J. Truelove

Acute Neurotoxicity of Domoic Acid in the Rat

A recent outbreak of human food poisoning, characterized by severe gastrointestinal and neurologic abnormalities, with a fatal outcome in 3 patients, was attributed to the consumption of poisonous mussels containing domoic acid at an abnormally high concentration. The purpose of the present study was to determine if domoic acid, a glutamate analogue extracted from poisonous mussel, was neurotoxic to rats. Groups of female Sprague-Dawley rats were dosed once intraperitoneally with 0, 1, 2, 4, or 7.5 mg domoic acid/kg of body weight and observed for a maximum period of 24 hr. Clinically, control rats and rats in the 1 mg/kg group were unremarkable. Seventy-five percent of the animals in the 2 mg/kg group had equivocal transient behavioral signs. One that was given 2 mg/kg and all rats given in excess of 4 mg/kg of body weight developed unequivocal behavioral and neurologic signs culminating in partial seizures and status epilepticus. Histopathologically, severely affected rats developed selective encephalopathy characterized by neuronal degeneration and vacuolation of the neuropil in the limbic and the olfactory systems, and retinopathy characterized by neuronal hydropic degeneration of the inner nuclear layer and vacuolation of the external plexiform layer. The results of this study suggest that domoic acid is excitotoxic and causes a characteristic syndrome with clinical signs and histopathologic lesions similar to those reported for kainic acid.

Domoic acid poisoning and mussel-associated intoxication: preliminary investigations into the response of mice and rats to toxic mussel extract.

Consumption of cultivated blue mussels from Prince Edward Island was recently associated with episodes of gastro-intestinal and neurological distress. Extracts of the toxic mussels, tested in the mouse bioassay for paralytic shellfish poison, caused an atypical response characterized by scratching, convulsions and death. The present investigation shows that the domoic acid present in toxic mussels can produce in mice and rats signs identical to those induced by mussel extracts. These studies, preliminary in nature by virtue of the scarcity of domoic acid, gave ip no-effect levels in mice of 0.59 mg/kg body weight based on the behavioural response (scratching) and 2.4 mg/kg for death. These levels correspond to levels of 24 and 94 ppm in mussels. When administered orally doses of between 35 and 70 mg domoic acid/kg body weight were required to produce toxicity in mice and rats. This reduced toxicity is consistent with a lack of absorption from the gastro-intestinal tract: faecal excretion accounted for 102 +/- 17% and 98 +/- 12% (mean +/- SE) of the domoic acid administered to mice and rats, respectively. Since human intoxication occurred at an estimated 1-5 mg domoic acid/kg body weight, susceptible individuals appear to be more sensitive than rodents to the oral toxicity of domoic acid.

Acute Parenteral Neurotoxicity of Domoic Acid in Cynomolgus Monkeys (M. fascicularis)

To study the CNS effects of domoic acid (D.A.), 6 adult Cynomolgus monkeys (M. fascicularis) were dosed intraperitoneally (4 mg/kg) or intravenously (0.025-0.5 mg/kg) with D.A. obtained from cultured mussels contaminated with this neurotoxin. Clinical signs of neurotoxicity were preceded by a short presymptomatic period (2-3 min) and an even shorter prodromal period (0.5-1 min). The symptomatic period proper was characterized by persistent chewing with frothing, varying degrees of gagging, and vomit. Monkeys in the higher dose regimen exhibited additional signs including abnormal head and body positions, rigidity of movements and loss of balance, and tremors. The duration of the symptomatic period was dose dependent. Excitotoxic lesions consisting of vacuolation of the neuropil, astrocytic swelling, and neuronal shrinkage and hyperchromasia were detected in the area postrema, the hypothalamus, the hippocampus, and the inner layers of the retina in monkeys given D.A. at 0.5 mg/kg intravenously and 4 mg/kg intraperitoneally. It was concluded that D.A., administered intravenously, is neuroexcitatory and a powerful emetic at doses of 0.025 to 0.2 mg/kg. At higher doses (0.5 mg/kg intravenously and 4 mg/kg intraperitoneally), D.A. is strongly excitotoxic.

Chronic Feeding Study of deoxynivalenol in B6C3F1 male and female mice

A 2 year feeding study was conducted with male and female B6C3F1 mice that consumed diets containing 0, 1, 5, or 10 ppm deoxynivalenol (DON). Survivability was good and, while the test animals gained less weight with increasing levels of DON in the diet, there were no consistent toxic manifestations associated with DON consumption. There was some evidence for an increase in serum IgA and IgG in females, and there were sporadic changes noted in the clinical chemistry and hematology parameters conducted at the terminal sacrifice. However, these changes were not considered to be biologically significant. The pathology results provided statistically significant dose-related evidence for a decrease in liver preneoplastic and neoplastic lesions as the dose level of DON increased. This negative trend probably results from the known positive correlation between body weight and the appearance of spontaneous hepatic neoplasms in this strain of mouse.

Subchronic toxicity study of domoic acid in the rat.

Male and female Sprague-Dawley rats were dosed by gavage for 64 days with 0, 0.1 or 5 mg/kg/day domoic acid. Treated animals showed no clinical abnormalities. Terminal values in haematology and clinical chemistry did not reveal differences between treated and control groups. Findings in histopathology and immunohistochemistry were unremarkable. The 24-hr urinary excretion rate for domoic acid determined at three time points was approximately 1.8% of the dose and remained unchanged during the study.

Experimental oral toxicity of domoic acid in cynomolgus monkeys (Macaca fascicularis) and rats.: Preliminary investigations☆

A recent outbreak of marine food poisoning in humans was attributed to the consumption of blue mussels (Mytilus edulis L.) contaminated with domoic acid (DA) that was produced by the diatom Nitzschia pungens. The clinical and morphological effects of single oral doses of extracts of mussels contaminated with DA or of DA isolated from toxic mussels were investigated in small groups (one to six) of cynomolgus monkeys (Macaca fascicularis; 0.5-10 mg DA/kg body weight) and of Sprague-Dawley rats (60 to 80 mg DA/kg body weight). Control animals were either given saline or were not treated. To test whether monosodium glutamate, present in the food consumed by some affected humans, and dimethylsulphoxide, suspected of being present in the plankton, enhanced the response, monosodium glutamate (at 0.25% of mussel extract bolus) or dimethylsulphoxide (at 1 g per bolus) were co-administered to two (one each) of the DA-treated monkeys. DA-treated monkeys developed transient excitation characterized by vomiting. DA-treated rats showed withdrawal followed by hyperexcitation and death (in one case). Mild to moderate central nervous system lesions consistent with neuroexcitation were present in both monkeys and rats. The addition of monosodium glutamate and dimethylsulphoxide had no significant effect on the appearance and severity of central nervous system clinical signs and lesions. The wide variations in the response of test animals to orally administered DA were attributed to the protective effect of vomiting, and to suspected incomplete or slow gastro-intestinal absorption of the toxic agent. The results reinforce the view that DA is an emetic and that under appropriate conditions may also inflict excitotoxic central nervous system damage.

Toxicological consequences of aroclor 1254 ingestion by female rhesus (macaca mulatta) monkeys. Part 1B. Prebreeding phase: Clinical and analytical laboratory findings

A group of 80 menstruating rhesus (Macaca mulatta) monkeys, with an average estimated age of 11.1 +/- 4.1 yr SD were first randomly allocated to four similar test rooms (20 monkeys/room), and then randomly allocated to one of five dose groups (four females/dose group/room). Each day, the monkeys self-ingested capsules containing doses of 0, 5, 20, 40 or 80 micrograms Aroclor 1254/kg body weight. After 25 months of daily dosing, approximately 90% of the treated females attained a qualitative pharmacokinetic steady state with respect to the concentration of polychlorinated biphenyl (PCB) in their adipose tissue. Subsequently, oestrogen and progesterone concentrations in serum were determined for one complete oestrous cycle and various immunological tests were conducted, while the monkeys continued to receive their daily dose of PCB. During the prebreeding phase of the study, blood for clinical and analytical monitoring including haematology, serum biochemistry, serum hydrocortisone, serum proteins (alpha 1, alpha 2, beta and gamma-globulins), serum immunoglobulins (A, G and M) and thyroid variables (thyroxine/triiodothyronine (T3) uptake ratio, percentage T3 uptake and free thyroxine index), were obtained monthly, as were specimens to ascertain the concentration of PCB in the blood, adipose tissue and faeces. Major findings among treated monkeys included the following: changes in haematology (decreased erythrocyte count, haematocrit, reticulocyte count, and mean platelet volume), serum biochemistry (decreased cholesterol and total bilirubin), immunotoxicity (decreased antibody production to sheep red blood cells and alterations in the percentage of T helper and T suppressor cells) and pathology (the number of regions of sebaceous gland lobules per unit of histological length was significantly reduced). These effects were observed at PCB doses lower than those previously reported for non-human primates.

Polychlorinated biphenyl toxicity in the pregnant cynomolgus monkey: A pilot study

Three pregnant cynomolgus monkeys (Macaca fascicularis) were dosed with 100 or 400 μg/kg/day of Aroclor® 1254 from approximately 60 days of gestation. One additional pregnant monkey was given dose vehicle only. The two monkeys dosed with 100 μg/kg/day delivered stillborn infants and the 400 μg/kg/day dosed monkey delivered a term infant that had impaired immunologic function compared with the control infant, and died at 139 days post partum. The three dams also had impaired immunologic capacity assessed at approximately 50 days post partum (148 days treatment). With the exception of loss of fingernails in two monkeys, no overt clinical signs of toxicity were observed in the adults. Polychlorinated biphenyl (PCB) concentrations and peak ratios in breast milk and tissues are reported.

30-day oral toxicity study of domoic acid in Cynomolgus monkeys: Lack of overt toxicity at doses approaching the acute toxic dose

Domoic acid was orally administered to 3 cynomolgus monkeys at doses of 0.5 mg/kg for 15 days and then at 0.75 mg/kg for another 15 days. After the 30-day dosing period, the treated monkeys were killed. Parameters monitored as markers for toxicity included body weight, food and water consumption, clinical observations, hematology, serum chemistry, light microscopy of all major organs (including brain and retina), and glial fibrillary acid protein immunohistochemistry. Domoic acid in serum and 24-hour urine samples was measured at several time points. All parameters measured remained unremarkable. Domoic acid concentrations measured in the 24-hour urine samples indicated that gastrointestinal absorption in the monkey was approximately 4-7 percent of the administered dose, which is at least twice that previously reported for the rat.

An 85-day study of butylated hydroxyanisole in the cynomolgus monkey

Groups of 8 cynomolgus monkeys (Macaca fascicularis) were given 500, 125 and 0 mg/kg body wt butylated hydroxyanisole (BHA) by gavage in corn oil 5 times/week for 20 days, after which the high dose was halved. No significant adverse clinical signs nor abnormal fibroscopic observations were noted before the experiment was terminated at 85 days. Blood levels of glucose, albumin, chloride, red blood cells (RBCs), hemoglobin (HGB), hematocrit (HCT) and mean corpuscular hemoglobin (MCH) were altered in a dose related manner but the altered values were well within normal ranges reported for this species. While histopathology showed no treatment related effects, the mitotic index was elevated 1.9-fold in the distal esophagus of monkeys in the high but not in the low dose group. Liver weights were increased in a dose related manner but liver monooxygenases, with the exception of decreased ethoxyresorufin deethylase activity, were all within normal limits. BHA given orally to monkeys at about the maximum tolerated dose failed to induce the massive changes noted with rats given 2% dietary BHA.