Paradoxical widespread c-Fos expression induced by a GABA agonist in the forebrain of transgenic mice with ectopic expression of the GABA(A) α6 subunit.

A GABA-site agonist gaboxadol (4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol) at 3 mg/kg induces strong anxiolytic response in a transgenic Thy1α6 mouse line ectopically expressing the GABA(A) receptor α6 subunit gene under the Thy-1.2 promoter. Now, we compared brain activation patterns between Thy1α6 and wild-type mice to identify brain structures potentially mediating this anxiolytic response. Acutely efficient anxiolytics such as benzodiazepines typically depress most brain regions while activating specifically neurons within the central extended amygdala. Gaboxadol treatment (3 mg/kg, i.p., 2 h) induced a significant increase in c-Fos expression selectively in many Thy1α6 brain regions including the limbic cortex, anterior olfactory nucleus, septal area and central and basolateral nuclei of amygdala. It failed to activate the lateral part of mediodorsal thalamic nucleus (MDL) in the Thy1α6 mice that was activated in the wild-type mice. Detailed mapping of the α6 subunit mRNA by in situ hybridization revealed expression in the middle layers of the isocortex, olfactory areas, hippocampal formation and basolateral nucleus of amygdala (BLA) in the Thy1α6 forebrain. The ligand autoradiographies (t-butylbicyclophosphoro[(35)S]thionate ([(35)S]TBPS) and [(3)H]Ro 15-4513) revealed high levels of pharmacologically active extrasynaptic α6ß and α6ßγ2 GABA(A) receptors in these same areas. However, c-Fos induction by gaboxadol treatment in Thy1α6 brain was not restricted to areas highly expressing the α6-containing GABA(A) receptors suggesting that indirect pathways lead to the paradoxically widespread activation. Interestingly, the activation pattern by gaboxadol at the dose that is anxiolytic in Thy1α6 mice resembled closely that observed after various fear- and stress-provoking challenges. However, our results are consistent with a recent observation that optogenetic activation of specific neuronal pathways in the extended amygdala mediates anxiolytic responses. Our results suggest that the widespread neuronal inhibition as typically associated with benzodiazepines is not the exclusive mechanism of anxiolysis.

Reversal of novelty-induced hyperlocomotion and hippocampal c-Fos expression in GluA1 knockout male mice by the mGluR2/3 agonist LY354740.

Dysfunctional glutamatergic neurotransmission has been implicated in schizophrenia and mood disorders. As a putative model for these disorders, a mouse line lacking the GluA1 subunit (GluA1-KO) of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate receptor displays a robust novelty-induced hyperlocomotion associated with excessive neuronal activation in the hippocampus. Agonists of metabotropic glutamate 2/3 receptors (mGluR2/3) inhibit glutamate release in various brain regions and they have been shown to inhibit neuronal activation in the hippocampus. Here, we tested a hypothesis that novelty-induced hyperlocomotion in the GluA1-KO mice is mediated via excessive hippocampal neuronal activation by analyzing whether an mGluR2/3 agonist inhibits this phenotypic feature. GluA1-KO mice and littermate wildtype (WT) controls were administered with (1S,2S,5R,6S)-2-aminobicyclo[3.1.0]hexane-2,6-dicarboxylic acid (LY354740) (15 mg/kg, i.p.) 30 min before a 2-h exposure to novel arenas after which c-Fos immunopositive cells were analyzed in the hippocampus. LY354740 (15 mg/kg) decreased hyperactivity in male GluA1-KO mice, with only a minimal effect in WT controls. This was observed in two cohorts of animals, one naïve to handling and injections, another pre-handled and accustomed to injections. LY354740 (15 mg/kg) also reduced the excessive c-Fos expression in the dorsal hippocampal CA1 pyramidal cell layer in maleGluA1-KO mice, while not affecting c-Fos levels in WT mice. In female mice, no significant effect for LY354740 (15 mg/kg) on hyperactive behavior or hippocampal c-Fos was observed in either genotype or treatment cohort. A higher dose of LY354740 (30 mg/kg) alleviated hyperlocomotion of GluA1-KO males, but not that of GluA1-KO females. In conclusion, the excessive behavioral hyperactivity of GluA1-KO mice can be partly prevented by reducing neuronal excitability in the hippocampus with the mGluR2/3 agonist suggesting that the hippocampal reactivity is strongly involved in the behavioral phenotype of GluA1-KO mice.

Similarities and differences between measured and predicted concentrations of pesticides in Dutch surface waters.

In order to have a thorough evaluation of the progress and effectiveness of Dutch crop protection policy, both model predictions and measured pesticide concentrations in surface waters are considered. To this purpose, monitoring data obtained by various water boards and other monitoring institutes were processed. Data were aggregated over a two year time period and over space (at 1x1 km-grid). A geographic view is given in the Dutch Pesticides Atlas (www.pesticidesatlas.nl). The model used for the predictions was the Dutch National Environmental Indicator NMI version 2 (www.nmi.alterra.nl) that has input data regarding spray drift data, crop interception, soil and climate and many more. Information on aggregation steps over time and space, grid sizes, information on crop areas was geared to one another for both instruments. Results on measured pesticide concentrations in surface waters and model predictions were compared to each other at the national scale. For this study, 10 different cases were selected covering a large range of pesticides' characteristics and pesticides' use. In 60% of the cases, the results were largely in agreement with each other when expressed as absolute numbers of measurements exceeding the environmental quality standard. This is very accurate and useful for policy purposes. Based on concentrations and on the order of magnitude, no significant agreement between measurements and model predictions was found. Differences were explained by various factors, and an overview of predominant systematic differences between the measurements and the model predictions was presented. Using both measurements and model predictions in supporting environmental policy evaluations is warranted, because of higher Weight-of-Evidence. Combining both can assist in optimizing the knowledge on pesticides behaviour, fate and ecological problems and therefore this is the preferred evaluation method.

Use of MGLUR2 and MGLUR3 knockout mice to explore in vivo receptor specificity of the MGLUR2/3 selective antagonist LY341495.

LY341495 is a metabotropic glutamate receptor (mGluR) antagonist showing selectivity to mGluR2/3 but having measurable antagonist efficacy across all mGluR subtypes at 10-1000 fold higher concentrations. In vivo in rodents it increases locomotor activity and wakefulness, enhances cognition and modulates emotions. It also induces widespread neuronal activation measured as c-Fos expression. To further investigate the receptor subtypes through which LY341495 might act in vivo we analyzed how its effects are altered in mGluR2-knockout (KO) and mGluR3-KO brains. In most regions, LY341495 (3 mg/kg, i.p., 2.5 h) -induced c-Fos expression was not altered in either KO brain. However, in mGluR3-KO mice, LY341495 was almost inactive in the central extended amygdala [central nucleus of the amygdala, lateral (CeL) and bed nucleus of the stria terminalis, laterodorsal (BSTLD)], suggesting that acute blockade of mGluR3 is activating these neurons in wildtype brain. In the ventrolateral nucleus of the thalamus (VL), LY341495 produced a significantly enhanced response in mGluR3-KO mice and attenuated response in mGluR2-KO mice. We also analyzed locomotion in familiar environment and found that locomotor activity was dose-dependently increased by LY341495 (1-30 mg/kg, i.p.) regardless of the genotype. In unfamiliar environment, both KO strains showed enhanced sensitivity to LY341495 in reducing locomotor habituation. Together our results indicate that certain effects of LY341495 may not be mediated by a blockade of either mGluR2 or mGluR3, but may involve other mGluR subtypes. Alternatively, functions of mGluR2 and mGluR3 may be redundant, resulting similar effects irrespective the receptor subtype being antagonized in vivo by LY341495.

Pesticides in groundwater and drinking water wells: overview of the situation in the Netherlands.

In the Netherlands, many of the fresh groundwater resources are vulnerable to pollution. Owing to high population densities and intensive farming practices, pesticide residues are found in groundwater at many places. Hence a number of drinking water abstraction wells contain pesticides residues, causing considerable costs for purification. The Water Framework Directive (WFD) requires countries to assess the chemical status of groundwater bodies and set up monitoring plans for groundwater quality, including pesticides. 771 groundwater samples were taken from monitoring wells in 2006 and analysed for a broad list of pesticides in order to fulfil these requirements. Pesticide were detected in 27% of samples, while in 11% the WFD limit of 0.1 microg/l was exceeded. In this paper, these and earlier measurements are evaluated further, considering also measurements in drinking water wells, information about the origin of measured pesticides and calculated trends in use and emissions. The measurements in the monitoring wells showed that where pesticides are used, 15-55% (minimal and maximal estimation) of the wells in shallow groundwater (1 to 20 m below soil surface) contain pesticides residues at concentrations above 0.1 microg/l. When the metabolites BAM and AMPA are excluded (as not relevant in human toxicological terms), the estimation range is 7-37%. These patterns observed in shallow groundwater are reflected by the occurrence of pesticides in vulnerable abstraction wells that are used for the production of drinking water. The WFD requires the determination of both status and trends. The design of current monitoring network is evaluated from this perspective. Several recommendations are made for more adequate and efficient monitoring.

Comparison of c-Fos induction in the brain by the mGlu2/3 receptor antagonist LY341495 and agonist LY354740: evidence for widespread endogenous tone at brain mGlu2/3 receptors in vivo.

LY341495 and LY354740 are potent and selective antagonist and agonist, respectively, for Group II metabotropic glutamate (mGlu2/3) receptors. Here we demonstrate that LY341495 (3 mg/kg) significantly increased c-Fos expression in almost all brain regions analyzed (44 out of 52 regions) in animals that were prehandled and kept in home-cage environment to minimize stress. Robust c-Fos induction was observed in all cortical regions, hippocampal CA1 and CA3 subregions, amygdala and several other subcortical nuclei. In contrast to LY341495, changes in c-Fos expression following LY354740 were more modest and not generally widespread (decreased in 1 region, dentate gyrus; and increased in 13 out of 52 regions). Interestingly, although LY354740 is anxiolytic in animals, LY341495 did not increase c-Fos expression in the paraventricular nucleus of the hypothalamus which is usually activated by stress/fear and several anxiogenic compounds. To further investigate the behavioral consequences of mGlu2/3 receptor antagonism, LY341495 was administered to prehandled animals that were placed in the elevated plus maze test under low light (low stress) conditions. Here LY341495 increased mouse elevated plus maze (EPM)-anxiety in a dose-dependent manner, significantly decreasing the time spent in open arms, but not affecting total ambulations. The behavioral consequences and associated widespread pattern of brain neuronal activations following blockade of mGlu2/3 receptors suggest that there is considerable endogenous glutamate tone throughout the brain at negative feedback peri-synaptic mGlu2/3 receptors, even under low stress conditions where synaptic glutamate release spillover would be expected to be minimized.

Anxiolytic activity of the MGLU2/3 receptor agonist LY354740 on the elevated plus maze is associated with the suppression of stress-induced c-Fos in the hippocampus and increases in c-Fos induction in several other stress-sensitive brain regions.

LY354740 is a potent and selective agonist for group II metabotropic glutamate (mGlu) receptors, mGlu2 and mGlu3 receptors, with anxiolytic activity in several animal models of anxiety, including the elevated plus maze (EPM) test. Here, we studied neuronal activation in mouse brain after EPM exposure in saline- and LY354740-treated mice using c-Fos immunoreactivity as a marker. The effect of LY354740 on c-Fos expression was also studied in cage control (no EPM) mice. Pretreatment with LY354740 (20 mg/kg, s.c.) produced robust anxiolytic behavior on the EPM. LY354740 administration decreased EPM-induced increases in c-Fos expression in the CA3 of the hippocampus, while having no significant effects on basal c-Fos expression in the hippocampus. LY354740 administration significantly increased c-Fos expression in specific limbic regions, including the lateral division of the central nucleus of the amygdala (CeL), lateral parabrachial nucleus, locus coeruleus, and Edinger-Westphal nucleus, whether or not animals were exposed to the EPM. Moreover, LY354740 administration per se significantly increased c-Fos expression in regions processing sensory information, including the paraventricular and lateral geniculate nucleus of the thalamus as well as the nucleus of the optic tract and superior colliculus. In particular, the suppression of fear-evoked neuronal activity in the hippocampus and drug-induced increases in neuronal activation in the CeL have been previously linked to the anxiolytic effects of clinically effective drugs such as benzodiazepines, and thus may contribute to anxiolytic actions of LY354740 in animal models and human anxiety patients.

Increased c-Fos expression in the centromedial nucleus of the thalamus in metabotropic glutamate 8 receptor knockout mice following the elevated plus maze test.

Ligands for metabotropic glutamate 8 (mGlu8) receptors, such as (S)-2-amino-4-phosphonobutanoic acid and (S)-3,4-dicarboxyphenylglycine suppress CNS excitability via presynaptic regulation of glutamate release and are anticonvulsant in mice. These observations suggest that mGlu8 receptors play a role in the regulation of neuronal excitability. To further characterize the role of mGlu8 receptors in vivo, the mGlu8 receptor knockout mouse was generated. Recently, we reported that mGlu8 receptor knockout mice showed increased anxiety in the elevated plus maze (EPM). Here, the pattern of c-Fos expression was studied in mGlu8 receptor knockout and wild-type mice after exposure to the EPM test for 5 min. The present study shows that the increased anxiety-related behavior of mGlu8 receptor knockout mice in the EPM was associated with a 2.3-fold higher (P<0.05) number of c-Fos positive cells in the centromedial nucleus of the thalamus compared with wild-type mice (when prehandled mice were used). The increased neuronal activity in the centromedial nucleus of the thalamus in the mGlu8 receptor knockout mouse was also observed in a separate experiment with naive mice (no prehandling). In these naive mGlu8 receptor knockouts, c-Fos expression was significantly induced by the EPM in the centrolateral nucleus of the thalamus, paraventricular nucleus of the hypothalamus, and granular cell layer of the dentate gyrus, but in naive wild-type mice c-Fos was significantly increased only in the piriform cortex. Basal c-Fos expression in the absence of EPM exposure did not differ between wild-type and mGlu8 receptor knockout mice in any brain region we examined. As the centromedial nucleus of the thalamus is important in regulating sensory information to higher brain regions, these results support the hypothesis that mGlu8 receptors are involved in the response to certain novel, aversive environments. In particular, the deletion of the mGlu8 receptor reduced the threshold of neuronal activation in stress-related brain regions such as the centromedial nucleus of the thalamus.

Field research for the authorisation of pesticides.

On request of the Dutch government a committee of the Health Council of the Netherlands has reviewed the role that results of field research in its broadest sense (i.e., including multi-species toxicity tests in the laboratory, research on model ecosystems et cetera) can play in ecotoxicological risk assessment for the authorisation of pesticides. The Committee believes that field research can provide valuable additional data about the exposure of non-target organisms and the resultant effects at population, community and ecosystem level. However, it frequently is unclear how these data might be used in reaching a decision about authorisation. To solve this problem, it is necessary to specify what is understood by "unacceptable damage". Both more clearly formulated protection goals of the government and a better understanding of the ecological significance of effects are needed to clarify this. Furthermore, the Committee points out that the statistical power of field trials must be sufficient to allow for the detection of changes that might be regarded as ecologically relevant. Finally, it recommends keeping a finger on the pulse in relation to authorised pesticides by monitoring their presence in environmental compartments and by investigating their role in suddenly occurring mortality among conspicuous animal species, such as birds, fish and honeybees. This kind of research forms a safety net for substances that have been wrongly authorised.

Excitatory actions of NMDA receptor antagonists in rat entorhinal cortex and cultured entorhinal cortical neurons.

We have characterized excitatory effects of non-competitive NMDA receptor antagonists MK-801, PCP, and ketamine in the rat entorhinal cortex and in cultured primary entorhinal cortical neurons using expression of immediate early gene c-fos as an indicator. NMDA receptor antagonists produced a strong and dose-dependent increase in c-fos mRNA and protein expression confined to neurons in the layer III of the caudal entorhinal cortex. Induction of c-fos mRNA is delayed and it is inhibited by antipsychotic drugs. Cultured entorhinal neurons are killed by high doses of MK-801 and PCP but c-fos expression is not induced in these neurons indicating that this in vitro model does not fully replicate the in vivo effects of PCP-like drugs in the entorhinal cortex. Excitatory effects of the NMDA receptor antagonists may be connected with the psychotropic side effects of these drugs and might become a useful model system to investigate neurobiology of psychosis.

EEG na enxaqueca: correlacao entre incidencia de anormalidades e formas clinicas, tempo de doenca e frequencia das crises. / EEG in migraine: correlation between abnormalities and clinical forms, lenght of the disease and crisis frequency

Os AA analisam os eletrencefalogramas de 344 pacientes com enxaqueca (119 com enxaqueca classica e 225 com enxaqueca nao classica). Maior percentual de anormalidades foi encontrado entre os pacientes com enxaqueca classica. Uma correlacao entre incidencia de anormalidades e tempo de doenca e frequencia das crises foi observada mas apenas entre os pacientes portadores de enxaqueca do tipo classica

Tumores cisticercoticos multiplos intracerebrais. Relato de um caso. / Multiple intracerebral cysticercoid tumors. Report of a case

Os AA. relatam o caso de um menor de 19 meses de idade, portador de crises convulsivas parciais e hipertensao intracraniana, consequentes a granulomas cisticercoticos multiplos intracerebrais. Dos quatro granulomas, tres foram extirpados cirurgicamente e o quarto nao removido, visto os AA. considerarem que pelo menos, no momento, ele nao estava provocando repercussoes locais ou funcionais

Aspectos atuais da terapeutica da enxaqueca. / Present aspects of the therapy of migraine

Os A.A. apoiados na experiencia obtida com o tratamento de 750 pacientes enxaquecosos, 576 dos quais tratados profilaticamente, analisam a terapeutica das crises e o tratamento profilatico com as modernas drogas atualmente disponiveis