Expression and role of serotonergic and nicotinic acetylcholine receptors in alveolar macrophages
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Alveolar macrophages (AM) hold a key position in initiating inflammatory responses in thelung by secreting TNFalpha and several additional cytokines and chemokines. Their activity iscontrolled by paracrine signals such as ATP which might either be actively release as asignaling molecule or liberated during cell death, thereby reflecting a danger signal. Thepresent study was aimed to determine to which extend, and via which receptor, serotonin (5-HT), an amine released from activated platelets, and nicotinic acetylcholine receptors(nAChR), a receptor class that has been identified in mediating anti-inflammatory effects, alsoparticipate in paracrine modulation of AM function.The expression of 5-HT type 2 receptors and the effects evoked by stimulation with 5-HTwere investigated in mouse AM. Expression of the 5-HT receptors was analyzed by RT-PCR.Alveolar macrophages expressed receptor subtypes 5-HT2A, 5-HT2B and 5-HT2C, with thestrongest and most consistent expression being noted for 5-HT2C receptors. In mouse AM, 5-HT (10-5 M) induced a rise in intracellular calcium concentration ([Ca2+]i) that was initiatedby release of Ca2+ from intracellular stores and depended on extracellular Ca2+ in a sustainedphase. This 5-HT-induced increase in [Ca2+]i was neither observed in AM treated with the 5-HT2C receptor-selective inhibitor RS102221 nor in AM derived from 5-HT2C receptordeficientmice. Alveolar macrophages stimulated with 5-HT (10-5 M) showed increasedproduction of CCL2, CCL5 and TNFalpha as determined by a dot-blot assay. These datademonstrate the presence of functional 5-HT2C receptors on AM and suggest a role of 5-HT asnovel modulator of AM function. Importantly, these effects are exclusively driven by the 5-HT2C receptor, thereby providing the potential for selective intervention.In addition, the nAChR inventory of freshly isolated rat and mouse AM was determined andthe cellular events evoked by stimulation with nicotine were investigated. Positive RT-PCRresults in rat AM were obtained for nAChR subunits alpha3, alpha5, alpha9, alpha10, beta1, and beta2, with moststable expression of subunits alpha9, alpha10, beta1, and beta2. Mouse AM expressed nAChR subunits alpha9,alpha10, beta2 and beta4. Notably, mRNA coding for subunit alpha7, which is proposed to convey thenicotinic anti-inflammatory response of macrophages from other sources than the lung, wasnot detected. RT-PCR data were supported by immunohistochemistry on rat AM isolated bylavage, as well as in lung tissue sections and by Western blotting. Measurements of [Ca2+]i inrat and mouse AM did not reveal any changes in response to nicotine. However, nicotine (10-4M), given 2 min prior to ATP and 5-HT, significantly reduced the agonist-induced rise in[Ca2+]i. This nicotinic effect was further studied in rat AM, where it was blocked by alpha-bungarotoxin and did not depend on the presence of extracellular calcium. These datademonstrate that AM are equipped with modulatory nAChR with properties distinct fromionotropic nAChR mediating synaptic transmission in the nervous system. Their stimulationwith nicotine dampens ATP-induced Ca2+-release from intracellular stores. Thus, the presentstudy identifies the first acute receptor-mediated nicotinic effect on AM with antiinflammatorypotential.Taken together, 5-HT, an amine released from activated platelets, and nAChR, a receptorclass that has been identified in mediating anti-inflammatory effects, participate in paracrinemodulation of AM function. Serotonergic stimulation of 5-HT2C receptors activates AM,whereas nAChR confer an inhibitory influence upon both serotonergic and purinergicactivation via pathways differing from those known from the nervous system. These datawarrant consideration during pharmacological modulation of cholinergic transmission and 5-HT metabolism and might offer an opportunity to modify AM function in vivo.Verknüpfung zu Publikationen oder weiteren Datensätzen
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Giessen : VVB Laufersweiler