Towards a Reconceptualization of Striatal Interactions Between Glutamatergic and Dopaminergic Neurotransmission and Their Contribution to the Production of Movements

According to the current model of the basal ganglia organization, simultaneous activation of the striato-nigral direct pathway by glutamatergic and dopaminergic neurotransmission should lead to a synergistic facilitatory action on locomotor activity, while in contrast activation of the indirect pathway by these two neurotransmittions should lead to antagonistic effects on locomotor activity. Based on published data, as a break with the current thinking, we propose a reconceptualization of functional interactions between dopaminergic and glutamatergic neurotransmission. In this model, dopaminergic neurotransmission is seen as a motor pacemaker responsible for the basal and primary activation of striatal output neurons and glutamate as a driver providing a multiple combination of tonic, phasic, facilitatory and inhibitory influxes resulting from the processing of environmental, emotional and mnesic stimuli. Thus, in the model, glutamate-coded inputs would allow tuning the intrinsic motor-activating properties of dopamine to adjust the production of locomotor activity into goal-oriented movements.Key Words: Dopamine-glutamate striatal interactions, locomotor activity, goal-oriented movements, nucleus accumbens, striatum.     

Lack of Maternal and Developmental Toxicity in Mice Given High Doses of Aluminium Hydroxide and Ascorbic Acid During Gestation

The present study was conducted to assess if the concurrent ingestion of high doses of aluminium hydroxide and ascorbic acid might result in maternal and developmental toxicity in mice. Three groups of pregnant Swiss mice were given by gavage daily doses of aluminium hydroxide (300 mg/kg), ascorbic acid (85 mg/kg), or aluminium hydroxide (300 mg/kg) concurrent with ascorbic acid 85 (mg/kg) on gestational days 6-15. A fourth group of animals received distilled water and served as control group. Dams were killed on gestation day 18 and foetuses were examined for external, internal, and skeletal abnormalities. The reproductive data did not show embryotoxic or foetotoxic effects in any group. No gross, internal, or skeletal malformations or variations related to the different treatments were found. There were no significant differences between control and treated groups on the aluminium levels in maternal liver and bone as well as in whole body foetuses, whereas aluminium concentrations were significantly higher in placenta and kidney of dams receiving aluminium hydroxide and aluminium hydroxide plus ascorbic acid than in those from the control group. Although in this study aluminium hydroxide was given at doses higher than those usually consumed by pregnant women, no signs of maternal or developmental toxicity were observed when the compound was given alone or concurrently with high doses of ascorbic acid.     

Degradation of Patulin in Pear Juice and Apple Juice by Ascorbic Acid and the Combination of Ascorbic Acid and Ferrous Iron

Patulin (PAT) is a toxic secondary metabolite produced by certain species of Penicillium sp. and Aspergillus sp. on apples and pears. In this study, we investigated the effects of ascorbic acid and the combination of ascorbic acid and ferrous iron on degradation of PAT in 100% pure pear juice and apple juice using high-performance liquid chromatography UV detector (HPLC-UVD). The addition of 2 different levels of ascorbic acid (143 or 286 μg/mL) into pear juice or apple juice containing 0.08 or 0.4 μg/mL of PAT showed 87.7–100% and 67.3–68.7% of PAT degradation rates, respectively, after 24 h incubation at 25 °C. Moreover, the addition of both ascorbic acid (143 or 286 μg/mL) and ferrous iron (0.033 or 0.11 μmol/mL) into pear juice or apple juice containing the same level of PAT exhibited higher PAT degradation rates (100 and 75–94%, respectively) than the addition of only ascorbic acid after 24 h incubation at 25 °C. Our data demonstrated that ascorbic acid plus ferrous iron as well as ascorbic acid were highly effective on degradation of PAT in pear juice and apple juice and that addition of both ascorbic acid and ferrous iron produced higher PAT degradation rates than addition of only ascorbic acid.     

Chemical Pathways of Peptide Degradation. V. Ascorbic Acid Promotes Rather than Inhibits the Oxidation of Methionine to Methionine Sulfoxide in Small Model Peptides

The effect of primary structure and external conditions on the oxidation of methionine to methionine sulfoxide by the ascorbate/Fe³⁺ system was studied in small model peptides. Degradation kinetics and yield of sulfoxide formation were dependent on the concentration of ascorbate and H⁺, with a maximum rate observed at pH 6–7. Phosphate buffer significantly accelerated the peptide degradation compared to Tris, HEPES, and MOPS buffers; however, the formation of sulfoxide was low. The oxidation could not be inhibited by the addition of EDTA. Other side products besides sulfoxide were observed, indicating the existence of various other pathways. The influence of methionine location at the C terminus, at the N terminus, and in the middle of the sequence was investigated. The presence of histidine in the sequence markedly increased the degradation rate as well as the sulfoxide production. The histidine catalysis of methionine oxidation occurred intramolecularly with a maximum enhancement of the oxidation rate and sulfoxide production when one residue was placed between the histidine and the methionine residue.     

Conjugation to Ascorbic Acid Enhances Brain Availability of Losartan Carboxylic Acid and Protects Against Parkinsonism in Rats

Identification of renin-angiotensin system in the interplay of hypertension and neurodegeneration has paved the way for the repurposing of antihypertensive drugs against Parkinsonism. Losartan carboxylic acid (LCA), the potent AT1 blocker metabolite of losartan, suffers from poor bioavailability and brain access. Since ascorbate transporters have earlier shown enough flexibility as carriers, we have conjugated losartan carboxylic acid to ascorbic acid with the aim of achieving higher oral/brain availability. Ester of LCA and ascorbic acid (FED) was developed keeping in view the substrate specificity of ascorbate transporters. Oral/brain bioavailability was assessed using in vivo pharmacokinetic model. Effect on central nervous system (CNS) and protection against Parkinsonism was evaluated using in vivo models. FED enhanced bioavailability of LCA. The higher brain availability of LCA enabled CNS protection as evident from the increase in locomotor activity, improved motor coordination, and protection against drug-induced catatonia. In conclusion, FED offers an approach to repurpose LCA against Parkinsonism. This can encourage further investigation to simultaneously address hypertension and neurodegeneration.     

Development of Tolerance in Mice to the Sedative Effects of the Neuroactive Steroid Minaxolone Following Chronic Exposure

Minaxolone is a potent ligand for the neurosteroid binding site of the GABA_A receptor. In radioligand binding studies to rat brain membranes, minaxolone caused a 69% increase in [³H]muscimol binding and a 25% increase in [³H]flunitrazepam binding and inhibited the binding of [³H]TBOB with an IC₅₀ of 1 μM. In mice, minaxolone (100 mg/kg, orally) had marked sedative effects as indicated by a reduction in locomotor activity. Chronic dosing with minaxolone (100 mg/kg, orally, once daily for 7 days) resulted in a loss of sedative response to an acute dose of the drug, indicating the development of tolerance. Chronic dosing with temazepam (10 mg/kg, orally, once daily for 7 days) resulted in the development of tolerance to an acute dose of temazepam; however, the two drugs did not appear to be cross-tolerant, indicating that they may have a different mechanism of action at the level of the GABA_A receptor.     

Adaptation to Ozone in Rats and Its Association with Ascorbic Acid in the Lung

Ozone (O₃) adaptation is a well-known, but poorly understoodphenomenon that has been demonstrated in humans and laboratoryanimals. This study examined pulmonary function and bronchoalveolarlavage fluid (BALF) parameters in O₃-adapted F-344 rats to explorepossible mechanisms of adaptation. Of particular interest wasascorbic acid (AA), an antioxidant reported to be protectiveagainst O₃ injury and found to be increased in O₃-adapted rats.Adaptation was induced by exposure to 0.25 ppm O₃, 12 hr/dayfor 6 or 14 weeks and evaluated with a challenge test, one thatreexposed rats to 1.0 ppm O₃ and measured attenuation in theO₃ effect on frequency of breathing. Pulmonary function wasassessed 1 day postexposure and adaptation and BALF were evaluated1, 3, and 7 days postexposure. Results showed that forced vitalcapacity increased over time but decreased due to exposure andthat the 14-week, O₃-exposed rats had an increase in forcedexpiratory flow rate. All of the O₃-exposed rats that were testeddemonstrated adaptation on Postexposure Days 1, 3, and 7, butit was diminished on Day 7. Adaptation was also more pronouncedin rats exposed for 14 weeks. Except for AA, BALF levels oftotal protein, potassium, lysozyme, uric acid, and -tocopherolwere unaffected by O₃ exposure. Lactic acid dehydrogenase, alkalinephosphatase, glucose-6-phosphate dehydrogenase, and total glutathionewere also assayed but were always below detectable limits. Ascorbicacid concentrations were elevated on Days 1, 3, and 7, showingpostexposure patterns similar to those found for adaptation.Significant correlation was found between AA concentration andthe magnitude of adaptation (r = 0.91, p < 0.002). We concludethat AA may play an important role in mechanisms associatedWith O₃ adaptation in rats.     

Adrenergic and Dopaminergic Modulation of Immunity in Multiple Sclerosis: Teaching Old Drugs New Tricks?

Multiple sclerosis (MS) is an autoimmune disorder of the CNS characterized by inflammation, demyelination and axonal loss. Classical evidence in experimental allergic encephalomyelitis, the animal model of MS, support the relevance of sympatoadrenergic as well as of dopaminergic mechanisms. In MS patients, dysregulation of adrenergic and dopaminergic pathways contribute to the disease in immune system cells as well as in glial cells. Available evidence is summarized and discussed also in the light of the novel role of dopamine, noradrenaline and adrenaline as transmitters in immune cells, providing a conceptual frame to exploit the potential of several dopaminergic and adrenergic agents, already in clinical use for non-immune indications and with a usually favourable risk-benefit profile, as add-on drugs to conventional immunomodulating therapies in MS.     

Role of Ascorbic Acid in Stratum Corneum Lipid Models Exposed to UV Irradiation

Purpose. The effects of ascorbic acid on Stratum corneum lipid models following ultraviolet irradiation were studied adding iron ions as transition metal catalysts. Methods. Lipid peroxidation was quantified by the thiobarbituric acid assay. The qualitative changes were studied on a molecular level by mass spectrometry. To elucidate the nature of free radical involvement we carried out electron paramagnetic resonance studies. The influence of ascorbic acid on the concentration of hydroxyl radicals was examined using the spin trapping technique. Moreover, we checked the vitamin's ability to react with stable radicals. Results. Ascorbic acid was found to have prooxidative effects in all lipid systems in a concentration dependent manner. The degradation products of ascorbic acid after its prooxidative action were detected. The concentration of the hydroxyl radicals in the Fenton assay was decreased by ascorbic acid. The quantification assay of 2,2-diphenyl-1-picrylhydrazyl hydrate showed reduced concentration levels of the stable radical caused by ascorbic acid. Conclusions. Considering human skin and its constant exposure to UV light and oxygen, an increased pool of iron ions in irradiated skin and the depletion of co-antioxidants, the administration of ascorbic acid in cosmetic formulations or in sunscreens could unfold adverse effects among the Stratum corneum lipids.     

Modulation of Immune Function in Mice Exposed to 0.8 PPM Ozone

Abstract

Ozone has been shown to both enhance and suppress local and systemic immune responses, depending upon the experimental conditions and the immune parameters under investigation. To gain insight into this apparent immunomodulation, mice were continuously exposed to 0.8 ppm of 0, for 1, 3, 7, and 74 days prior to assessment of their immune function. Lymphocytes from the mediastinal lymph nodes (MLN) and the spleen were examined in order to distinguish local and systemic immune compartments, respectively. Ozone exposure resulted in an initial decrease in MLN cell numbers and spleenlbody weight ratios, which then returned to normal and increased above control values following more prolonged exposure. SirnilarlF the lymphocyte responses to mitogen and the splenocyte tumoricidal activity were initially decreased by exposure to 0.8 ppm ozone but later returned to control levels. To evaluate the effect of O₃ on the specific immune response, mice were immunized parenterally with ovalbumin (OA) followed by an aerosol boost with OA at 14 days. A week later the animals were assessed for specific immune responses. Exposure to 0.8 ppm O₃ for 14 days prior to assay resulted in an increased MLN lymphocyte response to specific antigen, while the splenic (systemic) response was decreased. Specific IgG and IgA antibody titers to the antigen in the bronchoalveolar lavage fluid were significantly suppressed following 7 and 14 days of exposure to O₃ but were unaltered in serum. These data, in part, explain the divergent responses observed by others. Ozone exposure initially suppresses local and systemic aspects of innate immunity, which recover or adapt over more prolonged exposure. Specific immune responses, however, can be either suppressed or enhanced following long-term exposure, depending upon the anatomical site and biological end point measured.     

Reaction of 5-Aminosalicylic Acid with Peroxyl Radicals: Protection and Recovery by Ascorbic Acid and Amino Acids

Purpose The aims of the study are to analyze the interaction between 5-aminosalicylic acid (5-ASA) and peroxyl radicals and to evaluate the effect of some endogenous compounds such as ascorbic acid and amino acids on the oxidation of 5-ASA induced by 2,2′-azo-bis(2-amidinopropane) dihydrochloride. Methods The consumption and/or the recovery of 5-ASA (7.6 μM) exposed to a peroxyl radical source [2,2′-azo-bis(2-amidinopropane)] was followed by techniques such as spectrofluorescence, high-performance liquid chromatography, and differential pulse voltammetry. Results 5-Aminosalicylic acid was found to readily react with peroxyl radicals at micromolar concentrations and to protect c-Phycocyanin in a very similar fashion to that shown by Trolox. Exposure of 5-ASA to peroxyl radicals led to its oxidation into the corresponding quinone-imine. Disappearance of 5-ASA was prevented by tryptophan, cysteine, glutathione, and ascorbic acid. Furthermore, some of these compounds induced the partial (cysteine and glutathione) or total (ascorbic acid) recovery of 5-ASA when added after its almost total consumption. Conclusions 5-Aminosalicylic acid is a very efficient peroxyl radical scavenger. The 5-ASA oxidation by peroxyl radicals was prevented by ascorbic acid, cysteine, and glutathione. In addition, 5-ASA can be regenerated by these endogenous compounds, which would be a valuable mechanism to preserve 5-ASA in tissues undergoing oxidative stress conditions.     

Tolerance to haloperidol-induced increases in dopamine metabolites: fact or artifact?

Haloperidol increased 3,4-dihydroxyphenylacetic acid and homovanillic acid concentrations in the striatum, nucleus accumbens and olfactory tubercle of both drug-naive rats and rats pretreated with haloperidol (10 injections). The increases in metabolite concentrations were greater in all brain regions of the naive rats, suggesting that haloperidol pretreatment resulted in a decreased responsiveness to the drug (tolerance). However, subchronic haloperidol injections also resulted in decreased basal metabolite concentrations in rats killed 48 h after the last injection. While the response of drug-experienced rats to haloperidol was attenuated relative to that of drug-naive rats, this difference could be accounted for entirely by the decreased basal metabolite concentrations that occur after repeated haloperidol injections.     

Repeated Independent Exposures to Domoic Acid Do Not Enhance Symptomatic Toxicity in Outbred or Seizure-Sensitive Inbred Mice

Domoic acid (DA) is an environmental neurotoxin to humans. Thiswork examines whether repeated exposure to subsymptomatic orsymptomatic nonlethal doses of domoic acid leads to enhancedsymptomatic toxicity in ICR outbred and DBA inbred strains oflaboratory mice. A multiple independent exposure paradigm wasdesigned in which doses were administered intraperito neallyevery other day for 7 days to achieve four separate exposuresto domoic acid. We first examined the effect of repeated exposureon serum clearance of domoic acid. Serum domoic acid levelsdid not differ following a single or repeated exposure. We nextexamined the effect of repeated exposure on symptomatic toxicity.The mean toxicity scores did not show a significant differencebetween single and repeated exposures of either subsymptomatic(0.5 mg/ kg) or symptomatic sublethal (2.0 mg/kg) doses of domoicacid. We then examined the effects of repeated domoic acid exposureon a second strain of mouse. DBA mice were chosen based upontheir sensitivity to kainic acid-induced seizures; however,the ICR mice were more sensitive to low-dose domoic acid toxicity,particularly in terms of onset and duration of stereotypic scratchingbehavior. Our results indicate that both strains of mice havecomparable concentration-dependent toxic responses to domoicacid; however, differences exist in the magnitude of the responseand in specific symptoms. The mean toxicity scores did not showa significant difference when a single exposure (1.0 and 2.0mg/kg domoic acid) and repeated exposure of the same dose werecom pared in the DBA mice. This study provides no evidence thatshort-term repeated exposure to domoic acid in laboratory micealters domoic acid clearance from the serum, or leads to a moresensitive or a greater neurotoxic response.     

Effects of Subchronic Exposures to Concentrated Ambient Particles: VII. Degeneration of Dopaminergic Neurons in Apo E−/− Mice

Abstract

This study reports that subchronic exposure of Tuxedo, NY concentrated ambient particulates (CAPs) produces neuropathological damage in the brains of Apo E-deficient mice (Apo E^?/?). These genetically modified mice are characterized by elevated levels of oxidative stress (OS) in the brain. Microscopic examination of coronal sections of the brain, immunocytochemically stained for dopamineric neurons, indicated that neurons from the substantia nigral nucleus compacta were significantly reduced by 29% in CAPs-exposed Apo E^?/? mice relative to air-exposed Apo E^?/? controls. In addition, statistically significant increases (p < .05) in immunocytochemically stained astrocytes were noted. The dopaminergic neurons of the nucleus compacta are specifically targeted in Parkinson's disease. The present study expands the systems affected by particulate matter to include the brain, and supports an environmental role for the development of neurodegeneration in OS-susceptible individuals.     

Development of Acute Tolerance to Bumetanide: Bolus Injection Studies

Bumetanide was administered intravenously to four mongrel dogs, in a random crossover fashion, at doses of 0.05 mg/kg (I), 0.15 mg/kg (II), and 0.5 mg/kg (III) where urinary losses were replaced with lactated Ringer's solution at 1.5 ml/min (hydropenic conditions) or at a dose of 0.5 mg/kg (IV) where urinary losses were replaced with lactated Ringer's solution isovolumetrically (euvolemic conditions). Serial plasma and urine samples were assayed for bumetanide by high-performance liquid chromatography (HPLC) and for sodium by flame photometry. There were no significant differences in the pharmacokinetic parameters of bumetanide among Treatments I–IV. The dynamic parameters E _max (maximum effect attributable to the drug) and s (slope factor) were not different between treatments. However, a consistent, demonstrable increase in ER₅₀ (urinary excretion rate of drug producing 50% of E _max) was observed among Treatments I (2.34 µg/min), II (3.92 µg/min), and III (6.54 µg/min); also, a significant decrease in ER₅₀ was observed between Treatment III (6.54 µg/min) and Treatment IV (2.66 µg/min). These results show that hydration status has a marked effect on natriuretic and diuretic response and that tolerance can rapidly develop within a single intravenous dose of bumetanide.     

Development of Acute Tolerance to Bumetanide: Constant-Rate Infusion Studies

Bumetanide was administered intravenously to four mongrel dogs as a bolus of 8.7 µg/kg, immediately followed by a constant-rate infusion of 0.35 µg/min/kg at 0.036 ml/min. Treatment A consisted of a 90-min equilibration period and first hour (Phase I) of study in which animals were maintained under euvolemic conditions. During the subsequent 3 hr of Treatment A (Phase II), animals were maintained under hydropenic conditions. These experiments were then repeated 1 week later (Treatment B) with the temporal aspects of hydration reversed (Phase III, hydropenia; Phase IV, euvolemia). Serial plasma and urine samples were assayed for bumetanide by high-performance liquid chromatography (HPLC) and for sodium by flame photometry. The bumetanide excretion rate was not significantly different during the 4 hr of Treatment A, although minor differences were observed between Phase III and Phase IV of Treatment B. The sodium excretion rate showed significant differences between euvolemic and hydropenic conditions of both treatments. A two- to threefold difference in the sodium excretion rate persisted even when slight differences (<20%) in bumetanide excretion rates were taken into account. These results demonstrate that an acute tolerance does develop to constant-rate infusions of bumetanide when inadequate fluid and electrolyte replacement occurs and that this tolerance can be reversed by rehydration.     

Development and Validation of a HPTLC Method for Simultaneous Estimation of L-Glutamic Acid and γ-Aminobutyric Acid in Mice Brain

A new robust, simple and economic high performance thin layer chromatographic method was developed for simultaneous estimation of L-glutamic acid and γ-amino butyric acid in brain homogenate. The high performance thin layer chromatographic separation of these amino acid was achieved using n-butanol:glacial acetic acid:water (22:3:5 v/v/v) as mobile phase and ninhydrin as a derivatising agent. Quantitation of the method was achieved by densitometric method at 550 nm over the concentration range of 10-100 ng/spot. This method showed good separation of amino acids in the brain homogenate with R_f value of L-glutamic acid and γ-amino butyric acid as 21.67±0.58 and 33.67±0.58, respectively. The limit of detection and limit of quantification for L-glutamic acid was found to be 10 and 20 ng and for γ-amino butyric acid it was 4 and 10 ng, respectively. The method was also validated in terms of accuracy, precision and repeatability. The developed method was found to be precise and accurate with good reproducibility and shows promising applicability for studying pathological status of disease and therapeutic significance of drug treatment.     

Chronic Adolescent Exposure to Delta-9-Tetrahydrocannabinol in COMT Mutant Mice: Impact on Indices of Dopaminergic,Endocannabinoid and GABAergic Pathways

Cannabis use confers a two-fold increase in risk for psychosis, with adolescent use conferring an even greater risk. A high-low activity polymorphism in catechol-O-methyltransferase (COMT), a gene encoding the COMT enzyme involved in dopamine clearance in the brain, may interact with adolescent cannabis exposure to increase risk for schizophrenia. The impact of such an interaction on central neurotransmitter pathways implicated in schizophrenia is unknown. Male mice with knockout of the COMT gene were treated chronically with delta-9-tetrahydrocannabinol (THC) during adolescence (postnatal day 32–52). We measured the size and density of GABAergic cells and the protein expression of cannabinoid receptor 1 (CB1R) in the prefrontal cortex (PFC) and hippocampus (HPC) in knockout mice relative to heterozygous mutants and wild-type controls. Size and density of dopaminergic neurons was also assessed in the ventral tegmental area (VTA) across the genotypes. COMT genotype × THC treatment interactions were observed for: (1) dopaminergic cell size in the VTA, (2) CB1R protein expression in the HPC, and (3) parvalbumin (PV) cell size in the PFC. No effects of adolescent THC treatment were observed for PV and dopaminergic cell density across the COMT genotypes. COMT genotype modulates the effects of chronic THC administration during adolescence on indices of neurotransmitter function in the brain. These findings illuminate how COMT deletion and adolescent cannabis use can interact to modulate the function of neurotransmitters systems implicated in schizophrenia.     

Actinomycin D and Development of Tolerance to Morphine Analgesia in Rats

Abstract: Adult male (Wistar) rats were given an analgesic dose of morphine (10 mg/kg subcutaneously) with or without actinomycin D (10 μg/kg intraperitoneally) daily over a period of 3 or 4 days. The degree of analgesia was measured on the hot plate after each injection or after the last injection only. A high degree of tolerance to morphine analgesia was observed after only one or two doses of morphine whether actinomycin D was given or not. Tolerance developed at about the same rate whether the animals were tested each day on the hot plate or on the last day only. Actinomycin D had no analgesic action of its own but it reduced the analgesic action of morphine in animals tested on the first day.