Modulation of taurine release in glucose-free media by glutamate receptors in hippocampal slices from developing and adult mice

Taurine has been thought to protect neural cells against cell-damaging conditions to which the hippocampus is particularly vulnerable. We studied now how the release of preloaded [³H]taurine is regulated by glutamate receptors in glucose-free media in slices prepared from the mouse hippocampus from developing (7 days old) and young adult (3 months old) mice, using a superfusion system. The lack of glucose enhanced taurine release more from slices from developing mice than from slices from adults. At both ages ionotropic glutamate agonists significantly increased the release in a receptor-mediated manner. Of the metabotropic glutamate receptors those belonging to the group III were effective. The release was enhanced in adult mice but attenuated in developing mice. Both effects were blocked by the receptor antagonists. The results show that glutamate receptors affect taurine release in the absence of glucose in which condition taurine should be neuroprotective.     

Glucose‐based microbial production of the hormone melatonin in yeast Saccharomyces cerevisiae

Melatonin is a natural mammalian hormone that plays an important role in regulating the circadian cycle in humans. It is a clinically effective drug exhibiting positive effects as a sleep aid and a powerful antioxidant used as a dietary supplement. Commercial melatonin production is predominantly performed by complex chemical synthesis. In this study, we demonstrate microbial production of melatonin and related compounds, such as serotonin and N‐acetylserotonin. We generated Saccharomyces cerevisiae strains that comprise heterologous genes encoding one or more variants of an L‐tryptophan hydroxylase, a 5‐hydroxy‐L‐tryptophan decarboxylase, a serotonin acetyltransferase, an acetylserotonin O‐methyltransferase, and means for providing the cofactor tetrahydrobiopterin via heterologous biosynthesis and recycling pathways. We thereby achieved de novo melatonin biosynthesis from glucose. We furthermore accomplished increased product titers by altering expression levels of selected pathway enzymes and boosting co‐factor supply. The final yeast strain produced melatonin at a titer of 14.50 ± 0.57 mg L^?1 in a 76h fermentation using simulated fed‐batch medium with glucose as sole carbon source. Our study lays the basis for further developing a yeast cell factory for biological production of melatonin.     

Skills training groups for men with ADHD in compulsory care due to substance use disorder: a feasibility study

Dialectical behavior therapy (DBT)-based skills training has been developed and previously evaluated for adults with ADHD in a psychiatric outpatient context. The aim of the present study was to evaluate the feasibility of DBT-based skills training as a voluntary intervention for men with ADHD in compulsory care due to severe substance abuse. Forty sufficiently detoxified men with ADHD in compulsory care due to life-threatening substance use disorder (SUD) were included in DBT-based skills training groups. Self- and staff-rating scales were administered before and after the treatment. The refusal rate was 42.9 %. Of those who started the DBT-based skills training, 70 % completed the treatment (attendance at ≥75 % of the sessions). The treatment acceptability was good. Both ADHD and psychiatric symptoms decreased from pre- to post-intervention in self-ratings, but not in staff ratings. The patients reported improved general well-being. The correlation between self- and staff ratings was poor. Motivation for voluntary nonpharmacological treatment was low in a compulsory care context. However, the results indicate that a DBT-based skills training program for adults with ADHD may be feasible for some patients with ADHD in combination with SUD in compulsory care, provided that considerable resources are allocated with adjustments to the target group and compulsory care context.     

Strong genetic isolation despite wide distribution in a commercially exploited coastal shark

The common smoothhound, Mustelus mustelus, is an epibenthic species targeted by fisheries around the world driven by the increasing demand for shark products. Given the wide-spread occurrence of this species and corresponding lack of molecular data in many areas of said distribution, baseline molecular assessments of this commercially important shark may contribute to finer-scale analyses in areas in which this species is targeted. Therefore, population genetic analyses were conducted along the East Atlantic, from the Mediterranean Sea to the south-east coast of Africa, using microsatellite markers and the mitochondrial control region (mtCR). Overall, M. mustelus displayed low to moderate genetic diversity, with the Mediterranean populations appearing to exhibit the lowest mitochondrial diversity, and the west African populations displaying the lowest nuclear diversity. Microsatellite analysis indicated strong genetic differentiation between the three regions, with finer-scale population structure in each region, without correlation between genetic and geographical distance. For the mtCR sequences, a total of 18 haplotypes were identified, with a high degree of divergence discernable between the regions, largely in accordance with the microsatellite data. The study documents a remarkable level of population isolation across a vast area, suggesting little or no present-day connectivity among extant populations. The findings may serve as an essential baseline for global population management and commercial traceability of this threatened shark.

     

β-Alanine Release from the Adult and Developing Hippocampus Is Enhanced by Ionotropic Glutamate Receptor Agonists and Cell-Damaging Conditions

The release of the inhibitory amino acid -alanine was investigated in hippocampal slices from adult (3-month-old) and developing (7-day-old) mice, using a superfusion system. The release was enhanced by -alanine itself and the structural analogs taurine and -aminobutyrate. It was dependent on Na⁺, but independent of Ca²⁺ in both mature and immature hippocampus, being thus mostly mediated by uptake carriers operating in an outward direction. The release was potentiated in the developing mice, but not affected in the adults, by the ionotropic glutamate receptor agonists N-methyl-D-aspartate, kainate, 2-amino-3-hydroxy-5-methyl-4-isoxazolepropionate and tetrazolylglycine in a receptor-mediated manner. Cell-damaging conditions, including hypoxia, hypoglycemia, ischemia, oxidative stress and the presence of free radicals, greatly enhanced -alanine release at both ages, but more markedly in the adults. The great amounts of -alanine, together with the inhibitory amino acids taurine and -aminobutyrate, released simultaneously with the excitatory amino acids in the hippocampus may constitute an important protective mechanism against excitotoxicity, which leads to neuronal death.     

Ultrasound indentation of bovine knee articular cartilage in situ

We have earlier developed a handheld ultrasound indentation instrument for the diagnosis of articular cartilage degeneration. In ultrasound indentation, cartilage is compressed with the ultrasound transducer. Tissue thickness and deformation are calculated from the A-mode ultrasound signal and the stress applied is registered with the strain gauges. In this study, the applicability of the ultrasound indentation instrument to quantify site-dependent variation in the mechano-acoustic properties of bovine knee cartilage was investigated. Osteochondral blocks (n=6 per site) were prepared from the femoral medial condyle (FMC), the lateral facet of the patello-femoral groove (LPG) and the medial tibial plateau (MTP). Cartilage stiffness (dynamic modulus, E(dyn)), as obtained with the ultrasound indentation instrument in situ, correlated highly linearly (r=0.913, p<0.01) with the values obtained using the reference material-testing device in vitro. Reproducibility (standardized coefficient of variation) of the ultrasound indentation measurements was 5.2%, 1.7% and 3.1% for E(dyn), ultrasound reflection coefficient of articular surface (R) and thickness, respectively. E(dyn) and R were site dependent (p<0.05, Kruskall-Wallis H test). E(dyn) was significantly higher (p<0.05, Kruskall-Wallis Post Hoc test) in LPG (mean+/-SD: 10.1+/-3.1MPa) than in MTP (2.9+/-1.4MPa). In FMC, E(dyn) was 4.6+/-1.3MPa. R was significantly (p<0.05) lower at MTP (2.0+/-0.7%) than at other sites (FMC: 4.2+/-0.9%; LPG: 4.4+/-0.8%). Cartilage glycosaminoglycan concentration, as quantified with the digital densitometry, correlated positively with E(dyn) (r=0.678, p<0.01) and especially with the equilibrium Young's modulus (reference device, r=0.874, p<0.01) but it was not associated with R (r=0.294, p=0.24). We conclude that manual measurements are reproducible and the instrument may be used for detection of cartilage quality in situ. Especially, combined measurement of thickness, E(dyn) and R provides valuable diagnostic information on cartilage status.     

Taurine Release in the Developing and Adult Mouse Hippocampus: Involvement of Cyclic Guanosine Monophosphate

The inhibitory neuromodulator taurine is involved in osmoregulation and cell volume adjustments in the central nervous system. In addition, taurine protects neural cells from excitotoxicity and prevents harmful metabolic events evoked by cell-damaging conditions. The release of taurine in nervous cell preparations is greatly enhanced by glutamate receptor agonists and various cell-damaging conditions. NO-generating compounds also increase taurine release in the mouse hippocampus. The further involvement of the NO/cGMP pathway and protein kinases in preloaded [³H]taurine release from hippocampal slices from adult (3-month-old) and developing (7-day-old) mice in normoxia and in ischemia was now studied using a superfusion system. The release was enhanced by 8-Br-cGMP and the phosphodiesterase inhibitor 2-(2-propyloxyphenyl)-8-azapurin-6-one (zaprinast), particularly in the immature hippocampus, indicating that increased cGMP levels induce taurine release. The release was also increased by the inhibitor of soluble guanylyl cyclase, 1H-(1,2,4)oxadiazolo-(4,3a)quinoxalin-1-one (ODQ) and the protein kinase C activator 4-phorbol 12-myristate 13-acetate (PMA), but only in the adult hippocampus. The ischemia-induced release was also enhanced by increased cGMP levels in both adult and developing mice, whereas protein kinase inhibitors had no effects in any conditions. The results demonstrate that cGMP is able to modulate hippocampal taurine release in both adult and developing mice, the rise in cGMP levels evoking taurine release in normoxia and in ischemia. This could be part of the neuroprotective properties of taurine, being thus important particularly in cell-damaging conditions and in preventing excitotoxicity.     

Involvement of Amino-Acid Side Chains of Membrane Proteins in the Binding of Glutathione to Pig Cerebral Cortical Membranes

Glutathione (GSH), a general antioxidant and detoxifying compound, is the most abundant thiol-containing peptide in the central nervous system. It has been earlier shown to regulate the functions of glutamate receptors and to possess specific binding sites in both neurons and glial cells. The possible involvement of disulfide bonds, cysteinyl, arginyl, lysyl, glutamyl, and aspartyl residues in the binding of tritiated GSH to specific sites in pig cerebral cortical synaptic membranes was now studied after covalent modification of membrane proteins. Treatment of synaptic membranes with the thiol-modifying reagents 5,5-dithio-bis(2-nitrobenzoate) (DTNB) and 4,4-dithiodipyridine (DDP) dramatically enhanced the binding of [³H]GSH in a dose-dependent manner. Dithiothreitol (DTT) alone reduced the binding, but pretreatment of the membranes with DTT potentiated the enhancing effect of DTNB. On the other hand, when the modification with DTNB was followed by treatment with DTT, the enhancement by DTNB was completely reversed. N-ethylmaleimide, a thiol alkylating agent, and phenylisothiocyanate, a thiol- and amino-group modifying compound, reduced the binding, and their effects were additive. The guanidino-modifying agent phenylglyoxal reduced the binding but the carboxyl-modifying reagent 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide had no significant effect. The results indicate that cysteinyl side chains and disulfide bonds are essential in the binding of GSH to membrane proteins and that arginyl and lysyl side chains may also be directly involved in this process.     

Effects of gender,diet, exogenous melatonin and subchronic PCB exposure on plasma immunoglobulin G in mink

Effects of different fish-based diets (freshwater smelt, Baltic herring, marine herring/cod offal or their mixtures), gender, beta-glucan supplement, exogenous melatonin, and PCB exposure (Aroclor 1242((R)), 1 mg per animal per day in feed) on plasma immunoglobulin G (IgG) in the mink (Mustela vison) were studied. The aims of the study were to find out whether plasma IgG of the mink is affected by the subchronic PCB exposure, and whether biological, nutritional and hormonal effects are large enough to mask the possible IgG response. The concentration of IgG was determined using enzyme-linked immunosorbent assay (ELISA). Sexual dimorphism was detected, the males having higher levels of plasma IgG. In addition, melatonin tended to decrease IgG in females but not males. Diet also affected the humoral immune arm; the mixed-fish diets caused an unfavorable ratio of the oxidation products of lipids vs. vitamin E in liver, and resulted in low IgG concentration in plasma. In males fed Baltic herring, the beta-glucan supplement also lowered IgG levels. The PCBs failed to affect the plasma IgG of the smelt-fed female mink, and IgG concentration was not correlated with increased hepatic EROD activity or with the decreased total retinol in the liver of exposed mink. It is concluded that hormonal/seasonal and dietary factors affect the plasma IgG levels to such an extent that possible change in plasma IgG level due to PCBs in wild populations of mink is difficult to detect without a large amount of reference data.     

Identification of trypanosomes in wild animals from southern Cameroon using the polymerase chain reaction (PCR)

One possible explanation of the maintenance of many historical foci of sleeping sickness in Central Africa could be the existence of a wild animal reservoir. In this study, PCR was used to detect the different trypanosome species present in wild animal captured by hunters in the southern forest belt of Cameroon (Bipindi). Trypanosomes were also detected by a parasitological method (Quantitative buffy coat: QBC). Parasite could not be isolated in culture medium (Kit for in vitro isolation: KIVI). Specific primers of T. brucei s.l., T. congolense forest type, T. congolense savannah type, T. vivax, T. simiae and T. b. gambiense group 1 were used to identify parasites in the blood of 164 animals belonging to 24 different species including ungulates, rodents, pangolins, carnivores, reptiles and primates. Of the 24 studied species, eight were carrying T. b. gambiense group 1. Those parasites pathogenic to man were found in monkeys (Cercocebus torquatus and Cercopithecus nictitans), in ungulates (Cephalophus dorsalis and C. monticola), in carnivores (Nandinia binotata and Genetta servalina) and in rodents (Cricetomys gambianus and Atherurus africanus). 13 species (54%) were carrying T. brucei s.l. identified as non-gambiense group 1.