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

Abstract

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.

Introduction

Asthma is a chronic respiratory disease characterized by intermittent airway obstruction, eosinophilic inflammation, airway hyperreactivity, and high levels of IgE. Its underlying mechanisms are complex and may vary, but appear to be altogether caused by an intrapulmonary allergen-driven T helper type 2 (Th2) response characterized by increased secretion of cytokines (IL-4, IL-5, IL-13, and others) by CD4+ T lymphocytes (Sousa Mucida et al., 2003).

In asthmatic patients, the allergic reaction progresses in two distinct phases representing segments of the same phenomenon: an early response mediated by IgE and mast cells, and a late phase, during which Th2 lymphocytes, cytokines, and other cells build an inflammatory milieu that play a major role in the disease (Maddox and Schwartz, 2002).

Although the pathophysiology of asthma has been well characterized in humans and experimental models, little is known about its influence on brain activity and behavior. Descriptions of asthma triggered by artificial roses in allergic individuals, the ‘rose effect’, date to the 19 century (Mackenzie, 1886), and suggest an important role for immuneneuroendocrine interactions in asthma. Changes in emotional status and increased levels of anxiety are commonly associated with asthma crises (Lawrence, 2002, Lehrer et al., 1993); on the other hand, the distress associated with a recurrent disease can be responsible for generating pathological anxiety in patients suffering from long-term asthma (Dahlen and Janson, 2002, Rietveld et al., 2000).

Peripheral inflammation has long been implicated with changes in brain activity and behavior, thus influencing the outcome of several biological phenomena (Steinman, 2004). We previously reported some neural and behavioral correlates of allergic reactions using murine models of ovalbumin (OVA)-induced food allergy (Basso et al., 2003) and asthma (Costa-Pinto et al., 2005). Mice allergic to OVA avoided drinking an antigen-containing solution or entering a box compartment previously associated with OVA nebulization. Furthermore, OVA-sensitized animals displayed an increase in the activity of the paraventricular nucleus of the hypothalamus (PVN) and of the central nucleus of the amygdala (CeA) shortly after the OVA challenge (Basso et al., 2003, Costa-Pinto et al., 2005), represented by neurons stained for the Fos protein, a product of the immediate early gene and transcription factor c-fos (Sagar et al., 1988). These areas, the major collections of corticotropin-releasing factor (CRF)-expressing neurons in the central nervous system (CNS) (Gray, 1993), are involved in affective and emotional responses compatible to those of anxiety and avoidance in rodents (Menzaghi et al., 1993, Menzaghi et al., 1994).

The neural and behavioral responses associated with food allergy are IgE-dependent, since abrogation of anti-OVA IgE antibodies prevented the development of food aversion and the increase in the activity of the PVN and CeA previously observed in this model (Basso et al., 2003).

In an attempt to further dissect the pathways associated to the crosstalk between allergic reactions and the CNS, we employed neonatal capsaicin destruction of C-sensitive fibers and were able to show that these neurons are partially involved in signaling food allergy to the mouse brain (Basso et al., 2001). Indeed, activation of the PVN by the allergic response was totally blunted by capsaicin, unlike that of the CeA. We also found a partial inhibition of Fos-staining in the nucleus of the solitary tract (NTS), an important relay station conveying visceral sensory information to the brain (Basso et al., 2004). The suggestion that C-sensitive fibers play a role in transmitting signals derived from allergic responses within the gastrointestinal tract or the airways to the brain is strengthened by the already reported intimate anatomical contact between mast cells and the nerve endings of those afferents in mucosal surfaces (Bienenstock et al., 1988a, Bienenstock et al., 1991, Greene et al., 1988, Kiernan, 1990, Stead et al., 1989, Williams et al., 1997).

The relevance of both IgE-dependent mechanisms and C-sensitive fibers for signaling allergic responses triggered in the gastrointestinal tract, with consequent behavioral changes and brain activation, prompted us to ask whether asthma signaling to the mouse brain relies on IgE antibodies. Moreover, we focused on whether mast cell degranulation was crucial or not to these phenomena.

In this study we evaluated the activation of the PVN and CeA by Fos-staining (Sagar et al., 1988) and the development of aversion directed at the allergen, employing an adapted passive avoidance paradigm (Costa-Pinto et al., 2005), and asked whether asthma signaling to the mouse brain depends on IgE, employing a protocol of immunological tolerance that specifically blunts the IgE response (Russo et al., 2001), or depletion by a rat anti-IgE antibody (Haak-Frendscho et al., 1998). We also used an inhibitor of mast cell degranulation (Shishikura et al., 2006) to assess their relevance in these phenomena. Finally, using a mouse comparatively resistant to allergic airway inflammation (Brewer et al., 1999, Keane-Myers et al., 1998) we evaluated the role of pulmonary inflammatory cell infiltration on the neural and behavioral correlates of murine asthma.