Low levels of sarin affect the EEG in marmoset monkeys: A pilot study

The main purpose of this pilot study was to estimate the lowest observable adverse effect level (LOAEL) for the electroencephalogram (EEG) upon long-term, low-level exposure of vehicle-pretreated and pyridostigmine-pretreated marmoset monkeys to sarin vapour. This is the C·t value (t = 5 h) of exposure at which the EEG becomes significantly different from that resulting from air exposure of the same animals. The LOAELs for effects on the EEG in vehicle- and pyridostigmine-pretreated marmosets appeared to be 0.2 and 0.1 mg min m -3, respectively. Comparatively, the latter LOAEL values are at least an order of magnitude lower than the previously established LOAEL for miosis and only 2-5 times higher than the lowest observable effect level (LOEL) of bound sarin in blood. The second aim of the study was to analyse the EEG of the same marmosets again during a 5-h exposure to air 1 year after exposure to sarin vapour. All the marmosets still demonstrated significant (P < 0.05) EEG differences. In most vehicle-pretreated marmosets the energy (μV2) per EEG band was higher than that observed 1 year earlier, which might indicate that neurons had become more sensitive to excitation. This phenomenon was less pronounced in pyridostigmine-pretreated animals. Visual examination of the EEG records revealed clear bursts of alpha frequencies (ca. 9 Hz), resembling sleep-spindles, that were present more frequently in both groups of exposed marmosets than in naive animals. These late changes in spindle oscillation might be the result of changes in the cholinergic system due to exposure to sarin vapour 1 year previously. In conclusion, EEG abnormalities persisting for more than 1 year may occur in humans during long-term (5 h) exposure to subclinical levels of sarin that are not detectable by the currently fielded alarm systems. Copyright © 2004 John Wiley & Sons, Ltd

Long-term, low-level exposure of guinea pigs and marmosets to sarin vapor in air: Lowest observable effect level

Realistic scenarios for low-level exposure to nerve agents will often involve exposures over several hours to extremely low doses of agent. In order to expose animals to the lowest controllable concentrations of agent and to increase exposure times until a lowest observable effect level (LOEL) becomes measurable, a validated system was developed for exposing conscious animals to 0.05-1.0 μg/m3 (8-160 ppt) of sarin and other nerve agents. Based on cold trapping of sarin from the exposure air, the concentration could be measured semicontinuously, at 4-min time intervals by means of gas chromatography. We found that the LOEL upon a 5-h whole body exposure of guinea pigs and marmosets to sarin vapor corresponds with the measurement of an internal dose by means of fluoride-induced regeneration of sarin from phosphylated binding sites in plasma, mostly BuChE. For guinea pigs the LOEL was observed at Ct = 0.010 ± 0.002 mg/min/m3, whereas a Ct of 0.04 ± 0.01 mg/min/m3 was established for the LOEL in marmosets. These levels are several orders of magnitude lower than those based on classical measurement of depressed cholinesterase activities. At low exposure levels of guinea pigs and marmosets (≤1 μg/m3), a reasonable linearity was observed between exposure dose and internal dose. The data were addressed in the light of the recently recommended occupational exposure limits to sarin for workers without respiratory protection, which suggests that the exposure limits should be reconsidered if the slightest inhibition of cholinesterases should be prevented. © 2003 Elsevier Science (USA). All rights reserved

Intravenous toxicokinetics of sulfur mustard and its DNA-adducts in the hairless guinea pig and marmoset

ln order to provide a quantitative basis for pretreatment and therapy of intoxications with sulfur mustard the toxicokinetics of this agent as well as its major DNA-adducts are being studied in male hairless guinea pigs for the intravenous, respiratory and percutaneous routes. A highly sensitive method for bioanalysis of the intact agent in blood and tissues was developed, involving gas chromatography with automated thermodesorption injection and mass-spectrometric detection. Deuterated sulfur mustard is used as the internal standard. The absolute detection limit is 700 fg for sulfur mustard, which corresponds with a detection limit in blood of ca. 5 pg/ml. DNA-adducts are measured via the previously developed immuno-slot-blot method, using antibodies directed against the adduct of sulfur mustard to guanine. The intravenous 96-h LD50 of sulfur mustard in the hairless guinea pig was determined and appeared to be 8.2 mg/kg. The intravenous toxicokinetics of this dose in the hairless guinea pig are characterized by a very rapid distribution phase and a very slow elimination phase. A rapid adduct formation occurs in blood and lung, and subsequently in other organs. The adduct levels in lung were remarkably high. A considerable repair of the adducts is observed within 6 h. However, at 2 days after administration of sulfur mustard adducts are still detectable in most of the organs studied. The intravenous toxicokinetics of sulfur mustard were also studied in the marmoset at a dose corresponding with 1 LD50 in the hairless guinea pig. The results obtained sofar will be discussed

Intravenous and inhalation toxicokinetics of sarin stereoisomers in atropinized guinea pigs

We report the first toxicokinetic studies of (±)-sarin. The toxicokinetics of the stereoisomers of this nerve agent were studied in anesthetized, atropinized, and restrained guinea pigs after intravenous bolus administration of a dose corresponding to 0.8 LD50 and after nose-only exposure to vapor concentrations yielding 0.4 and 0.8 LCt50 in an 8-min exposure time. During exposure the respiratory minute volume and frequency were monitored. Blood samples were taken for gas chromatographic analysis of the nerve agent stereoisomers and for measurement of the activity of blood acetylcholinesterase (ACHE). In all experiments, the concentration of (+)-sarin was below the detection limit <5 pg/ml). The concentration-time profile of the toxic isomer, i.e., (-)-sarin, after an intravenous bolus was adequately described with a two-exponential equation. (-)-Sarin is distributed ca. 10-fold faster than C(-)P(-)-soman, whereas its elimination proceeds almost 10-fold slower. During nose-only exposure to 0.4 and 0.8 LCt50 of (±)-sarin in 8 min, (-)-sarin appeared to be rapidly absorbed. The blood AChE activity decreased during the exposure period to ca. 15 and 70% of control activity, respectively. There were no effects on the respiratory parameters. A significant nonlinearity of the toxico-kinetics with dose was observed for the respiratory experiments. © 2000 Academic Press

Whole body exposure of marmoset monkeys to low levels of sarin: Lowest observable effect level in blood, and early effects on pupil size and EEG

Realistic scenarios for low-level exposure to nerve agents will often involve exposures over several hours to extremely low doses of agent. The purpose of this pilot study was to indicate for sarin: (i) the lowest observable effect level (LOEL), i.e., the C.t-value (t = 5 h) of exposure at which an internal dose (fluoride-regenerated sarin from blood BuChE) becomes measurable; (ii) The lowest observable adverse effect level (LOAEL) for miosis and EEG-abnormalities in marmosets. For that purpose a validated system was developed for exposing conscious animals to the lowest controllable concentrations of sarin (8-80 ppt: 0.05-0.5μg.m-3), and to increase concentrations (7-150 μg.m-3) and exposure times (t ≤ 5 h) until observable adverse effect levels become measurable. The ratio of pupil and iris diameters, measured on digital photographs taken on-line during exposure, was calculated as a measure for miosis. EEG-signals were recorded telemetrically; the averaged amounts of energy (μV2) per EEG-band of air-exposed animals were compared to the amounts of energy of the corresponding bands of sarin-exposed animals. The LOEL was observed at Ct = 0.04 ± 0.01 mg.min.m-3. This level is several orders of magnitude lower than that based on classical measurement of depressed ChE activity. Pupil size was significantly (p < 0.05) decreased at a dose of 2.5 ± 0.8 mg.min.m-3. The earliest significant (p < 0.05) changes on the EEG-bands were observed at a dose of 0.2 mg.min.m-3. The latter LOAEL-value is an order of magnitude lower than that for miosis, and only 2-5 times higher than the LOEL. One year after the exposure to sarin vapor all marmosets still demonstrated significant (p < 0.05) EEG differences. In most marmosets the energy (μV2) per EEG-band was higher than that observed one year earlier, which might indicate that neurons had become more sensitive to excitation. Visual examination of the EEG-records revealed clear bursts of alpha frequencies (around 9 Hz), resembling sleep-spindles, which were more frequently present in exposed than in naive animals. These late changes in spindle oscillation might be the result of persistent changes in the cholinergic system due to exposure to sarin vapor one year before. In conclusion, LOEL levels, miosis, and EEG-abnormalities, may occur in humans during long term (5 h) exposure to levels of sarin which are not detectable by the currently fielded alarm systems

Low level exposure to GB vapor in air: Diagnosis/dosimetry, lowest observable effect level (LOEL) and lowest observable adverse effect level (LOAEL)

The purpose of the present study was to establish for both saline-pretreated and pyridostigmine-pretreated conscious guinea pigs whole-body exposed to low levels of GB vapor: (i) The Lowest Observable Effect Level (LOEL) for GB, i.e., the C.t-value (t = 5-h) of exposure at which an internal dose (fluoride-regenerated GB from blood ChE) becomes measurable. (ii) The Lowest Observable Adverse Effect Level (LOAEL) for GB, i.e., the C.t-value (t = 5-h) of exposure at which effects start to affect performance. (iii) Whether unexpected adverse effects on performance emerge through the combination of pyridostigmine-pretreatment and GB exposure. Read-out parameters reflecting adverse effects on performance during exposure were pupil size (miosis), EEG, visually evoked response (VER), and 1-h after exposure startle-response and shuttle-box active avoidance behavior. The lowest exposure concentration of GB which resulted in significant (p < 0.05) effects on these parameters by the end of a 5-h exposure period was taken for calculating the LOAEL (C.t-value) for each of these parameters. The averaged LOEL for saline-pretreated guinea pigs was 0.010 ± 0.002.0 mg.min.m-3 (t = 84 ± 17 min) which did not differ significantly from that of pyridostigmine-pretreated animals (0.014 ± 0.002.0 mg.min.m-3). These data were addressed in the light of the recommended occupational limits for GB vapor in air