Proposal to Eliminate Urethane-Treated Positive Control Dose Groups in 26-Week Tg.rasH2 Carcinogenicity Studies.

Short-term (26 weeks) Tg.rasH2 mouse carcinogenicity studies have been conducted as an alternative model to the conventional 2-year mouse carcinogenicity studies, using urethane as a positive control material. In these studies, urethane was used at a dose of 1,000 mg/kg/dose, administered intraperitoneally on days 1, 3, and 5. Urethane consistently produces lung adenomas and carcinomas and hemangiosarcomas of the spleen, proving validity of the assay. We conducted 3 pilot studies at 3 different sites of Charles River Laboratories using a lower dose of urethane (500 mg/kg/dose), administered on days 1, 3, and 5, followed by a 12-week observation period. Our results demonstrate that a lower dose can be used successfully with fewer number of animals per sex to prove the validity of the assay. However, based on our cumulative experience with this model, we propose to eliminate positive control dose groups in future Tg.rasH2 carcinogenicity studies.

Effects of exposure to chronic ergot alkaloids on phenylephrine contractile response of maternal pedal artery, umbilical artery, and umbilical vein in pregnant ewes.

Ergot alkaloids are known to cause devastating effects in livestock. The objectives of this study were to evaluate the effects of prolonged ergot exposure on selected vasculature in pregnant sheep and to examine the role of alpha-1 adrenergic receptors in mediating these effects. Twelve 60-day pregnant sheep were randomly placed into control and exposure groups (n = 6/group). Ergot-contaminated feed pellets were given orally once a day for 45 d to the exposure group [46.6 µg/kg body weight (BW) total ergot alkaloids]. The control group (Ctl) received ergot-free pellets. The maternal pedal artery from the left hindlimb and the umbilical artery and vein were dissected and mounted in a tissue bath. The vascular contractile response to a cumulatively increasing dose of phenylephrine (PE) in the exposure group (Exp) was compared with the control groups. Chronic exposure to ergot alkaloids resulted in a 70.6% and 91.3% increase in PE contractile response in the umbilical artery (Ctl EC50 = 3.962 × 10-6; Exp EC50 = 1.161 × 10-6, P < 0.0001) and the umbilical vein (Ctl EC50 = 7.889 × 10-6; Exp EC50 = 6.801 × 10-7, P < 0.0001), respectively, but there was no increase in the pedal artery (P = 0.3927), when compared to the control group. Fetal weight in the ergot-exposed group was significantly lower than in the control group (Ctl 3.3 ± 0.17 kg; Exp 2.07 ± 0.13 kg, P = 0.0002). The increase in contractile response in the umbilical vein may result in decreased blood supply to the fetus causing decreased fetal weight. Negative impact was seen at significantly lower levels of ergot alkaloids than what is currently allowed by Canadian standards, which suggests that these limits should be reevaluated to ensure livestock safety.

Les alcaloïdes de l'ergot sont connus pour avoir des effets dévastateurs sur le bétail. Les objectifs de cette étude étaient d'évaluer les effets d'une exposition prolongée à l'ergot sur une vascularisation sélectionnée chez les brebis gestantes et d'examiner le rôle des récepteurs alpha-1 adrénergiques dans la médiation de ces effets. Douze brebis gestantes de 60 jours ont été placées au hasard dans des groupes témoins et exposés (n = 6/groupe). Des granulés alimentaires contaminés par l'ergot ont été administrés par voie orale une fois par jour pendant 45 jours au groupe exposé [46,6 µg/kg de poids corporel (PC) d'alcaloïdes totaux de l'ergot]. Le groupe témoin (Ctl) a reçu des granulés sans ergot. L'artère pédieuse maternelle du membre postérieur gauche et l'artère et la veine ombilicales ont été disséquées et montées dans un bain de tissu. La réponse contractile vasculaire à une dose cumulative croissante de phényléphrine (PE) dans le groupe exposé (Exp) a été comparée aux groupes témoins. L'exposition chronique aux alcaloïdes de l'ergot a entraîné une augmentation de 70,6 % et de 91,3 % de la réponse contractile de l'PE dans l'artère ombilicale (Ctl EC50 = 3,962 × 10−6; Exp EC50 = 1,161 × 10−6, P < 0,0001) et la veine ombilicale (Ctl EC50 = 7,889 × 10−6; Exp EC50 = 6,801 × 10−7, P < 0,0001), respectivement, mais il n'y a pas eu d'augmentation dans l'artère pédieuse (P = 0,3927), par rapport au groupe témoin. Le poids foetal du groupe exposé à l'ergot était significativement plus faible que celui du groupe témoin (Ctl 3,3 ± 0,17 kg; Exp 2,07 ± 0,13 kg, P = 0,0002). L'augmentation de la réponse contractile dans la veine ombilicale peut entraîner une diminution de l'apport sanguin au foetus, ce qui entraîne une diminution du poids foetal. Un impact négatif a été observé à des niveaux significativement plus faibles d'alcaloïdes de l'ergot que ce qui est actuellement autorisé par les normes canadiennes, ce qui suggère que ces limites devraient être réévaluées pour assurer la sécurité du bétail.(Traduit par Docteur Serge Messier).

Prolonged absorption and susceptibility to enterohepatic circulation after oral administration of ergot alkaloids in ewes.

The objective of this study was to evaluate the pharmacokinetics profile of ergot alkaloids when administered to sheep orally. Although ergot alkaloids frequently contaminate animal feed, current understanding of their pharmacokinetics in animals cannot adequately predict toxicity. Blood samples were collected from ewes at 0.5, 1, 3, 5, and 12 h after oral exposure to 4 ergot alkaloids: ergocornine, ergocristine, ergocryptine, and ergosine, followed by serum analysis of these alkaloids using high performance liquid chromatography and tandem mass spectrometry. The alkaloids showed extended absorption time, in addition to clear signs of enterohepatic circulation. This pharmacokinetic profile suggests potential enhanced toxicity in animals with disorders related to secretion of bile acid. It may also explain the high susceptibility of sheep to ergot poisoning compared to other species. An extended sampling protocol (> 12 h) is necessary, however, to identify the pharmacokinetic properties of ergot alkaloids in ewes. In conclusion, ewes exposed to ergot alkaloids showed a prolonged absorption phase and enterohepatic circulation, which is in contrast with human ergot pharmacokinetics.

L'objectif de cette étude était d'évaluer le profil pharmacocinétique des alcaloïdes de l'ergot lorsqu'ils sont administrés à des moutons par voie orale. Bien que les alcaloïdes de l'ergot contaminent fréquemment les aliments pour animaux, la compréhension actuelle de leur pharmacocinétique chez les animaux ne permet pas de prédire de manière adéquate la toxicité. Des échantillons de sang ont été prélevés chez les brebis à 0,5, 1, 3, 5 et 12 h après exposition orale à quatre alcaloïdes de l'ergot : ergocornine, ergocristine, ergocryptine et ergosine, suivi d'une analyse sérique de ces alcaloïdes par chromatographie liquide à haute performance et spectrométrie de masse en tandem. Les alcaloïdes ont montré un temps d'absorption prolongé, en plus de signes évidents de circulation entérohépatique. Ce profil pharmacocinétique suggère une toxicité potentiellement accrue chez les animaux présentant des troubles liés à la sécrétion d'acide biliaire. Cela peut également expliquer la forte sensibilité des moutons à l'empoisonnement par l'ergot par rapport aux autres espèces. Un protocole de prélèvement étendu (> 12 h) est cependant nécessaire pour identifier les propriétés pharmacocinétiques des alcaloïdes de l'ergot chez les brebis. En conclusion, les brebis exposées aux alcaloïdes ont montré une phase d'absorption prolongée et une circulation entérohépatique, ce qui contraste avec la pharmacocinétique de l'ergot chez l'humain.(Traduit par Docteur Serge Messier).

Vasoactive Effects of Acute Ergot Exposure in Sheep.

Ergotism is a common and increasing problem in Saskatchewan's livestock. Chronic exposure to low concentrations of ergot alkaloids is known to cause severe arterial vasoconstriction and gangrene through the activation of adrenergic and serotonergic receptors on vascular smooth muscles. The acute vascular effects of a single oral dose with high-level exposure to ergot alkaloids remain unknown and are examined in this study. This study had two main objectives; the first was to evaluate the role of α1-adrenergic receptors in mediating the acute vasocontractile response after single-dose exposure in sheep. The second was to examine whether terazosin (TE) could abolish the vascular contractile effects of ergot alkaloids. Twelve adult female sheep were randomly placed into control and exposure groups (n = 6/group). Ergot sclerotia were collected and finely ground. The concentrations of six ergot alkaloids (ergocornine, ergocristine, ergocryptine, ergometrine, ergosine, and ergotamine) were determined using HPLC/MS at Prairie Diagnostic Services Inc., (Saskatoon, SK, Canada). Each ewe within the treatment group received a single oral treatment of ground ergot sclerotia at a dose of 600 µg/kg BW (total ergot) while each ewe in the control group received water. Animals were euthanized 12 h after the treatment, and the pedal artery (dorsal metatarsal III artery) from the left hind limb from each animal was carefully dissected and mounted in an isolated tissue bath. The vascular contractile response to phenylephrine (PE) (α1-adrenergic agonist) was compared between the two groups before and after TE (α1-adrenergic antagonist) treatment. Acute exposure to ergot alkaloids resulted in a 38% increase in vascular sensitivity to PE compared to control (Ctl EC50 = 1.74 × 10-6 M; Exp EC50 = 1.079 × 10-6 M, p = 0.046). TE treatment resulted in a significant dose-dependent increase in EC50 in both exposure and control groups (p < 0.05 for all treatments). Surprisingly, TE effect was significantly more pronounced in the ergot exposed group compared to the control group at two of the three concentrations of TE (TE 30 nM, p = 0.36; TE 100 nM, p < 0.001; TE 300 nM, p < 0.001). Similar to chronic exposure, acute exposure to ergot alkaloids results in increased vascular sensitivity to PE. TE is a more potent dose-dependent antagonist for the PE contractile response in sheep exposed to ergot compared to the control group. This study may indicate that the dry gangrene seen in sheep, and likely other species, might be related to the activation of α1-adrenergic receptor. This effect may be reversed using TE, especially at early stages of the disease before cell death occurs. This study may also indicate that acute-single dose exposure scenario may be useful in the study of vascular effects of ergot alkaloids.