Voluntary oral dosing for precise experimental compound delivery in adult rats.

Preclinical drug studies routinely administer experimental compounds to animal models with the goal of minimizing potential adverse events from the procedure. In this study, we assessed the ability to train adult male Long Evans rats to accept daily voluntarily syringe feedings of l-3,4-dihydroxyphenylalanine (L-DOPA) compared to intraperitoneal (IP) injections. Rats were trained to become familiar with the syringe and then fed a training solution that did not contain the experimental compound. If the rat was compliant during the training phase, the dilution of training solution was continuously decreased and replaced with the experimental solution. Voluntary oral dosing compliance was recorded and quantified throughout the study. To assess drug activity within the drug-targeted tissues, the striatum and retina were collected and analyzed for L-DOPA, dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) levels by high performance liquid chromatography (HPLC). Drug delivery efficiency by oral dosing was directly compared to IP injection by collecting plasma and analyzing L-DOPA levels with HPLC. Adult male rats had high compliance for voluntary oral dosing. HPLC showed that oral administration of the compound at the same dose as IP injection yielded significantly lower plasma levels, and that higher oral L-DOPA doses yield higher plasma L-DOPA content. This study describes detailed methodology to train adult rats to syringe feed experimental compounds and provides important preclinical research on drug dosing and drug delivery to the striatum and retina.

Effect of Elicitation with (+)-Usnic Acid on Accumulation of Phenolic Acids and Flavonoids in Agitated Microshoots of Eryngium alpinum L.

The present work was aimed at studying the potential of elicitation on the accumulation of phenolic compounds in in vitro shoot cultures of Eryngium alpinum L., a protected plant from the Apiaceae family. The study examined the influence of (+)-usnic acid on the biomass growth as well as on the biosynthesis of the desired flavonoids and phenolic acids in the cultured microshoots. The phenolic compound content was determined by HPLC-DAD. The flavonoid of the highest concentration was isoquercetin, and the phenolic acids of the highest amount were rosmarinic acid, caffeic acid and 3,4-dihydroxyphenylacetic acid, both in the non-elicited and elicited biomass. Isoquercetin accumulation was efficiently increased by a longer elicitation with a lower concentration of lichenic compound (107.17 ± 4.67 mg/100 g DW) or a shorter elicitation with a higher concentration of acid (127.54 ± 11.34 and 108.37 ± 12.1 mg/100 g DW). Rosmarinic acid production generally remained high in all elicited and non-elicited microshoots. The highest content of this acid was recorded at 24 h of elicitation with 3.125 µM usnic acid (512.69 ± 4.89 mg/100 g DW). The process of elicitation with (+)-usnic acid, a well-known lichenic compound with allelopathic nature, may therefore be an effective technique of enhancing phenolic compound accumulation in alpine eryngo microshoot biomass.

Effects of a high-fat-diet supplemented with probiotics and ω3-fatty acids on appetite regulatory neuropeptides and neurotransmitters in a pig model.

The pig is a valuable animal model to study obesity in humans due to the physiological similarity between humans and pigs in terms of digestive and associated metabolic processes. The dietary use of vegetal protein, probiotics and omega-3 fatty acids is recommended to control weight gain and to fight obesity-associated metabolic disorders. Likewise, there are recent reports on their beneficial effects on brain functions. The hypothalamus is the central part of the brain that regulates food intake by means of the production of food intake-regulatory hypothalamic neuropeptides, as neuropeptide Y (NPY), orexin A and pro-opiomelanocortin (POMC), and neurotransmitters, such as dopamine and serotonin. Other mesolimbic areas, such as the hippocampus, are also involved in the control of food intake. In this study, the effect of a high fat diet (HFD) alone or supplemented with these additives on brain neuropeptides and neurotransmitters was assessed in forty-three young pigs fed for 10 weeks with a control diet (T1), a high fat diet (HFD, T2), and HFD with vegetal protein supplemented with Bifidobacterium breve CECT8242 alone (T3) or in combination with omega-3 fatty acids (T4). A HFD provoked changes in regulatory neuropeptides and 3,4-dihydroxyphenylacetic acid (DOPAC) in the hypothalamus and alterations mostly in the dopaminergic system in the ventral hippocampus. Supplementation of the HFD with B. breve CECT8242, especially in combination with omega-3 fatty acids, was able to partially reverse the effects of HFD. Correlations between productive and neurochemical parameters supported these findings. These results confirm that pigs are an appropriate animal model alternative to rodents for the study of the effects of HFD on weight gain and obesity. Furthermore, they indicate the potential benefits of probiotics and omega-3 fatty acids on brain function.

Physiological responses during acute stress recovery depend on stress coping style in European sea bass, Dicentrarchus labrax.

Individual stress coping style (reactive, intermediate and proactive) was determined in 3 groups of 120 pit tagged European seabass using the hypoxia avoidance test. The same three groups (no change in social composition) were then reared according to the standards recommended for this species. Then, 127 days later, individuals initially characterized as reactive, intermediate or proactive were submitted to an acute confinement stress for 30 min. Blood samples were taken to measure plasma cortisol levels 30 min (Stress30) or 150 min (Stress150) after the end of the confinement stress. Individuals were then sacrificed to sample the telencephalon in order to measure the main monoamines and their catabolites (at Stress30 only). Individuals from Stress150 were sampled for whole brain for a transcriptomic analysis. The main results showed that reactive individuals had a lower body mass than intermediate individuals which did not differ from proactive individuals. The physiological cortisol response did not differ between coping style at Stress30 but at Stress150 when intermediate and proactive individuals had recovered pre stress levels, reactive individuals showed a significant higher level illustrating a modulation of stress recovery by coping style. Serotonin turnover ratio was higher in proactive and reactive individuals compared to intermediate individuals and a significant positive correlation was observed with cortisol levels whatever the coping style. Further, the confinement stress led to a general increase in the serotonin turnover comparable between coping styles. Stress150 had a significant effect on target mRNA copy number (Gapdh mRNA copy number decreased while ifrd1 mRNA copy number increased) and such changes tended to depend upon coping style.

Anxiolytic effect of a novel 9,10-dihydrophenanthrene, juncuenin H, is associated with metabolic changes in cortical serotonin/dopamine levels in mice.

Two novel phenanthrenoids, juncuenin H (1) and dijuncuenin B (2), together with eight known phenanthrenoids, effusol (3), dehydroeffusol (4), juncusol (5), dehydrojuncusol (6), juncuenin B (7), dehydrojuncuenin B (8), juncuenin A (9), and dehydrojuncuenin A (10), were isolated from the underground parts of Juncus setchuenensis. The structures of the compounds were determined by 1D and 2D NMR and mass spectroscopy. The anxiolytic activities of compounds 1, 6, 9, and 10 were evaluated. In order to explore the mechanisms underlying their anxiolytic activities, the levels of serotonin (5-HT), dopamine (DA), and their metabolites in the cerebral cortex and hippocampus of mice treated with compound 1 were determined by quantitative mass spectrometry. The mice treated with compound 1 had significantly lower levels of 5-HT, 3-methoxytyramine (3-MT), 5-hydroxyindole-3-acetic acid (5-HIAA), homovanillic acid (HVA), and 3, 4-dihydroxyphenylacetic acid (DOPAC) in the cerebral cortex than those of the vehicle control-treated mice. The levels of HVA and 5-HIAA in the hippocampus were also significantly lower in the mice treated with compound 1 than in the control group mice. These results suggest that the metabolic changes, reflected in the levels of DA and/or 5-HT, may contribute to the anxiolytic activity of the phenanthrenoids studied herein.

Analysis of Catecholamine and Their Metabolites in Mice Brain by Liquid Chromatography-Mass Spectrometry Using Sulfonated Mixed-mode Copolymer Column.

In this study, a simultaneous assay for catecholamines and their metabolites in the brain was established using liquid chromatography-mass spectrometry (LC-MS). To achieve complete separation, a cation-exchange/reversed-phase mixed-mode copolymer resin column containing 0.81 wt% sulfo groups was used for the simultaneous LC-MS assay. The analyzed catecholamines were dopamine (DA), norepinephrine (NE), and epinephrine (E), while the metabolites lacking amino groups were 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and 3-methoxy-4-hydroxyphenylglycol (MHPG). The metabolites were separated and detected using LC-MS, on columns with and without sulfo groups. However, we could not achieve adequate separation of catecholamines on both columns using a gradient elution of 0 - 50 (v/v)% methanol containing 0.1 (v/v)% formic acid (FA). When volatile ion-pairing reagents were added to the mobile phase, they improved the retention and detection of catecholamines on the sulfonated mixed-mode column. Under optimized elution conditions, which involved a linear gradient elution of water containing 0.1 (v/v)% FA to 50 (v/v)% acetonitrile in 50 mM ammonium formate at 40°C and a 0.20 mL/min rate, all six target molecules were simultaneously detected within 25 min, when using negative mode LC-MS on a sulfonated mixed-mode column. The limits of detection (LODs) for DA, NE, E, DOPCA, HVA, and MHPG were determined to be 20.7, 12.6, 74.6, 1110, 18.7, and 3196 nM, respectively. Moreover, the established LC-MS assay allowed the detection of endogenous DA, NE, and HVA, in normal mouse brain samples at concentrations higher than 20, 9, and 4 pmol/mg, respectively.

Neuropharmacological effects of camel milk related to modulation of biogenic amines in experimental animals.

Camel milk is reported as anti-diabetic, hepato-protective, anticancer, antioxidant, antiviral and neuroprotectant in numerous studies. Based on its neuroprotective profile, camel milk is investigated for its possible beneficial effect in treating anxiety and depression and its effect on brain biogenic amines in the present study. Head dip, cage crossing, stationary rod, elevated plus-maze, open field, light & dark box and forced swim tests were used to measure change in rodents' behavior after camel milk administration. Any possible change in brain biogenic amines level after camel milk treatment was evaluated using High Performance Liquid Chromatography (HPLC) technique. Camel milk administration resulted in significant increase (p<0.001) in exploratory and locomotor activity and showing anxiolytic behavior in rodents. In depression-like model, rats showed significant increase (p<0.001) in struggling time after 30-days administration of camel milk. HPLC detection of brain biogenic amines revealed significant increase (p<0.001) in norepinephrine, insignificant increase in 5-hydroxytryptamine and significant decrease (p<0.001) in dopamine, 3,4-dihydroxyphenylacetic acid, homovanillic acid and 5-hydroxyindoleacetic acid in camel milk treated group. Based on above findings, camel milk is suggested as anxiolytic and antidepressant in the administered doses. However, further experimental and clinical investigations are required to authenticate the same at different doses.

The effects of electrical stimulation of the peripheral vestibular system on neurochemical release in the rat striatum.

For over a century, it has been speculated that the vestibular system transmits information about self-motion to the striatum. There have been inconsistent reports of such a connection, and interest in the subject has been increased by the experimental use of galvanic vestibular stimulation in the treatment of Parkinson's Disease patients. Nonetheless, there are few data available on the effects of vestibular stimulation on neurochemical changes in the striatum. We used in vivo microdialysis to analyse changes in the extracellular levels of amino acids and monoamines in the rat striatum, following electrical vestibular stimulation. Stimulation caused a significant decrease in serine and threonine, compared to the no-stimulation controls (P ≤ 0.005 and P ≤ 0.01, respectively). The ratio of DOPAC:dopamine, decreased on the ipsilateral side following stimulation (P ≤ 0.005). There was a significant treatment x side x intensity interaction for taurine levels (P ≤ 0.002), due to a decrease on the contralateral side in stimulated animals, which varied as a function of current. These results show that peripheral vestibular stimulation causes some neurochemical changes in the striatum and support the view that activaton of the vestibular system exerts effects on the function of the striatum.

Intrinsic exercise capacity in rats influences dopamine neuroplasticity induced by physical training.

The study evaluates whether the intrinsic capacity for physical exercise influences dopamine neuroplasticity induced by physical training. Male rats were submitted to three progressive tests until fatigue. Based on the maximal time of exercise (TE), rats were considered as low performance (LP), standard performance (SP) or high performance (HP) to exercise. Eight animals from each group (LP, SP, and HP) were randomly subdivided in sedentary (SED) or trained (TR). Physical training was performed for 6 wk. After that, concentrations of dopamine (DA), serotonin (5-HT), and their metabolites and mRNA levels of D1 receptor ( Drd1), D2 receptor ( Drd2), dopamine transporter ( Dat), tyrosine hydroxylase ( Th), glia cell line neurotrophic factor ( Gdnf), and brain-derived neurotrophic factor ( Bdnf) were determined in the caudate-putamen (CPu). TE was increased with training in all performance groups. However, the relative increase was markedly higher in LP rats, and this was associated with a training-induced increase in dopaminergic activity in the CPu, which was determined by the 3,4-dihydroxyphenylacetic acid (DOPAC)/DA ratio. An opposite monoamine response was found in HP-TR rats, in which physical training decreased the DOPAC/DA ratio in the CPu. Moreover, LP-SED rats displayed higher levels of Drd2 in the CPu compared with the other SED groups, and this higher expression was decreased by physical training. Physical training also decreased Dat and increased Gdnf in the CPu of LP rats. Physical training decreased Bdnf in the CPu only in HP rats. Thus, we provide evidence that the intrinsic capacity to exercise affects the neuroplasticity of the dopaminergic system in response to physical training. NEW & NOTEWORTHY The findings reported reveal that dopaminergic neuroplasticity in caudate-putamen induced by physical training is influenced by the intrinsic capacity to exercise in rats. To evaluate the dopaminergic neuroplasticity, we analyzed mRNA levels of D1 receptor, D2 receptor, dopamine transporter, tyrosine hydroxylase, glia cell line neurotrophic factor, and brain-derived neurotrophic factor as well as concentrations of dopamine, serotonin, and their metabolites. These results expand our knowledge about the interrelationship between genetic background, physical training, and dopaminergic neuroplasticity.

Assessment of gait dynamics in rotenone-induced rat model of Parkinson's disease by footprint method.

Rotenone (organic pesticide and inhibitor of mitochondrial complex I) is used to generate an experimental model of Parkinson's disease (PD). In the present study, we investigated rotenone-induced locomotor deficits, gait dynamics and muscular weakness in rats. The study also determined dopamine (DA) and dihydroxyphenylacetic acid (DOPAC) levels following rotenone administration. In the study, adult male rats were administered subcutaneously (s.c.) with rotenone (1.5 mg/kg/day) for 8 days. Motor activities were monitored by the Kondziela's inverted screen test, beam walking test and footprint test. Animals were decapitated after behavioral analysis and brains were dissected out for neurochemical estimation. Results showed that the levels of DA and DOPAC were significantly decreased, which further supported by significant impaired motor coordination in rotenone treated rats. In conclusion, the behavioral and neurochemical findings of our study further strengthen the previous report and emphasizes on short term administration of rotenone producing PD-like symptoms in rats.

Effects of Mitochondrial Antioxidant SkQ1 on Biochemical and Behavioral Parameters in a Parkinsonism Model in Mice.

According to one hypothesis, Parkinson's disease pathogenesis is largely caused by dopamine catabolism that is catalyzed on mitochondrial membranes by monoamine oxidase. Reactive oxygen species are formed as a byproduct of these reactions, which can lead to mitochondrial damage followed by cell degeneration and death. In this study, we investigated the effects of administration of the mitochondrial antioxidant SkQ1 on biochemical, immunohistochemical, and behavioral parameters in a Parkinson-like condition caused by protoxin MPTP injections in C57BL/6 mice. SkQ1 administration increased dopamine quantity and decreased signs of sensory-motor deficiency as well as destruction of dopaminergic neurons in the substantia nigra and ventral tegmental area in mice with the Parkinson-like condition.

Determination of dopamine, serotonin, biosynthesis precursors and metabolites in rat brain microdialysates by ultrasonic-assisted in situ derivatization-dispersive liquid-liquid microextraction coupled with UHPLC-MS/MS.

This paper, for the first time, reported a simple, rapid, sensitive and environmental friendly ultrasonic-assisted in situ derivatization-dispersive liquid-liquid microextraction (in situ UAD-DLLME) method followed by ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) for the simultaneous determination of dopamine (DA), serotonin (5-HT) and their biosynthesis precursors and metabolites in rat brain microdialysates. In this work, a commercial reagent, Lissamine rhodamine B sulfonylchloride (LRSC), was proposed as a derivatization reagent. The ionization efficiency of neurotransmitters was greatly enhanced through the introduction of a permanent charged moiety of LRSC into their derivatives during electrospray ionization MS (ESI-MS) analysis. Parameters of in situ UAD-DLLME and UHPLC-MS/MS conditions were all optimized in detail. The optimum conditions of in situ UAD-DLLME were found to be as follows: a mixture of 150µL of acetonitrile (dispersant) containing LRSC (derivatization reagents) and 50µL of low toxic bromobenzene (extractant) was rapidly injected into an aqueous sample containing 30µL of microdialysate and 800µL of NaHCO3-Na2CO3 buffer solution (pH 10.5) at 37°C. After ultrasonication for 3min and centrifuging for 2min, the sedimented phase was conveniently injected for UHPLC-MS/MS analysis. Under the optimized conditions, good linearity was observed with the limits of detection (LODs, S/N>3) and limits of quantification (LOQs, S/N>10) in the range of 0.002-0.008 and 0.015-0.040nmol/L, respectively. Meanwhile, it also brought good results of precision (3.2-13.0%, peak area RSDs %), accuracy (86.4-112%), recovery (73.9-105%), matrix effect (86.2-105%), and stability (3.1-8.8%, peak area RSDs %). The developed method was successfully applied for the simultaneous determination of multiple neurotransmitters, their precursors and metabolites in brain microdialysates of normal and L-DOPA induced dyskinesias (LID) rats.

Multiple cone pathways are involved in photic regulation of retinal dopamine.

Dopamine is a key neurotransmitter in the retina and plays a central role in the light adaptive processes of the visual system. The sole source of retinal dopamine is dopaminergic amacrine cells (DACs). We and others have previously demonstrated that DACs are activated by rods, cones, and intrinsically photosensitive retinal ganglion cells (ipRGCs) upon illumination. However, it is still not clear how each class of photosensitive cells generates light responses in DACs. We genetically isolated cone function in mice to specifically examine the cone-mediated responses of DACs and their neural pathways. In addition to the reported excitatory input to DACs from light-increment (ON) bipolar cells, we found that cones alternatively signal to DACs via a retrograde signalling pathway from ipRGCs. Cones also produce ON and light-decrement (OFF) inhibitory responses in DACs, which are mediated by other amacrine cells, likely driven by type 1 and type 2/3a OFF bipolar cells, respectively. Dye injections indicated that DACs had similar morphological profiles with or without ON/OFF inhibition. Our data demonstrate that cones utilize specific parallel excitatory and inhibitory circuits to modulate DAC activity and efficiently regulate dopamine release and the light-adaptive state of the retina.

Convergent evidence from alcohol-dependent humans and rats for a hyperdopaminergic state in protracted abstinence.

A major hypothesis in addiction research is that alcohol induces neuroadaptations in the mesolimbic dopamine (DA) system and that these neuroadaptations represent a key neurochemical event in compulsive drug use and relapse. Whether these neuroadaptations lead to a hypo- or hyperdopaminergic state during abstinence is a long-standing, unresolved debate among addiction researchers. The answer is of critical importance for understanding the neurobiological mechanism of addictive behavior. Here we set out to study systematically the neuroadaptive changes in the DA system during the addiction cycle in alcohol-dependent patients and rats. In postmortem brain samples from human alcoholics we found a strong down-regulation of the D1 receptor- and DA transporter (DAT)-binding sites, but D2-like receptor binding was unaffected. To gain insight into the time course of these neuroadaptations, we compared the human data with that from alcohol-dependent rats at several time points during abstinence. We found a dynamic regulation of D1 and DAT during 3 wk of abstinence. After the third week the rat data mirrored our human data. This time point was characterized by elevated extracellular DA levels, lack of synaptic response to D1 stimulation, and augmented motor activity. Further functional evidence is given by a genetic rat model for hyperdopaminergia that resembles a phenocopy of alcohol-dependent rats during protracted abstinence. In summary, we provide a new dynamic model of abstinence-related changes in the striatal DA system; in this model a hyperdopaminergic state during protracted abstinence is associated with vulnerability for relapse.

Hydrophilic interaction chromatography combined with dispersive liquid-liquid microextraction as a preconcentration tool for the simultaneous determination of the panel of underivatized neurotransmitters in human urine samples.

A simple and sensitive method using dispersive liquid-liquid microextraction (DLLME) followed by liquid chromatography coupled to mass spectrometry (LC-MS) with a hydrophilic interaction chromatography (HILIC) column was developed for the simultaneous determination of 13 compounds of different polarities, comprising monoamine neurotransmitters (dopamine, norepinephrine, epinephrine and serotonin) along with their respective precursors and metabolites, in human urine samples. The microextraction procedure was based on the fast injection of a mixture of ethanol (disperser solvent) and dichloromethane (extraction solvent) into a human urine sample, forming a cloudy solution in the Eppendorf tube. After centrifugation, the sedimented phase was collected and subsequently analyzed by LC-HILIC-MS in about 12min without a derivatization step. The separation was performed on an XBridge Amide™ BEH column 3.0×100mm, 3.5mm and the mobile phase consisted of phase A: 10mM ammonium formate buffer in water pH 3.0 and phase B: 10 mM ammonium formate buffer in acetonitrile, under gradient program elution. Tyrosine, tryptophan, 5-hydroxytryptophan, dopamine, epinephrine, norepinephrine, serotonin, 3-methoxytyramine, 5-hydroxyindole-3-acetic acid, 3,4-dihydroxy-l-phenylalanine and norvaline (internal standard) were detected in the positive ionization mode. While vanillylmandelic acid, homovanillic acid, 3,4-dihydroxyphenylacetic acid and 3,4-dihydroxybenzylamine (internal standard) were detected in the negative ionization mode. Parameters influencing DLLME and LC-HILIC-MS were investigated. Under the optimum conditions, the proposed method exhibited a low detection limit (5-10ngmL(-1)), and good linearity with R between 0.9991 and 0.9998. The recoveries in human urine samples were 99.0%±3.6%. for the 13 studied biogenic amines with intra- and inter-day RSDs of 0.24-9.55% and 0.31-10.0%, respectively. The developed DLLME-LC-MS method could be successfully applied for the determination of trace amounts of polar endogenous compounds, such as neurotransmitters, in human urine samples, including samples with a reduced volume obtained from pediatric patients.

Amperometric detection in the presence of carbon nanotubes dispersed in background electrolyte: Evaluating its suitability for capillary electrokinetic chromatography separations of polyphenolic compounds.

This work reports on the positive effects observed upon both the separation and analytical signals in electrophoretic separations of selected phenolic compounds when using aqueous BGE containing carbon nanotubes (CNTs) in connection to electrochemical detection (ECD). The influence of the presence of surfactant-coated CNTs in the BGE upon the amperometric response of probe compounds was evaluated under hydrodynamic regime in capillary flow injection experiments as well as electrophoretic separations. Among the surfactants employed to disperse CNTs within BGE, SDS shows the best results in terms of dispersion stability and degree of dispersion of the CNTs. ECD allows working with BGEs containing CNTs concentrations of, at least, 24.0 mg/L without increasing of baseline noise, on the opposite to that reported when using UV-visible detection, and the presence of CNTs in the BGE improves the electrochemical response of some of the tested compounds. These benefits were reflected in higher sensibility in the electrochemical signal and additional improved resolution in the electrophoretic separation of (±)-catechin and sinapic acid when using these BGE containing CNTs.

Atrazine is primarily responsible for the toxicity of long-term exposure to a combination of atrazine and inorganic arsenic in the nigrostriatal system of the albino rat.

Chronic and simultaneous exposure to a variety of chemicals present in the environment is an unavoidable fact. However, given the complexity of studying chemical mixtures, most toxicological studies have focused on the effects of short-term exposure to single substances. The aim of this study was to evaluate the effects on the nigrostriatal system of the chronic, simultaneous exposure to two widely distributed substances that have been identified as potential dopaminergic system toxicants, inorganic arsenic (iAs) and atrazine (ATR). Six groups of rats were treated daily for one year with atrazine (10mg ATR/kg), inorganic arsenic (0.5 or 50mgiAs/L of drinking water), or a combination of ATR+0.5mgiAs/L or ATR+50mgiAs/L. The 50mgiAs/L group showed locomotor hypoactivity, while all treatments decreased motor coordination in contrast no effects of treatment were found on the place and response learning tasks. Regarding markers for liver and muscle damage, there were no differences between groups in creatine kinase (CK) or aspartate transaminase (AST) activities, while decreases in lactate dehydrogenase (LDH) levels were found in some exposed groups. The striatal DA content was significantly reduced in ATR, 0.5mgiAs/L, ATR+0.5mgiAs/L, and ATR+50mgiAs/L groups, in comparison to the control group. The number of mesencephalic tyrosine hydroxylase positive cells decreased in the ATR and ATR+0.5mgiAs/L groups compared to the control. In contrast, immunoreactivity to cytochrome oxidase was reduced compared to the control in all treated groups, except for the group treated with 0.5iAsmg alone. Our results indicate that ATR has deleterious effects on dopaminergic neurons and that the combination of ATR and iAs does not exacerbate these effects.

Sex-dependent neurochemical effects of environmental enrichment in the visual system.

Sex differences in the visual system have been reported in aspects of human vision, such as color perception, peripheral vision and even in the activation of the primary visual cortex. Similarly sex differences have been identified in the visual system of laboratory animals such as monkeys and rats. On the other hand, environmental enrichment (EE) has long been known to affect visual tissues. Taking into consideration the variation in the experimental approaches concerning EE and the sex differences in the visual system, we investigated in male and female rats the serotonergic and dopaminergic effects of EE in the retina and the visual cortex at different time points (i.e. P0-25, P0-P90 and P90-P150). Early EE in adulthood increased the serotonergic activity of the male visual cortex and the female retina (P0-P90). In addition early enrichment (P0-P90) increased dopaminergic activity in the female retina and in the visual cortex of both sexes. Late enrichment increased the serotonergic activity in the retina and visual cortex of both sexes (P90-P150), but increased the dopaminergic activity in the visual cortex only in male animals. In the present study we expose marked sex differences in the neurochemistry of visual tissues and we demonstrate for the first time that EE can in fact modify the serotonergic and dopaminergic neurotransmission in the retina and visual cortex. Overall, the present study underpins the sex-dependent neurochemical status of the visual system and provides insights into the different mechanisms underlying visual processing in the two sexes.

Enhanced potentiometry by metallic nanoparticles.

Measuring the oxidation-reduction potential (Eh) requires an interface that is not selective toward specific species but exchanges electrons with all redox couples in the solution. Sluggish electron transfer (ET) kinetics with the species will not reflect the "true" Eh of the solution. Here, we present a novel approach by which adsorbed metal nanoparticles (NPs) are used for enhancing ET exchange rates between redox species and electrode surface and therefore affect significantly the measurement of the open circuit potential (OCP) and cyclic voltammetry (CV). The OCP and CV of various organic and inorganic species such as l-dopa, dopac, iron(II), and iodide are measured by bare stainless steel and by stainless steel modified by either Pt or Au NPs. We study the effect of the surface coverage of the stainless steel surface by NPs on the electrochemical response. Moreover, the stainless steel electrode was modified simultaneously by Au and Pt nanoparticles. This improved concurrently the stainless steel response (CV and potentiometry) toward two different species; l-dopa, which shows fast electron transfer on Pt, and catechol, which exhibits fast electron transfer on Au. We believe that this approach could be a first step toward developing a superior electrode for measuring the "true" Eh of complex aquatic systems.

Analysis of glutamate, GABA, noradrenaline, dopamine, serotonin, and metabolites using microbore UHPLC with electrochemical detection.

The applicability of microbore ultrahigh performance liquid chromatography (UHPLC) with electrochemical detection for offline analysis of a number of well-known neurotransmitters in less than 10 µL microdialysis fractions is described. Two methods are presented for the analysis of monoamine or amino acid neurotransmitters, using the same UHPLC instrument. Speed of analysis of noradrenaline (NA), dopamine (DA), serotonin (5-HT), and the metabolites homovanillic acid (HVA), 5-hydroxyindole aceticacid (5-HIAA), and 3,4-dihydroxyphenylacetic acid (DOPAC) was predominated by the retention behavior of NA, the nonideal behavior of matrix components, and the loss in signal of 5-HT. This method was optimized to meet the requirements for detection sensitivity and minimizing the size of collected fractions, which determines temporal resolution in microdialysis. The amino acid neurotransmitters glutamate (Glu) and γ-aminobutyric acid (GABA) were analyzed after an automated derivatization procedure. Under optimized conditions, Glu was resolved from a number of early eluting system peaks, while the total runtime was decreased to 15 min by a 4-fold increase of the flow rate under UHPLC conditions. The detection limit for Glu and GABA was 10 nmol/L (15 fmol in 1.5 µL); the monoamine neurotransmitters had a detection limit between 32 and 83 pmol/L (0.16-0.42 fmol in 5 µL) in standard solutions. Using UHPLC, the analysis times varied from 15 min to less than 2 min depending on the complexity of the samples and the substances to be analyzed.