To define the role of nontypeable Haemophilus influenzae (NTHI) lipooligosaccharide (LOS) in the inductionConsiderable evidence indicates that endotoxin or its subcomponent lipopolysaccharide (LPS) or lipooligosaccharide (LOS), each an integral component of the outer membrane of gram-negative bacteria, is refractile and present in a large percentage of middle ear effusions from children with otitis media (OM) (10,14). Endotoxin is a potent stimulant of proinflammatory reactions of the host's innate immune response (22). The inflammatory cytokines are thought to be of central importance in the pathogenesis and regulation of proliferation, chemotaxis, and activation of inflammatory cells during the course of middle ear infections (21, 29). A previous report from our laboratory indicates that there is a significant correlation between endotoxin, tumor necrosis factor alpha (TNF-␣), and interleukin 1␤ (IL-1␤) concentrations in middle ear effusions from children with OM (35). Recent experimental studies, moreover, demonstrate that formalin-killed nontypeable Haemophilus influenzae (NTHI) stimulates the release of IL-1␤, IL-6, IL-8, and TNF-␣ from the middle ear mucosa in the guinea pig OM model (30). It has been proposed that the initial interaction of endotoxin with middle ear epithelium leads to the production and release of inflammatory cytokines in the middle ear (2,5,8). This is followed by recruitment of neutrophils, eruption of a cytokine-mediated inflammatory cascade, and neutrophil activation, resulting in the release of inflammatory molecules which participate directly in middle ear inflammation (27,37). To date, reports on the effects of NTHI LOS on induction of cytokine gene expression by human cells have focused primarily on stimulation of monocytes (34) and bronchial and tracheal epithelial cells (7,16,24). The effects of NTHI LOS on cytokine and chemokine gene expression by cultured human middle ear epithelial (HMEE) cells have never been reported.Major advances have also been made recently in identifying the molecular mechanisms underlying LPS or endotoxin recognition and signaling for LPS-induced cell activation. It is believed that membrane-bound CD14 (mCD14), a glycosyl phosphatidylinositol-anchored protein expressed on myeloid cells, binds to LPS and initiates its signaling (33). Toll-like receptors (TLRs) have been identified as the signaling molecules and may act as transmembrane coreceptors with CD14 in LPS activation of different cell populations (15). The potential role of NTHI LOS in the activation of HMEE cells via these pathways has also not been explored.It is now possible to define the role of NTHI LOS in the pathogenesis of OM by using mutants with various LOS gene disruptions as tools. A recent study from our laboratory indicates that disruption of the NTHI htrB and rfaD LOS genes does not impact the ability of NTHI to colonize the nasopharynx; however, it induces marked differences in the abilities of

Recently we reported that rat taste receptor cells respond to the neurotransmitter serotonin with an inhibition of a calcium-activated potassium current [17]. In the present study, this observation is confirmed and extended by studying the effects of an array of serotonergic agonists on membrane properties, calcium-activated potassium current, and voltage-dependent sodium current in taste receptor cells using the patch-clamp recording technique in the whole-cell configuration. Serotonergic inhibition of calcium-activated potassium current was mimicked by the agonists N-(3-trifluoromethylphenyl)piperazine and by (+/-)-2-dipropylamino-8-hydroxy-1,2,3,4-tetrahydronaphthalene. Both produced reversible inhibition of K(Ca) as well as significantly increasing the input resistance of the cell. The agonists 1-(1-naphthyl)piperazine and buspirone (both serotonin receptor 1A agonists) were similarly effective in reducing K(Ca). Outward current was unaffected by application of phenylbiguanide, a serotonin receptor 3 agonist, though current was affected by subsequent application of (+/-)-2-dipropylamino-8-hydroxy-1,2,3, 4-tetrahydronaphthalene. Two agonists-N-(3-trifluoromethylphenyl)piperazine and (+/-)-2-dipropylamino-8-hydroxy-1,2,3,4-tetrahydronaphthalene-were also tested on voltage-dependent sodium currents; both were effective and reversible in reducing its magnitude at a variety of applied potentials. These data are consistent with the notion that serotonin effects in rat taste receptor cells are mediated by serotonin 1A receptors, though other receptor subtypes may be additionally expressed. Serotonin may affect the taste cell electrical properties during active stimulation in a paracrine fashion.

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