The noble gas xenon provides protection and trophic stimulation to midbrain dopamine neurons.

Despite its low chemical reactivity, the noble gas xenon possesses a remarkable spectrum of biological effects. In particular, xenon is a strong neuroprotectant in preclinical models of hypoxic-ischemic brain injury. In this study, we wished to determine whether xenon retained its neuroprotective potential in experimental settings that model the progressive loss of midbrain dopamine (DA) neurons in Parkinson's disease. Using rat midbrain cultures, we established that xenon was partially protective for DA neurons through either direct or indirect effects on these neurons. So, when DA neurons were exposed to l-trans-pyrrolidine-2,4-dicarboxylic acid so as to increase ambient glutamate levels and generate slow and sustained excitotoxicity, the effect of xenon on DA neurons was direct. The vitamin E analog Trolox also partially rescued DA neurons in this setting and enhanced neuroprotection by xenon. However, in the situation where DA cell death was spontaneous, the protection of DA neurons by xenon appeared indirect as it occurred through the repression of a mechanism mediated by proliferating glial cells, presumably astrocytes and their precursor cells. Xenon also exerted trophic effects for DA neurons in this paradigm. The effects of xenon were mimicked and improved by the N-methyl-d-aspartate glutamate receptor antagonist memantine and xenon itself appeared to work by antagonizing N-methyl-d-aspartate receptors. Note that another noble gas argon could not reproduce xenon effects. Overall, present data indicate that xenon can provide protection and trophic support to DA neurons that are vulnerable in Parkinson's disease. This suggests that xenon might have some therapeutic value for this disorder.

Metabolomics-Based Screening of the Malaria Box Reveals both Novel and Established Mechanisms of Action.

High-throughput phenotypic screening of chemical libraries has resulted in the identification of thousands of compounds with potent antimalarial activity, although in most cases, the mechanism(s) of action of these compounds remains unknown. Here we have investigated the mode of action of 90 antimalarial compounds derived from the Malaria Box collection using high-coverage, untargeted metabolomics analysis. Approximately half of the tested compounds induced significant metabolic perturbations in in vitro cultures of Plasmodium falciparum In most cases, the metabolic profiles were highly correlated with known antimalarials, in particular artemisinin, the 4-aminoquinolines, or atovaquone. Select Malaria Box compounds also induced changes in intermediates in essential metabolic pathways, such as isoprenoid biosynthesis (i.e., 2-C-methyl-d-erythritol 2,4-cyclodiphosphate) and linolenic acid metabolism (i.e., traumatic acid). This study provides a comprehensive database of the metabolic perturbations induced by chemically diverse inhibitors and highlights the utility of metabolomics for triaging new lead compounds and defining specific modes of action, which will assist with the development and optimization of new antimalarial drugs.

Prefrontal cortical kappa-opioid receptor modulation of local neurotransmission and conditioned place aversion.

Kappa-opioid receptors (KORs) are important for motivation and other medial prefrontal cortex (mPFC)-dependent behaviors. Although KORs are present in the mPFC, their role in regulating transmission in this brain region and their contribution to KOR-mediated aversion are not known. Using in vivo microdialysis in rats and mice, we demonstrate that intra-mPFC administration of the selective KOR agonist U69,593 decreased local dopamine (DA) overflow, while reverse dialysis of the KOR antagonist nor-Binaltorphimine (nor-BNI) enhanced mPFC DA overflow. Extracellular amino-acid levels were also affected by KORs, as U69,593 reduced glutamate and GABA levels driven by the glutamate reuptake blocker, l-trans-pyrrolidine-2,4-dicarboxylate. Whole-cell recordings from mPFC layer V pyramidal neurons revealed that U69,593 decreased the frequency, but not amplitude, of glutamatergic mini EPSPs. To determine whether KOR regulation of mPFC DA overflow was mediated by KOR on DA terminals, we utilized a Cre recombinase-driven mouse line lacking KOR in DA neurons. In these mice, basal DA release or uptake was unaltered relative to controls, but attenuation of mPFC DA overflow by local U69,593 was not observed, indicating KOR acts directly on mPFC DA terminals to locally inhibit DA levels. Conditioning procedures were then used to determine whether mPFC KOR signaling was necessary for KOR-mediated aversion. U69,593-mediated conditioned place aversion was blocked by intra-mPFC nor-BNI microinjection. These findings demonstrate that mPFC KORs negatively regulate DA and amino-acid neurotransmission, and are necessary for KOR-mediated aversion.

D,L-cis-2,3-Pyrrolidine dicarboxylate alters [3H]-L-glutamate binding and induces convulsions in mice.

This study investigated whether D,L-cis-2,3-Pyrrolidine dicarboxylate (D,L-cis-2,3-PDC), a new glutamate analogue, alters glutamate binding to cerebral plasma membranes and whether N-methyl-D-aspartate (NMDA) receptors are involved in the convulsant effect of this compound. D,L-cis-2,3-PDC reduced sodium-independent [3H]-L-glutamate binding to lysed membrane preparations from adult rat cortex and had no effect on sodium-dependent glutamate binding. Intracerebroventricular administration of D,L-cis-2,3-PDC (7.5-25 nmol/5 microl) induced generalized tonic-clonic convulsions in mice in a dose-dependent manner. The coadministration of MK-801 (7 nmol/2.5 microl), with D,L-cis-2,3-PDC (16.5 nmol/2.5 microl), fully protected the animals against D,L-cis-2,3-PDC-induced convulsions, while the coadministration of DNQX (10 nmol/2.5 microl) increased the latency to convulsions but did not alter the percentage of animals that had convulsions. These results suggest that D,L-cis-2,3-PDC-induced effects are mediated predominantly by NMDA receptors.

Oxidative neuronal death caused by glutamate uptake inhibition in cultured hippocampal neurons.

Glutamate transporters are coupled with cystine/glutamate antiporters to supply cystine as a component of glutathione, an important antioxidant. We sought evidence that L-trans-pyrrolidine-2,4-dicarboxylate (PDC) enhances glutamate-induced neuronal damage not only via the N-methyl-D-aspartate (NMDA) receptor mediated pathway, but also through induction of oxidative stress. Cultured hippocampal cells were exposed to glutamate (100 microM) for 5 min, washed and incubated for 18 hr with PDC (200 microM). PDC, increasing the neuronal death to 147% of that induced by glutamate alone, depleted glutathione in the culture, and produced dichloro-dihydro-fluorescein-diacetate-positive reactive oxygen species in neurons. N-acetylcysteine (2 mM) not only reduced PDC-enhanced neuronal death but also recovered glutathione and abolished the reactive oxygen species in these neurons. Threo-beta-benzyloxyaspartate, another type of glutamate transporter inhibitor, also induced glutathione depletion in the glutamate-preloaded cells, suggesting the involvement of glutamate transporter blocking in glutathione depletion. The NMDA receptor antagonist MK-801, although partially effective in reducing PDC toxicity, slightly recovered glutathione level but did not reduce the reactive oxygen species even at a high concentration (100 microM). N-acetylcysteine, dimethylsulfoxide, alpha-phenyl-N-butyl nitrone and glutathione ethylester prevented neuronal death enhanced by PDC, but superoxide dismutase and catalase did not. Our study provides evidence that the block of glutamate uptake by PDC exerts toxicity on glutamate-pretreated neurons not only through the accumulation of extracellular glutamate and subsequent activation of the NMDA receptor but also through depletion of glutathione and generation of reactive oxygen species.

[The role of transaminases and mitochondrial anion porters in the energetics of animals of different ages]. / Rol' transaminaz i anionperenoschikov mitokhondrii v énergetike zhivotnykh raznogo vozrasta.

Aminohydroxyacetate, an inhibitor of aminotransferases, decreases the rate of oxygen consumption by 1-day-old young rats by 35% 10 min after intraperitoneal injection, whereas for 20-day-old rats, the inhibitory effect is 56%, and for adult mice, it is 83%. More prolonged exposure to aminohydroxyacetate leads to death of the animal. One-day-old rats die 90 min after the injection, 20-day-old rats die 30 min after the injection, and adult mice die 15 min after the injection. Butylmalonate (an inhibitor of mitochondrial dicarboxylate translocator) decreases the rate of energy metabolism to a lower extent, by 38% in adult mice and by 18% in 1-day-old rats. The animals generally remain alive after the exposure to this compound. 1,2,3-benzyltricarboxylate, an inhibitor of tricarboxylate transport, shows only a weak effect on energy metabolism of animals of any age. These results provide evidence that the role of the transaminase system in energy metabolism increases with age. Mechanisms underlying weak sensitivity of newborn animals to these inhibitors are discussed.

Pharmacological dissociation of glutamatergic metabotropic signal transduction pathways in cortical astrocytes.

Using cultured cortical astrocytes we demonstrate differential activation of metabotropic signal transduction pathways with 1-aminocyclopentane-trans-1S3R-dicarboxylic acid (1S3R-ACPD) and the glutamate transport inhibitor trans-2,4-pyrrolidine dicarboxylic acid (trans-2,4-PDC). Phosphoinositide hydrolysis was more potently stimulated by 1S3R-ACPD than by L-trans-2,4-PDC; however, L-trans-2,4-PDC was far more efficacious than 1S3R-ACPD at inhibiting cyclic AMP accumulation. The metabotropic receptor antagonist (+)-alpha-methyl-4-carboxyphenylglycine ((+)-MCPG) inhibited 1S3R-ACPD stimulation of phosphoinositide hydrolysis but not its ability to inhibit cyclic AMP accumulation thereby demonstrating a means to pharmacologically dissociate these two metabotropic signal transduction pathways in astrocytes. (+)-MCPG produced similar antagonism of the metabotropic agonist properties of L-trans-2,4-PDC. The metabotropic effects of L-trans-2,4-PDC could not be reduced with enzymatic treatment of the cultures to remove extracellular glutamate, suggesting that these effects are not secondary to the ability of this compound to inhibit glutamate uptake. Taken together the findings indicate the presence of multiple glutamatergic signal transduction pathways in astrocytes and suggest a similarity in the pharmacophores for metabotropic receptors and glutamate transporters.

Secretion and contraluminal uptake of dicarboxylic acids in the proximal convolution of rat kidney.

The transport of dicarboxylic acids in the proximal convolution was investigated by measuring: a) the zero net flux transtubular concentration difference of DL-methyl-succinate, b) its 2-s influx from the interstitium into tubular cells, and c) its 3.5-s efflux from the tubular lumen. With the first method a luminal concentration exceeding the peritubular concentration was observed, thus indicating a net active transtubular secretion of this slowly metabolized substance. All transport steps, luminal and contraluminal , as well as the overall transport, were Na+-dependent and inhibited by lithium (apparent Ki approximately equal to 1.8 mmol/l). The overall transport of methylsuccinate , as well as the contraluminal influx into proximal tubular cells, could be inhibited by paraaminohippurate and H2-DIDS with an apparent Ki of approximately equal to 1.8 mmol/l, by taurocholate with an apparent Ki approximately equal to 3.1 mmol/l and by pyruvate with an apparent Ki approximately equal to 5 mmol/l, but not by sulfate, thiosulfate, L-lactate, oxalate and urate. As judged from the inhibition of contraluminal methylsuccinate influx by 48 dicarboxylic acids (aliphatic and aromatic), a specificity pattern was observed similar to that of inhibition of luminal efflux of 2-oxoglutarate [22]: a preference of dicarboxylates in the transconfiguration with a chain length of 4-5 carbons; little change in the inhibitory potency with CH3-, OH-, SH- and O=, but strong reduction with a NH3+ in the 2 position; only a small reduction of inhibitory potency with 2,3 disubstituted SH and OH analogs; preference of the dicarboxylic benzene in the 1,4 position and of the diacetyl benzene in the 1,2 position. The data indicate a Na+-dependent dicarboxylic transport system at the contraluminal cell side of the proximal tubule which is very similar to the luminal transport system for dicarboxylic acids.