NMDA-mediated modulation of dopamine release is modified in rat prefrontal cortex and nucleus accumbens after chronic nicotine treatment

In this study, we investigate the effects of chronic administration of (−)nicotine on the function of the NMDA-mediated modulation of [³H]dopamine (DA) release in rat prefrontal cortex (PFC) and nucleus accumbens (NAc). In the PFC synaptosomes NMDA in a concentration-dependent manner evoked [³H]DA release in rats chronically treated with vehicle (14 days) with an EC₅₀ of 13.1 ± 2.0 μM. The NMDA-evoked overflow of the [³H]DA in PFC nerve endings of rats treated with (−)nicotine was significantly lower (−43%) than in vehicle treated rats. The EC₅₀ was 9.0 ± 1.4 μM. Exposure of NAc synaptosomes of rats treated with vehicle to NMDA produced an increase in [³H]DA overflow with an EC₅₀ of 14.5 ± 5.5 μM. This effect was significantly enhanced in chronically treated animals. The EC₅₀ was 10.5 ± 0.5 μM. The K⁺-evoked release of [³H]DA was not modified by the (−)nicotine administration. Both the changes of the NMDA-evoked [³H]DA overflow in the NAc and PFC disappeared after 14 days withdrawal. The results show that chronic (−)nicotine differentially affects the NMDA-mediated [³H]DA release in the PFC and NAc of the rat.     

HISTAMINE-SENSITIVE ADENYLATE CYCLASE IN HYPOTHALAMUS OF RAT BRAIN: H1 AND H2 RECEPTORS

Abstract— In in vitro experiments on rat hypothalamic homogenates the effects of biogenic amines such as histamine (HA), noradrenaline (NA), dopamine (DA), serotonin (5-HT) and drugs such as isoprenaline (ISP), 2-(2-pyridyl)ethylamine (H₂ stimulant—H_ls), 4-methyl-histamine (H₂ stimulant H_2s), mepyramine (H₁ antagonistp H_la), cimetidine (H₂ antagonist—H_2a) were tested on adenylate cyclase activity. HA possessed a powerful stimulating effect on hypothalamic adenylate cyclase activity, higher than that shown by the other substances.
The stimulating effect of HA was greatest in hypothalamic tissue from male rats, while tissue from females showed only a modest stimulation. H_2s, induced a greater stimulation of adenylate cyclase than H_ls. On the other hand, the H_2a inhibited HA stimulation to a greater extent than the H_la, H_la and H_2a, when used together, completely inhibited the HA stimulation. HA may have a neurotrans-mitter role in the hypothalamus, and in this area there appears to be a mixed population of H₁ and H₂ receptors, with a majority of H₂ receptors.     

Long-term treatment with fluoxetine induces desensitization of 5-HT4 receptor-dependent signalling and functionality in rat brain

The mode of action of antidepressant drugs may be related to mechanisms of monoamines receptor adaptation, including serotonin 5-HT₄ receptor subtypes. Here we investigated the effects of repeated treatment with the selective serotonin reuptake inhibitor fluoxetine for 21 days (5 and 10 mg/kg, p.o., once daily) on the sensitivity of 5-HT₄ receptors by using receptor autoradiography, adenylate cyclase assays and extracellular recording techniques in rat brain. Fluoxetine treatment decreased the density of 5-HT₄ receptor binding in the CA1 field of hippocampus as well as in several areas of the striatum over the doses of 5–10 mg/kg. In a similar way, we found a significant lower response to zacopride-stimulated adenylate cyclase activity in the fluoxetine 10 mg/kg/day treated group. Furthermore, post-synaptic 5-HT₄ receptor activity in hippocampus-measured as the excitatory action of zacopride in the pyramidal cells of CA1 evoked by Schaffer collateral stimulation was attenuated in rats treated with both doses of fluoxetine. Taken together, these results support the concept that a net decrease in the signalization pathway of 5-HT₄ receptors occurs after chronic selective serotonin reuptake inhibitor treatment: this effect may underlie the therapeutic efficacy of these drugs.     

Dopaminergic Regulation of Septohippocampal Cholinergic Neurons

Abstract: The extent to which acetylcholine (ACh) release in the hippocampus is regulated by dopaminergic mechanisms was assessed using in vivo microdialysis in freely moving rats. Systemic administration of the dopamine (DA) receptor agonist apomorphine (1.0 mg/kg) or the specific D₁ agonist CY 208–243 (1.0 mg/kg) increased microdialysate concentrations of ACh in the hippocampus. The D₂ receptor agonist quinpirole (0.5 mg/kg) produced a small but statistically significant decrease in hippocampal ACh release. d -Amphetamine (2.0 mg/kg) increased ACh release, an effect that was blocked by the D₁ receptor antagonist SCH 23390 (0.3 mg/kg) but not by the D₂ antagonist raclopride (1.0 mg/kg). These findings suggest that endogenous DA stimulates septo-hippocampal cholinergic neurons primarily via actions at D₁ receptors. In addition, these results are similar to previous findings regarding the dopaminergic regulation of cortical ACh release, and suggest that the anatomical continuum formed by basal forebrain cholinergic neurons that project to the cortex and hippocampus acts as a functional unit, at least with respect to its regulation by DA.     

Population differences in water-use efficiency of Eucalyptus microtheca seedlings under different watering regimes

Gas exchange, water-use efficiency (WUE), carbon isotope composition ( Δ ¹³C) and growth traits were compared among 5 populations of Eucalyptus microtheca F. Muell. Seedlings grown from seed collected across the natural distribution of the species were maintained under water-stressed and well-watered conditions. Gas exchange was measured in terms of net photosynthesis (A) and transpiration (E); WUE was measured in terms of instantaneous water-use efficiency (WUE_i) and transpiration efficiency (WUE_T); growth traits were measured in terms of total biomass (TB), root/shoot ratio (RS), and specific leaf area density (DEN). Significant differences in all traits were detected among the populations. Overall population variation was 1.68–2.50 and 1.48–2.26 μmol CO₂ uptake per mmol H₂O transpired (WUE_i), 1.97–3.04 and 1.64–2.36 g dry matter accumulation per kg water transpired (WUE_T), and Δ ¹³C was −28.81 to −26.75‰ and −30.56 to −30.04‰ under the water-stressed and well-watered conditions, respectively. In addition, WUE_i, WUE_T and Δ ¹³C were significantly correlated with A, E, RS, DEN and TB. The study indicated that measurement of WUE may be a useful trait for selecting genotypes with improved drought adaptation and biomass productivity under different environmental conditions.     

The roles of membrane estrogen receptor subtypes in modulating dopamine transporters in PC-12 cells

The effects of 17β-estradiol (E₂) on dopamine (DA) transport could explain gender and life-stage differences in the incidence of some neurological disorders. We tested the effects of E₂ at physiological concentrations on DA efflux in nerve growth factor-differentiated rat pheochromocytoma cells that express estrogen receptors (ER) α, ERβ, and G-protein coupled receptor 30 (GPR30), and DA transporter (DAT). DAT efflux was determined as the transporter-specific loss of ³H-DA from pre-loaded cells; a 9–15 min 10⁻⁹ M E₂ treatment caused maximal DA efflux. Such rapid estrogenic action suggests a non-genomic response, and an E₂-dendrimer conjugate (limited to non-nuclear actions) caused DA efflux within 5 min. Efflux dose–responses for E₂ were non-monotonic, also characteristic of non-genomic estrogenic actions. ERα siRNA knockdown abolished E₂-mediated DA efflux, while ERβ knockdown did not, and GPR30 knockdown increased E₂-mediated DA efflux (suggesting GPR30 is inhibitory). Use of ER-selective agonists/antagonists demonstrated that ERα is the predominant mediator of E₂-mediated DA efflux, with inhibitory contributions from GPR30 and ERβ. E₂ also caused trafficking of ERα to the plasma membrane, trafficking of ERβ away from the plasma membrane, and unchanged membrane GPR30 levels. Therefore, ERα is largely responsible for non-genomic estrogenic effects on DAT activity.     

Implications of water stress-induced changes in the levels of endogenous ascorbic acid and hydrogen peroxide in Vigna seedlings

Vigna cutjang Endl. cv. Pusa Barsati seedlings, subjected to increasing degrees of water stress (−0.5, −1.0, −1,5 MPa), produced an approximately proportional increase in glycolate oxidase activity, hydrogen peroxide (H₂O₂) and proline content but a decrease in catalase activity, ascorbic acid and protein content. Leaf water potential (leaf ψ) and relative water content (RWC) were also lowered with increasing stress. Pretreatment with l -cysteine and reduced glutathione (10-3 M) decreased glycolate oxidase activity, H₂O₂ content, ascorbic acid oxidase activity, proline content and also slightly improved the water status of leaves stressed (−1.0 MPa) for 2 days. Pretreatment of non-stressed seedlings with these antioxidants had little or no effect. These studies indicate that treatment with antioxidants makes the plant tolerant against water stress by modulating the endogenous levels of H₂O₂ and ascorbic acid in stressed tissue.     

Effects of the Dopamine D3 Antagonist PD 58491 and Its Interaction with the Dopamine D3 Agonist PD 128907 on Brain Dopamine Synthesis in Rat

Abstract: The dopamine (DA) D₃ receptor antagonist PD 58491 {3-[4-[1-[4-[2-[4-(3-diethylaminopropoxy)phenyl]-benzoimidazol-1-yl-butyl]-1 H -benzoimidazol-2-yl]-phenoxy]propyl]diethylamine} bound with high affinity and selectivity to recombinant human DA D₃ versus D_2L and D_4.2 receptors transfected into Chinese hamster ovary cells: K _i values of 19.5 n M versus 2,362 and >3,000 n M , respectively. In contrast, the putative DA D₃ receptor antagonist (+)-AJ76 displayed low affinity and selectivity for D₃ versus D_2L and D_4.2 receptors (91 n M vs. 253 and 193 n M , respectively). In vitro, PD 58491 (1 n M −1µ M ) exhibited D₃ receptor antagonist activity, reversing the quinpirole (10 n M )-induced stimulation of [³H]thymidine uptake in D₃ CHOpro-5 cells, but did not have any significant intrinsic activity by itself in this assay. PD 58491 did not decrease the γ-butyrolactone-induced increase in DA synthesis ( l -3,4-dihydroxyphenylalanine accumulation) in rat striatum, indicating that the compound possessed no in vivo DA D₂/D₃ receptor agonist action at DA autoreceptors. PD 58491 (3–30 mg/kg, i.p.) generally did not alter DA or serotonin synthesis in either the striatum or mesolimbic region of rat brain. The D₃-preferring agonist PD 128907 decreased DA synthesis in striatum and mesolimbic regions, and this effect was attenuated by pretreatment with PD 58491. These findings support the hypothesis that DA D₃ autoreceptors may in part modulate the synthesis and release of DA in striatum and mesolimbic regions.     

Soils contain two different activities for oxidation of hydrogen

Abstract Hydrogen oxidation rates were measured in a neutral compost soil and an acidic sandy loam at H₂ mixing ratios of 0.01 to 5000 ppmv. The kinetics were biphasic showing two different K _m values for H₂, one at about 10–40 nM dissolved H₂, the other at about 1.2–1.4 μM H₂. The low- K _m activity was less sensitive to chloroform fumigation than the high- K _m activity. If sterile soil was amended with Paracoccus denitrificans or a H₂-oxidizing strain isolated from compost soil, it exhibited only a high- K _m (0.7–0.9 μM) activity. It also failed to utilize H₂ mixing ratios below a threshold of 1.6–3.0 ppmv H₂ (160–300 mPa). A similar result was obtained when fresh soil samples were suspended in water, and H₂ oxidation was determined from the decrease of dissolved H₂. However, H₂ was again utilized to mixing ratios lower than 0.05 ppmv, if the supernatant of the soil suspension or the settled soil particles were dried onto sterile soil or purified quarz sand. Obviously, soils contain two different activities for oxidation of H₂: (1) a high- K _m, high-threshold activity which apparently is due to aerobic H₂-oxidizing bacteria, and (2) a low- K _m, low-threshold activity whose origin is unknown but presumably is due to soil enzymes.     

Protective effect of salicylates against hydrogen peroxide stress in yeast

Aims:  To investigate the effects of salicylates in Saccharomyces cerevisiae exposed to oxidative stress induced by hydrogen peroxide (H₂O₂).
Methods and Results:  Saccharomyces cerevisiae was cultured through to the postlogarithmic phase of growth. Stress was induced by the addition of 1·5 mmol l⁻¹ H₂O₂ for 1 h, while N-acetyl-l-cysteine (NAC) and glutathione (GSSG) were used as control agents that affect the redox balance. Sodium salicylate, at 0·01–10 mmol l⁻¹or acetylsalicylic acid, at 0·02–2·5 mmol l⁻¹ was administered at various times before hydrogen peroxide stress. Both agents conferred resistance to a subsequent hydrogen peroxide stress, similarly to the induction of the adaptive response observed upon pretreatment with NAC and GSSG. Sodium salicylate was more potent as a short-term, but not as a long-term pretreatment agent, compared to acetylsalicylic acid.
Conclusions:  Pharmacological pretreatment with salicylates resulted in dose related increases in cell survival, indicating the induction of the protective response in yeast.
Significance and Impact of the study:  The possible role of salicylates in the modulation of the hydrogen peroxide stress response in eukaryotic cells address questions on the effects of these commonly used therapeutic agents in a number of disorders exhibiting an oxidative stress component.     

Effects of 1-Methyl-4-Phenylpyridinium on Isolated Rat Brain Mitochondria: Evidence for a Primary Involvement of Energy Depletion

Abstract: The effects of 1-methyl-4-phenylpyridinium (MPP⁺) on the oxygen consumption, ATP production, H₂O₂ production, and mitochondrial NADH-CoQ₁ reductase (complex I) activity of isolated rat brain mitochondria were investigated. Using glutamate and malate as substrates, concentrations of 10–100 µ M MPP⁺ had no effect on state 4 (−ADP) respiration but decreased state 3 (+ADP) respiration and ATP production. Incubating mitochondria with ADP for 30 min after loading with varying concentrations of MPP⁺ produced a concentration-dependent decrease in H₂O₂ production. Incubation of mitochondria with ADP for 60 min after loading with 100 µ M MPP⁺ caused no loss of complex I activity after washing of MPP⁺ from the mitochondrial membranes. These data are consistent with MPP⁺ initially binding specifically to complex I and inhibiting both the flow of reducing equivalents and the production of H₂O₂ by the mitochondrial respiratory chain, without irreversibly damaging complex I. However, mitochondria incubated with H₂O₂ in the presence of Cu²⁺ ions showed decreased complex I activity. This study provides additional evidence that cellular damage initiated by MPP⁺ is due primarily to energy depletion caused by specific binding to complex I, any increased damage due to free radical production by mitochondria being a secondary effect.     

Effect of dilution on methanogenesis, hydrogen turnover and interspecies hydrogen transfer in anoxic paddy soil

Abstract Dilution of anoxic slurries of paddy soil resulted in a proportional decrease of the rates of total methanogenesis and the rate constants of H₂ turnover per gram soil. Dilution did not affect the fraction of H₂/CO₂-dependent methanogenesis which made up 22% of total CH₄ production. However, dilution resulted in a ten fold decrease of the H₂ steady state partial pressure from approximately 4 to 0.4 Pa indicating that H₂/CO₂-dependent methanogenesis was more or less independent of the H₂ pool. The rates of H₂ production calculated from the H₂ turnover rate constants and the H₂ steady state partial pressures accounted for only < 5% of H₂/CO₂-dependent methanogenesis in undiluted soil slurries and for even less after dilution. Upon dilution, the Gibbs free energy available for H₂/CO₂-dependent methanogenesis decreased from −28.4 to only −5.6 kJ per mol. The results indicate that methane was mainly produced from interspecies H₂ transfer within syntrophic bacterial associations and was not significantly affected by the outside H₂ pool.     

Bactericidal activity of catechin-copper (II) complexes on Escherichia coli ATCC11775 in the absence of hydrogen peroxide

Washed Escherichia coli ATCC11775 cells were killed by (–)-epigallocatechin (EGC) in the presence of a non- lethal concentration of Cu²⁺ (1 μmol l⁻¹) without additional H₂O₂, but not by (–)-epicatechin (EC). EGC alone (< 0·1 mmol l⁻¹) did not reduce the viability of the cells. The survival curve obtained in the presence of EGC and Cu²⁺ was similar to that obtained in the presence of (–)-adrenaline (EN) and Cu²⁺.     

Manipulation of fatty acid amide hydrolase functional activity alters sensitivity and dependence to ethanol

The aim of this study was to examine the role of fatty acid amide hydrolase (FAAH) on ethanol sensitivity, preference, and dependence. The deletion of FAAH gene or the inhibition of FAAH by carbamoyl-biphenyl-3-yl-cyclohexylcarbamate (URB597) (0.1 mg/kg) markedly increased the preference for ethanol. The study further reveals that URB597 specifically acts through FAAH and that cannabinoid-1 (CB₁) receptor is critical for N -arachidonoyl ethanolamide (AEA) mediated ethanol-reinforced behavior as revealed by lack of URB597 effect in both FAAH and CB₁−/− mice compared with vehicle-treated −/− mice. The FAAH −/− mice displayed a lower sensitivity to hypothermic and sedative effects to acute ethanol challenge. The FAAH −/− mice also exhibited a reduction in the severity of handling-induced convulsions following withdrawal from chronic ethanol exposure. The CB₁ receptor and proenkephalin gene expressions, and CB₁ receptor and μ-opioid (MO) receptor-mediated G-protein activation were found to be significantly lower in the caudate-putamen, nucleus accumbens core and shell of FAAH −/− than +/+ mice. Interestingly, the MO receptor-stimulated G-protein signaling was greater in the striatum of FAAH −/− than +/+ mice following voluntary ethanol consumption. These findings suggest that an elevation in the AEA content and its action on the limbic CB₁ receptor and MO receptor might contribute to ethanol-reinforced behavior. Treatment with drugs that decrease AEA tone might prove useful in reducing excessive ethanol consumption.     

A strict anaerobic extreme thermophilic hydrogen-producing culture enriched from digested household waste

Aims:  The aim of this study was to enrich, characterize and identify strict anaerobic extreme thermophilic hydrogen (H₂) producers from digested household solid wastes.
Methods and Results:  A strict anaerobic extreme thermophilic H₂ producing bacterial culture was enriched from a lab-scale digester treating household wastes at 70°C. The enriched mixed culture consisted of two rod-shaped bacterial members growing at an optimal temperature of 80°C and an optimal pH 8·1. The culture was able to utilize glucose, galactose, mannose, xylose, arabinose, maltose, sucrose, pyruvate and glycerol as carbon sources. Growth on glucose produced acetate, H₂ and carbon dioxide. Maximal H₂ production rate on glucose was 1·1 mmol l⁻¹ h⁻¹ with a maximum H₂ yield of 1·9 mole H₂ per mole glucose. 16S ribosomal DNA clone library analyses showed that the culture members were phylogenetically affiliated to the genera Bacillus and Clostridium. Relative abundance of the culture members, assessed by fluorescence in situ hybridization, were 87 ± 5% and 13 ± 5% for Bacillus and Clostridium , respectively.
Conclusions:  An extreme thermophilic, strict anaerobic, mixed microbial culture with H₂-producing potential was enriched from digested household wastes.
Significance and Impact of the Study:  This study provided a culture with a potential to be applied in reactor systems for extreme thermophilic H₂ production from complex organic wastes.     

Activation of Endogenous Antioxidant Defenses in Neuronal Cells Prevents Free Radical-Mediated Damage

Abstract: Dopamine (DA) is oxidized to the neurotoxic prooxidant species H₂O₂, OH^•, and DA quinones. We tested whether dimethyl fumarate (DMF), an electrophile shown to induce a pleiotropic antioxidant response in nonneuronal cells, could reduce the toxicity of DA metabolites in neural cells. Treatment of the N18-RE-105 neuroblastoma-retina hybridoma cell line with 30–150 µ M dopamine led to cell death within 24 h, which increased steeply with dose, decreased with higher plating density, and was blocked by the H₂O₂-metabolizing enzyme catalase. Pretreatment with DMF (30 µ M , 24 h) significantly attenuated DA and H₂O₂ toxicity (40–60%) but not that caused by the calcium ionophore ionomycin. DMF treatment also elevated total intracellular GSH and increased activities of the antioxidant enzymes quinone reductase (QR), glutathione S -transferase (GST), glutathione reductase, and the pentose phosphate enzyme glucose-6-phosphate dehydrogenase. To assess the protective efficacy of QR and GST, a stable cell line was constructed in which these enzymes were overexpressed. Cell death in the overexpressing line was not significantly different from that in a cell line expressing normal QR and GST activities, indicating that these two enzymes alone are insufficient for protection against DA toxicity. Although the relative importance of a single antioxidant enzyme such as QR or GST may be small, antioxidant inducers such as DMF may prove valuable as agents that elicit a broad-spectrum neuroprotective response.     

Serotonergic Regulation of Acetylcholine Release in Rat Frontal Cortex

Abstract: The extent to which serotonin regulates the activity of cortically projecting cholinergic neurons was studied using in vivo microdialysis to monitor interstitial concentrations of acetylcholine in the frontal cortex of freely moving rats. Systemic administration of the serotonin release-inducing agent fenfluramine (3 or 10 mg/kg, i.p.) increased acetylcholine release by 110–130%. The fenfluramine-induced increase in acetylcholine release was significantly attenuated by pretreatment with the selective serotonin uptake inhibitor fluoxetine (10 mg/kg, i.p.). Pretreatment with the selective dopamine D₁ receptor antagonist SCH-23390 (0.3 mg/kg, s.c.) failed to prevent the fenfluramine-induced increase in acetylcholine release. In contrast, the serotonin 5-HT_2A receptor antagonist ketanserin (5 mg/kg, i.p.) blocked fenfluramine-induced increases in acetylcholine release. In contrast to previous studies that have concluded that serotonin has inhibitory actions on cortical acetylcholine release, the present results indicate that fenfluramine increases cortical acetylcholine release in vivo by its ability to enhance serotonin transmission and that serotonin produces these effects at least in part via actions at serotonin 5-HT_2A receptors.     

Vertical distribution of methane metabolism in microbial mats of the Great Sippewissett Salt Marsh

Methane metabolism was investigated with respect to depth in intertidal microbial mats of the Great Sippewissett Salt Marsh, Massachusetts. Although sulfate-reducing organisms dominate anaerobic carbon consumption in marine microbial mats, methanogens persist and their activity varies vertically and temporally in the mat system. In the Sippewissett mats, potential methane production for all mat layers was higher in the spring (17.2 ± 4.5 nmol CH₄ cm⁻² day⁻¹) than in the fall (3.0 ± 1.1 nmol CH₄ cm⁻² day⁻¹) and maximal rates were consistently observed in proximity to the chemocline (5–10 mm depth). The methane flux from the mat surface did not vary appreciably over time due to the ability of methanotrophic activity to limit net methane production. Evidence indicates that both aerobic and anaerobic oxidation of methane occurs in this system. The importance of H₂ as a substrate for methanogenesis appeared to be the greatest at the mat surface (0–10 mm), and the proportion of methylotrophic methanogens generally increased with depth. These results suggest that both non-equilibrium H₂ dynamics and the use of non-competitive substrates permit coexistence of methanogens and sulfate-reducing organisms in the mat system.