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The assessment of the environmental impact of pollutants at the ecosystem level raises theoretical and methodological problems of substantial magnitude. Despite the usefulness of a more reductionist approach related to the measure of the effects of pollutant on populations, such as specific toxicity testing or--regarding the assessment of impact of chronic exposure--the use of various biomarkers, these measures do not provide data relevant to conclusions at the community and ecosystem level. The assessment of effect on entire ecosystems requires two kinds of information: (1) What are the consequences of pollutant exposure on the community structure, and (2) what is its impact on fundamental ecological processes that control the ecosystem functioning? Disturbances of structure may be appraised in terms of the major parameters that currently describe it at the community level, such as species richness, ecological diversity, and dominance. The assessment of pollutant effect on ecosystem biodiversity, though underestimated for a long time, is a major issue in applied ecotoxicological research. Accurate attention must be given to keystone species, which in both terrestrial and aquatic ecosystems control the diversification of the whole trophic web in a community. Therefore, if induced by a given pollutant, the decline of the populations of a keystone species may lead to a major destructing of the community. The second major issue in ecotoxicological research is assessment of damage to ecosystem functioning. Because productivity is of such practical importance, appraising changes in primary and secondary productivity is acutely needed. Indeed, chronic pollutant exposure is a permanent threat to biological natural resources as it impedes their production and renewal. Another major point in the assessment of effects at the ecosystem level is the effect of pollution on natural biogeochemical cycles, as well as the study of the ways through which a number of various and often common contaminants, some of them acting at global scale, interact with such cycles. It is also fundamental to assess the effects of pollutants on decomposer activity in both soil and natural waters and on its interaction with biogeochemical processes such as element recycling. In a larger context, though still almost unexplored, ecotoxicology is related to assessment of effects on complex ecosystem assemblages on a regional scale (i.e., landscape ecology and its relationship to functional ecotoxicology).

Outdoor artificial ponds (mesocosms) of 12 m 3 were designed for long-term ecotoxicological studies. Sediment, macrophytes ( Typha angustifolia and Elodea canadensis ), and free and caged freshwater snails [ Lymnaea palustris (Muller)] and wood lice ( Asellus aquaticus L.) were collected in nearby natural ecosystems and introduced in the mesocosms. Sixty goldfish ( Carassius auratus L.) were caged in each pond. Introduced species developed and reproduced in every mesocosm. Animal species (mainly insects and amphibians) spontaneously colonized the ponds, developed, and reproduced. The resulting communities qualitatively resemble those living in natural lentic systems in the surrounding area. Homogenity in physical and chemical conditions and in abundance of phytoplanktonic, periphytic, and macroinvertebrate communities between the different mesocosms was assessed during the stabilization period (8 months). Except for periphyton biomass, no divergent evolution was observed between the ponds. Mesocosm water was slightly eutrophic, alkaline (mean pH: 8.47 ± 0.09), and moderately hard and mineralized. The homogenous and realistic environmental conditions and high ecological representativity of the outdoor experimental ponds were suitable for extensive ecotoxicological studies. Considerations on the choice and origin of introduced species and on possible interactive effects of the transfer of organisms from natural environments, maintainance conditions, and pollutant exposure are discussed.

Common terns (Sterna hirundo) exhibit a remarkable range of variation in reproductive success. Several factors are known to contribute to reproductive failure either before hatching or between the time of hatching and fledging : predation pressure, food availability, flooding, competition for nesting sites, and toxic chemicals. Contaminants such as organochlorine pesticides, polychlorinated biphenyls (PCBs), polychlorodibenzodioxins (PCDDs), polychlorodibenzofurans (PCDFs), mercury and selenium were proved to significantly impair tern reproduction. During the reproductive period of 1988, an important mortality of common terns was observed in french colonies around the river Rhine. Approximately 50% of the young died a few days after hatching. The objective of the present study was to determine whether the intoxication by chlorinated compounds could have been responsible for the observed reproductive failure. 25 refs., 1 tab.

Functional development of the adrenocorticotropic axis was inferred from plasma ACTH and corticosterone levels in intact and embryonically bursectomized (BFX) embryos and chicks from 8 days before to 56 days after hatch. Bursectomy was surgically made at 80 h of incubation and resulted in various alterations in developing adrenocorticotropic axis: ether stress-induced hormonal stimulation could be detected more precociously in BFX (day-6) than in intact embryos; the non stress-responsive period of newly hatched controls did not appear in BFX chicks; BFX young adult chicken exhibited quite smaller responses to stress than controls. In ovo injection of bursin (Lys-His-Gly-NH2) to 6- and 9-days old BFX embryos could restore normal adrenocorticotropic development provided convenient doses of tripeptide were used: administration of 100 fg or 100 pg of bursin was effective to restore normal hormonal levels at all stages studied whereas 100 micrograms was effective at embryonic stages only. The tripeptide Lys-His-Gly-NH2 is suggested as a possible signal from the immune B system directed at the hypothalamo-hypophysial-adrenocortical axis.

Plasma corticosterone levels were determined by protein-binding assay at 2- and 5-min intervals after systemic or neurogenic stress. The same corticosterone (B) profile was observed after hemispherectomy as in intact pigeons, exhibiting episodic increase including several successive peaks after stress application. Hypothalamic stimulation of ACTH injection resulted in a simpler adrenocortical response, analogous to the first peak of the response to stress. Postero-lateral deafferentation of the hypothalamus unabled neurogenic stress to promote any plasma B increase and significantly altered the response to systemic stress since only the first peak of B occurred. Such a single peak response to both systemic and neurogenic stresses was observed after surgical interruption of neural connections between hypothalamic and thalamic structures without interferring with posterior afferents to the basal hypothalamus. The role of thalamic-hypothalamic interrelationships in modulating the stress-induced adrenocorticotropic activity is discussed.

In this paper are discussed the major ecotoxicological criteria that are presently in current use for the detection of environmental pollution and in the assessment of its biological impact. Additionally to the widespread use of bioindicator species, more recent criteria relying on biocoenotic indexes have been proposed. Other methods, more complex are intended to compare the effects of a given pollution on communities structure. They stem from the computation of importance value curves or even on the factorial analysis of correspondence. Ultimately, the determination of the variations in primary and (on) secondary productivity may be achieved in order to appraise accurately the impact of a given ambient contamination on the productivity of the affected communities.

Xenopus spawns were immersed in 0.5, 1, and 2 ppm Lindane (γ-BHC) in tap water. The highest concentration caused a decrease of hatching rate. Tadpole mortality was investigated until metamorphosis. Close to zero through the first week, it showed a slow increase correlated with Lindane concentration throughout the period preceding metamorphosis. The development of surviving tadpoles from hatching to metamorphosis was 4 weeks longer than in controls. Moreover the average weight was lower in the treated groups. Exposed animals frequently displayed morphological abnormalities: lateral deviation of the body axis and a dorsal bump of the cranial area were observed in tadpoles, whereas darkening of pigmentation appeared in tadpoles as well as in surviving adults. The symptoms of intoxication are not specific to Lindane. Neurotoxic pesticides produce the same kind of effects. Our results suggest: a changed endocrine function of the hypothalamo-hypophyseal axis which regulates growth and metamorphosis, and a dysfunction of the intermediate lobe of the hypophysis which controls pigmentation.

La symptomatologie de l'intoxication par les insecticides a surtout fait l'objet de descriptions. Des resultats plus precis et plus inTEressants peuvent etre obtenus par l'emploi de sysTEmes actographiques perfectionnes. Nous presentons ici des donnees relatives a l'intoxication de la Mouche domestique par divers insecticides de contact: DDT, Lindane, parathion, Dipterex®, fenthion, Baygon®, carbaryl, Mesurol®, Temik®. Les enregistrements ont eTE obtenus avec un actographe electro-acoustique de grande sensibiliTE. Les resultats montrent que les symptomes de l'intoxication par des insecticides appartenant a des familles chimiques differentes se deroulent selon des modaliTEs caracTEristiques, ce qui doit etre en rapport avec des voies ou modes d'action sensiblement differents.

The corpora allata of adult and 5th instar nymphs of the migratory locust exhibit several ultrastructural alterations after lindane intoxication. The type of cell degeneration observed here is very similar to ultrastructural changes obtained after the administration of precocene.

Brain homogenates ofLocusta migratoria are found to possess enzyme capable of catalyzing the N-acetylation of tryptamine. A main product of the enzymatic reaction is isolated and identified as N-acetyltryptamine by chromatography analyse. Both insecticides, chlordimeform and lindane, inhibit enzyme activity. A direct correlation between the degree of intoxication and acetylation of tryptamine is described.

Acute intoxication of adult female rats by an injection of 15 mg of Lindane or 25 mg of 2-4-5 T induces, in one hour, as decrease in the adrenal epinephrine level and an increase in the urinary excretion of this hormone. These modifications are absent after chronic intoxication by these pesticides added in the food (60 and 100 ppm respectively), under normal, hypo- and hyperlipidic diet.

It can be assumed from previous data that the stress-induced polyphasic adrenocortical response involves two phenomenons. First, direct hypothalamic afferents stimulate the adrenocorticotropic axis resulting in a rapid increase in plasma corticosterone levels up to a maximum of about 40 ng . ml-1 at 15 min. Then, a thalamic-hypothalamic loop generates the late rebounding components. Intravenous injection of muscimol (0.5 ml . kg-1) produces only slight and short duration (45 min) disturbances (piloerection; panting; corticosteronemia). Stress application, 1 hr after muscimol administration, elicits only the first peak of corticosterone with magnitude and delay similar to that in control. However, the late rebounding component is no more visible. Thus, GABA-ergic stimulation leads to a situation similar to that seen after section of neural connections between thalamic nuclei and infundibular complex. The hypothalamic-adrenocorticotropic reactivity itself is not affected whereas the function of the regulating thalamo-hypothalamic loop is markedly impaired.

The effect of bursa of Fabricius on the endocrine function of the chick testes was studied in vivo by comparing plasma testosterone levels from 48 h before hatch to 16 weeks of age in both intact and bursectomized chicken. Early bursectomy was performed at 80 h of incubation. Post surgery survival was low (12% at 1 week). In controls, plasma testosterone levels were found to be low (100-200 pg. ml-1) from 48 h before to 48 h after hatch, then to raise up to a plateau (2,200 pg. ml-1) at 6 weeks. After bursectomy, values were first higher than in intact (210-440 pg. ml-1 from 48 h before/after hatch and 515 vs 300 pg.ml-1 at 3 days) but no difference could be further detected from 1 to 16 weeks of age. It is suggested that, in addition to the effect of androgen on bursa of Fabricius, the later reciprocally influences the gonadotropic axis during the early stage of development.

Kinetic evaluation of the adrenocortical activation induced after hypothalamic stimulation was obtained by determining plasma corticosterone levels (protein binding assay) at 2, then 5 and 10 min intervals for 100 min. Chronic catheterization and an electronic miniature device for hypothalamic telestimulation allowed for avoiding any environmental disturbance. Plasma corticosterone was rapidly increasing up to a peak (x 4 basal initial level) at 14 min after stimulating the hypothalamus for one sec. Corticosteronemia was then decreasing to resting level within one hour. This response was approximately the same in intact as in thalamic pigeons.

Early embryonic bursectomy (BFX) disturbed the adrenocortical functioning. The stress-unresponsive period that occurred in controls, and lasted for 2-3 weeks after hatching, no longer appeared in BFX chicks. In contrast, the magnitude of the stress-induced hypercorticosteronemia was much lower in BFX than in sham-operated 5 week-old chicken. It was assumed that such adrenocortical dysfunction was due to bursal deprivation, since grafting bursal buds onto the chorio-allantoic membrane of BFX embryos restored all the parameters under study, i.e., the post-hatching stress unresponsive period and the high magnitude of stress-induced responses in adults. Factor(s) involved in such interregulation are not known but do not seem to affect directly adrenocortical cells because intramuscular injection of a moderate dose of ACTH resulted in the same hypercorticosteronemia whether 3 day-old and 5 week-old chicks had been bursectomized or sham-operated.

Plasma levels of both adrenocorticotropic hormone (ACTH) and corticosterone (B) were determined in embryos (day 15 of incubation), chicks (day 3 after hatch) and young chickens (8 weeks). Experimental animals were bursectomized at 80 hr of incubation, i.e., before any anlage of the bursa of Fabricius could develop. Bursectomized (BFX) animals were compared to sham-operated controls (T), in basal, resting condition and 7 (ACTH) or 14 min (B) after ether stress was delivered for 30 sec. Basal B and ACTH levels seemed not to be significantly modified in BFX embryos, chicks and chickens. Hypophysial and adrenocortical response to stress appeared more precociously in BFX embryos (day 15 of incubation) than in intact ones (day 19). The non stress-responsive period that was observed for one week after hatch of T birds did not appear in 3-day-old BFX chicks whose both B and ACTH stress-induced levels were as high as in intact adults. In contrast, adrenocortical and pituitary corticotropic responses to stress were markedly impaired (by 50%) in adult BFX chickens as compared to intact controls.

The thalamic pigeon was taken as a model to investigate mechanisms of adaptation to chronic intermittent stress. Muscimol was infused into the 3rd ventricle at the rate of 0.25 microgram.hr-1 by means of an osmotic minipump. In controls, pumps were filled in with saline. Animals were placed under constant light regimen and electrical foot shocks were delivered, thrice a day, at fixed time interval following a short dark period (3 min). In controls, adaptation of the adrenocortical response to chronic intermittent stress was achieved within 7 days, including both attenuation of the post-stress hypercorticosteronemia and occurrence of a pre-stress conditioned peak of corticosterone. The anticipating conditioned constituent subsisted in muscimol treated animals whereas no attenuation of the post-stress peak could be detected. GABAergic chronic treatment also resulted in decreased basal plasma corticosterone levels. In controls, extinction of the conditioned endocrine response was not obtained after conditioning stimulus (dark period) was presented alone for 2 weeks.

Stress application elicited successive attenuating peaks of plasma corticosterone level occurring 14, 35, 60 and 90 min. later. Late peaks were suppressed after bilateral electrolytic lesions were placed in the nuclei dorsomedialis anterior thalami, resulting in a single peak (the first one) of corticosteronemia after stress application. It is suggested that reverberating adrenocortical response to stress depends upon interrelationships between the adrenocorticotropic hypothalamus and the anterior medio-dorsal thalamus.

The functioning of the hypothalamic pituitary-adrenocortical system was tested in young pigeons before and after hatching, by radiocompetitive assay of plasma corticosterone (B) at rest and under stress. 1 degree B was expressed in ng/ml. Resting adult values (8-10) could not be observed before the birds were 6 week-old. B levels were very low 36 h before and 24 and 48 h after hatching (3-5). Then, they progressively increased. 2 degrees B concentrations soared during the 24 h immediately preceding hatching to rise to 45 ng/ml at the time of hatching. After that, corticosteronemia rapidly decreased. 3 degrees Neurogenic stress was applied by means of electrical stimulation of the led skin for 15 sec. Ten to 12 min later, B rose markedly (30 ng/ml) in 6 week-old pigeons as well as in adult ones. No response could be observed from 36 h before to 24 h after hatching. Significant adrenocortical activation was obtained in 3-7 day old squabs (18 ng/ml). 4 degrees Systemic stress (ether vapor) led to significant hypercorticosteronemia (30-40 ng) in pre-hatching embryos. No response could be observed for 24 h after hatching. Ether-induced adrenocortical activation appeared again in 2-3 day-old pigeons. 5 degrees The development of both resting and stress-altered corticosteronemia did not seem to be related to the increase in body weight of young birds. One can present the hypothesis that a strongly stressfull situation occurring during the period which immediately precedes hatching, results in a rapid activation of the adrenocortex. The period following hatching corresponds to the "non stress responsive period" of mammals and could be explained by some feedback mechanisms.

Chronic catheterization and miniature recording device allowed plasma corticosterone (B) and hypothalamic multiunit activity (MUA) to be simultaneously obtained from freely behaving, awake pigeons, before and for 4 hrs after intravenous injection of metyrapone. Injection of vehicle (tartric acid : 100 mg/4 ml/kg) led to MUA and B profiles quite similar to stress-induced responses, i.e., a rapid and sustained rebounding increase in hypothalamic firing rate and, shifted by 5-10 min, in plasma B. These responses were progressively attenuating for 90-120 mn. Metyrapone administration induced first a rapid and short MUA and B increase. Then both parameters drastically decreased near zero for about 2 hrs and were slowly restored to initial values within 3-4 hrs. It is suggested that metyrapone treatment inhibited both peripheral (B synthesis) and central (hypothalamic neurons) levels of the corticotropic axis.

Plasma thyroxine and corticosterone levels were determined by competitive protein binding assay, at 3 hr intervals, throughout the photoperiod. Pigeons were kept in controlled environment (21 +/- 1 degree C; 14L6-20: 10D). Intact controls exhibited low thyroxine (T4) and corticosterone (B) levels for the light phase of the photoperiod. Values were rising during the night, up to a peak at 03 hr. Electrolytic lesions were placed bilaterally in either the nucleus anterior medialis hypothalami or the n. preopticus, or the n. supraopticus. Circadian rhythms of both T4 and B were markedly altered in all lesioned pigeons, with a shift of very high T4 values to the morning times and a complete disorganization of B patterns, with very heterogeneous values. The possibility is raised that anterior hypothalamic formations participate in the endogenous oscillator circuitry in birds.

A short and moderate increase in plasma ACTH and corticosterone levels was observed after i.v. injection of the GABA-agonist, muscimol as a bolus of 0.5 mg . kg-1. Doses lower than 0.3 mg . kg-1 were ineffective. 6 hrs. after i.v. injection of 1 ml of an undiluted immune serum raised against oCRF41, resting hormone levels were markedly diminished, stress-induced responses were almost completely suppressed, whereas i.v. injection of muscimol still resulted in an unaltered adrenocorticotropic stimulation. Adenohypophysectomized animals displayed a low basal adrenal function, no stress-induced response and interestingly enough no longer responded to i.v. muscimol injection. Consequently, acute GABA-ergic stimulation appears to act directly onto the pituitary corticotroph cells. It cannot be inferred so far whether this action is of physiological importance but it can at least contribute to elucidate some controversial data prevailing in this field.

Biochemical markers are early warning systems of the exposition of organisms to pollutants. Among them, the induction of the fish EROD activity was successfully used in many studies. However, many confounding factors may affect the EROD response. In addition, a better knowledge of EROD baseline levels is necessary so as to assess to which extent an impact occurred. The measurement of the fish EROD induction is a suitable biomarker even if complementary studies are necessary for a better intepretation of the results., La mesure de l'induction de l'activité EROD chez les poissons est désormais bien validée et utilisée largement. Ce biomarqueur permet de mettre en évidence des situations polluées variées. Cependant il s'agit de bien prendre en compte les facteurs naturels de variabilité de cette réponse biologique (maturité sexuelle, _). Pour autant, il est possible d'engager la normalisation de cette méthode.

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The use of biochemical biomarkers among fishes to monitor the contamination and its impact on aquatic hydrosystems is an essential complement of chemical supervision programs. The induction measurement of the EROD activity among fishes has been widespread for about twenty years and has allowed to highlight various types of pollution. However, the natural variability factors of this biological response must be taken into account. Finally, the studies comparing this early response to other biological variables are to be pursued. It is urgent to perform as soon as possible studies of intercalibration of the method in order to favour its development in pre-routine use., L'utilisation de biomarqueurs biochimiques chez les poissons pour un suivi de la contamination et de ses impacts sur les hydrosytèmes aquatiques est un complément indispensable des programmes de surveillance chimique. La mesure de l'induction de l'activité EROD chez les poissons est désormais répandue depuis une vingtaine d'année et a permis de mettre en évidence des situations polluées variées. Cependant il s'agit de bien prendre en compte les facteurs naturels de variabilité de cette réponse biologique. Enfin les études comparant cette réponse précoce à d'autres variables biologiques sont à poursuivre. Pour autant, il est urgent de conduire dès à présent des études d'intercalibration de la méthode afin de favoriser son développement en utilisation pré-routinière.

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chap. 10; National audience

chap. 12; National audience

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chap. 2; National audience