Evaluation of neurobehavioral abnormalities and immunotoxicity in response to oral imidacloprid exposure in domestic chickens (Gallus gallus domesticus)

ABSTRACT

Domestic chickens (Gallus gallus domesticus) were exposed to imidacloprid by gavage once daily for 7 consecutive days at 0, 0.03, 0.34, 3.42, 10.25, and 15.5 mg/kg/day (n = 20 per group; 5 6-week-old males, 5 6-week-old females, 5 9-week-old males, and 5 9-week-old females). The severity and duration of neurobehavioral abnormalities were recorded. Components of the innate and adaptive immune system were assessed with 7 standard functional assays. Temporary neurobehavioral abnormalities were observed in a dose-dependent manner, including muscle tremors, ataxia, and depressed mentation. Based upon mean clinical severity scores, the no observed adverse effect level (NOAEL) was 3.42 mg/kg/day, and the lowest observed adverse effect level (LOAEL) was 10.25 mg/kg/day. The effective dose value for the presence of any neurobehavioral abnormalities in 50% of the test group (ED₅₀) was 4.62 ± 0.98 mg/kg/day. The ED₅₀ for an adjusted score that included both severity and duration of neurobehavioral abnormalities was 11.24 ± 9.33 mg/kg/day. These ED₅₀ values are equivalent to a 1 kg bird ingesting 29 or 70 imidacloprid treated soybean seeds respectively. Immunotoxicity was not documented, possible causes include the assays were insensitive, relevant immune functions were not examined, or imidacloprid is not immunotoxic at this dosing schedule in this species. Neurobehavioral abnormalities were a more sensitive indicator of the sublethal effects of imidacloprid than immunotoxicity.     

Learning to Observe Relationships and Coping

Milieu relationships provide the critical background presence to staff's attempts to motivate, regulate, and teach patients how to cope with stress. Forging a connection with hospitalized children and adolescents demands attention to how they respond to adults and engage with staff around milieu expectations. Assessment guides that deal with these issues are presented. Important aspects of children's relatedness are presented in the context of their working models of adults and the influence of these representations on their response to staff. Coping skills are explained with particular emphasis on behavioral coping strategies. Tied to the assessment process are interventions that emphasize staff's role in helping patients manage strong affects and avoid the use of nonproductive behavior regulation strategies.     

Follistatin and activin A production by the male reproductive tract

Follistatin is a binding protein for the activin and inhibin family of hormones, regulating their biological activity. In the male reproductive tract, the interaction of these factors is likely to be involved in the regulation of the proliferation of several cell types. We have investigated the presence of follistatin and activin A in seminal plasma using specific immunoassays and have localized follistatin and activin/inhibin subunits in the adult human testis, prostate and seminal vesicle to establish their likely sources. High concentrations of immunoreactive follistatin were present in seminal plasma in normal men (mean 97.9 ng/ml; 1.43 ng/ml in peripheral plasma) and were similar in men with oligo/azoospermia and following vasectomy. Follistatin immunoreactivity was localized to both Leydig and Sertoli cells of the testis, and to epithelial cells of the prostate gland and seminal vesicle, which are likely to be the predominant sources of the hormone in seminal plasma. Activin A was also present in seminal plasma in normal men but was undetectable following vasectomy, thus deriving from the testis. Consistent with this finding, the betaA-subunit was immunolocalized in Sertoli and Leydig cells but was not present in seminal vesicle or prostate gland. The functional significance of the high concentrations of follistatin secreted into seminal plasma by the prostate gland and/or seminal vesicle is uncertain, but they may regulate the biological activity of testis-derived activin A and inhibin B.      

Differential regulation of cytokine genes in gingival epithelial cells challenged by Fusobacterium nucleatum and Porphyromonas gingivalis

IL-8 mRNA in human gingival epithelial cells (HGECs) is up-regulated by Fusobacterium nucleatum, and up-/down-regulated by Porphyromonas gingivalis in a complex interaction in the early stages (< or = 4 h) after infection. The mechanisms involved in this regulation in response to F. nucleatum and/or P. gingivalis infection, and identification of co-regulated cytokine genes, are the focus of this investigation. Heat, formalin or protease treatment of F. nucleatum cells attenuated the IL-8 mRNA up-regulation. NF-kappaB, mitogen-activated protein kinase (MAPK) p38 and MAPK kinase/extracellular signal-regulated kinase (MEK/ERK) pathways were involved in IL-8 mRNA induction by F. nucleatum. Pretreatment of P. gingivalis with heat, formalin or protease enhanced IL-8 mRNA induction. NF-kappaB, MARK p38, and MEK/ERK pathways were also involved in this induction. In contrast, down-regulation of IL-8 mRNA by P. gingivalis involved MEK/ERK, but not NF-kappaB or MAPK p38 pathways. cDNA arrays analysis revealed that mRNA down-regulation by P. gingivalis is a specific reaction that only a number of genes, e.g. IL-1beta, IL-8, macrophage inflammatory protein-2alpha, and migration inhibitory factor-related protein-14, are affected based on examination of 278 cytokine/receptor genes. These data indicate that F. nucleatum and P. gingivalis trigger specific and differential gene regulation pathways in HGECs.     

Mycoplasma pulmonis Inhibits Electrogenic Ion Transport across Murine Tracheal Epithelial Cell Monolayers

Murine chronic respiratory disease is characterized by persistent colonization of tracheal and bronchial epithelial cell surfaces by Mycoplasma pulmonis, submucosal and intraluminal immune and inflammatory cells, and altered airway activity. To determine the direct effect of M. pulmonis upon transepithelial ion transport in the absence of immune and inflammatory cell responses, primary mouse tracheal epithelial cell monolayers (MTEs) were apically infected and assayed in Ussing chambers. M. pulmonis-infected MTEs, but not those infected with a nonmurine mycoplasma, demonstrated reductions in amiloride-sensitive Na⁺ absorption, cyclic AMP, and cholinergic-stimulated Cl⁻ secretion and transepithelial resistance. These effects were shown to require interaction of viable organisms with the apical surface of the monolayer and to be dependent upon organism number and duration of infection. Altered transport due to M. pulmonis was not merely a result of epithelial cell death as evidenced by the following: (i) active transport of Na⁺ and Cl⁻, albeit at reduced rates; (ii) normal cell morphology, including intact tight junctions, as demonstrated by electron microscopy; (iii) maintenance of a mean transepithelial resistance of 440 Ω/cm²; and (iv) lack of leakage of fluid from the basolateral to the apical surface of the monolayer. Alteration in epithelial ion transport in vitro is consistent with impaired pulmonary clearance and altered airway function in M. pulmonis-infected animals. Furthermore, the ability of M. pulmonis to alter transport without killing the host cell may explain its successful parasitism and long-term persistence in the host. Further study of the MTE-M. pulmonis model should elucidate the molecular mechanisms which mediate this reduction in transepithelial ion transport.Mycoplasmas, the smallest free-living prokaryotes, continue to be a significant cause of respiratory infections in a variety of animals, including humans (44). Their limited biosynthetic capability dictates that these organisms must both colonize and parasitize epithelial cell surfaces. It is therefore surprising that mycoplasmas produce diseases that are slowly progressing and chronic and yet often clinically inconspicuous. The molecular mechanisms responsible for this tenuous truce between the pathogen and the host cell have not yet been identified. Murine respiratory mycoplasmosis, a naturally occurring respiratory disease in laboratory rats and mice caused by Mycoplasma pulmonis (6, 8) and characterized by chronic, often lifelong tracheitis, bronchopneumonia, and bronchiectasis (7, 23, 28), would seem to be an ideal model with which to study these mechanisms. Although M. pulmonis-infected animals generate intense local and systemic immune responses (24, 33, 42, 43), they are incapable of eliminating the organism from the respiratory epithelium. Remodeling of the airways typically observed in infected animals, impairment of pulmonary clearance, and accumulation of mucus (23) suggest that M. pulmonis may compromise the ability of the epithelial cells to absorb and secrete fluid and electrolytes.Airway epithelial cells possess two major active transport processes, Na⁺ absorption and Cl⁻ section. Water, in turn, osmotically follows the transepithelial movement of these ions, thereby providing a fluid film between the mucus layer and the epithelial cell surface. The depth and composition of this fluid microenvironment must be carefully regulated to allow the exchange of gases and the humidification of the airway epithelium and to ensure proper mucociliary clearance (47, 49). The consequences of impaired fluid and electrolyte transport in the airways are strikingly apparent in cases of cystic fibrosis, a recessive genetic disease resulting from the loss of epithelial chloride channels (31, 48). The studies reported here were designed to determine whether M. pulmonis infection alters electrolyte transport across the murine tracheal epithelium.The effect of bacteria upon mammalian epithelia in the absence of immune and inflammatory cells can be systematically evaluated by using the Ussing chamber model. Short-circuit current (I_sc) studies have helped determine the mechanism by which cholera toxin (18) and other enterotoxins affect the intestinal mucosa (17, 35). Study of various types of epithelial cells by using the Ussing chamber model has also resulted in the identification of numerous microbial substances capable of directly altering the ion transport capacity of the airway epithelium (3, 20, 41).In the present study, control and M. pulmonis-infected mouse tracheal epithelial cells (MTEs) were evaluated in Ussing chambers. The results clearly show that M. pulmonis infection directly alters epithelial ion transport and that this alteration is species specific, dose and time related, and dependent upon the association of viable organisms with the apical cell surface.