Nicotinic drug treatment can affect the expression of neuronal nicotinic acetylcholine receptors ( nAChR) both in vivo and in vitro through molecular mechanisms not fully understood. The present study investigated the effect of the novel cytisine dimer 1,2-bisN-cytisinylethane (CC4) on nAChR natively expressed by SH-SY5Y neuroblastoma cells in culture. 2 CC4 lacked the agonist properties of cytisine and was a potent antagonist (IC50 220 nM) on nAChRs. Chronic treatment of SH-SY5Y cells with 1 mM CC4 for 48 h increased the expression of H-3-epibatidine (H-3-Epi; 3-4-fold) or I-125-alpha-bungarotoxin (I-125-alpha Bgtx; 1.2-fold) sensitive receptors present on the cell membrane and in the intracellular pool. Comparable data were obtained with nicotine or cytisine, but not with carbamylcholine, d-tubocurarine, di-hydro-beta-erythroidine or hexametonium. 3 Immunoprecipitation and immunopurification studies showed that the increase in H-3-Epi- binding receptors was due to the enhanced expression of alpha 3 beta 2 and alpha 3 beta 2 beta 4 subtypes without changes in subunit mRNA transcription or receptor half-life. The upregulation was not dependent on agonist/antagonist properties of the drugs, and did not concern muscarinic or serotonin receptors. 4 Whole-cell patch clamp analysis of CC4-treated cells demonstrated larger nicotine-evoked inward currents with augmented sensitivity to the blockers alpha-conotoxin MII or methyllycaconitine. 5 In conclusion, chronic treatment with CC4 increased the number of nAChRs containing beta 2 and alpha 7 subunits on the plasma membrane, where they were functionally active. In the case of beta 2-containing receptors, we propose that CC4, by binding to intracellular receptors, triggered a conformational reorganisation of intracellular subunits that stimulated preferential assembly and membrane-directed trafficking of beta 2-containing receptor subtypes
Long-term exposure to the new nicotinic agonist 1,2-bis-cytisinylethane upregulates nicotinic receptor subtypes of SH-SY5Y human neuorblastoma cells.
Clementi F;Gotti C
2005
Abstract
Nicotinic drug treatment can affect the expression of neuronal nicotinic acetylcholine receptors ( nAChR) both in vivo and in vitro through molecular mechanisms not fully understood. The present study investigated the effect of the novel cytisine dimer 1,2-bisN-cytisinylethane (CC4) on nAChR natively expressed by SH-SY5Y neuroblastoma cells in culture. 2 CC4 lacked the agonist properties of cytisine and was a potent antagonist (IC50 220 nM) on nAChRs. Chronic treatment of SH-SY5Y cells with 1 mM CC4 for 48 h increased the expression of H-3-epibatidine (H-3-Epi; 3-4-fold) or I-125-alpha-bungarotoxin (I-125-alpha Bgtx; 1.2-fold) sensitive receptors present on the cell membrane and in the intracellular pool. Comparable data were obtained with nicotine or cytisine, but not with carbamylcholine, d-tubocurarine, di-hydro-beta-erythroidine or hexametonium. 3 Immunoprecipitation and immunopurification studies showed that the increase in H-3-Epi- binding receptors was due to the enhanced expression of alpha 3 beta 2 and alpha 3 beta 2 beta 4 subtypes without changes in subunit mRNA transcription or receptor half-life. The upregulation was not dependent on agonist/antagonist properties of the drugs, and did not concern muscarinic or serotonin receptors. 4 Whole-cell patch clamp analysis of CC4-treated cells demonstrated larger nicotine-evoked inward currents with augmented sensitivity to the blockers alpha-conotoxin MII or methyllycaconitine. 5 In conclusion, chronic treatment with CC4 increased the number of nAChRs containing beta 2 and alpha 7 subunits on the plasma membrane, where they were functionally active. In the case of beta 2-containing receptors, we propose that CC4, by binding to intracellular receptors, triggered a conformational reorganisation of intracellular subunits that stimulated preferential assembly and membrane-directed trafficking of beta 2-containing receptor subtypesI documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
