Neural and behavioral correlates of food allergy.

Food allergy accounts for a great number of reactions leading to diminished quality of life in western countries. There has been an abundance of reports of behavioral changes, as well as psychiatric conditions associated with food allergy over the past decades. Most of this field inspired little medical attention for its lack of a solid scientific ground. We review the literature on the association of food allergy and brain activity, leading to changes in emotion and behavior. Moreover, we describe an experimental paradigm employed to dissect the biological relevance of this association. Mice allergic to ovalbumin avoid a palatable sweet solution in order to escape contact with antigen. This choice is associated with increased levels of anxiety, compatible with a conflicting situation. These responses are associated with increased activity in brain areas associated with emotional and affective behavior, which are also important for anxiety and stress responses. Higher levels of corticosterone accompany these changes in behavior. These responses are mediated by specific antibodies and prevented by depletion or immunological tolerance. They are also partially mediated by C-sensitive afferents and mast cells. Far from anecdote, neural repercussions of food allergy should be considered when planning a therapeutic strategy in affected individuals.

Food aversion: a critical balance between allergen-specific IgE levels and taste preference.

Animals sensitized to allergens change their feeding behavior and avoid drinking the otherwise preferred sweetened solutions containing the allergens. This phenomenon, known as food aversion, appears to be mediated by allergen-specific IgE antibodies. Here we investigated food aversion in BALB/c and C57BL/6 mice, which differ in their allergic responses to the allergen ovalbumin as well as in their preference for sweet taste. BALB/c mice present higher levels of IgE and a natural lower preference for sweet flavors when compared to C57BL/6 mice. Specifically, we studied a conflicting situation in which animals simultaneously experienced the aversive contact with the allergen and the attractive sweet taste of increasing concentrations of sucrose. We found that BALB/c mice were more prone to develop food aversion than C57BL/6 mice and that this aversive behavior could be abolished in both strains by increasing the palatability of the solution containing the allergen. In both strains food aversion was positively correlated with the levels of allergen-specific IgE antibodies and inversely correlated with their preference for sucrose sweetened solutions.

Behavior: a relevant tool for brain-immune system interaction studies.

Neuroimmunomodulation describes the field focused on understanding the mechanisms by which the central nervous system interacts with the immune system, potentially leading to changes in animal behavior. Nonetheless, not many articles dealing with neuroimmunomodulation employ behavior as an analytical endpoint. Even fewer papers deal with social status as a possible modifier of neuroimmune phenomena. In the described sets of experiments, we tackle both, using a paradigm of social dominance and subordination. We first review data on the effects of different ranks within a stable hierarchical relationship. Submissive mice in this condition display more anxiety-like behaviors, have decreased innate immunity, and show a decreased resistance to implantation and development of melanoma metastases in their lungs. This suggests that even in a stable, social, hierarchical rank, submissive animals may be subjected to higher levels of stress, with putative biological relevance to host susceptibility to disease. Second, we review data on how dominant and submissive mice respond differentially to lipopolysaccharide (LPS), employing a motivational perspective to sickness behavior. Dominant animals display decreased number and frequency in several aspects of behavior, particularly agonistic social interaction, that is, directed toward the submissive cage mate. This was not observed in submissive mice that maintained the required behavior expected by its dominant mate. Expression of sickness behavior relies on motivational reorganization of priorities, which are different along different social ranks, leading to diverse outcomes. We suggest that in vitro assessment of neuroimmune phenomena can only be understood based on the behavioral context in which they occur.

Behavioral effects of LPS in adult, middle-aged and aged mice.

Acute infections lead to alterations in behavior, collectively known as sickness behavior, which includes reduction in locomotion, food ingestion, sexual and social behavior, environmental exploration, and sleep profile. Although generally seen as undesired, sickness behavior represents a conserved strategy for animals to overcome disease. Aging process is associated with a variety of changes in immunity, which are referred to as immunosenescence, and include higher mortality by infectious diseases. Few works studied sickness behavior display in old animals. Thus, we sought to investigate the display of sickness related behaviors on aged mice. Adult (3-6 months old), middle-aged (12-15 m) and aged mice (18-22 m) were treated with i.p. LPS (200 microg/kg) and their behaviors were assessed in the open field and in the elevated plus-maze. Exploratory activity was similar in aged mice treated or not with LPS in both apparati. In the open field, locomotion remained at baseline levels; in the elevated plus-maze, there was a time-dependent decrease in motor activity.

Pfaffia paniculata (Brazilian ginseng) methanolic extract reduces angiogenesis in mice.

Pfaffia paniculata (Brazilian ginseng) roots have been indicated for the treatment of several diseases. Our studies have shown that P. paniculata roots present antineoplastic effects and cancer chemopreventive activity in a mouse hepatocarcinogenesis model. The purpose of this study was to investigate the effects of the Brazilian ginseng on corneal angiogenesis in mice. We first conducted a toxicological study employing 250, 500, or 1000mg/kg/day of the methanolic extract of P. paniculata roots by gavage to BALB/c mice. Animals did not lose weight during the treatment nor presented histopathological alterations. The effect of this root on angiogenesis in the cornea of BALB/c mice was then assessed. Male mice were treated, by gavage, once a day, with doses of 250, 500, or 1000mg/kg of methanolic extract of P. paniculata powdered root for 10 days; filtered water was used as control. Corneal cauterization was accomplished by the contact of a silver nitrate crystal on the central area of the cornea, in the 5th day of treatment with P. paniculata, which continued thereafter; the animals were euthanized on the 6th day after cauterization. Newly formed blood vessels were filled with India ink, and the corneas were routinely processed. Blood vessels were quantified in an image analysis system. A smaller total area of neovascularization in the mouse cornea was observed in animals treated with 1000mg/kg of the methanolic extract of P. paniculata. These results indicate an antiangiogenic effect of this extract. The mechanisms of this antiangiogenic activity of P. paniculata should be further investigated.

Hepatic granulomas induced by Schistosoma mansoni in mice deficient for connexin 43 present lower cell proliferation and higher collagen content.

Granuloma formation involves a coordinated interaction between monocytes and macrophages, epithelioid cells, lymphocytes, eosinophils, neutrophils and fibroblasts. It has been established that extracellular communication via cytokines is important for the assembly of granulomas. However, the importance of gap junctions and intercellular communication to granuloma formation and development had never been assessed. Connexins are proteins that form gap junctions, and connexin 43 (Cx43) is present in macrophages, lymphoid cells, myelogenous cells, fibroblasts and others. We analyzed the effect of heterologous deletion of Gja1 (Cx43 gene) on the formation and development of hepatic granulomas induced by Schistosoma mansoni eggs. Heterozygous (Cx43(+/-)) and wild-type (Cx43(+/+)) mice were infected subcutaneously with S. mansoni cercarie and evaluated after 6, 8 and 12 weeks. Granuloma cells express Cx43, as revealed by real-time PCR in isolated granulomas, and by immunohistochemistry. Cx43 expression was reduced in Cx43(+/-) mice, as expected. No differences in the average area of granulomas or number of cells per granuloma were observed between mice of different genotypes. However, granuloma cells from Cx43(+/-) mice displayed a reduced index of the proliferating cell nuclear antigen (PCNA) labeling at 8 and 12 weeks post-infection. Moreover, Cx43(+/-) granulomas unexpectedly presented a higher degree of fibrosis, quantified by morphometric analysis in Sirius Red-stained slides. Our results indicate that the deletion of one allele of the Cx43 gene, and possibly the reduced gap junction intercellular communication capacity (GJIC), may impair the interactions between granuloma cells, reducing their proliferation and increasing their collagen content, thereby modifying the characteristics of S. mansoni granuloma in mice.

Role of mast cell degranulation in the neural correlates of the immediate allergic reaction in a murine model of asthma.

Experimental airway allergy in mice leads to increased activity in specific hypothalamic and amygdaloid nuclei, and behavioral changes. The experiments described here were designed to determine the role of anaphylactic antibodies, mast cell degranulation, and lung inflammation in the neural and behavioral correlates of an experimental murine asthma-like response. Animals were sensitized intraperitoneally with ovalbumin adsorbed to alum, and challenged by intranasal ovalbumin instillation or aerosol. To induce immunological tolerance, animals were fed ovalbumin in the drinking water for 5 consecutive days, along with primary sensitization. Depletion of IgE was also accomplished with a non-anaphylactic anti-IgE antibody. Mast cell degranulation was inhibited by cromolyn. In addition to BALB/c animals, C3H/HeJ mice were used for their relative resistance to lung allergic inflammation. We confirmed that ovalbumin challenge in allergic mice leads to increased activity in the paraventricular nucleus of the hypothalamus and central nucleus of the amygdala, and avoidance behavior towards an allergen-associated compartment. Moreover, these responses were precluded by oral tolerance or anti-IgE treatment, even in the presence of IgG1. Cromolyn abrogates both responses in the presence of anaphylactic antibodies. Finally, although sensitized C3H/HeJ mice did not develop airway inflammation, they exhibited brain and behavioral changes similar to BALB/c animals. The repercussions of murine allergic asthma on brain and behavior are IgE-dependent, mediated by mast cell degranulation, and do not require a pulmonary inflammatory infiltrate, suggesting that the early phase of this immediate allergic response suffices for the brain activation associated with avoidance behavior towards exposure to the allergen.

Neural correlates of IgE-mediated allergy.

Although many authors have considered a direct interaction between allergic reactions and behavioral changes, supporting evidence has been elusive. In this series of studies we show that after oral or nasal ovalbumin (OVA) challenge, allergic mice present increased Fos expression in the paraventricular nucleus of the hypothalamus (PVN) and in the central nucleus of the amygdala (CeA). Mice with food allergy display higher levels of anxiety and increased serum corticosterone levels, and allergy-activated neurons express corticotropin-releasing factor (CRF) in the PVN and CeA. OVA-allergic mice develop aversion to an antigen-containing solution, and also avoid a dark compartment previously associated with nebulized OVA. Results on brain Fos expression and behavioral data seem compatible with adaptive responses. Removal of IgE by either antibody depletion or the development of oral tolerance precluded all responses analyzed here. C-sensitive fiber destruction by neonatal capsaicin inhibited the activation in the PVN, but not in the CeA, and decreased the magnitude of food aversion. Cromolyn, a mast cell stabilizer, completely blocked Fos expression in the PVN and CeA, and precluded the development of aversion to the dark compartment associated with nebulized OVA. Employing mice that do not develop an important inflammatory infiltrate following nasal OVA challenge, we found that inflammatory cells are not required at the site of challenge in order to trigger neural or behavioral correlates of murine experimental asthma. Altogether, we have built a solid foundation for understanding neuroimmune interactions during allergic responses that may contribute to the comprehension of psychological disorders associated with allergy.

Microscopical study of experimental wound healing in Notothenia coriiceps (Cabeçuda) at 0 degrees C.

Notothenia coriiceps (Cabeçuda) is an Antarctic benthic fish frequently found with lesions in the tegument caused by seal predation. We have investigated epidermal repair in these animals by means of a microscopic study of experimental wound healing at 0 degrees C. At 24--48 h after wound induction, mucous exudate and necrotic lining cells covered the wound. At 7--14 days, an epidermal "tongue" could be discerned, folded at the tip, with intercellular oedema between the tip and the wound border. After 23--30 days, the wound was completely closed and the migrating epidermis, with intercellular oedema, was reduced. By 45--90 days, melanocytes progressively increased in the epidermis but no scales were formed. The inflammatory infiltrate was mainly composed of neutrophils after 7 days, at which time they were mostly replaced by macrophages; lymphocytes and plasma cells were also present. The border epidermis slid towards the centre, folding at the tip and finally fusing to form a diaphragm. The cells of the epidermis began to multiply only after complete closure of the wound. The lack of scale formation on induced and naturally found wounds, even after 90 days, suggests that different mechanisms in wound repair occur at 0 degrees C from those in fish from temperate and tropical environment. This is the first report of successful wound repair at polar temperatures, indicating the adaptation of N. coriiceps to the Antarctic environment.

Avoidance behavior and neural correlates of allergen exposure in a murine model of asthma.

Allergic asthma is characterized by intermittent airway obstruction, inflammation, airway hyperreactivity, and increased production of IgE. The pathophysiology of asthma is well understood but little is known about its influences on brain activity and behavior. We recently described the neural correlates of food allergy and its associated modulation of behavior using an experimental model that also generates a T helper type 2 (Th2)-skewed response, with high levels of IgE. Here we show that mice allergic to ovalbumin (OVA) have an increase in the activity of the paraventricular nucleus of the hypothalamus (PVN) and in the central nucleus of the amygdala (CeA) following a single nasal OVA challenge. Moreover, we adapted a classical passive avoidance test using an OVA aerosol as the aversive stimulus. We found that allergic mice avoid entering the dark compartment of the apparatus that had been previously associated with nebulization of the allergen, while their non-immunized controls still move into the dark side of the test box. Thus, allergic mice have increased activity in areas of the CNS commonly associated with emotionality-related behavioral responses, such as the avoidance of a context previously associated with an unpleasant or harmful situation. Moreover, our findings on the avoidance test illustrate that previous experience with an airborne allergen can modify behavior.

Neural pathways involved in food allergy signaling in the mouse brain: role of capsaicin-sensitive afferents.

There is increasing evidence supporting the notion that brain-gut communication is crucial for the manifestation of functional gastrointestinal (GI) disorders. Employing denervation by neonatal capsaicin treatment, we investigated here the role of unmyelinated C-fibers in food allergy signaling in the brain. We found that 90 min after oral ovalbumin (OVA) challenge, allergic mice present increased c-fos expression in emotionality-related brain areas such as the paraventricular nucleus of the hypothalamus (PVN) and the central nucleus of the amygdala (CeA). Food allergy also induced enhanced Fos immunoreactivity in the nucleus of tractus solitarii (NTS) of OVA-immunized animals. We also show that while the degree of Fos staining in the NTS of allergic mice was only diminished by neonatal capsaicin, it was completely blocked in the PVN. However, capsaicin did not modify food allergy-induced c-fos expression in the CeA. In conclusion, this study provides evidence showing that unmyelinated C-fibers are part of the neural pathways involved in food allergy-induced activation of specific brain areas, particularly the PVN and to a lesser extent the NTS.

Inflammatory infiltrate, VEGF and FGF-2 contents during corneal angiogenesis in STZ-diabetic rats.

Angiogenesis is a key mechanism that influences several physiological and pathological processes, including wound healing. During the past decades, many groups have shown that controlling angiogenesis might be an answer to overcome pathological situations when this process is out of control. Many altered metabolic states exert considerable influence on the development of angiogenesis. We have chosen diabetes as a model of a progressive metabolic disease with many associated conditions, including an alteration of wound healing dynamics described elsewhere. To evaluate the growth of newly formed blood vessels during diabetes, we induced corneal angiogenesis through silver nitrate cauterization in streptozotocin-induced diabetic rats, always comparing to control non-diabetic or insulin-treated diabetic rats. Computer-aided analysis showed that both the percentage of area taken by vessels on the cornea and their average length were decreased in diabetic animals; furthermore, this diminishment was prevented by insulin treatment in previously diabetic rats. Immunohistochemical staining of neutrophils and macrophages (EDI clone) did not show any differences on number of migrating cells in the cornea. Immunolocalization of vascular endothelial growth factor and basic fibroblast growth factor did not differ considerably among groups either. These results support previous findings that angiogenesis is decreased due to the development of diabetes mellitus but contrasts to descriptions from other investigators in regard to the inflammatory infiltrate and production of growth factors. In our experimental conditions, the cause of the decreased angiogenesis in diabetic rats remains for further elucidation.