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 ...

The synthesis and release of non-neuronal acetylcholine, a widely expressed signaling molecule, were investigated in the human placenta. This tissue is free of cholinergic neurons, i.e. a contamination of neuronal acetylcholine can be excluded. The villus showed a choline acetyltransferase (ChAT) activity of 0.65 nmol/mg protein per h and contained 500 nmol acetylcholine/g dry weight. In the absence of cholinesterase inhibitors the release of acetylcholine from isolated villus pieces amounted to 1.3 nmol/g wet weight per 10 min corresponding to a fractional release rate of 0.13% per min. The following substances did not significantly modify the release of acetylcholine: oxotremorine (1 microM), scopolamine (1 microM), (+)-tubocurarine (30 microM), forskolin (30 microM), ouabain (10 microM), 4alpha-phorbol 12,13-didecanoate (1 microM) and tetrodotoxin (1 microM). Removal of extracellular calcium, phorbol 12,13-dibutyrate (1 microM) and colchicine (100 microM) reduced the acetylcholine release between 30% and 50%. High potassium chloride (54 mM and 108 mM) increased the acetylcholine release slightly (by about 30%). A concentration of 10 microM nicotine was ineffective, but 100 microM nicotine enhanced acetylcholine release gradually over a 50-min period without desensitization of the response. The facilitatory effect of nicotine was prevented by 30 microM (+)-tubocurarine. Inhibitors of cholinesterase (physostigmine, neostigmine; 3 microM) facilitated the efflux of acetylcholine about sixfold, and a combination of both (+)-tubocurarine (30 microM) and scopolamine (1 microM) halved the enhancing effect. In conclusion, release mechanisms differ between non-neuronal and neuronal acetylcholine. Facilitatory nicotine receptors are present which are activated by applied nicotine or by blocking cholinesterase. Thus, cholinesterase inhibitors increase assayed acetylcholine by two mechanisms, protection of hydrolysis and stimulation of facilitatory nicotine receptors.

The presence of the Cavβ2 is essential for the structural integrity and function of the rod photoreceptor synapse. The Cavβ2 is less essential for the morphology of cone and bipolar cell ribbon synapses, although the impaired photopic electroretinogram suggests a functional alteration also of the cone-mediated signaling in Cavβ2-deficient retinas.

1. The depletion of both norepinephrine (NE) and acetylcholine (ACh) in the visual cortex can decrease plasticity. This decrease in plasticity, although dramatic under some circumstances, fails to occur under others. 2. We depleted cortical NE and ACh in 35- to 42-day-old kittens by making a lesion of the white matter behind the cingulate gyrus. One eye was sutured on the day of the lesion. We recorded from the visual cortex 7 days or 2-3 mo later and used the influence of the deprived eye on the cortical cells as a measure of plasticity. 3. We measured NE content by high-pressure liquid chromatography (HPLC) and inferred ACh depletion from depletion of choline acetyltransferase (ChAT) activity. NE depletion averaged 60% in the successfully depleted animals. Depletion of ChAT activity was consistent with NE depletion. 4. When recording occurred 7 days after the lesion and the sutured eye was contralateral to the lesion, plasticity was decreased on the side with the lesion; 70% of the cells were driven by the deprived eye. On the control, uninjured side only 15% of the cells were driven by the deprived eye. 5. In two circumstances the lesion did not cause a decrease in plasticity. In animals with suture ipsilateral to the lesion, the cortex remained plastic. In these animals only 26% of the cells in the hemisphere with the lesion were driven by the deprived eye. The cortex also retained its plasticity if the contralateral eye remained sutured for several months after the lesion, even though there was no recovery from NE and ACh depletion. 6. We conclude that depletion of NE and ACh does decrease plasticity; that is, it protects the deprived eye from losing its ability to drive cortical cells, at least for a short period of time. Depletion protects only the normally dominant contralateral pathway; the ipsilateral visual pathway remains plastic. 7. Perhaps the importance of the side of the deprived eye can be explained by assuming that depletion of NE and ACh removes facilitatory input. This would decrease the ability of cortical cells on the side with lesion to potentiate the input from the nondeprived eye relative to the deprived eye; that is, it would decrease the molecular deprivation (MD) effect. A removal of facilitation would also increase the visual input required to drive cells.(ABSTRACT TRUNCATED AT 400 WORDS)

BoCo showed significantly better hemostatic effect than PECo and OxCe.