Pharmacological Characteristics Of Lymphocytic Non-neuronal Acetylcholine Receptors

The discovery of non-neuronal AChRs in lymphocytes raised a new concern about apossible interaction between mAChRs and nAChRs in the immune system just likein neuronal system. Despite that smoking and betel nut produce adverse effects onneuronal system, nicotine and arecoline targeting the lymphocytic non-neuronalnAChRs and mAChRs, respectively, should be taken into serious consideration. Inaddition, the widespread expression of AChRs on lymphocytes would increase therisk of cholinergic drugs that cause neuronal dysfunctions.Although there have been great progresses on the biological characteristics ofAChRs on lymphocytic non-neuronal cholinergic system, little is known about themodulation of these receptors. Moreover, the physiological role of cholinergicsystem in lymphocytes has not been elucidated in detail. For this reason, the aim ofthe study is to assess the effects of cholinergic agonists and antagonists on immunefunction, which will be of great value to understand the pharmacologicalcharacteristic of lymphocytic non-neuronal AChRs.1. Effects of cholinergic modulators on the splenocytes proliferation in mice.The effects of AChRs' agonists and antagonists on the proliferation of micesplenocytes stimulated by concanavalin-A (con-A) were measured by MTTreduction assay in vitro. Oxotremorine or pilocarpine (muscarinic agonists)increase ConA-induced splenocytes proliferation with no relation with nAChRs.In contrast, nicotine (a nAChRs agonist) reduced the proliferation of splenocytesinduced by ConA. The latter was related in part withα7-AChRs-mediatedpathway. Supporting this outcome, we observed the inhibition of splenocytesproliferation exposed toα7-AChRs selective agonist choline without relationwith mAChRs. Carbachol showed two different effects, related to non-neuronalmAChRs or nAChRs, according to the concentration used in splenocytes. An inhibition of splenocytes proliferation was observed when clinical dose ofatropine, anisodamine and scopolamine (mAChRs antagonists) wereadministered. Neither others mAChRs antagonist (pirenzepine and 4-DAMP)nor nAChRs antagonists (mecamylamine,α-BTX and MLA) reversed theConA-induced splenocytes proliferation at any doses tested. These resultssuggested that the synthesis and release of Ach from lymphocytes act as anauto/paracrine factor that regulates the proliferation of splenocytes throughmAChRs and nAChRs. The induction of neuronal dysfunction by nicotine andcholinergic drugs could also affect lymphocytic proliferation throughnon-neuronal AChRs.2. Immune responses in mice to cholinergic agonists and antagonists.Arecoline (2 mg/kg), nicotine (1.5 mg/kg), anisodamine (2 mg/kg) andscopolamine (0.2 mg/kg) were administered subcutaneously to four groups for 4weeks to evaluate the immunological function in vitro and in vivo. The followingindexes for immunological activation were assessed: spleen index, serumhemolysin levels, interleukin-2 levels and splenocytes proliferation. Furthermore,the results revealed that arecoline and nicotine depress immune function vianon-neuronal cholinergic mAChRs-and nAChRs-mediated pathways,respectively. Moreover, the data indicate that different aspects of the immuneresponse can be simultaneously reduced by systemic administration of arecolineand nicotine (cholinergic agonists), and anisodamine and scopolamine(cholinergic antagonists).3. Effect of nicotine on the expression ofα7-AChRs and M3-AChRs(1) RT-PCR was used to elucidate the expression ofα7-AChRs andM3-AChRs in mice splenocytes stimulated by nicotine in vitro. After 48-h,nicotine did not alter mRNA level ofα7-AChRs and M3-AChRs.(2) Mice were treated with 1.5 mg/kg (s.c) of nicotine once a day for 2, 3 and 4weeks. RNA was extracted from spleen samples and converted to double-stranded DNA. Nicotine (nAChRs agonist) reduced the expressionofα7-AChRs mRNA after 4 weeks of treatment. No differences wereobserved inα7-AChRs mRNA after 2 and 3 weeks of treatment. Inaddition, nicotine did not alter M3-AChRs at any time of treatment. Theseoutcomes indicate that expression ofα7-AChRs mRNA in lymphocytesdiffer from neuronal cells. Also, the structures ofα7-AChRs innon-excitative and excitative cells are different.