1 The release of acetylcholine was investigated in the human placenta villus, a useful model for the characterization of the non-neuronal cholinergic system. 2 Quinine, an inhibitor of organic cation transporters (OCT), reduced acetylcholine release in a reversible and concentration-dependent manner with an IC 50 value of 5 mM. The maximal e ect, inhibition by 99%, occurred at a concentration of 300 mM. 3 Procaine (100 mM), a sodium channel blocker, and vesamicol (10 mM), an inhibitor of the vesicular acetylcholine transporter, were ine ective. 4 Corticosterone, an inhibitor of OCT subtype 1, 2 and 3 reduced acetylcholine in a concentrationdependent manner with an IC 50 value of 2 mM. 5 Substrates of OCT subtype 1, 2 and 3 (amiloride, cimetidine, guanidine, noradrenaline, verapamil) inhibited acetylcholine release, whereas carnitine, a substrate of subtype OCTN2, exerted no e ect. 6 Long term exposure (48 and 72 h) of villus strips to anti-sense oligonucleotides (5 mM) directed against transcription of OCT1 and OCT3 reduced the release of acetylcholine, whereas OCT2 antisense oliogonucleotides were ine ective. 7 It is concluded that the release of non-neuronal acetylcholine from the human placenta is mediated via organic cation transporters of the OCT1 and OCT3 subtype.
IntroductionAcetycholine has been demonstrated in the vast majority of human non-neuronal cells, for example epithelial, endothelial, mesothelial and immune cells as well as smooth muscle ®bres (Grando, 1997;Wessler et al., 1998;Kawashima & Fujii, 2000). In addition, nicotinic and muscarinic receptors are widely expressed on these cells (Brunner & Kukovetz, 1986;Grando et al., 1995;Maus et al., 1998;Costa et al., 1988;Kawashima & Fujii, 2000). Non-neuronal acetylcholine can act as a local cell molecule via paracrine and autocrine mechanisms to control basic cell functions such as proliferation, di erentiation and cell ± cell contact (Grando, 1997;Wessler et al., 1998;Kawashima & Fujii, 2000). The release mechanisms by which non-neuronal cells, for example epithelial cells of the placenta, liberate acetylcholine into the extracellular space, are unknown. The human placenta is not innervated by extrinsic or intrinsic cholinergic neurons (Sastry & Sadavongvivad, 1979;Sastry, 1997). Thus, released acetylcholine is not contaminated by neuronal acetylcholine and can be used as a model for studying the release mechanisms of non-neuronal acetylcholine. Acetylcholine, an organic cation, may be a substrate for organic cation transporters (OCT). OCTs are most widely expressed in di erent cell types including the human placenta (Koepsell, 1998;Dresser et al., 1999). Three organic cation transporters have been cloned which represent high capacity non-neuronal monoamine transporters, OCT subtype 1 (GruÈ ndemann et al., 1994;Nagel et al., 1997), subtype 2 (Okuda et al., 1996 GruÈ ndemann et al., 1997; and subtype 3, the latter also being known as extraneuronal monoamine transporter uptake 2 (GruÈ ndemann et al., 1998;Kekuda et al., 1998;Wu et al., 1998). To ...
