Monitoring of pesticides in ambient air: Prioritization of substances.

Despite the richness of data collected on pesticide concentrations in ambient air in France, knowledge on this topic remains partial and heterogeneous in the absence of specific regulations. The population exposure remains thus difficult to estimate; therefore it was necessary to define modalities for implementing national monitoring of pesticides in ambient air in metropolitan France and in the overseas territories. The objective of this work was to identify which active substances (a.s.) have to be monitored in priority. As part of a collective expertise, a group of multidisciplinary experts has developed a method to rank active substances authorised as plant protection products, biocides and antiparasitic agents, which were available on the French market in 2015. A 3-steps approach has been developed. The first step consisted of a theoretical approach based on a hierarchy of substances according to four criteria: (a) national uses, (b) emission potential to the air, (c) persistence in the air, and (d) chronic toxicity. The three first criteria give information on their potential to be present in the atmosphere, and the fourth criterion allows to consider their potential of hazard. The second step was an observational approach based on existing database on pesticide air measurements in France. In the third step, both approaches were combined using decision trees to select priority pesticides. Among the 1316 a.s. first identified from the EU Pesticides database, 90 were selected, among which 43 required metrological and/or analytical development. The experts recommended confirming the relevance of performing a longer term monitoring of these a. s. after a one-year exploratory campaign. The proposed method is reproduceable, transparent, easy to update (e.g. in the light of a change in product authorization), and can be adapted to other agricultural and geographical conditions, and objectives (e.g. monitoring of the ecotoxicological effects of pesticides).

In vivo D-serine hetero-exchange through alanine-serine-cysteine (ASC) transporters detected by microelectrode biosensors.

D-serine, a co-agonist of N-methyl D-aspartate (NMDA) receptors, has been implicated in neurological and psychiatric disorders such as cerebral ischemia, lateral amyotrophic sclerosis, or schizophrenia. D-serine signaling represents an important pharmacological target for treating these diseases; however, the biochemical mechanisms controlling extracellular D-serine levels in vivo are still unclear. D-serine heteroexchange through small neutral amino acid transporters has been shown in cell cultures and brain slices and could provide a biochemical mechanism for the control of D-serine extracellular concentration in vivo. Alternatively, exocytotic D-serine release has also been proposed. In this study, the dynamics of D-serine release and clearance were explored in vivo on a second-by-second time scale using microelectrode biosensors. The rate of D-serine clearance in the rat frontal cortex after a microionophoretic injection revealed a transporter-mediated uptake mechanism. D-serine uptake was blocked by small neutral l-amino acids, implicating alanine-serine-cysteine (ASC) transporters, in particular high affinity Asc-1 and low affinity ASCT2 transporters. Interestingly, changes in alanine, serine, or threonine levels resulted in D-serine release through ASC transporters. Asc-1, but not ASCT2, appeared to release D-serine in response to changes in amino acid concentrations. Finally, neuronal silencing by tetrodotoxin increased D-serine extracellular concentration by an ASC-transporter-dependent mechanism. Together, these results indicate that D-serine heteroexchange through ASC transporters is present in vivo and may constitute a key component in the regulation of D-serine extracellular concentration.

d-Serine diffusion through the blood-brain barrier: effect on d-serine compartmentalization and storage.

d-Serine is a co-agonist of N-methyl-d-aspartate (NMDA) receptors. It has been implicated in the etiology of schizophrenia and has shown efficacy as an adjuvant to reduce positive and negative symptoms of schizophrenia. In addition, d-serine can modulate cognition in animals when administered alone. However, the neurochemical effects of exogenous d-serine on extra- and intra-cellular d-serine brain levels are poorly understood. In this study, we used both high performance liquid chromatography (HPLC) and enzyme-based microelectrode biosensors to quantify d-serine in the rat brain. We demonstrated levels of 2.3-2.8µM in the extracellular medium, 4µM in plasma and 188pmol/mg in brain tissue samples. An intraperitoneal (i.p.) d-serine injection (1g/kg) produced a slow increase in extracellular d-serine concentration in the cortex despite a surge in d-serine up to 13mM in the plasma, indicating poor diffusion through the blood-brain barrier. Using the respective volume fractions of blood, extracellular and intracellular spaces published in the literature, we estimated that d-serine intracellular stores represented more than 99% of total d-serine. These intracellular stores almost doubled 3h after d-serine administration. Overall, our data indicate that d-serine administration increases brain extra- and intra-cellular concentrations despite weak diffusion through the blood-brain barrier. These results pave the way for a better understanding of the neurochemical mechanisms by which d-serine administration modulates cognition.

Characterization of a yeast D-amino acid oxidase microbiosensor for D-serine detection in the central nervous system.

d-Serine is an endogenous ligand for N-methyl-d-aspartate (NMDA) receptors, and alterations in its concentration have been related to several brain disorders, especially schizophrenia. It is therefore an important target neuromodulator for the pharmaceutical industry. To monitor d-serine levels in vivo, we have developed a microbiosensor based on cylindrical platinum microelectrodes, covered with a membrane of poly-m-phenylenediamine (PPD) and a layer of immobilized d-amino acid oxidase from the yeast Rhodotorula gracilis (RgDAAO). By detecting the hydrogen peroxide produced by enzymatic degradation of d-serine, this microbiosensor shows a detection limit of 16 nM and a mean response time of 2 s. Interferences by ascorbic acid, uric acid, l-cysteine, and by biogenic amines and their metabolites are rejected at more than 97% by the PPD layer. Although several d-amino acids are potential substrates for RgDAAO, d-serine was the only endogenous substrate present in sufficient concentration to be detected by our microbiosensor in the central nervous system. When implanted in the cortex of anesthetized rats, this microbiosensor detected the increase in concentration of d-serine resulting from its diffusion across the blood-brain barrier after an intraperitoneal injection. This new device will make it possible to investigate in vivo the variations in d-serine concentrations occurring under normal and pathological conditions and to assess the pharmacological potency of new drugs designed to impact d-serine metabolism.