Search

A series of experiments were conducted to investigate the elimination of swainsonine in the milk of lactating ruminants following a single dose oral exposure to swainsonine (locoweed; Oxytropis sericea) and to assess subsequent subclinical effects on the mothers and their nursing young. In a preliminary experiment, lactating ewes were gavaged with locoweed providing 0.8 mg swainsonine/kg BW (n = 4; BW = 75.8 +/- 3.6 kg; lactation = d 45) and lactating cows were offered up to 2.0 mg swainsonine/kg BW free choice (n = 16; BW = 389.6 +/- 20.9 kg; lactation = d 90). Serum and milk were collected at h 0 (before treatment), 3, 6, 12, and 24 for ewes, and h 0 (before treatment), 6, 12, 18, and 24 for cows. Swainsonine was highest (P < 0.05) by h 6 in the serum and milk of ewes. Consumption of at least 0.61 mg swainsonine/kg BW induced consistent (> 0.025 microg/mL) appearance of swainsonine in cow serum and milk. In response to the results obtained in the preliminary experiment, a subsequent experiment utilizing lactating ewes (n = 13; BW = 74.8 +/- 6.4 kg; lactation = d 30) and cows (n = 13; BW = 460.8 +/- 51.9 kg; lactation = d 90) was conducted. Each lactating ruminant was gavaged with a locoweed extract to provide 0 (control), 0.2, or 0.8 mg swainsonine/kg BW and individually penned with her nursing young. Serum and milk from the mothers and serum from the nursing young were collected at h 0 (before treatment), 3, 6, 9, 12, 24 and 48 (an additional sample was obtained at h 72 for ewes and lambs). Serum and milk swainsonine was higher (P < 0.05) in the 0.8 mg treated groups and maximal (P < 0.05) concentrations occurred from h 3 to 6 for ewes and h 6 to 12 h for cows (P < 0.05). Rises in alkaline phosphatase activity indicated subclinical toxicity in the treated ewes (P < 0.05). Following a single dose oral exposure to 0.2 and 0.8 mg swainsonine/kg BW provided by a locoweed extract, swainsonine was detected in the serum and milk of lactating ewes and cows, and rises in serum alkaline phosphatase activity were observed in the ewes. Neither swainsonine nor changes in alkaline phosphatase activity was detected in the serum of the lambs and calves nursing the ewes and cows dosed with swainsonine.

The effects of ionophore supplementation on selected serum constituents of sheep consuming locoweed were investigated. Sixteen sheep were allotted by weight to a 2x2 factorial arrangement of treatments: 1) no locoweed, no lasalocid, 2) no locoweed, 0.75 mg lasalocid/kg BW, 3) 0.5 mg swainsonine/kg BW, no lasalocid, 4) 0.5 mg swainsonine/kg BW, 0.75 mg lasalocid/kg BW. Swainsonine was provided by locoweed (Oxytropissericea), and sheep were fed a blue grama based diet at 2.5% BW for a 35 d treatment period. Diets were formulated to be isocaloric and isonitrogenous. Blood samples were collected on d 1, 7,14, 21, 31 and 35 to determine serum swainsonine concentration, alkaline phosphatase, total iron, aspartate aminotransferase, g-glutamyltransferase, and lactate dehydrogenase activity and total cholesterol, and triglyceride concentrations. No lasalocid by locoweed interaction (P > 0.4) was noted for any response variable measured. Average daily gains (P = 0.4) and orts (P = 0.7) were not affected by the treatments. No lasalocid treatment (P = 0.7) or day (P = 0.1) effect of serum swainsonine was observed. A locoweed by day interaction (P < 0.0001) of serum alkaline phosphatase was detected. Alkaline phosphatase levels were elevated (P < 0.01) for locoweed treated sheep at 24 h following initial exposure and remained elevated throughout the trail. Total iron was suppressed (P < 0.08) in locoweed fed sheep. A day effect (P < 0.02) was observed for serum iron. However, no linear, quadratic, or cubic effects of day were noted (P >0.2). A locoweed by day interaction (P < 0.0001) of serum aspartate aminotransferase and g-glutamyltransferase was detected. Aspartate aminotransferase levels were elevated (P < 0.0001) by d 7 for locoweed treated animals and remained elevated throughout the trial. g--Glutamyltransferase levels were suppressed (P < 0.0001) by day 7 for locoweed treated animals and remained suppressed throughout the trial. A locoweed by day interaction (P = 0.06) of serum cholesterol was detected. However, no linear, quadratic, or cubic effects of day were detected (P = 0.2). Lasalocid treatment had no effect on any serum constituent measured. Use of lasalocid in grazing animals should not increase the likelihood of locoweed intoxication.

Peripheral blood lymphocytes (PBL) were obtained from heifers and wethers to assess the in vitro effects of swainsonine. Stimulated and unstimulated PBL (2x10(5)/well; 96 well plates) were incubated in the presence of swainsonine (2, .2, .02, .002, and 0 microg/mL; n = 32) for 72 h. Mitogens used to stimulate PBL were concanavalin A (ConA; 5 microg/mL), phytohemagglutinin-P (PHA-P; 5 microg/mL), phytohemagglutinin-M (PHA-M; 5 microg/mL), pokeweed mitogen (PWM; 5 microg/mL), and lipopolysaccharide (LPS; 10 microg/mL). Controls received no swainsonine or mitogen. Effects of swainsonine are expressed as a percentage of stimulation relative to control (stimulation index; SI). Unstimulated bovine PBL treated with 2 microg/mL of swainsonine exhibited a depressed (P<.001; SI of 87 and 100 for 2 microg/mL and control, respectively; SE = 2.7) SI. At concentrations of .2 and 2 microg/mL, swainsonine inhibited (P<.001; SI of 351, 310, and 464 for .2, 2, and 0 microg/mL, respectively; SE = 27.1) bovine PBL proliferative response to PHA-P. Swainsonine had a mitogenic effect on unstimulated ovine PBL at .2 and 2 microg/mL (P<.05; SI of 118, 113, and 100 for .2 and 2 microg/mL and control, respectively; SE = 4.4). Swainsonine inhibited ovine proliferative responses to PHA-P at .2 and 2 microg/mL (P<.005; SI of 190, 178, and 228 for .2, 2, and 0 microg/mL, respectively; SE = 9.4) and to PHA-M at .002 and 2 microg/mL (P<.03; SI of 165, 167, and 192 for .002, 2, and 0 microg/mL, respectively; SE = 7.9). The opposing responses of unstimulated ovine (mitogenic) and bovine (antiproliferative) PBL to swainsonine indicates a differential species response. Swainsonine suppression of PHA-P-induced proliferation would indicate a negative effect on ovine and bovine T-cell function.

The objective of this research was to characterize the adrenergic receptor response in veins of cattle that grazed tall fescue (Festuca arundinacea Schreb.), with (E+) and without (E-) fungal endophyte (Neotyphodium coenophialum) infection. Measurement of contractile response of lateral saphenous veins to selective adrenergic receptor agonists (alphal, phenylephrine; alpha2, BHT-920) revealed enhanced reactivity (greater contractile response) only for the alpha2 adrenergic receptor in E+ pastured cattle. This response was consistent over a 3-yr grazing period. Significant contractility differences in response to BHT-920 occurred at concentrations of 3x10(-6) M and greater. Early research has reasoned that the enhanced reactivity is due to endophyte toxin effect on intracellular signaling systems. The enhanced contractile effect would alter blood flow dynamics and contribute to clinical signs and pathologic change in animals. The alpha2-adrenergic receptor is also involved in the control of many metabolic reactions. Thus, for drug therapy to be successful in reversing the adverse effects of E+ tall fescue exposure in cattle, effects on the alpha2-adrenergic tissue receptors must be neutralized.

We and others have recently shown that specific fragments of cross-linked fibrin affect cell behavior. In order to develop a facile method for the preparative scale purification of fibrin fragment D dimer, a simple gradient generating system for conventional chromatography was developed and validated, and methods of fibrin fragment D dimer purification were compared. The experimentally measured salt concentration/time relationship fell directly on the model-predicted line. Model-predicted changes in the reservoir volume and/or salt concentration in the limit buffer affected both the initial slope and the shape of the concentration/time relationship. This gradient generation method was used to separate the D domains of fibrin(ogen) from the amino terminal region E domain using anion-exchange chromatography. While the predicted salt gradient was achieved, a salt-dependent separation was found to be less optimal than that of a pH-dependent separation, as validated by Coomassie-stained SDS-PAGE and by immunoblotting. In conclusion, a facile, user-friendly, computer-based method to predict and generate salt gradients was written and validated by direct experimentation. While fibrinogen fragment purification was acceptable using this system, both separation and yields of fibrinogen and fibrin fragments were superior using a pH-based separation technique., (Copyright 1998 Academic Press.)

Two replicate trials determined the effects of dietary supplementation on snakeweed toxicity in beef cows of poor condition. Cows were stratified by weight and randomly assigned to 3 dietary treatments. Dietary treatments were control (n = 3/trial; medium quality hay, 9.49% crude protein), corn supplementation (n = 3/trial; control diet + 628 g cracked corn), and protein supplementation (n = 3/trial; control diet + 800 g 42% protein supplement). Corn and protein dietary treatments were fed to be isocaloric. Each trial consisted of 2 phases (68 days/phase). Phase 1 consisted of dietary treatments without snakeweed. In phase 2 dietary treatments contained snakeweed as 10% of the dry matter. Phase 1 and 2 dietary treatments were isocaloric/isonitrogenous. Dry matter intake of the control diet was limited to 1.3% of body weight/d. Body condition score and back fat were measured on days 0, 21 and 68 of each phase. Serum samples were collected at the onset of each trial and on days 28, 42 and 56 of each phase. Serum bromosulphthalein (BSP) elimination half life (t1/2) was estimated during week 6 of each phase. Serum BSP elimination t1/2 was higher for the control diet versus corn and protein treatments. Increased blood urea nitrogen (BUN) was found by day 28 of phase 2. Serum total bilirubin increased by day 28 in phase 2 compared to baseline for the control and corn dietary treatments. Additionally, serum indirect bilirubin was higher by day 28 in phase 2. Likewise, serum direct bilirubin increased during phase 2 on day 28 in the corn diet, but decreased by day 28 for the protein diet. Alkaline phosphatase levels were higher (P < 0.05) in the controls by day 28, but lower in the protein treatment by day 28 in phase 2. Changes were noted during phase 2 for some of the serum clinical profiles; however, these changes appear due to dietary restriction. In contrast, changes during phase 2 point to possible hepatotoxic and renal toxic effects of snakeweed. Phase 2 data suggest a benefit of protein supplementation for improving animal tolerance to snakeweed.

The objective of these experiments was to test the hypothesis that the major alkaloid classes found in endophyte-infected tall fescue could act as growth promoters for vascular smooth muscle. Bovine vascular smooth muscle cells (VSMC) from the dorsal metatarsal artery were grown in vitro and exposed to five concentrations (10(-6), 10(-8), 10(-9), 10(-11) and 0 M) of ergonovine, alpha-ergocryptine, ergovaline, and N-acetyl loline for 48 h. The mitogenic potential of the alkaloids was tested on both actively growing cells (serum fed, 10% fetal bovine serum) and quiescent cells (serum starved, 0.1% serum) to assess the need for serum constituents for alkaloid induced growth or toxicity. Ergonovine stimulated (P < .07) VSMC growth at 10(-6) M concentration in growing and quiescent culturs and at 10(-8) M concentration in quiescent cultures. alpha-Ergocryptine stimulated (P < .01) growth at 10(-6) M concentration in growing cultures and at 10(-8) and 10(-9) M concentrations in quiescent cultures. Ergovaline exhibited a dual activity on the growth of VSMC in culture, stimulating (P = .06) growth of quiescent cells at 10(-9) M concentration but inhibiting (P < .05) growth of growing cultures at concentrations of 10(-6) and 10(-9) M. This duality of activity was also noted for N-acetyl loline: N-acetyl loline stimulated (P < .05) growth of quiescent cultures at concentrations of 10(-8), 10(-9), and 10(-11) M but inhibited (P < .05) growth of growing cultures at concentrations of 10(-8) and 10(-9) M. The growth effects of the alkaloids in vitro on VSMC support in part the hypothesis that the alkaloids may contribute to the vascular complications noted in cattle grazing endophyte-infected tall fescue through hyperplasia of the intima. This would result in a decreased luminal diameter of the blood vessels and a resultant decrease in blood flow to the afflicted tissues. The diminished blood flow to tissues would result in tissue death and reduced ability to dissipate heat.

Gravid mares grazing endophyte-infested (E+) tall fescue exhibit increased gestation lengths, agalactia, foal and mare mortality, tough and thickened placentas, weak and dysmature foals, increased sweating during warm weather, reduced serum prolactin and progesterone, and increased serum estradiol-17 beta levels. Also, E+ tall fescue hay is less digestible than endophyte-free (E-) hay. Unlike many other species, horses consuming E+ tall fescue do not exhibit increased body temperature. Young horses consuming only E+ pasture do not gain as well as those consuming E- pasture. There is little difference in gain when the pasture is supplemented with enough concentrate to meet NRC requirements for growth. Neither selenium injections nor supplementing with corn at 50% of the NRC requirements for energy reduces the effects of toxic tall fescue on reproduction and lactation in gravid mares. It seems that the alkaloids of E+ tall fescue are serving as D2 dopamine receptor agonists. This activity would explain their prolactin-lowering effect. Domperidone, a dopamine receptor antagonist, is effective in preventing the signs of tall fescue toxicosis in horses without neuroleptic side effects.

We have studied the relationship between patient movement and changes in the auditory steady-state evoked potential, upper facial muscle electromyogram (FEMG), electroencephalographic-zero crossing frequency (EEG-ZCF) and heart rate during emergence from anaesthesia. Twelve healthy patients underwent surgery during stable isoflurane-nitrous oxide-oxygen anaesthesia without neuromuscular block. After skin closure, anaesthesia was discontinued abruptly while mechanical ventilation was continued until the patient moved. The magnitude of change in each physiological signal was evaluated in decibels (dB). Both the auditory steady state evoked potential and FEMG showed significant increases in amplitude during the last 5-min period before movement (6.1 and 10.7 dB, respectively). EEG-ZCF increased rapidly after anaesthesia was discontinued (2.5 dB) but there was no further increase in activity before movement. Heart rate did not change before movement. The use of the decibel transformation offers a promising method of displaying and interpreting changes in physiological variables during anaesthesia.

Cultured rat pituitary cells were studied to: determine the effects of ergovaline and loline on in vitro prolactin release; delineate the agonistic activity of these alkaloids at the D2 dopamine receptor, using 2 selective D2 dopamine receptor antagonists; and compare the efficacy of 2 dopamine receptor antagonists in reversing effects of the treatments on in vitro prolactin secretion. Ergovaline reduced in vitro prolactin release by at least 40% (P < 0.05) at concentrations of 10(-4), 10(-6), and 10(-8) M. However, loline reduced (P < 0.05) prolactin release only at the highest concentration, 10(-4) M. Two standard dopamine agonists, dopamine and alpha-ergocryptine, were used to verify that the inhibitory control mechanisms of in vitro prolactin release were intact. Both compounds reduced prolactin release by at least 40% for concentrations of 10(-4), 10(-6), or 10(-8) M. Selective D2 dopamine receptor antagonists (10(-6) M), domperidone and sulpiride, reversed (P < 0.05) the effect of loline on in vitro prolactin release. However, only domperidone (10(-6) M) was able to reverse (P < 0.05) the effect of ergovaline and only at the lowest ergovaline concentration (10(-8) M). Domperidone was more effective (P < 0.05) in reversing the prolactin-suppressing effect of alpha-ergocryptine than was sulpiride. The dose-response curve for domperidone (cubic fit, P < 0.0001) indicated a threshold concentration (10(-7) M) for reversal of alpha-ergocryptine's (10(-8) M) effect on prolactin release. However, at similar concentration of sulpiride (quadratic fit, P < 0.007), a threshold level was not obtained.(ABSTRACT TRUNCATED AT 250 WORDS)

We evaluated the effectiveness of 2 dopamine antagonists as treatments for fescue toxicosis in horses. Sixteen gravid mares were assigned by breed and expected foaling date to 1 of 3 treatment groups: endophyte-infested control; 1.1 mg of domperidone/kg of body weight/d; and 3.3 mg of sulpiride/kg/d. Mares were pastured on endophyte-infected fescue and received 0.454 kg of a corn and dried molasses carrier containing the drug treatment. Treatment started 30 days prior to expected foaling date and continued until parturition. Blood samples were collected, and mammary gland scores were recorded every 5 days. Body weight and body condition scores were obtained every 28 days. Serum was analyzed for prolactin, progesterone, and estradiol-17 beta concentrations. Domperidone-treated mares had shorter (P = 0.09) gestation duration and foaled closer (P = 0.07) to their expected parturition date than did control mares. Mammary gland scores were higher (P < 0.05) for domperidone-treated mares than for control mares. By 4 and 9 days after the start of treatment, serum prolactin concentration was higher (P < 0.05) in domperidone-treated mares and sulpiride-treated mares, respectively, than in control mares. Domperidone- and sulpiride-treated mares had higher (P < 0.05) serum progesterone and lower (P < 0.01) estradiol-17 beta concentrations than did control mares. These results indicate that domperidone may offer considerable potential as a treatment for fescue toxicosis in horses.

Treatment of bovine lateral saphenous vein (cranial branch) and dorsal metatarsal artery with lysergamide (lysergic acid amide), an alkaloid in abundance in tall fescue, resulted in vasoconstriction similar to that previously shown for the ergot alkaloids ergonovine and ergotamine. Preincubation of tissues with lysergamide resulted in partial inhibition of the contractile response induced by the selective adrenergic agents phenylephrine and BHT-920 (P < .05), indicating partial agonist or antagonist activity of lysergamide at these receptors. Bovine vessels were strongly contracted by serotonin, and the response was markedly inhibited (P < .01) when tissues were preincubated with lysergamide. Studies with selective serotonergic agents indicated that lysergamide may have predilection for serotonin-2 receptors (5-HT2). Thus, lysergamide by itself has vasoconstrictor activity and acts as a partial agonist or antagonist at adrenergic and serotonergic receptors. Further studies are needed to establish the exact receptor effects of lysergamide. However, it is evident that this important alkaloid found in tall fescue infested with the endophytic fungus Acremonium coenophialum should be given consideration in studies designed to alleviate the fescue toxicosis syndrome in cattle.

Studies of fescue toxicosis have identified several classes of plant/fungal alkaloids that may be responsible for the toxicosis problem of endophyte (Acremonium coenophialum) infected tall fescue (Festuca arundinacea). This review explores the interaction of these alkaloids with the animal's system and attempts to explain the various anomalies of fescue toxicosis through discussion of both in vitro and in vivo experiments.

The objective of this research was to measure the effects of endophyte-infected tall fescue seed extract and various alkaloids associated with the endophyte on in vitro prolactin secretion by rat hemipituitaries. Rat anterior pituitaries (AP) were dissected into halves and placed in temperature-controlled culture chambers (37 degrees C). The tissue was perfused with culture media at a flow rate of 12 mL/h. After perfusion for at least 90 min with control media, AP halves were exposed to their respective treatments for 15 min before they were returned to the control media. The treatments for Exp. 1 were .01 micrograms of alpha-ergocryptine/mL of culture medium, .01 microgram of ergonovine/mL of culture medium, .01 gram-equivalents of endophyte-infected tall fescue seed/mL of culture medium, and .01 gram-equivalents of endophyte free tall fescue seed/mL of culture medium. Treatments for Exp. 2 consisted of 10(-4), 10(-6), and 10(-8) M concentrations of perloline, N-formyl loline, N-acetyl loline, N-methyl loline, and alpha-ergocryptine. alpha-Ergocryptine suppressed (P less than .10) prolactin secretion in both experiments. Ergonovine and perloline both stimulated (P less than .10) prolactin secretion. The loline alkaloids (N-formyl loline, N-acetyl loline, N-methyl loline) had no effect on prolactin secretion. The endophyte-infected seed extract treatment suppressed (P less than .10) prolactin secretion. The endophyte-free seed extract treatment had no effect on prolactin secretion. In Exp. 2, prolactin secretion from AP responded to alpha-ergocryptine treatment in a dose-dependent fashion.(ABSTRACT TRUNCATED AT 250 WORDS)