Cina M. Mack

Diurnal variation in thermoregulatory response to chlorpyrifos and carbaryl in the rat

Time of day of exposure is rarely considered in the study of insecticide toxicology. It would be expected that the circadian temperature rhythm (CTR) as well as the circadian rhythms of other physiological processes would affect the efficacy of anticholinesterase (antiChE) insecticides. The ability of antiChE insecticides to alter core temperature (T(c)) could be affected by time of exposure in relation to the CTR. To this end, we assessed time of exposure on the efficacy of the antiChE insecticides chlorpyrifos (CHP) and carbaryl (CAR) to alter T(c) in the rat. T(c) and motor activity (MA) were monitored by radiotelemetry. Rats were dosed orally with 0, 30, and 50 mg/kg CHP or 0, 25 and 75 mg/kg CAR at 09:00 and 15:00 h. Both insecticides caused an acute decrease followed by a delayed increase in T(c) by 24-48 h post-exposure. The temperature index (TI) (area under curve of DeltaT(c) with time) was significantly greater when CHP was given at 15:00 h as compared with 09:00 h. The maximum decrease in T(c) was similar for morning and afternoon CHP. The TI following CAR was similar for morning and afternoon exposure. CHP suppressed the 24 h MA equally when given in the morning and afternoon. CAR was more effective in reducing MA when given in the morning as compared with the afternoon. The T(c) increase measured 24 h after dosing was greater when CHP was given in the morning. Overall, time of day affected the thermoregulatory toxicity of CHP but not CAR. Another experiment showed that the hypothermic efficacy of oxotremorine, a muscarinic agonist, was greater when injected at 09:00 h as compared with 15:00 h. Hence, cholinergic stimulation is probably not the only mechanism to explain the effects of the chronotoxicogical effects of some antiChE insecticides.

A multi-laboratory evaluation of microelectrode array-based measurements of neural network activity for acute neurotoxicity testing ☆

HIGHLIGHTSFour laboratories tested 6 compounds for neuroactivity using microelectrode arrays.All four laboratories were able to correctly identify the three neurotoxic compounds.Three non‐neuroactive compounds were correctly identified 10/12 times.Despite methodological differences, results were consistent across laboratories.These results support use of microelectrode arrays for neurotoxicity screening. ABSTRACT There is a need for methods to screen and prioritize chemicals for potential hazard, including neurotoxicity. Microelectrode array (MEA) systems enable simultaneous extracellular recordings from multiple sites in neural networks in real time and thereby provide a robust measure of network activity. In this study, spontaneous activity measurements from primary neuronal cultures treated with three neurotoxic or three non‐neurotoxic compounds was evaluated across four different laboratories. All four individual laboratories correctly identifed the neurotoxic compounds chlorpyrifos oxon (an organophosphate insecticide), deltamethrin (a pyrethroid insecticide) and domoic acid (an excitotoxicant). By contrast, the other three compounds (glyphosate, dimethyl phthalate and acetaminophen) considered to be non‐neurotoxic (“negative controls”), produced only sporadic changes of the measured parameters. The results were consistent across the different laboratories, as all three neurotoxic compounds caused concentration‐dependent inhibition of mean firing rate (MFR). Further, MFR appeared to be the most sensitive parameter for effects of neurotoxic compounds, as changes in electrical activity measured by mean frequency intra burst (MFIB), and mean burst duration (MBD) did not result in concentration‐response relationships for some of the positive compounds, or required higher concentrations for an effect to be observed. However, greater numbers of compounds need to be tested to confirm this. The results obtained indicate that measurement of spontaneous electrical activity using MEAs provides a robust assessment of compound effects on neural network function.

Glufosinate binds N-methyl-D-aspartate receptors and increases neuronal network activity in vitro.

Glufosinate (GLF) at high levels in mammals causes convulsions and amnesia through a mechanism that is not completely understood. The structural similarity of GLF to glutamate (GLU) implicates the glutamatergic system as a target for GLF neurotoxicity. The current work examined in vitro GLF interaction with N-methyl-D-aspartate subtype GLU receptors (NMDARs) and GLT-1 transporters via [(3)H]CGP 39653 binding experiments and [(3)H]GLU uptake assays, respectively. GLF effects on neuronal network activity were assessed using microelectrode array (MEA) recordings in primary cultures of cortical neurons. GLF and its primary metabolite N-acetylglufosinate (NAcGLF) bind to the NMDAR; the IC50 value for GLF was 668 μM and for NAcGLF was about 100 μM. Concentrations of GLF greater than 1000 μM were needed to decrease GLU uptake through GLT-1. In MEA recordings from networks of rat primary cortical neurons, the concentration-responses for NMDA, GLF and NAcGLF on network mean firing rates (MFR) were biphasic, increasing at lower concentrations and decreasing below control levels at higher concentrations. Increases in MFR occurred between 3-10 μM NMDA (290% control, maximum), 100-300 μM NAcGLF (190% control, maximum) and 10-1000 μM GLF (340% control, maximum). The NMDAR antagonist MK801 attenuated both NMDA and GLF increases in MFR. The GLF concentration required to alter GLU transport through GLT-1 is not likely to be attained in vivo, and therefore not relevant to the neurotoxic mode of action. However, toxicokinetic data from reports of intentional human poisonings indicate that GLF concentrations in the CNS after acute exposure could reach levels high enough to lead to effects mediated via NMDARs. Furthermore, the newly characterized action of NAcGLF at the NMDAR suggests that both the parent compound and metabolite could contribute to neurotoxicity via this pathway.