Cocaine Directly Inhibits Alpha 6-Containing Nicotinic Acetylcholine Receptors In Human Sh-Ep1 Cells And Mouse Vta Da Neurons

Alpha6-containing nicotinic acetylcholine receptors are primarily found in neurons of the midbrain dopaminergic (DA) system, suggesting these receptors are potentially involved in drug reward and dependence. Here, we report a novel effect that cocaine directly inhibits alpha 6N/alpha 3C beta 2 beta 3-nAChR (alpha 6*-nAChRs) function. Human alpha 6*-nAChRs were heterologously expressed within cells of the SH-EP1 cell line for functional characterization. Mechanically dissociated DA neurons from mouse ventral tegmental area (VTA) were used as a model of presynaptic alpha 6*-nAChR activation since this method preserves terminal boutons. Patch-clamp recordings in whole-cell configuration were used to measure alpha 6*-nAChR function as well as evaluate the effects of cocaine. In SH-EP1 cells containing heterologously expressed human alpha 6*-nAChRs, cocaine inhibits nicotine-induced inward currents in a concentration-dependent manner with an IC50 value of 30 mu M. Interestingly, in the presence of 30 mu M cocaine, the maximal current response of the nicotine concentration-response curve is reduced without changing nicotine's EC50 value, suggesting a noncompetitive mechanism. Furthermore, analysis of whole-cell current kinetics demonstrated that cocaine slows nAChR channel activation but accelerates whole-cell current decay time. Our findings demonstrate that cocaine-induced inhibition occurs solely with bath application, but not during intracellular administration, and this inhibition is not use-dependent. Additionally, in Xenopus oocytes, cocaine inhibits both alpha 6N/alpha 3C beta 2 beta 3-nAChRs and alpha 6M211L/alpha 3IC beta 2 beta 3-nCAhRs similarly, suggesting that cocaine may not act on the alpha 3 transmembrane domain of chimeric alpha 6N/alpha 3C beta 2 beta 3-nAChR. In mechanically isolated VTA DA neurons, cocaine abolishes alpha 6*-nAChR-mediated enhancement of spontaneous inhibitory postsynaptic currents (sIPSCs). Collectively, these studies provide the first evidence that cocaine directly inhibits the function of both heterologously and naturally expressed alpha 6*-nAChRs. These findings suggest that alpha 6*-nAChRs may provide a novel pharmacological target mediating the effects of cocaine and may underlie a novel mechanism of cocaine reward and dependence.