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Acute poisoning with OPs may lead to a range of neurological effects, which cannot be explained by AChE inhibition alone. Several OPs interact directly with cholinergic receptors in mammals, but such data does not exist for invertebrates. The aim of current study was to investigate the direct and indirect effects of diazinon on the contractions of rat ileum and to compare those effects on the nervemuscle preparation of the Ascaris suum. In the presence of increasing concentrations of diazinon (3, 10 and 30 nM), EFS-induced ileal contractions were increased significantly. In the same preparation, diazinon 3 nM, significantly increased contractions induced by EFS, but did not affect the contractions caused by 5MFI. Contrarily, 1 mu M of diazinon significantly and reversibly inhibited the EFS-induced ileal contractions. Diazinon exhibited competitive and non-competitive inhibitions of 5MFI induced contractions. The control EC50 of 5MFI was 2.48 mu M with R-max = 1.88 g. In the presence of diazinon, EC50 was 12.45 mu M, while R-max was reduced to 0.43 g. After washing, the EC50 and R-max values were again closer to the control level (3.80 mu M and 1.04 g). Diazinon 1 mu M did not inhibit Ascaris suum contractions caused by ACh, but it increased the R-max. Diazinon in our study exhibits two opposite effects on the motor activity of the ileum. In low nanomolar concentrations the dominat is its effect on AChE and the stimulation of contractions. Furthermore, in concentrations that approach micromolar values diazinon has a direct inhibitory effect on muscarinic receptors. The direct inhibitory effect of diazinon on A. suum contractions was not found.

We present an overview of the toxicological profile of the fast-acting, lipophilic macrocyclic imine toxins, an emerging family of organic compounds associated with algal blooms, shellfish contamination and neurotoxicity. Worldwide, shellfish contamination incidents are expanding; therefore, the significance of these toxins for the shellfish food industry deserves further study. Emphasis is directed to the dinoflagellate species involved in their production, their chemical structures, and their specific mode of interaction with their principal natural molecular targets, the nicotinic acetylcholine receptors, or with the soluble acetylcholine-binding protein, used as a surrogate receptor model. The dinoflagellates Karenia selliformis and Alexandrium ostenfeldii / A. peruvianum have been implicated in the biosynthesis of gymnodimines and spirolides, while Vulcanodinium rugosum is the producer of pinnatoxins and portimine. The cyclic imine toxins are characterized by a macrocyclic skeleton comprising 14-27 carbon atoms, flanked by two conserved moieties, the cyclic imine and the spiroketal ring system. These phycotoxins generally display high affinity and broad specificity for the muscle type and neuronal nicotinic acetylcholine receptors, a feature consistent with their binding site at the receptor subunit interfaces, composed of residues highly conserved among all nAChRs, and explaining the diverse toxicity among animal species. This is an article for the special issue XVth International Symposium on Cholinergic Mechanisms.

Acute poisoning with OPs may lead to a range of neurological effects, which cannot be explained by AChE inhibition alone. Several OPs interact directly with cholinergic receptors in mammals, but such data does not exist for invertebrates. The aim of current study was to investigate the direct and indirect effects of diazinon on the contractions of rat ileum and to compare those effects on the nervemuscle preparation of the Ascaris suum. In the presence of increasing concentrations of diazinon (3, 10 and 30 nM), EFS-induced ileal contractions were increased significantly. In the same preparation, diazinon 3 nM, significantly increased contractions induced by EFS, but did not affect the contractions caused by 5MFI. Contrarily, 1 mu M of diazinon significantly and reversibly inhibited the EFS-induced ileal contractions. Diazinon exhibited competitive and non-competitive inhibitions of 5MFI induced contractions. The control EC50 of 5MFI was 2.48 mu M with R-max = 1.88 g. In the presence of diazinon, EC50 was 12.45 mu M, while R-max was reduced to 0.43 g. After washing, the EC50 and R-max values were again closer to the control level (3.80 mu M and 1.04 g). Diazinon 1 mu M did not inhibit Ascaris suum contractions caused by ACh, but it increased the R-max. Diazinon in our study exhibits two opposite effects on the motor activity of the ileum. In low nanomolar concentrations the dominat is its effect on AChE and the stimulation of contractions. Furthermore, in concentrations that approach micromolar values diazinon has a direct inhibitory effect on muscarinic receptors. The direct inhibitory effect of diazinon on A. suum contractions was not found.

Monoaminergic dysregulation is implicated in attention-deficit/hyperactivity disorder (ADHD), and methylphenidate and amphetamines are the most frequently prescribed pharmacological agents for treating ADHD. However, it has recently been proposed that the core symptoms of the disorder might be due to an imbalance between monoaminergic and cholinergic systems. In this study, we used fibroblast cell homogenates from boys with and without ADHD as an extraneural cell model to examine the cholinergic receptor density, that is, muscarinic acetylcholine receptors (mAChRs). We found that the binding capacity (Bmax) of [³H] Quinuclidinyl benzilate (³H-QNB) to mAChRs was decreased by almost 50 % in the children with ADHD (mean = 30.6 fmol/mg protein, SD = 25.6) in comparison with controls [mean = 63.1 fmol/mg protein, SD = 20.5, p ≤ 0.01 (Student's unpaired t test)]. The decreased Bmax indicates a reduced cholinergic receptor density, which might constitute a biomarker for ADHD. However, these preliminary findings need to be replicated in larger ADHD and comparison cohorts

Monoaminergic dysregulation is implicated in attention-deficit/hyperactivity disorder (ADHD), and methylphenidate and amphetamines are the most frequently prescribed pharmacological agents for treating ADHD. However, it has recently been proposed that the core symptoms of the disorder might be due to an imbalance between monoaminergic and cholinergic systems. In this study, we used fibroblast cell homogenates from boys with and without ADHD as an extraneural cell model to examine the cholinergic receptor density, that is, muscarinic acetylcholine receptors (mAChRs). We found that the binding capacity (Bmax) of [³H] Quinuclidinyl benzilate (³H-QNB) to mAChRs was decreased by almost 50 % in the children with ADHD (mean = 30.6 fmol/mg protein, SD = 25.6) in comparison with controls [mean = 63.1 fmol/mg protein, SD = 20.5, p ≤ 0.01 (Student's unpaired t test)]. The decreased Bmax indicates a reduced cholinergic receptor density, which might constitute a biomarker for ADHD. However, these preliminary findings need to be replicated in larger ADHD and comparison cohorts

Monoaminergic dysregulation is implicated in attention-deficit/hyperactivity disorder (ADHD), and methylphenidate and amphetamines are the most frequently prescribed pharmacological agents for treating ADHD. However, it has recently been proposed that the core symptoms of the disorder might be due to an imbalance between monoaminergic and cholinergic systems. In this study, we used fibroblast cell homogenates from boys with and without ADHD as an extraneural cell model to examine the cholinergic receptor density, that is, muscarinic acetylcholine receptors (mAChRs). We found that the binding capacity (Bmax) of [³H] Quinuclidinyl benzilate (³H-QNB) to mAChRs was decreased by almost 50 % in the children with ADHD (mean = 30.6 fmol/mg protein, SD = 25.6) in comparison with controls [mean = 63.1 fmol/mg protein, SD = 20.5, p ≤ 0.01 (Student's unpaired t test)]. The decreased Bmax indicates a reduced cholinergic receptor density, which might constitute a biomarker for ADHD. However, these preliminary findings need to be replicated in larger ADHD and comparison cohorts

Monoaminergic dysregulation is implicated in attention-deficit/hyperactivity disorder (ADHD), and methylphenidate and amphetamines are the most frequently prescribed pharmacological agents for treating ADHD. However, it has recently been proposed that the core symptoms of the disorder might be due to an imbalance between monoaminergic and cholinergic systems. In this study, we used fibroblast cell homogenates from boys with and without ADHD as an extraneural cell model to examine the cholinergic receptor density, that is, muscarinic acetylcholine receptors (mAChRs). We found that the binding capacity (Bmax) of [³H] Quinuclidinyl benzilate (³H-QNB) to mAChRs was decreased by almost 50 % in the children with ADHD (mean = 30.6 fmol/mg protein, SD = 25.6) in comparison with controls [mean = 63.1 fmol/mg protein, SD = 20.5, p ≤ 0.01 (Student's unpaired t test)]. The decreased Bmax indicates a reduced cholinergic receptor density, which might constitute a biomarker for ADHD. However, these preliminary findings need to be replicated in larger ADHD and comparison cohorts

Monoaminergic dysregulation is implicated in attention-deficit/hyperactivity disorder (ADHD), and methylphenidate and amphetamines are the most frequently prescribed pharmacological agents for treating ADHD. However, it has recently been proposed that the core symptoms of the disorder might be due to an imbalance between monoaminergic and cholinergic systems. In this study, we used fibroblast cell homogenates from boys with and without ADHD as an extraneural cell model to examine the cholinergic receptor density, that is, muscarinic acetylcholine receptors (mAChRs). We found that the binding capacity (Bmax) of [³H] Quinuclidinyl benzilate (³H-QNB) to mAChRs was decreased by almost 50 % in the children with ADHD (mean = 30.6 fmol/mg protein, SD = 25.6) in comparison with controls [mean = 63.1 fmol/mg protein, SD = 20.5, p ≤ 0.01 (Student's unpaired t test)]. The decreased Bmax indicates a reduced cholinergic receptor density, which might constitute a biomarker for ADHD. However, these preliminary findings need to be replicated in larger ADHD and comparison cohorts

Cevimeline, a therapeutic drug for xerostomia, is an agonist of muscarinic acetylcholine receptors (mAChRs), and directly stimulates the peripheral mAChRs of the salivary glands. Since cevimeline is distributed in the brain after its oral administration, it is possible that it affects the central nervous system. However, it is unknown how cevimeline affects the superior salivatory (SS) neurons, which control submandibular salivation. In the present study, we examined the effects of cevimeline on the SS neurons using the whole-cell patch-clamp technique in brain slices. In Wistar rats (6-10 days), the SS neurons were retrogradely labeled by Texas Red applied to the chorda-lingual nerve. Two days after injection, whole-cell recordings were obtained from the labeled cells, and miniature excitatory postsynaptic currents (mEPSCs) were examined. Cevimeline induced the inward currents dose-dependently and increased the frequency of mEPSCs. Therefore, it is suggested that cevimeline enhances the excitability via post- and presynaptic muscarinic receptors in the rat SS neurons. In conclusion, cevimeline may enhance the excitability of the SS neurons.

The phosphodiesterase (PDE) 5 inhibitor sildenafil has been shown to display psychotropic actions in humans and animals, and has been used for the treatment of antidepressant-associated erectile dysfunction. However, its effects on the neurobiology of depression are unknown. Nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) inhibition is anti-depressant in animals, and increasing cGMP with sildenafil is anxiogenic in rodents. Substantial cholinergic-nitrergic interaction exists in the brain, while sildenafil shows modulatory actions on cholinergic transmission. Depression is also associated with increased cholinergic drive. Here we report that sildenafil increases muscarinic acetylcholine receptor (mAChR) signaling in human neuroblastoma cells. We also show that fluoxetine (20 mg/kg/day × 7 days), as well as a combination of sildenafil (10 mg/kg/day × 7 days) plus the antimuscarinic atropine (1 mg/kg/day × 7 days) demonstrates significant, comparable antidepressant-like effects in the rat forced swim test (FST) and also reduces cortical β-adrenergic receptor (β-AR) density, while sildenafil or atropine alone did not. Importantly, sildenafil did not modify fluoxetine’s response. Sildenafil thus demonstrates antidepressant-like effects but only after central muscarinic receptor blockade, providing evidence for cholinergic-nitrergic interactions in the neurobiology of depression

The phosphodiesterase (PDE) 5 inhibitor sildenafil has been shown to display psychotropic actions in humans and animals, and has been used for the treatment of antidepressant-associated erectile dysfunction. However, its effects on the neurobiology of depression are unknown. Nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) inhibition is anti-depressant in animals, and increasing cGMP with sildenafil is anxiogenic in rodents. Substantial cholinergic-nitrergic interaction exists in the brain, while sildenafil shows modulatory actions on cholinergic transmission. Depression is also associated with increased cholinergic drive. Here we report that sildenafil increases muscarinic acetylcholine receptor (mAChR) signaling in human neuroblastoma cells. We also show that fluoxetine (20 mg/kg/day × 7 days), as well as a combination of sildenafil (10 mg/kg/day × 7 days) plus the antimuscarinic atropine (1 mg/kg/day × 7 days) demonstrates significant, comparable antidepressant-like effects in the rat forced swim test (FST) and also reduces cortical β-adrenergic receptor (β-AR) density, while sildenafil or atropine alone did not. Importantly, sildenafil did not modify fluoxetine’s response. Sildenafil thus demonstrates antidepressant-like effects but only after central muscarinic receptor blockade, providing evidence for cholinergic-nitrergic interactions in the neurobiology of depression

The expression of mRNA coding for m2 subtype of muscarinic cholinergic receptors was assessed in dorsal root ganglia (DRG) of 15-day post-natal rats. Northern blot analysis on total RNA using a mixture of two different oligonucleotide probes, indicated the presence of a single prominent band of approximately 6.5 kb in rat DRG; a band of the same size was observed both in brainstem and cortex taken as positive controls. Analysis by RT-PCR of the mRNA coding for a region of the third cytoplasmic loop of m2 receptor showed a single signal both in rat DRG and hippocampus. In situ hybridization was then used to identify the neuronal subpopulations expressing the mRNA for M2. The transcripts were preferentially localized in medium-small neurons of the ganglion as well as in satellite cells surrounding the neuron cell body. Large neurons were usually negative. Finally, competition binding experiments, performed in the presence of [3H]-quinuclidinyl benzilate (QNB) and methoctramine (a selective competitor for M2 receptors), demonstrated the presence of M2 receptor protein (K(i) = 100 nM), as previously observed in chick DRG. The preferential localization of M2 in medium-small neurons of the ganglion suggests the involvement of this receptor subtype in the transduction of nociceptive stimuli.

type:text, This investigation confirmed decreases in muscarinic acetylcholine receptor (mAChR) concentration in the hippocampus of patients with senile dementia of Alzheimer-type (SDAT) using a binding assay with homogenate. We performed quantitative in vitro autoradiography of mAChR in the hippocampus obtained at autopsy from non-demented controls and SDAT patients, who had a marked memory disturbance since the early stage. 3H-QNB (1.09nM) and 3H-pirenzepine (18.0nM) were used as ligands to label total mAChR and Ml receptors, respectively. Total mAChR and Ml receptor densities showed a decrease by about 20% and 25%, respectively, in the CA1 area of SDAT cases, when compared with that of normal control cases. No changes were observed in the dentate gyrus, CA3, subiculum or entorhinal cortex. These observations are thought to be partly causative of the marked memory deficits which appeared in these cases.

This investigation confirmed decreases in muscarinic acetylcholine receptor (mAChR) concentration in the hippocampus of patients with senile dementia of Alzheimer-type (SDAT) using a binding assay with homogenate. We performed quantitative in vitro autoradiography of mAChR in the hippocampus obtained at autopsy from non-demented controls and SDAT patients, who had a marked memory disturbance since the early stage. 3H-QNB (1.09nM) and 3H-pirenzepine (18.0nM) were used as ligands to label total mAChR and Ml receptors, respectively. Total mAChR and Ml receptor densities showed a decrease by about 20% and 25%, respectively, in the CA1 area of SDAT cases, when compared with that of normal control cases. No changes were observed in the dentate gyrus, CA3, subiculum or entorhinal cortex. These observations are thought to be partly causative of the marked memory deficits which appeared in these cases.

This investigation confirmed decreases in muscarinic acetylcholine receptor (mAChR) concentration in the hippocampus of patients with senile dementia of Alzheimer-type (SDAT) using a binding assay with homogenate. We performed quantitative in vitro autoradiography of mAChR in the hippocampus obtained at autopsy from non-demented controls and SDAT patients, who had a marked memory disturbance since the early stage. 3H-QNB (1.09nM) and 3H-pirenzepine (18.0nM) were used as ligands to label total mAChR and Ml receptors, respectively. Total mAChR and Ml receptor densities showed a decrease by about 20% and 25%, respectively, in the CA1 area of SDAT cases, when compared with that of normal control cases. No changes were observed in the dentate gyrus, CA3, subiculum or entorhinal cortex. These observations are thought to be partly causative of the marked memory deficits which appeared in these cases.

This investigation confirmed decreases in muscarinic acetylcholine receptor (mAChR) concentration in the hippocampus of patients with senile dementia of Alzheimer-type (SDAT) using a binding assay with homogenate. We performed quantitative in vitro autoradiography of mAChR in the hippocampus obtained at autopsy from non-demented controls and SDAT patients, who had a marked memory disturbance since the early stage. 3H-QNB (1.09nM) and 3H-pirenzepine (18.0nM) were used as ligands to label total mAChR and Ml receptors, respectively. Total mAChR and Ml receptor densities showed a decrease by about 20% and 25%, respectively, in the CA1 area of SDAT cases, when compared with that of normal control cases. No changes were observed in the dentate gyrus, CA3, subiculum or entorhinal cortex. These observations are thought to be partly causative of the marked memory deficits which appeared in these cases.