An optimized method for adult zebrafish brain-tissue dissociation that allows access mitochondrial function under healthy and epileptic conditions.

Mitochondria play key roles in brain metabolism. Not surprisingly, mitochondria dysfunction is a ubiquitous cause of neurodegenerative diseases. In turn, acquired forms of epilepsy etiology is specifically intriguing since mitochondria function and dysfunction remain not completely enlightened. Investigation in the field includes models of epileptic disorder using mainly rodents followed by mitochondrial function evaluation, which in general evidenced controversial data. So, we considered the efforts and limitations in this research field and we took into account that sample preparation and quality are critical for bioenergetics investigation. For these reasons the aim of the present study was to develop a thorough protocol for adult zebrafish brain-tissue dissociation to evaluate oxygen consumption flux and reach the bioenergetics profile in health and models of epileptic disorder in both, in vitro using pentylenetetrazole (PTZ) and N-methyl-D-Aspartic acid (NMDA), and in vivo after kainic acid (KA)-induced status epilepticus. In conclusion, we verify that fire-polished glass Pasteur pipette is eligible to brain-tissue dissociation and to study mitochondrial function and dysfunction in adult zebrafish. The results give evidence for large effect size in increase of coupling efficiency respiration (p/O2) correlated to treatment with PTZ and spare respiratory capacity (SRC) in KA-induced model indicating oxidative phosphorylation (OXPHOS) variable alterations. Further investigation is needed in order to clarify the bioenergetics role as well as other mitochondrial functions in epilepsy.

Lipid-lowering effect of Pinus sp. sawdust and Pycnoporus sanguineus mycelium in streptozotocin-induced diabetic rats.

The purpose of this study was to investigate the effect of diet supplementations on biochemical, hematological, and redox metabolism parameters in streptozotocin-induced diabetic rats. Healthy male Wistar rats and streptozotocin-induced diabetic rats were provided diets supplemented with 20% of Pinus sp. sawdust or Pycnoporus sanguineus mycelium for 4 weeks. Diabetic rats treated with both Pinus sp. sawdust- and P. sanguineus mycelium-supplemented diets presented a significant decrease in non-HDL cholesterol of 38.43% and 33.53% and triglycerides of 70.03% and 69.81%, respectively, compared to diabetic control. As far as we know, this is the first report of a significant decrease in serum lipids attributed to these supplementations. Even though with the alterations in hematological and redox metabolism parameters related to these diet treatments, our data suggest that Pinus sp. sawdust and Pycnoporus sanguineus mycelium could be a useful a diet supplement to control diabetic dyslipidemia in animals. PRACTICAL APPLICATIONS: Pinus sp. sawdust is a residue from the wood industry that can be reused as a substrate to cultivate Pycnoporus sanguineus mycelium. Both species have specific and rich natural compounds. The results of the present study surprisingly showed that diets supplemented with the isolated substrate (Pinus sp. sawdust) and Pycnoporus sanguineus mycelium significantly decrease non-HDL cholesterol and triglycerides in induced diabetic animal model. These new natural approaches can be interesting to develop a nutraceutical to treat dyslipidemia.

Grape seed proanthocyanidins improves mitochondrial function and reduces oxidative stress through an increase in sirtuin 3 expression in EA.hy926 cells in high glucose condition.

Proanthocyanidins are phenolic compounds abundant in the diet, commonly found in grapes and derivatives, foods known for their health-promoting benefits. There is previous evidence showing the antidiabetic activity of proanthocyanidins, however, their mechanisms of action have not been fully elucidated. This study evaluated the capacity of grape seed proanthocyanidins extract (GSPE) to modulate oxidative stress, nitric oxide levels, mitochondrial dysfunction, apoptosis, and sirtuin expression in endothelial cells EA.hy926 under high glucose condition. In addition, the possible toxic effects of GSPE was evaluated in a zebrafish embryos model. The results showed that GSPE was able to enhance cell viability and avoid the disturbance in redox metabolism induced by high glucose. Moreover, GSPE was able to avoid mitochondria dysfunction and the increased in p53 and poly-(ADP-ribose) polymerase expression induced by high glucose exposition. These effects were attributed to the increase in expression of sirtuin 3, a protein able to regulate mitochondrial function. GSPE in an effective concentration did not show toxic effects in zebrafish embryos model. Taken together, these data elucidate the key molecular target of GSPE for future pharmacological interventions in diabetic patients.

Fetal alcohol spectrum disorders model alters the functionality of glutamatergic neurotransmission in adult zebrafish.

Fetal alcohol spectrum disorders (FASD) describe a wide range of ethanol-induced developmental disabilities, including craniofacial dysmorphology, and neurochemical and behavioral impairments. Zebrafish has become a popular animal model to evaluate the long-lasting effects of, both, severe and milder forms of FASD, including alterations to neurotransmission. Glutamate is one of the most affected neurotransmitter systems in ethanol-induced developmental disabilities. Therefore, the aim of the present study was to evaluate the functionality of the glutamatergic neurotransmitter system in an adult zebrafish FASD model. Zebrafish larvae (24 h post-fertilization) were exposed to ethanol (0.1 %, 0.25 %, 0.5 %, and 1%) for 2 h. After 4 months, the animals were euthanized and their brains were removed. The following variables were measured: glutamate uptake, glutamate binding, glutamine synthetase activity, Na+/K + ATPase activity, and high-resolution respirometry. Embryonic ethanol exposure reduced Na+-dependent glutamate uptake in the zebrafish brain. This reduction was positively modulated by ceftriaxone treatment, a beta-lactam antibiotic that promotes the expression of the glutamate transporter EAAT2. Moreover, the 0.5 % and 1% ethanol groups demonstrated reduced glutamate binding to brain membranes and decreased Na+/K + ATPase activity in adulthood. In addition, ethanol reduced glutamine synthetase activity in the 1% EtOH group. Embryonic ethanol exposure did not alter the immunocontent of the glutamate vesicular transporter VGLUT2 and the mitochondrial energetic metabolism of the brain in adulthood. Our results suggest that embryonic ethanol exposure may cause significant alterations in glutamatergic neurotransmission in the adult zebrafish brain.