells. In this study, magnetic iron nanowires with an iron oxide layer are coated with (3-Aminopropyl)triethoxysilane (APTES), and subsequently labeled with a fluorogenic pH-dependent dye pHrodo™ Red, covalently bound to the aminosilane surface. Time-lapse live imaging of human colon carcinoma HCT 116 cells interacting with the labeled iron nanowires is performed for 24 hours. As the pHrodo™ Red conjugated nanowires are non-fluorescent outside the cells but fluoresce brightly inside, internalized nanowires are distinguished from non-internalized ones and their behavior inside the cells can be tracked for the respective time length. A machine learning-based computational framework dedicated to automatic analysis of live cell imaging data, Cell Cognition, is adapted and used to classify cells with internalized and non-internalized nanowires and subsequently determine the uptake percentage by cells at different time points. An uptake of 85 % by HCT 116 cells is observed after 24 hours incubation at NW-to-cell ratios of 200. While the approach of using pHrodo™ Red for internalization studies is not novel in the literature, this study reports for the first time the utilization of a machine-learning based time-resolved automatic analysis pipeline for quantification of nanowire uptake by cells. This pipeline has also been used for comparison studies with nickel nanowires coated with APTES and labeled with pHrodo™ Red, and another cell line derived from the cervix carcinoma, HeLa. It has thus the potential to be used for studying the interaction of different types of nanostructures with potentially any live cell types.