nvir'onrnental EAr.hives of
Arch. Environ. Contam. Toxicol. 22, 36--40(1992)
c
ontamination and
|oxi~31ogy
Effect of Deoxynivalenol on Neurotransmitters in Discrete Regions of Swine Brain D. B. Prelusky l'z, J. M. Yeung 3, B. K. T h o m p s o n 4, and H. L. Trenholm I Agriculture Canada, Government of Canada, Ottawa, Ontario, K1A 0C6, Canada and Medical College of Pennsylvania, Philadelphia, Pennsylvania 19129, USA Abstract. The effect of deoxynivalenol (DON, vomitoxin) on brain amine levels was investigated in swine. DON, a trichothecene'mycotoxin, causes suppression of feed intake (anorexia) in susceptible species. Following acute administration of DON to pigs (0.25 mg/kg, IV), concentrations of endogenous catecholamines norepinephrine (NE), dopamine (DA), 3,4-dihydroxyphenyl-acetic acid (DOPAC), and homovanillic acid (HVA), and the indoleamines, 5-hydroxytryptamine (5HT, serotonin) and 5-hydroxyindoleacetic acid (5HIAA) were determined in five brain regions, periodically during the 24 h post-dosing. Analysis was carried out by high performance liquid chromatography, using electrochemical detection. Effects of DON in the swine brain were transmitter, time and region-specific. It was observed that levels of the major transmitters (NE, DA and 5HT) were statistically different from controls in the hypothalamus (Hypo), frontal cortex (FCX) and cerebellum (Cb) up to 8 h post-dosing. Overall, DON administration elevated NE and depressed DA concentrations in these regions, and levels of 5HT which increased initially in Hypo (1 h), had dropped significantly below controls in both Hypo and FCX at 8 h. These alterations, however, were not indicative of known neurochemical changes associated with chemical-induced anorexia. Instead, this data suggested that the neurochemical effects of acute DON exposure might be due to peripheral toxicological events (i.e., vomiting), which overwhelmed its more subtie feed refusal activity.
As mycotoxin contamination of feed and feed products becomes more evident, there is an increasing interest concerning its toxicological impact. Deoxynivalenol (DON; vomi-
I Animal Research Centre; Contribution No. 1743. 2 To whom correspondence should be addressed. 3 Department of Psychiatry; current address: Bureau of Chemical Safety, Health and Welfare Canada, Ottawa, Ontario, Canada K1A 0L2. 4 Research Program Service; Contribution No. R082.
toxin) is one of numerous trichothecene mycotoxins produced by various strains of the F u s a r i u m fungus, a common contaminant of grain grown in temperate climates. While associated with a number of trichothecene-induced symptoms, DON is considered one of the least potent in this class of toxin (Trenholm et al. 1986; Ueno 1986). Still, in animal studies DON produces two characteristic actions: reduction of feed consumption (anorexia) when present at low levels in the diet, and vomiting (emesis) at higher, acute doses (Forsyth et al. 1977; Vesonder and Hesseltine 1981). Other trichothecenes (T-2 toxin, fusaranon-x, nivalenol, etc.) are also capable of producing these effects with varied potencies (Ueno 1977). The mechanism behind DON's anorexic activity is not clear, although recent studies have attempted to link it to alterations in neurotransmission. Important aspects of feeding behavior in animals are mediated by the action of multiple neurochemicals. These compounds, however, include a wide range of catecholamines, amino-acids, and neuropeptides as possible transmitters, whose mode-of-action in this regard are generally not well defined (Leibowitz 1985). Several investigators have reported that changes in brain concentrations of certain catecholamines (monoamines) do occur in trichothecene treated animals, depending on the toxin and the species involved. It appeared that species (i.e., poultry) relatively tolerant to the acute effects of toxins (DON, T-2 toxin, cyclopiazonic acid) were less likely to show significant alterations in brain amines compared to species more susceptible (i.e., swine, rats) (Ahmed and Singh 1984; Boyd et al. 1988; Chi e t al. 1981; Fitzpatrick e t al, 1988a, 1988b; MacDonald e t al. 1988; Porter et al. 1988). It is difficult, however, to establish whether a substance alters feeding behavior by acting directly on central mechanisms which specifically control hunger/satiety, or by acting peripherally by inducing a malaise associated with non-specific toxicoses.
The present study was undertaken to examine the neurochemical effects in swine following acute intravenous (IV) administration of DON. Animals were evaluated for biogenic amine and metabolite concentrations at various times postdosing, in five selected brain regions with recognized catecholaminergic and serotonergic activity.
Effect of Deoxynivalenol on Neurotransmitters
Methods
Chemicals Deoxynivalenol was provided by Dr. J. D. Miller, Agriculture Canada, Ottawa, biosynthetically produced and purified according to a published method (Miller and Arnison 1986). Purity of the toxin (>96%) was confirmed by reversed phase high performance liquid chromatography (HPLC). The neurotransmitter standards norepinephrine (NE), dopamine (DA), 3,4-dihydroxyphenyl-acetic acid (DOPAC), homovanillic acid (HVA), 5-hydroxytyramine (5HT, serotonin), and 5-hydroxyindole-acetic acid (5HIAA) were obtained from Sigma Chemical Co. (St. Louis, MO), as were the sodium meta-bisulphite, EDTA (disodium salt), and octylsulphate.
Animals and Treatment
37
Statistical Analysis Analyses of variance were applied to the monoamine concentrations, looking specifically for trends across elapsed time postdosing. The model included terms for time and DON effects and their interaction. Because the experiment was conducted over two days, an additional term was added to the model representing day differences. Owing to operational requirements, time of dosage was restricted to specific times of the day. No evidence was found to suggest that the time of dosing seriously impacted on the results. There were a few extreme values obtained in the experiment. The analyses of variance were calculated with and without these data. In cases where these values had a serious impact on the results, the results with the values excluded are reported in order to avoid placing too much weight on one or two observations.
Results Animals
Forty healthy Yorkshire barrows, bred at the Animal Research Centre farm under pathogen-free conditions and 10-13 weeks of age (15-23 kg), were used. Following a short acclimatization period in group pens (8 pigs/pen), animals were randomly allocated to treatments, and fasted for 4 h prior to DON administration. For dosing, DON was dissolved in 10% aqueous ethanol, filter sterilized (0.22 ~m filter units, Millipore Products, Bedford, MA), and injected into the ear vein at a dose of 0.25 mg/kg body weight. Control animals received the same amount of vehicle solution (300 ~1) as the toxintreated pigs. Animals within a group were dosed during a 40 min time period in the day: 8-h group, 0800-0840 h; 0.33-h group, 08500930 h; 24-h group, 1000-1040 h; 3-h group, 1200-1240 h; 1-h group, 1250-1330 h. The pigs were not fed for the duration of the study although water was provided ad libitum. Each group of eight pigs (4 treated, 4 control) were killed at each of five different time intervals after dosing (0.33, 1, 3, 8, and 24 h). To sacrifice, animals were administered T-61 followed quickly by exsanguination. Brains were removed immediately and the frontal cortex (FCX), cerebellum (Cb), hypothalamus (Hypo), hippocampus (Hip), and pons/medulla (P/M) regions were dissected. Tissues were frozen immediately in liquid N2 and stored at -70°C until analyzed.
A m i n e and Metabolite Determinations The amines and the metabolites were analyzed according to a modification of the procedure of Wester et al. (1987). Briefly, on the day of assay, 80 mg of tissue was sonicated in 200 I~1of 0.1 M perchloric acid containing 0.3 mM Na2EDTA and 0.5 mM sodium metabisulphite. Isoproterenol (30 ng in 30 ~1) was added to each tube prior to sonication as an internal standard. Tissue was sonicated on ice for 15 sec using a Fisher sonic dismembrator at 50% relative output. Homogenates were centrifuged at 48,000 x g for 15 min at 4°C. The supernatant was injected directly onto a HPLC with amperometric detection (LC-4B, Bioanalytical System Inc., West Lafayette, IN). A C18 ODS, 250 x 46 mm, 5 ixm column (Biophase, Bioanalytical Systems Inc., West Lafayette, IN) was used for the separation of amines and their metabolites. The mobile phase consisted of 100 mM citrate buffer including 0.3 mM Na:EDTA, 6% (v/v) acetonitrile and 0.334 mM octylsulphate at a pH of 2.33. The flow rate was 1 ml/min and the detector was set at +0.8V. The peak-area ratios of the amine or metabolite to internal standard in each sample were compared to the peak-area ratios of standard concentrations of these substances and the internal standard. Quantitations were done by linear regression. Results were expressed in ng/g tissue.
Following injection of pigs with D O N , the earliest o v e r t signs of toxicity were o b s e r v e d within 3 min. T h e s e consisted of chewing, grinding of teeth, and increasing salivation, with subsequent onset of vomiting by individual animals 7-22 min post-dosing. E m e s i s subsided in all animals by 90 min post-dosing. N o control animals receiving only the vehicle (10% ethanol/water) displayed any s y m p t o m s of toxicosis.
Catecholamine Profiles The levels in m o n o a m i n e concentrations in selected brain regions following D O N administration are p r o v i d e d in Table 1. Depending on the transmitter m e a s u r e d , there w e r e considerable differences b e t w e e n dose and control in all regions except the P/M where only a single significant variation (P < 0.05) was noted ( H V A , 1 h). O t h e r regional alterations w e r e manifest more. The c o n c e n t r a t i o n of N E had i n c r e a s e d quickly (20 min) in the H y p o (101%), Cb (19%) and F C X (28%), remaining elevated for the initial 3 h post-dosing period, after which time values in all three areas returned to control levels by 8 h and for the remainder of the study. In comparison, D A levels were reduced substantially (43--47%) in the same brain regions, and r e m a i n e d depressed also up to 3 h, depending on the region (Hypo, 20 min; Cb, 1 h; F C X , 3 h). The effect of D O N on endogenous levels of the D A metabolites, D O P A C and H V A p r o d u c e d a m o r e variable response. Both amines b e c a m e highly elevated in the Hip for the initial 3 h, although there w e r e no significant changes (P > 0.05) of either D A or N E in this region. In the F C X , though, while both metabolites displayed an initial d e c r e a s e ( D O P A C , 57%; H V A , 42%) at 20 min (as did DA), the levels rebounded a b o v e baseline concentrations by 1 and 3 h, suggesting increased metabolism or utilization of D A which also concurrently increased b e t w e e n 1-3 h post-dosing. D O P A C levels in the hypothalamus and c e r e b e l l u m w e r e unaffected, whereas hypothalmic H V A s h o w e d a v e r y significant (P < 0.001) initial d e c r e a s e which quickly returned to control levels by 1 h. Cerebellum H V A showed a singular increase at 3 h (P < 0.01) which faded by 8 h.
38
D . B . Prelusky et al.
Table 1. Effect of deoxynivalenol on pig brain amine levels in selected regions a Brain region Time (h)
Neurotransmitter and metabolite levels b NE
DA
DOPAC
HVA
5HT
5H1AA
377.6 758.4 ***e 302.4 462.5** 272.5 567.7*** 429.9 502.8 365.6 230.9 37.2
179.3 101.3"~ 153.3 167.7 191.0 190.4 164.6 169.9 258.4 228.8 25.0
200.8 144.1 165~6 221.1 177.0 233.4 178.3 248.7 144.5 180.9 24.1
683.5 259.6ttt 521.8 522.7 530.8 428.6 528.5 536.1 250.0 382.7 58.8
218.8 225.6 170.0 269.1"** 227.7 216.7 283.2 172.0t~t 285.6 232.3 17.9
290.0 284.9 268.3 314.8 325.5 396.2 382.6 360.8 258.4 188.2 35.5
Hypothalamus .33 ~ a 1c d 3c a 8¢ a 24 ~ d SEM f Cerebellum .33 c o 1c d 3~ o 8c o 24 e d SEM Frontal cortex .33 c a 1c a 3~ a 8c a 24 c a SEM Hippocampus .33 ¢ d 1c d 3c d 8c a 24 ~ d SEM Ports/Medulla .33 c a 1~ a 3c a 8c a 24 ~ d SEM
154.3 183.8** 134.9 152.0" 132.9 160.8"* 146.3 135.6 170.1 152.3 8.9
5.1 2.7tt 5.5 2.5t? 5.7 4.0 5.0 7.2* 4.4 5.0 0.80
4.9 6.2 3.7 2.7 3.6 4.8 5.2 5.6 4.2 4.8 1.1
17.8 21.1 17.5 22.3 20.0 29.8* 29.0 23.9 24.8 21.3 2.8
169.1 216.2'* 159.7 198.5"* 171.6 223.1" 182.4 165.3 170.9 152.6 12.5
8.0 4.2t 11.9 6.5t 13.7 7.8tt" 9.3 10.0 10.4 7.4 1.5
18.5 7.9tt 14.4 25.4** 11.3 19.6" 12.4 12.9 15.9 17.8 2.1
47.0 27. l~t 39.5 54.8* 37.1 54.3* 52.8 35.6 35.4 26.9 5.3
80.1 66.0 76.4 74.9 77.3 75.3 93.7 58.2tt'~ 71.3 56.2 6.5
82.9 47.4ttt 73.7 72.9 63.6 76.3 83.8 80.2 72.1 66.3 6.2
10.6 9.7 7.2 15.5"** 9.5 16.8"** 11.4 9.6 10.3 9.8 0.98
51.8 81.3"* 39.0 79.8*** 54.0 95.3*** 67.1 66.4 53.5 62.4 13.5
50.5 38.5 50.1 66.5 66.3 73.2 83.0 91.5 60.2 37.8t 5.9
105.4 146.5 148.2 184.1 188.4 156.5 163.2 t53.5 153.7 136.9 28.8
46.4 37.5 29.7 43.8 39.7 38.0 25.9 21.0 28.7 24.1 6.2
66.4 72.2 56.7 88.3* 73.9 90.2 63.4 45.6 56.6 57.2 12.2
126.2 155.7 119.6 150.8 136.3 137.0 158.3 145.9 139.7 120.1 10.8 254.3 214.4 196.2 270.6 267.1 238.6 239.2 168.7 184.3 193.5 35.3
5.7 7.3 6.0 8.2 9.2 8.0 9.5 7.4 8.9 7.6 0.88 18.2 16.8 17.5 24.6 16.4 19.1 18.7 19.8 17.3 24.5 3.3
nd g
197.9 179.5 189.0 199.1 211.3 208.7 201.0 135.8 183.9 134.5 32.9
26.5 20.2tt 26.5 18.8tt 24.8 19.3t 29.2 33.3 17.8 19.9 2.1
181.0 147.3 154.2 140.5 152.9 174.5 164.5 128.5 130.1 106.6 37.7
a IV dose, 0.25 mg DON/kg body weight b n g amine/g brain tissue, mean, n = 3 or 4 animals (duplicate determinations). N E = norepinephrine; DA = dopamine; DOPAC = 3,4-dihydroxyphenyl-acetic acid; H V A = homovanillic acid; 5HT = 5-hydroxytryptamine; 5 H I A A = 5-hydroxyindoleacetic acid c Control animals d Dosed animals Significantly different from the control group: increase; *P < 0.05; **P < 0.01 ; ***P < 0.001: decrease; t P < 0.05; ~tP < 0.01 ; t t t P < 0.001 f SEM = standard error of mean g n d = not detected
Effect of Deoxynivalenol on Neurotransmitters Indoleamine Profiles
The effects of DON on indolamine levels were also region specific (Table 1). Hypothalamic serotonin (5HT) concentrations showed a large increase (58%) at 1 h post-dosing (P < 0.001), which subsequently dropped below control levels (39%) by 8 h (P < 0.001). At 8 h, there was also a corresponding decrease (38%) in cortical 5HT levels (P < 0.001), which like Hypo 5HT returned to control levels by 24 h. There was essentially no change from 5HT control values in the hippocampus and pons/medulla. The 5HT levels in the cerebellum were too low to quantitate. The 5HIAA, a major metabolite of 5HT, showed an initial rapid decrease in both the Cb and FCX, but while cortical 5HIAA quickly rebounded to control values, levels in the cerebellum remained depressed for the 3 h post-dosing. There were no significant (P > 0.05) changes measured in 5HIAA concentrations in the hypo, hip, or P/M regions.
Discussion This study investigated the effects of DON on brain biogenic amine levels following a single IV administration of the toxin to swine, which are considered a species highly sensitive to the feed refusal and emetic activities of trichothecenes. The effects observed varied markedly with time and brain region for the different transmitter measured. It was evident that there were some fluctuations in baseline (control) metabolite concentrations as samples were collected throughout the day (0910-1640 h). With most transmitters the change during this period was moderate (up to ---20% average daily levels), the one exception being hypothalmic HVA in which a very large variation was noted (250683 ng/g). These data suggested circadian rhythm in brain amine concentrations occurs and/or variation induced by individual animals' response to experimental stress (i.e., handling, injection, crowding), both of which have been shown to influence turnover rates of specific catecholamines or indoleamines. Considering the multiple mechanisms which are involved in maintaining CNS function, various authors have reported neurotransmitter systems are subject to biorhythms, and that these patterns can be altered by disruption of normal behaviour, (i.e., stress, fasting, light/dark cycle, exercise, etc.) (Davis 1989; Karczmar 1978). In several studies investigating the effects of mycotoxins on biogenic amines, the data has also shown that control levels varied with time, depending on species and compound measured (Ahmed and Singh 1984; Boyd et al. 1988; Chi et al. 1981; Fitzpatrick et al. 1988b; Porter et al. 1988). In the current study, no statistical evidence was found to suggest that the time of dosing seriously impacted on the results. In rats it has been shown that there is rapid depletion of NE in most brain regions accompanying both physical and psychological stress (Iimori et al. 1982). In the present study, a similar trend was observed in control animals following injection of the vehicle only (although the effect of circadian rhythm still could not be ruled out). Ball (1988) found that pigs judged to be more excited or stressed just prior to slaughter had significantly lower hypothalamic concentrations of all the neurochemicals measured (NE, 5HT, 5HIAA, HVA), and Hallberg et al. (1983) noted that stress-
39 susceptible pigs had lower brain amine levels compared to more stress-tolerant strains. Although available data indicate that certain neuroregulators are involved in the feeding process, information is still too limited and variable to predict the effects of certain anorexic-inducing xenobiotics on neurochemical patterns. These chemicals can reduce feed intake by a variety of different mechanisms, affecting feeding time, intervals between meals, intake amount, rate, and meal preference (i.e., protein, carbohydrate, fat, etc.) (Davis 1989). Prior evidence has suggested that important aspects of feeding may be controlled by 5HT in the brain and periphery; elevated levels causing a loss of appetite (Shor-Posner et al. 1986). This was possibly reflected in the current study where hypothalamic 5HT exhibited a large increase at 1 h post-dosing, but did not account for its subsequent depletion in both the hypotbalamus and cortex at 8 h. It was not readily apparent whether this abrupt decrease was due to the increased utilization of 5HT or to a negative feedback in 5HT synthesis in response to its earlier enhanced release. However, the ratio of 5HIAA/5HT which can be used as an index for 5HT metabolism or utilization, was increased in both the Hypo (57%) and FCX (53%) at 8 h. This indicated an enhanced utilization of 5HT which could have an anorexic consequence. Changes in NE and DA levels following DON administration, though, were not consistent with current opinion on anorexia (Hoebel et al. 1989; Leibowitz 1989). Both elevated NE and depressed DA levels are associated with stimulation of the feeding behaviour, opposite to the effect expected with DON. Consequently, it is probable that the catecholamine imbalances observed here were not due to the anorexic activity of DON, but instead were secondary to other toxicological responses produced by the acute dose of the mycotoxin, possibly its emetic activity. Smith and MacDonald (1987) demonstrated that acute administration of the Fusarium toxin, fusaric acid, to swine caused significant increases in hypothalamic NE and 5HT, and cortical NE; the same pattern as reported here for DON. The function of neuroregulators in the vomiting action is not well understood. It has been reported that both NE and 5HT probably have a strong role in relaying information during the process (Carpenter et al. 1983; Schwartz et al. 1986), and dopamine, an emetic itself, appears to have the greater involvement in initiating the emetic reflex. Similarly, DA has been implicated as a mediator of learned aversion. Hoebel et al. (1989) reported that brain DA levels decreased when rats were exposed to substances perceived to cause illness. Thus, in the current study, decreased DA levels which occurred early in pigs may reflect a negative response to DONinduced nausea and malaise. Possibly this is accomplished through enhanced metabolism of DA to N E and other metabolites (DOPAC, HVA). The absence of longer-term effects may reflect a short toxicological half-life for DON. It has been shown that up to 75% of DON administered IV to swine was eliminated within 6 h (Prelusky et al. 1988), and essentially all regional amine levels in the present study (except for 5HT), had returned to control levels by 8 h post-dosing (Table 1). This showed that a single sublethal dose of the toxin, although quickly symptomatic, was limited in duration. In comparison, Fitzpatrick et al. (1988a) reported that 5HT and 5HIAA levels were still elevated in all brain regions of the rat 24 h after oral DON
40
administration, but the dosage used was 10 times that in the current study. Similarly, prolonged alteration in various monoamine concentrations have also been reported in rats and chickens following oral administration of lethal amounts of T-2 toxin, a related but more acutely toxic trichotecene (Boyd et al. 1988; Chi et al. 1981; MacDonald et al. 1988). In these latter studies, however, doses were sufficient to produce mortality, and the effects on brain amine levels were probably due to the overall chemical assault rather than any selective anorexic activity. It has been postulated that the effect of D O N and other trichothecenes on feeding behaviour is regulated through alterations in the availability of specific brain neurotransmitters. In the current study, it was shown that D O N administered IV to swine caused a marked change in certain regional monoamine concentrations, but this data, in conjunction with other published studies clearly demonstrates the difficulties in trying to establish a relationship between the neurochemical profile and the feed refusal activity of xenobiotics. Numerous internal, as well as environmental factors can influence the signals mediating behavior, and feeding can be either enhanced or inhibited. For example, Ball et al. (1988) showed that while it was possible to elevate hypothalamic 5HT levels in pigs with a tryptophan supplemented diet, which should accordingly decrease appetite, it, in fact, had no significant effect on feed intake or daily weight gain. Further studies are needed to fully understand whether the anorectic mechanism of action of D O N is neurotransmitter and/or second messenger-specific, or a secondary consequence of other pharmacological effects.
References
Ahmed N, Singh US (1984) Effect of aflatoxin B1 on brain serotonin and catecholamines in chickens. Toxicol Lett 21:365-367 Ball RO (1988) Effects of stress at slaughter on brain neurotransmitters in market pigs. Ontario Swine Research Review, Ont Agric College Publ No 0689, ISSN 0842-9839, pp 77-78 Ball RO, Lawson SL, Moore H (1988) Effect of dietary tryptophan concentration and feeding period on serotonin concentration in hypothalamus of market pigs. Ontario Swine Research Review, Ont Agric College Publ No 0689 ISSN 0842-9839. pp 78-80 Boyd KE, Fitzpatrick DW, Wilson JR, Wilson LM (1988) Effect of T-2 toxin on brain biogenic monoamines in rats and chickens. Can J Vet Res 52:181-185 Carpenter DO, Briggs DB, Strominger N (1983) Response of neurons of canine area postrema to neurotransmitters and peptides. Cell Mol Neurobiol 3:113-126 Chi MS, E1-Halawani ME, Waibel PE, Mirocha CJ (1981) Effects of T-2 toxin on brain catecholamines and selected blood components in growing chickens. Poult Sci 60:137-141 Davis JD (1989) The microstructure of ingestive behavior In: Schneider LH, Cooper SJ, Haimi KA (eds) The psychobiology of human eating disorders: Preclinical and clinical perspectives. Vo1575, Annals New York Academy Sciences, NY pp 106--121 Fitzpatrick DW, Boyd KE, Watts BM (1988a) Comparison of the trichothecenes deoxynivalenol and T-2 toxin for their effects on brain biogenic monoamines in the rat. Toxicol Lett 40:241-245 Fitzpatrick DW, Boyd KE, Wilson LM, Wilson JR (1988b) Effect of the trichothecene deoxynivalenol in brain biogenic monoamine concentrations in rats and chickens. J Environ Sci Health B23:159-170 Forsyth DM, Yoshizawa T, Morooka N, Tuite J (1977) Emetic and refusal activity of deoxynivalenol to swine. Appl Environ Microbiol 34:547-552
D.B. Prelusky et al. Hallberg JW, Draper DD, Topel DG, Altrogge DM (1983) Neural catechoalmine deficiencies in the porcine stress syndrome. Am J Vet Res 44:368-371 Hoebel BG, Hernandez L, Schwartz DH, Mark GP, Hunter GA (1989) Microdialysis studies of brain norepinephrine, serotonin, and dopamine release during ingestive behavior In: Schneider LH, Cooper SJ, Haimi KA (eds) The psychobiology of human eating disorders: Preclinical and clinical perspectives. Annals New York Academy Sciences, NY, pp 221-233 Iimori K, Tanaka M, Kohno Y, Ida Y, Nakagawa R, Hoaki Y, Tsuda A, Nagasaki N (1982) Psychological stress enhances noradrenaline turnover in specific brain regions in rats. Pharmacol Biochem Behav 16:637-640 Karczmar AG (1978) Multitransmitter mechanisms underlying selected function, particularly aggression, learning, and sexual behavior In: Deniker P, Radouco-Thomas C, Villeneuve A (eds) Neuropsychopharmacology, Pergamon Press, Oxford. pp 581608 Leibowitz SF (1985) Brain neurotransmitters and appetite regulation. Psychopharmacol Bull 21:412-418 - (1989) Hypothalamic neuropeptide Y, galanin, and amines: Concept of coexistence in relation to feeding behavior. In: Schneider LH, Cooper SJ, Haimi KA (eds) The psychobiology of human eating disorders: Preclinical and clinical perspectives. Annals New York Academy of Sciences, NY, pp 171-191 MacDonald EJ, Cavan KR, Smith TK (1988) Effect of acute oral doses of T-2 toxin on tissue concentrations of biogenic amines in the rat. J Anim Sci 66:434-441 Miller JD, Arnison PG (1986) Degradation of deoxynivalenol by suspension cultures of the Fusarium head blight resistant wheat cultivar Frontana. Can J Plant Pathol 8:147-150 Porter JK, Norred WP, Cole RJ, Dorner JW (1988) Neurochemical effects of cyclopiazonic acid in chickens. Proc Soc Exp Biol Med 187:335-340 Prelusky DB, Hartin KE, Trenholm HL, Miller JD (1988) Pharmacokinetic fate of 14C-labelled deoxynivalenol in swine. Fundam Appl Toxicol 10:276-286 Schwartz J-C, Agid Y, Bouthenet M-L, Javoy-Agid F, LlorensCortes C, Martres M-P, Pollard H, Sales N, Taquet H (1986) Neurochemical investigations into the human area postrema In: Davis CJ, Lake-Bakaar GV, Grahame-Smith DE (eds) Nausea and vomiting: Mechanism and treatment. Springer-Verlag, NY, pp 18-30 Shor-Posner G, Grinker JA, Marinescu C, Brown O, Leibowitz SF (1986) Hypothalamic serotonin in the control of meal patterns and macronutrient selection. Brain Res Bull 17:663-671 Smith TK, MacDonald EJ (1987) Effect of an acute dose of fusaric acid on brain neurochemistry in swine. Can J Anim Sci 67:1182 (abstract) Trenholm HL, Friend DW, Hamilton RMG, Thompson BK, Hartin KE (1986) Incidence and toxicology of deoxynivalenol as an emerging mycotoxin problem. Proc VI Int Conf Mycotoxicosis, pp 76-82 Ueno Y (1986) Trichothecenes as environmental toxicants. Reviews Environ Toxicol 2:303-341 - (1977) Mode of action of trichothecenes. Pure Appl Chem 49:1737-1745 Vesonder RF, Hesseltine CW (1981) Vomitoxin: Natural occurrence on cereal grains and significance as a refusal and emetic factor to swine. Proc Biochem 16:12-15 Wester P, Gottfries J, Winblad B (1987) Simultaneous liquid chromatographic determination of seventeen of the major monoamine neurotransmitters, precursors and metabolites. II. Assessment of human brain and cerebrospinal fluid concentrations. J Chromatogr 415:275-288 Manuscript received May 16, 1991 and in revised form August 28, 1991.
Arch. Environ. Contam. Toxicol. 22, 36--40(1992)
c
ontamination and
|oxi~31ogy
Effect of Deoxynivalenol on Neurotransmitters in Discrete Regions of Swine Brain D. B. Prelusky l'z, J. M. Yeung 3, B. K. T h o m p s o n 4, and H. L. Trenholm I Agriculture Canada, Government of Canada, Ottawa, Ontario, K1A 0C6, Canada and Medical College of Pennsylvania, Philadelphia, Pennsylvania 19129, USA Abstract. The effect of deoxynivalenol (DON, vomitoxin) on brain amine levels was investigated in swine. DON, a trichothecene'mycotoxin, causes suppression of feed intake (anorexia) in susceptible species. Following acute administration of DON to pigs (0.25 mg/kg, IV), concentrations of endogenous catecholamines norepinephrine (NE), dopamine (DA), 3,4-dihydroxyphenyl-acetic acid (DOPAC), and homovanillic acid (HVA), and the indoleamines, 5-hydroxytryptamine (5HT, serotonin) and 5-hydroxyindoleacetic acid (5HIAA) were determined in five brain regions, periodically during the 24 h post-dosing. Analysis was carried out by high performance liquid chromatography, using electrochemical detection. Effects of DON in the swine brain were transmitter, time and region-specific. It was observed that levels of the major transmitters (NE, DA and 5HT) were statistically different from controls in the hypothalamus (Hypo), frontal cortex (FCX) and cerebellum (Cb) up to 8 h post-dosing. Overall, DON administration elevated NE and depressed DA concentrations in these regions, and levels of 5HT which increased initially in Hypo (1 h), had dropped significantly below controls in both Hypo and FCX at 8 h. These alterations, however, were not indicative of known neurochemical changes associated with chemical-induced anorexia. Instead, this data suggested that the neurochemical effects of acute DON exposure might be due to peripheral toxicological events (i.e., vomiting), which overwhelmed its more subtie feed refusal activity.
As mycotoxin contamination of feed and feed products becomes more evident, there is an increasing interest concerning its toxicological impact. Deoxynivalenol (DON; vomi-
I Animal Research Centre; Contribution No. 1743. 2 To whom correspondence should be addressed. 3 Department of Psychiatry; current address: Bureau of Chemical Safety, Health and Welfare Canada, Ottawa, Ontario, Canada K1A 0L2. 4 Research Program Service; Contribution No. R082.
toxin) is one of numerous trichothecene mycotoxins produced by various strains of the F u s a r i u m fungus, a common contaminant of grain grown in temperate climates. While associated with a number of trichothecene-induced symptoms, DON is considered one of the least potent in this class of toxin (Trenholm et al. 1986; Ueno 1986). Still, in animal studies DON produces two characteristic actions: reduction of feed consumption (anorexia) when present at low levels in the diet, and vomiting (emesis) at higher, acute doses (Forsyth et al. 1977; Vesonder and Hesseltine 1981). Other trichothecenes (T-2 toxin, fusaranon-x, nivalenol, etc.) are also capable of producing these effects with varied potencies (Ueno 1977). The mechanism behind DON's anorexic activity is not clear, although recent studies have attempted to link it to alterations in neurotransmission. Important aspects of feeding behavior in animals are mediated by the action of multiple neurochemicals. These compounds, however, include a wide range of catecholamines, amino-acids, and neuropeptides as possible transmitters, whose mode-of-action in this regard are generally not well defined (Leibowitz 1985). Several investigators have reported that changes in brain concentrations of certain catecholamines (monoamines) do occur in trichothecene treated animals, depending on the toxin and the species involved. It appeared that species (i.e., poultry) relatively tolerant to the acute effects of toxins (DON, T-2 toxin, cyclopiazonic acid) were less likely to show significant alterations in brain amines compared to species more susceptible (i.e., swine, rats) (Ahmed and Singh 1984; Boyd et al. 1988; Chi e t al. 1981; Fitzpatrick e t al, 1988a, 1988b; MacDonald e t al. 1988; Porter et al. 1988). It is difficult, however, to establish whether a substance alters feeding behavior by acting directly on central mechanisms which specifically control hunger/satiety, or by acting peripherally by inducing a malaise associated with non-specific toxicoses.
The present study was undertaken to examine the neurochemical effects in swine following acute intravenous (IV) administration of DON. Animals were evaluated for biogenic amine and metabolite concentrations at various times postdosing, in five selected brain regions with recognized catecholaminergic and serotonergic activity.
Effect of Deoxynivalenol on Neurotransmitters
Methods
Chemicals Deoxynivalenol was provided by Dr. J. D. Miller, Agriculture Canada, Ottawa, biosynthetically produced and purified according to a published method (Miller and Arnison 1986). Purity of the toxin (>96%) was confirmed by reversed phase high performance liquid chromatography (HPLC). The neurotransmitter standards norepinephrine (NE), dopamine (DA), 3,4-dihydroxyphenyl-acetic acid (DOPAC), homovanillic acid (HVA), 5-hydroxytyramine (5HT, serotonin), and 5-hydroxyindole-acetic acid (5HIAA) were obtained from Sigma Chemical Co. (St. Louis, MO), as were the sodium meta-bisulphite, EDTA (disodium salt), and octylsulphate.
Animals and Treatment
37
Statistical Analysis Analyses of variance were applied to the monoamine concentrations, looking specifically for trends across elapsed time postdosing. The model included terms for time and DON effects and their interaction. Because the experiment was conducted over two days, an additional term was added to the model representing day differences. Owing to operational requirements, time of dosage was restricted to specific times of the day. No evidence was found to suggest that the time of dosing seriously impacted on the results. There were a few extreme values obtained in the experiment. The analyses of variance were calculated with and without these data. In cases where these values had a serious impact on the results, the results with the values excluded are reported in order to avoid placing too much weight on one or two observations.
Results Animals
Forty healthy Yorkshire barrows, bred at the Animal Research Centre farm under pathogen-free conditions and 10-13 weeks of age (15-23 kg), were used. Following a short acclimatization period in group pens (8 pigs/pen), animals were randomly allocated to treatments, and fasted for 4 h prior to DON administration. For dosing, DON was dissolved in 10% aqueous ethanol, filter sterilized (0.22 ~m filter units, Millipore Products, Bedford, MA), and injected into the ear vein at a dose of 0.25 mg/kg body weight. Control animals received the same amount of vehicle solution (300 ~1) as the toxintreated pigs. Animals within a group were dosed during a 40 min time period in the day: 8-h group, 0800-0840 h; 0.33-h group, 08500930 h; 24-h group, 1000-1040 h; 3-h group, 1200-1240 h; 1-h group, 1250-1330 h. The pigs were not fed for the duration of the study although water was provided ad libitum. Each group of eight pigs (4 treated, 4 control) were killed at each of five different time intervals after dosing (0.33, 1, 3, 8, and 24 h). To sacrifice, animals were administered T-61 followed quickly by exsanguination. Brains were removed immediately and the frontal cortex (FCX), cerebellum (Cb), hypothalamus (Hypo), hippocampus (Hip), and pons/medulla (P/M) regions were dissected. Tissues were frozen immediately in liquid N2 and stored at -70°C until analyzed.
A m i n e and Metabolite Determinations The amines and the metabolites were analyzed according to a modification of the procedure of Wester et al. (1987). Briefly, on the day of assay, 80 mg of tissue was sonicated in 200 I~1of 0.1 M perchloric acid containing 0.3 mM Na2EDTA and 0.5 mM sodium metabisulphite. Isoproterenol (30 ng in 30 ~1) was added to each tube prior to sonication as an internal standard. Tissue was sonicated on ice for 15 sec using a Fisher sonic dismembrator at 50% relative output. Homogenates were centrifuged at 48,000 x g for 15 min at 4°C. The supernatant was injected directly onto a HPLC with amperometric detection (LC-4B, Bioanalytical System Inc., West Lafayette, IN). A C18 ODS, 250 x 46 mm, 5 ixm column (Biophase, Bioanalytical Systems Inc., West Lafayette, IN) was used for the separation of amines and their metabolites. The mobile phase consisted of 100 mM citrate buffer including 0.3 mM Na:EDTA, 6% (v/v) acetonitrile and 0.334 mM octylsulphate at a pH of 2.33. The flow rate was 1 ml/min and the detector was set at +0.8V. The peak-area ratios of the amine or metabolite to internal standard in each sample were compared to the peak-area ratios of standard concentrations of these substances and the internal standard. Quantitations were done by linear regression. Results were expressed in ng/g tissue.
Following injection of pigs with D O N , the earliest o v e r t signs of toxicity were o b s e r v e d within 3 min. T h e s e consisted of chewing, grinding of teeth, and increasing salivation, with subsequent onset of vomiting by individual animals 7-22 min post-dosing. E m e s i s subsided in all animals by 90 min post-dosing. N o control animals receiving only the vehicle (10% ethanol/water) displayed any s y m p t o m s of toxicosis.
Catecholamine Profiles The levels in m o n o a m i n e concentrations in selected brain regions following D O N administration are p r o v i d e d in Table 1. Depending on the transmitter m e a s u r e d , there w e r e considerable differences b e t w e e n dose and control in all regions except the P/M where only a single significant variation (P < 0.05) was noted ( H V A , 1 h). O t h e r regional alterations w e r e manifest more. The c o n c e n t r a t i o n of N E had i n c r e a s e d quickly (20 min) in the H y p o (101%), Cb (19%) and F C X (28%), remaining elevated for the initial 3 h post-dosing period, after which time values in all three areas returned to control levels by 8 h and for the remainder of the study. In comparison, D A levels were reduced substantially (43--47%) in the same brain regions, and r e m a i n e d depressed also up to 3 h, depending on the region (Hypo, 20 min; Cb, 1 h; F C X , 3 h). The effect of D O N on endogenous levels of the D A metabolites, D O P A C and H V A p r o d u c e d a m o r e variable response. Both amines b e c a m e highly elevated in the Hip for the initial 3 h, although there w e r e no significant changes (P > 0.05) of either D A or N E in this region. In the F C X , though, while both metabolites displayed an initial d e c r e a s e ( D O P A C , 57%; H V A , 42%) at 20 min (as did DA), the levels rebounded a b o v e baseline concentrations by 1 and 3 h, suggesting increased metabolism or utilization of D A which also concurrently increased b e t w e e n 1-3 h post-dosing. D O P A C levels in the hypothalamus and c e r e b e l l u m w e r e unaffected, whereas hypothalmic H V A s h o w e d a v e r y significant (P < 0.001) initial d e c r e a s e which quickly returned to control levels by 1 h. Cerebellum H V A showed a singular increase at 3 h (P < 0.01) which faded by 8 h.
38
D . B . Prelusky et al.
Table 1. Effect of deoxynivalenol on pig brain amine levels in selected regions a Brain region Time (h)
Neurotransmitter and metabolite levels b NE
DA
DOPAC
HVA
5HT
5H1AA
377.6 758.4 ***e 302.4 462.5** 272.5 567.7*** 429.9 502.8 365.6 230.9 37.2
179.3 101.3"~ 153.3 167.7 191.0 190.4 164.6 169.9 258.4 228.8 25.0
200.8 144.1 165~6 221.1 177.0 233.4 178.3 248.7 144.5 180.9 24.1
683.5 259.6ttt 521.8 522.7 530.8 428.6 528.5 536.1 250.0 382.7 58.8
218.8 225.6 170.0 269.1"** 227.7 216.7 283.2 172.0t~t 285.6 232.3 17.9
290.0 284.9 268.3 314.8 325.5 396.2 382.6 360.8 258.4 188.2 35.5
Hypothalamus .33 ~ a 1c d 3c a 8¢ a 24 ~ d SEM f Cerebellum .33 c o 1c d 3~ o 8c o 24 e d SEM Frontal cortex .33 c a 1c a 3~ a 8c a 24 c a SEM Hippocampus .33 ¢ d 1c d 3c d 8c a 24 ~ d SEM Ports/Medulla .33 c a 1~ a 3c a 8c a 24 ~ d SEM
154.3 183.8** 134.9 152.0" 132.9 160.8"* 146.3 135.6 170.1 152.3 8.9
5.1 2.7tt 5.5 2.5t? 5.7 4.0 5.0 7.2* 4.4 5.0 0.80
4.9 6.2 3.7 2.7 3.6 4.8 5.2 5.6 4.2 4.8 1.1
17.8 21.1 17.5 22.3 20.0 29.8* 29.0 23.9 24.8 21.3 2.8
169.1 216.2'* 159.7 198.5"* 171.6 223.1" 182.4 165.3 170.9 152.6 12.5
8.0 4.2t 11.9 6.5t 13.7 7.8tt" 9.3 10.0 10.4 7.4 1.5
18.5 7.9tt 14.4 25.4** 11.3 19.6" 12.4 12.9 15.9 17.8 2.1
47.0 27. l~t 39.5 54.8* 37.1 54.3* 52.8 35.6 35.4 26.9 5.3
80.1 66.0 76.4 74.9 77.3 75.3 93.7 58.2tt'~ 71.3 56.2 6.5
82.9 47.4ttt 73.7 72.9 63.6 76.3 83.8 80.2 72.1 66.3 6.2
10.6 9.7 7.2 15.5"** 9.5 16.8"** 11.4 9.6 10.3 9.8 0.98
51.8 81.3"* 39.0 79.8*** 54.0 95.3*** 67.1 66.4 53.5 62.4 13.5
50.5 38.5 50.1 66.5 66.3 73.2 83.0 91.5 60.2 37.8t 5.9
105.4 146.5 148.2 184.1 188.4 156.5 163.2 t53.5 153.7 136.9 28.8
46.4 37.5 29.7 43.8 39.7 38.0 25.9 21.0 28.7 24.1 6.2
66.4 72.2 56.7 88.3* 73.9 90.2 63.4 45.6 56.6 57.2 12.2
126.2 155.7 119.6 150.8 136.3 137.0 158.3 145.9 139.7 120.1 10.8 254.3 214.4 196.2 270.6 267.1 238.6 239.2 168.7 184.3 193.5 35.3
5.7 7.3 6.0 8.2 9.2 8.0 9.5 7.4 8.9 7.6 0.88 18.2 16.8 17.5 24.6 16.4 19.1 18.7 19.8 17.3 24.5 3.3
nd g
197.9 179.5 189.0 199.1 211.3 208.7 201.0 135.8 183.9 134.5 32.9
26.5 20.2tt 26.5 18.8tt 24.8 19.3t 29.2 33.3 17.8 19.9 2.1
181.0 147.3 154.2 140.5 152.9 174.5 164.5 128.5 130.1 106.6 37.7
a IV dose, 0.25 mg DON/kg body weight b n g amine/g brain tissue, mean, n = 3 or 4 animals (duplicate determinations). N E = norepinephrine; DA = dopamine; DOPAC = 3,4-dihydroxyphenyl-acetic acid; H V A = homovanillic acid; 5HT = 5-hydroxytryptamine; 5 H I A A = 5-hydroxyindoleacetic acid c Control animals d Dosed animals Significantly different from the control group: increase; *P < 0.05; **P < 0.01 ; ***P < 0.001: decrease; t P < 0.05; ~tP < 0.01 ; t t t P < 0.001 f SEM = standard error of mean g n d = not detected
Effect of Deoxynivalenol on Neurotransmitters Indoleamine Profiles
The effects of DON on indolamine levels were also region specific (Table 1). Hypothalamic serotonin (5HT) concentrations showed a large increase (58%) at 1 h post-dosing (P < 0.001), which subsequently dropped below control levels (39%) by 8 h (P < 0.001). At 8 h, there was also a corresponding decrease (38%) in cortical 5HT levels (P < 0.001), which like Hypo 5HT returned to control levels by 24 h. There was essentially no change from 5HT control values in the hippocampus and pons/medulla. The 5HT levels in the cerebellum were too low to quantitate. The 5HIAA, a major metabolite of 5HT, showed an initial rapid decrease in both the Cb and FCX, but while cortical 5HIAA quickly rebounded to control values, levels in the cerebellum remained depressed for the 3 h post-dosing. There were no significant (P > 0.05) changes measured in 5HIAA concentrations in the hypo, hip, or P/M regions.
Discussion This study investigated the effects of DON on brain biogenic amine levels following a single IV administration of the toxin to swine, which are considered a species highly sensitive to the feed refusal and emetic activities of trichothecenes. The effects observed varied markedly with time and brain region for the different transmitter measured. It was evident that there were some fluctuations in baseline (control) metabolite concentrations as samples were collected throughout the day (0910-1640 h). With most transmitters the change during this period was moderate (up to ---20% average daily levels), the one exception being hypothalmic HVA in which a very large variation was noted (250683 ng/g). These data suggested circadian rhythm in brain amine concentrations occurs and/or variation induced by individual animals' response to experimental stress (i.e., handling, injection, crowding), both of which have been shown to influence turnover rates of specific catecholamines or indoleamines. Considering the multiple mechanisms which are involved in maintaining CNS function, various authors have reported neurotransmitter systems are subject to biorhythms, and that these patterns can be altered by disruption of normal behaviour, (i.e., stress, fasting, light/dark cycle, exercise, etc.) (Davis 1989; Karczmar 1978). In several studies investigating the effects of mycotoxins on biogenic amines, the data has also shown that control levels varied with time, depending on species and compound measured (Ahmed and Singh 1984; Boyd et al. 1988; Chi et al. 1981; Fitzpatrick et al. 1988b; Porter et al. 1988). In the current study, no statistical evidence was found to suggest that the time of dosing seriously impacted on the results. In rats it has been shown that there is rapid depletion of NE in most brain regions accompanying both physical and psychological stress (Iimori et al. 1982). In the present study, a similar trend was observed in control animals following injection of the vehicle only (although the effect of circadian rhythm still could not be ruled out). Ball (1988) found that pigs judged to be more excited or stressed just prior to slaughter had significantly lower hypothalamic concentrations of all the neurochemicals measured (NE, 5HT, 5HIAA, HVA), and Hallberg et al. (1983) noted that stress-
39 susceptible pigs had lower brain amine levels compared to more stress-tolerant strains. Although available data indicate that certain neuroregulators are involved in the feeding process, information is still too limited and variable to predict the effects of certain anorexic-inducing xenobiotics on neurochemical patterns. These chemicals can reduce feed intake by a variety of different mechanisms, affecting feeding time, intervals between meals, intake amount, rate, and meal preference (i.e., protein, carbohydrate, fat, etc.) (Davis 1989). Prior evidence has suggested that important aspects of feeding may be controlled by 5HT in the brain and periphery; elevated levels causing a loss of appetite (Shor-Posner et al. 1986). This was possibly reflected in the current study where hypothalamic 5HT exhibited a large increase at 1 h post-dosing, but did not account for its subsequent depletion in both the hypotbalamus and cortex at 8 h. It was not readily apparent whether this abrupt decrease was due to the increased utilization of 5HT or to a negative feedback in 5HT synthesis in response to its earlier enhanced release. However, the ratio of 5HIAA/5HT which can be used as an index for 5HT metabolism or utilization, was increased in both the Hypo (57%) and FCX (53%) at 8 h. This indicated an enhanced utilization of 5HT which could have an anorexic consequence. Changes in NE and DA levels following DON administration, though, were not consistent with current opinion on anorexia (Hoebel et al. 1989; Leibowitz 1989). Both elevated NE and depressed DA levels are associated with stimulation of the feeding behaviour, opposite to the effect expected with DON. Consequently, it is probable that the catecholamine imbalances observed here were not due to the anorexic activity of DON, but instead were secondary to other toxicological responses produced by the acute dose of the mycotoxin, possibly its emetic activity. Smith and MacDonald (1987) demonstrated that acute administration of the Fusarium toxin, fusaric acid, to swine caused significant increases in hypothalamic NE and 5HT, and cortical NE; the same pattern as reported here for DON. The function of neuroregulators in the vomiting action is not well understood. It has been reported that both NE and 5HT probably have a strong role in relaying information during the process (Carpenter et al. 1983; Schwartz et al. 1986), and dopamine, an emetic itself, appears to have the greater involvement in initiating the emetic reflex. Similarly, DA has been implicated as a mediator of learned aversion. Hoebel et al. (1989) reported that brain DA levels decreased when rats were exposed to substances perceived to cause illness. Thus, in the current study, decreased DA levels which occurred early in pigs may reflect a negative response to DONinduced nausea and malaise. Possibly this is accomplished through enhanced metabolism of DA to N E and other metabolites (DOPAC, HVA). The absence of longer-term effects may reflect a short toxicological half-life for DON. It has been shown that up to 75% of DON administered IV to swine was eliminated within 6 h (Prelusky et al. 1988), and essentially all regional amine levels in the present study (except for 5HT), had returned to control levels by 8 h post-dosing (Table 1). This showed that a single sublethal dose of the toxin, although quickly symptomatic, was limited in duration. In comparison, Fitzpatrick et al. (1988a) reported that 5HT and 5HIAA levels were still elevated in all brain regions of the rat 24 h after oral DON
40
administration, but the dosage used was 10 times that in the current study. Similarly, prolonged alteration in various monoamine concentrations have also been reported in rats and chickens following oral administration of lethal amounts of T-2 toxin, a related but more acutely toxic trichotecene (Boyd et al. 1988; Chi et al. 1981; MacDonald et al. 1988). In these latter studies, however, doses were sufficient to produce mortality, and the effects on brain amine levels were probably due to the overall chemical assault rather than any selective anorexic activity. It has been postulated that the effect of D O N and other trichothecenes on feeding behaviour is regulated through alterations in the availability of specific brain neurotransmitters. In the current study, it was shown that D O N administered IV to swine caused a marked change in certain regional monoamine concentrations, but this data, in conjunction with other published studies clearly demonstrates the difficulties in trying to establish a relationship between the neurochemical profile and the feed refusal activity of xenobiotics. Numerous internal, as well as environmental factors can influence the signals mediating behavior, and feeding can be either enhanced or inhibited. For example, Ball et al. (1988) showed that while it was possible to elevate hypothalamic 5HT levels in pigs with a tryptophan supplemented diet, which should accordingly decrease appetite, it, in fact, had no significant effect on feed intake or daily weight gain. Further studies are needed to fully understand whether the anorectic mechanism of action of D O N is neurotransmitter and/or second messenger-specific, or a secondary consequence of other pharmacological effects.
References
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D.B. Prelusky et al. Hallberg JW, Draper DD, Topel DG, Altrogge DM (1983) Neural catechoalmine deficiencies in the porcine stress syndrome. Am J Vet Res 44:368-371 Hoebel BG, Hernandez L, Schwartz DH, Mark GP, Hunter GA (1989) Microdialysis studies of brain norepinephrine, serotonin, and dopamine release during ingestive behavior In: Schneider LH, Cooper SJ, Haimi KA (eds) The psychobiology of human eating disorders: Preclinical and clinical perspectives. Annals New York Academy Sciences, NY, pp 221-233 Iimori K, Tanaka M, Kohno Y, Ida Y, Nakagawa R, Hoaki Y, Tsuda A, Nagasaki N (1982) Psychological stress enhances noradrenaline turnover in specific brain regions in rats. Pharmacol Biochem Behav 16:637-640 Karczmar AG (1978) Multitransmitter mechanisms underlying selected function, particularly aggression, learning, and sexual behavior In: Deniker P, Radouco-Thomas C, Villeneuve A (eds) Neuropsychopharmacology, Pergamon Press, Oxford. pp 581608 Leibowitz SF (1985) Brain neurotransmitters and appetite regulation. Psychopharmacol Bull 21:412-418 - (1989) Hypothalamic neuropeptide Y, galanin, and amines: Concept of coexistence in relation to feeding behavior. In: Schneider LH, Cooper SJ, Haimi KA (eds) The psychobiology of human eating disorders: Preclinical and clinical perspectives. Annals New York Academy of Sciences, NY, pp 171-191 MacDonald EJ, Cavan KR, Smith TK (1988) Effect of acute oral doses of T-2 toxin on tissue concentrations of biogenic amines in the rat. J Anim Sci 66:434-441 Miller JD, Arnison PG (1986) Degradation of deoxynivalenol by suspension cultures of the Fusarium head blight resistant wheat cultivar Frontana. Can J Plant Pathol 8:147-150 Porter JK, Norred WP, Cole RJ, Dorner JW (1988) Neurochemical effects of cyclopiazonic acid in chickens. Proc Soc Exp Biol Med 187:335-340 Prelusky DB, Hartin KE, Trenholm HL, Miller JD (1988) Pharmacokinetic fate of 14C-labelled deoxynivalenol in swine. Fundam Appl Toxicol 10:276-286 Schwartz J-C, Agid Y, Bouthenet M-L, Javoy-Agid F, LlorensCortes C, Martres M-P, Pollard H, Sales N, Taquet H (1986) Neurochemical investigations into the human area postrema In: Davis CJ, Lake-Bakaar GV, Grahame-Smith DE (eds) Nausea and vomiting: Mechanism and treatment. Springer-Verlag, NY, pp 18-30 Shor-Posner G, Grinker JA, Marinescu C, Brown O, Leibowitz SF (1986) Hypothalamic serotonin in the control of meal patterns and macronutrient selection. Brain Res Bull 17:663-671 Smith TK, MacDonald EJ (1987) Effect of an acute dose of fusaric acid on brain neurochemistry in swine. Can J Anim Sci 67:1182 (abstract) Trenholm HL, Friend DW, Hamilton RMG, Thompson BK, Hartin KE (1986) Incidence and toxicology of deoxynivalenol as an emerging mycotoxin problem. Proc VI Int Conf Mycotoxicosis, pp 76-82 Ueno Y (1986) Trichothecenes as environmental toxicants. Reviews Environ Toxicol 2:303-341 - (1977) Mode of action of trichothecenes. Pure Appl Chem 49:1737-1745 Vesonder RF, Hesseltine CW (1981) Vomitoxin: Natural occurrence on cereal grains and significance as a refusal and emetic factor to swine. Proc Biochem 16:12-15 Wester P, Gottfries J, Winblad B (1987) Simultaneous liquid chromatographic determination of seventeen of the major monoamine neurotransmitters, precursors and metabolites. II. Assessment of human brain and cerebrospinal fluid concentrations. J Chromatogr 415:275-288 Manuscript received May 16, 1991 and in revised form August 28, 1991.
