Epilepsy is characterized by recurrent seizures due to a perturbed balance between glutamate and GABA neurotransmission. Our goal is to reveal the molecular mechanisms of the changes upon repeated challenges of this balance, suggesting knowledge-based neuroprotection. To address this goal, a set of metabolic indicators in the post-seizure rat brain cortex is compared before and after pharmacological kindling with pentylenetetrazole (PTZ). Vitamins B1 and B6 supporting energy and neurotransmitter metabolism are studied as neuroprotectors. PTZ kindling increases the seizure severity (1.3 fold, p < 0.01), elevating post-seizure rearings (1.5 fold, p = 0.03) and steps out of the walls (2 fold, p = 0.01). In the kindled vs. non-kindled rats, the post-seizure p53 level is increased 1.3 fold (p = 0.03), reciprocating a 1.4-fold (p = 0.02) decrease in the activity of 2-oxoglutarate dehydrogenase complex (OGDHC) controlling the glutamate degradation. Further, decreased expression of deacylases SIRT3 (1.4 fold, p = 0.01) and SIRT5 (1.5 fold, p = 0.01) reciprocates increased acetylation of 15 kDa proteins 1.5 fold (p < 0.01). Finally, the kindling abrogates the stress response to multiple saline injections in the control animals, manifested in the increased activities of the pyruvate dehydrogenase complex, malic enzyme, glutamine synthetase and decreased malate dehydrogenase activity. Post-seizure animals demonstrate correlations of p53 expression to the levels of glutamate (r = 0.79, p = 0.05). The correlations of the seizure severity and duration to the levels of GABA (r = 0.59, p = 0.05) and glutamate dehydrogenase activity (r = 0.58, p = 0.02), respectively, are substituted by the correlation of the seizure latency with the OGDHC activity (r = 0.69, p < 0.01) after the vitamins administration, testifying to the vitamins-dependent impact of the kindling on glutamate/GABA metabolism. The vitamins also abrogate the correlations of behavioral parameters with seizure duration (r 0.53–0.59, p < 0.03). Thus, increased seizures and modified post-seizure behavior in rats after PTZ kindling are associated with multiple changes in the vitamin-dependent brain metabolism of amino acids, linked to key metabolic regulators: p53, OGDHC, SIRT3 and SIRT5.

Fine scale spatial structure (FSSS) of cytoplasmic genes in plants is thought to be generated via founder events and can be amplified when seeds germinate close to their mother. In gynodioecious species these processes are expected to generate FSSS in sex ratio because maternally inherited cytoplasmic male sterility genes partially influence sex expression. Here we document a striking example of FSSS in both mitochondrial genetic markers and sex in roadside populations of Silene vulgaris. We show that in one population FSSS of sexes influences relative fruit production of females compared to hermaphrodites. Furthermore, FSSS in sex ratio is expected to persist into future generations because offspring sex ratios from females are female-biased whereas offspring sex ratios from hermaphrodites are hermaphrodite-biased. Earlier studies indicated that pollen limitation is the most likely mechanism underlying negative frequency dependent fitness of females. Our results support the theoretical predictions that FSSS in sex ratio can reduce female fitness by decreasing the frequency at which females experience hermaphrodites. We argue that the influence of FSSS on female fitness is complementary to the influence of larger scale population structure on female fitness, and that population structure at both scales will act to decrease female frequencies in gynodioecious species. Better comprehension of the spatial structure of genders and genes controlling sex expression at a local scale is required for future progress toward understanding sex ratio evolution in gynodioecious plants.

Coupling glycolysis and mitochondrial tricarboxylic acid cycle, pyruvate dehydrogenase (PDH) complex (PDHC) is highly responsive to cellular demands through multiple mechanisms, including PDH phosphorylation. PDHC also produces acetyl-CoA for protein acetylation involved in circadian regulation of metabolism. Thiamine (vitamin B1) diphosphate (ThDP) is known to activate PDH as both coenzyme and inhibitor of the PDH inactivating kinases. Molecular mechanisms integrating the function of thiamine-dependent PDHC into general redox metabolism, underlie physiological fitness of a cell or an organism. Here, we characterize the daytime- and thiamine-dependent changes in the rat brain PDHC function, expression and phosphorylation, assessing their impact on protein acetylation and metabolic regulation. Morning administration of thiamine significantly downregulates both the PDH phosphorylation at Ser293 and SIRT3 protein level, the effects not observed upon the evening administration. This action of thiamine nullifies the daytime-dependent changes in the brain PDHC activity and mitochondrial acetylation, inducing diurnal difference in the cytosolic acetylation and acetylation of total brain proteins. Screening the daytime dependence of central metabolic enzymes and proteins of thiol/disulfide metabolism reveals that thiamine also cancels daily changes in the malate dehydrogenase activity, opposite to those of the PDHC activity. Correlation analysis indicates that thiamine abrogates the strong positive correlation between the total acetylation of the brain proteins and PDHC function. Simultaneously, thiamine heightens interplay between the expression of PDHC components and total acetylation or SIRT2 protein level. These thiamine effects on the brain acetylation system change metabolic impact of acetylation. The changes are exemplified by the thiamine enhancement of the SIRT2 correlations with metabolic enzymes and proteins of thiol-disulfide metabolism. Thus, we show the daytime- and thiamine-dependent changes in the function and phosphorylation of brain PDHC, contributing to regulation of the brain acetylation system and redox metabolism. The daytime-dependent action of thiamine on PDHC and SIRT3 may be of therapeutic significance in correcting perturbed diurnal regulation.

The disturbed metabolism of vitamins B1 or B6, which are essential for neurotransmitters homeostasis, may cause seizures. Our study aims at revealing therapeutic potential of vitamins B1 and B6 by estimating the short- and long-term effects of their combined administration with the seizure inductor pentylenetetrazole (PTZ). The PTZ dose dependence of a seizure and its parameters according to modified Racine’s scale, along with delayed physiological and biochemical consequences the next day after the seizure are assessed regarding sexual dimorphism in epilepsy. PTZ sensitivity is stronger in the female than the male rats. The next day after a seizure, sex differences in behavior and brain biochemistry arise. The induced sex differences in anxiety and locomotor activity correspond to the disappearance of sex differences in the brain aspartate and alanine, with appearance of those in glutamate and glutamine. PTZ decreases the brain malate dehydrogenase activity and urea in the males and the phenylalanine in the females. The administration of vitamins B1 and B6 24 h before PTZ delays a seizure in female rats only. This desensitization is not observed at short intervals (0.5–2 h) between the administration of the vitamins and PTZ. With the increasing interval, the pyridoxal kinase (PLK) activity in the female brain decreases, suggesting that the PLK downregulation by vitamins contributes to the desensitization. The delayed effects of vitamins and/or PTZ are mostly sex-specific and interacting. Our findings on the sex differences in sensitivity to epileptogenic factors, action of vitamins B1/B6 and associated biochemical events have medical implications.

Function of brain amino acids as neurotransmitters or their precursors implies changes in the amino acid levels and/or metabolism in response to physiological and environmental challenges. Modelling such challenges by pregnancy and/or hypoxia, we characterize the amino acid pool in the rat cerebellum, quantifying the levels and correlations of 15 amino acids and activity of 2-oxoglutarate dehydrogenase complex (OGDHC). The parameters are systemic indicators of metabolism because OGDHC limits the flux through mitochondrial TCA cycle, where amino acids are degraded and their precursors synthesized. Compared to non-pregnant state, pregnancy increases the cerebellar content of glutamate and tryptophan, decreasing interdependence between the quantified components of amino acid metabolism. In response to hypoxia, the dependence of cerebellar amino acid pool on OGDHC and the average levels of arginine, glutamate, lysine, methionine, serine, phenylalanine, and tryptophan increase in non-pregnant rats only. This is accompanied by a higher hypoxic resistance of the non-pregnant vs. pregnant rats, pointing to adaptive significance of the hypoxia-induced changes in the cerebellar amino acid metabolism. These adaptive mechanisms are not effective in the pregnancy-changed metabolic network. Thus, the cerebellar amino acid levels and OGDHC activity provide sensitive markers of the physiology-dependent organization of metabolic network and its stress adaptations.

Specific inhibitors of mitochondrial 2-oxoglutarate dehydrogenase (OGDH) are administered to animals to model the downregulation of the enzyme as observed in neurodegenerative diseases. Comparison of the effects of succinyl phosphonate (SP, 0.02 mmol/kg) and its uncharged precursor, triethyl succinyl phosphonate (TESP, 0.02 and 0.1 mmol/kg) reveals a biphasic response of the rat brain metabolism and physiology to increasing perturbation of OGDH function. At the low (TE)SP dose, glutamate, NAD⁺, and the activities of dehydrogenases of 2-oxoglutarate and malate increase, followed by their decreases at the high TESP dose. The complementary changes, i.e., an initial decrease followed by growth, are demonstrated by activities of pyruvate dehydrogenase and glutamine synthetase, and levels of oxidized glutathione and citrulline. While most of these indicators return to control levels at the high TESP dose, OGDH activity decreases and oxidized glutathione increases, compared to their control values. The first phase of metabolic perturbations does not cause significant physiological changes, but in the second phase, the ECG parameters and behavior reveal decreased adaptability and increased anxiety. Thus, lower levels of OGDH inhibition are compensated by the rearranged metabolic network, while the increased levels induce a metabolic switch to a lower redox state of the brain, associated with elevated stress of the animals.