Radiotherapy is an important therapeutic approach against cervical cancer but associated with adverse effects including vaginal fibrosis and dyspareunia. We here assessed the immunological and oxidative responses to cervical irradiation in an animal model for radiation-induced cervicitis. Rats were sedated and either exposed to 20 Gy of ionising radiation given by a linear accelerator or only sedated (controls) and euthanized 1-14 days later. The expressions of toll-like receptors (TLRs) and coupled intracellular pathways in the cervix were assessed with immunohistofluorescence and western blot. Expression of cytokines were analysed with the Bio-Plex Suspension Array System (Bio-Rad). We showed that TLRs 2-9 were expressed in the rat cervix and cervical irradiation induced up-regulation of TLR5, TRIF and NF-κB. In the irradiated cervical epithelium, TLR5 and TRIF were increased in concert with an up-regulation of oxidative stress (8-OHdG) and antioxidant enzymes (SOD-1 and catalase). G-CSF, M-CSF, IL-10, IL- 17A, IL-18 and RANTES expressions in the cervix decreased two weeks after cervical irradiation. In conclusion, the rat uterine cervix expresses the TLRs 2-9. Cervical irradiation induces immunological changes and oxidative stress, which could have importance in the development of adverse effects to radiotherapy.

• MeSH
• Adaptor Proteins, Vesicular Transport immunology   MeSH
• Cervix Uteri immunology   pathology   MeSH
• Cytokines immunology   MeSH
• Radiation Injuries, Experimental immunology   pathology   MeSH
• Rats MeSH
• NF-kappa B immunology   MeSH
• Oxidative Stress immunology   radiation effects   MeSH
• Rats, Sprague-Dawley MeSH
• Gene Expression Regulation immunology   radiation effects   MeSH
• Toll-Like Receptors immunology   MeSH
• Uterine Cervicitis immunology   pathology   MeSH
• Gamma Rays adverse effects   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Currently, we have assessed the neuronal control of the urinary bladder in radiation cystitis and whether interstitial cells contribute to the condition. Fourteen days after bladder irradiation (20 Gy), rats were sedated and the urinary bladder was cut into two sagittal halves where electrical field stimulation (EFS; 5-20 Hz) was applied on the pelvic nerve afferents or stretch (80 mN) on one-half of the bladder, while contractions were registered on the contralateral half of the bladder in the absence and presence of increasing doses of imatinib (1-10 mg/kg; inhibitor of c-kit-positive interstitial cells), atropine (1 mg/kg; to block muscarinic M3 receptors), or pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (2 mg/kg; P2X1 purinoceptor antagonist). Urinary bladders were also excised for organ bath experiments, Western blot, quantitative polymerase chain reaction, and immunohistochemistry. In vivo, EFS contractions were enhanced after irradiation, and imatinib (1-10 mg/mg) inhibited contractions by EFS and stretched dose-dependently in controls but not in irradiated bladders. In the irradiated bladder in vitro, atropine resistance was increased and imatinib (100 µM) inhibited contractions by EFS and agonists (ATP, methacholine) in irradiated bladders and controls. The urinary bladder expressions of P2X1 purinoceptors, muscarinic M3 receptor, choline acetyltransferase, c-kit, and the agonist of c-kit, stem cell factor, were not changed by irradiation. In conclusion, bladder irradiation affects several levels of neuronal control of the urinary bladder. Interstitial cells may contribute to some of the symptoms associated with radiation cystitis.

• MeSH
• Cystitis pathology   physiopathology   MeSH
• Rats MeSH
• Urinary Bladder pathology   physiopathology   MeSH
• Disease Models, Animal MeSH
• Neurons pathology   MeSH
• Muscle Contraction MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

BACKGROUND: Pharmaceuticals with targets in the cholinergic transmission have been used for decades and are still fundamental treatments in many diseases and conditions today. Both the transmission and the effects of the somatomotoric and the parasympathetic nervous systems may be targeted by such treatments. Irrespective of the knowledge that the effects of neuronal signalling in the nervous systems may include a number of different receptor subtypes of both the nicotinic and the muscarinic receptors, this complexity is generally overlooked when assessing the mechanisms of action of pharmaceuticals. METHODS: We have search of bibliographic databases for peer-reviewed research literature focused on the cholinergic system. Also, we have taken advantage of our expertise in this field to deduce the conclusions of this study. RESULTS: Presently, the life cycle of acetylcholine, muscarinic receptors and their effects are reviewed in the major organ systems of the body. Neuronal and non-neuronal sources of acetylcholine are elucidated. Examples of pharmaceuticals, in particular cholinesterase inhibitors, affecting these systems are discussed. The review focuses on salivary glands, the respiratory tract and the lower urinary tract, since the complexity of the interplay of different muscarinic receptor subtypes is of significance for physiological, pharmacological and toxicological effects in these organs. CONCLUSION: Most pharmaceuticals targeting muscarinic receptors are employed at such large doses that no selectivity can be expected. However, some differences in the adverse effect profile of muscarinic antagonists may still be explained by the variation of expression of muscarinic receptor subtypes in different organs. However, a complex pattern of interactions between muscarinic receptor subtypes occurs and needs to be considered when searching for selective pharmaceuticals. In the development of new entities for the treatment of for instance pesticide intoxication, the muscarinic receptor selectivity needs to be considered. Reactivators generally have a muscarinic M2 receptor acting profile. Such a blockade may engrave the situation since it may enlarge the effect of the muscarinic M3 receptor effect. This may explain why respiratory arrest is the major cause for deaths by esterase blocking.

• MeSH
• Cholinesterase Inhibitors pharmacology   MeSH
• Receptor Cross-Talk drug effects   MeSH
• Humans MeSH
• Receptors, Muscarinic drug effects   MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Review MeSH

Organophosphorus nerve agents inhibit acetylcholinesterase (AChE) which causes the breakdown of the transmitter acetylcholine (ACh) in the synaptic cleft. Overstimulation of cholinergic receptors (muscarinic and nicotinic) by excessive amounts of ACh causes several health problems and may even cause death. Reversible AChE inhibitors play an important role in prophylaxis against nerve agents. The presented study investigated whether 7-methoxytacrine (7-MEOTA) and 7-MEOTA-donepezil derivatives can act as central and peripheral reversible AChE inhibitors and simultaneously antagonize muscarinic and nicotinic receptors. The possible mechanism of action was studied on cell cultures (patch clamp technique, calcium mobilization assay) and on isolated smooth muscle tissue (contraction study). Furthermore, the kinetics of the compounds were also examined. CNS availability was predicted by determining the passive blood-brain barrier penetration estimated via a modified PAMPA assay. In conclusion, this study provides promising evidence that the new synthesized 7-MEOTA-donepezil derivatives have the desired anticholinergic effect; they can inhibit AChE, and nicotinic and muscarinic receptors in the micromolar range. Furthermore, they seem to penetrate readily into the CNS. However, their real potency and benefit must be verified by in vivo experiments.

• MeSH
• Muscarinic Antagonists administration & dosage   MeSH
• Cholinergic Antagonists administration & dosage   MeSH
• Indans administration & dosage   MeSH
• Muscle Fibers, Skeletal drug effects   metabolism   MeSH
• Rats MeSH
• Cells, Cultured MeSH
• Nicotinic Antagonists administration & dosage   MeSH
• Piperidines administration & dosage   MeSH
• Rats, Wistar MeSH
• Tacrine administration & dosage   analogs & derivatives   MeSH
• Dose-Response Relationship, Drug MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Obidoxime, a weak acetylcholine-esterase (AChE) inhibitor, exerts muscarinic receptor antagonism with a significant muscarinic M2 receptor selective profile. The current examinations aimed to determine the functional significance of muscarinic M2 receptors in the state of AChE inhibition, elucidating muscarinic M2 and M3 receptor interaction. In the in vitro examinations, methacholine evoked concentration-dependent bladder contractile and atrial frequency inhibitory responses. Although atropine abolished both, methoctramine (1 μmol/L) only affected the cholinergic response in the atrial preparations. However, in the presence of methoctramine, physostigmine, an AChE inhibitor, increased the basal tension of the bladder strip preparations (+68%), as well as the contractile responses to low concentrations of methacholine (< 5 μmol/L; +90-290%). In contrast to physostigmine, obidoxime alone raised the basal tension (+58%) and the responses to low concentrations of methacholine (< 5 μmol/L; +80-450%). Physostigmine concentration-dependently increased methacholine-evoked responses, similarly to obidoxime at low concentrations. However, at large concentrations (> 5 μmol/L), obidoxime, because of its unselective muscarinic receptor antagonism, inhibited the methacholine bladder responses. In conclusion, the current results show that muscarinic M2 receptors inhibit muscarinic M3 receptor-evoked contractile responses to low concentrations of acetylcholine in the synaptic cleft. The muscarinic M2 and M3 receptor crosstalk could be a counteracting mechanism in the treatment of AChE inhibition when using reactivators, such as obidoxime.

• MeSH
• Muscarinic Antagonists pharmacology   MeSH
• Atropine pharmacology   MeSH
• Cholinesterase Inhibitors pharmacology   MeSH
• Diamines pharmacology   MeSH
• Physostigmine pharmacology   MeSH
• Receptor Cross-Talk MeSH
• Rats MeSH
• Methacholine Chloride pharmacology   MeSH
• Urinary Bladder drug effects   enzymology   metabolism   MeSH
• Obidoxime Chloride pharmacology   MeSH
• Receptor, Muscarinic M2 antagonists & inhibitors   MeSH
• Receptor, Muscarinic M3 antagonists & inhibitors   MeSH
• Heart Atria drug effects   enzymology   metabolism   MeSH
• Muscle Contraction drug effects   MeSH
• In Vitro Techniques MeSH
• Dose-Response Relationship, Drug MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Nitric oxide (NO) has pivotal roles in cyclophosphamide- (CYP-) induced cystitis during which mucosal nitric oxide synthase (NOS) and muscarinic M5 receptor expressions are upregulated. In cystitis, urothelial muscarinic NO-linked effects hamper contractility. Therefore we wondered if a blockade of this axis also affects the induction of cystitis in the rat. Rats were pretreated with saline, the muscarinic receptor antagonist 4-DAMP (1 mg/kg ip), or the NOS inhibitor L-NAME (30 mg/kg ip) for five days. 60 h before the experiments the rats were treated with saline or CYP. Methacholine-, ATP-, and adenosine-evoked responses were smaller in preparations from CYP-treated rats than from saline-treated ones. Pretreatment with 4-DAMP did not change this relation, while pretreatment with L-NAME normalized the responses in the CYP-treated animals. The functional results were strengthened by the morphological observations; 4-DAMP pretreatment did not affect the parameters studied, namely, expression of muscarinic M5 receptors, P1A1 purinoceptors, mast cell distribution, or bladder wall enlargement. However, pretreatment with L-NAME attenuated the differences. Thus, the current study provides new insights into the complex mechanisms behind CYP-induced cystitis. The NO effects coupled to urothelial muscarinic receptors have a minor role in the development of cystitis. Inhibition of NOS may prevent the progression of cystitis.

• MeSH
• Adenosine pharmacology   MeSH
• Adenosine Triphosphate pharmacology   MeSH
• Cyclophosphamide MeSH
• Cystitis drug therapy   pathology   physiopathology   MeSH
• Immunohistochemistry MeSH
• Enzyme Inhibitors pharmacology   therapeutic use   MeSH
• Mast Cells drug effects   pathology   MeSH
• Methacholine Chloride pharmacology   MeSH
• Urinary Bladder drug effects   pathology   physiopathology   MeSH
• Rats, Sprague-Dawley MeSH
• Receptors, Purinergic metabolism   MeSH
• Receptors, Muscarinic metabolism   MeSH
• Muscle Contraction drug effects   MeSH
• Nitric Oxide Synthase antagonists & inhibitors   metabolism   MeSH
• In Vitro Techniques MeSH
• Animals MeSH
• Check Tag
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Alzheimer´s disease (AD) is a progressive neurodegenerative dementia which currently represents one of the biggest threats for the human kind. The cure is still unknown and various hypotheses (cholinergic, amyloidal, oxidative, vascular etc.) are investigated in order to understand the pathophysiology of the disease and on this basis find an effective treatment. Tacrine, the first approved drug for the AD disease treatment, has been reported to be a multitargeted drug, however it was withdrawn from the market particularly due to its hepatotoxicity. Its derivative 7-methoxytacrine (7- MEOTA) probably due to the different metabolization does not exert this side effect. The aim of our study was to compare these two cholinesterase inhibitors from various, mainly cholinergic, points of view relevant for a potential AD drug. We found that 7-MEOTA does not fall behind its more well-known parent compound - tacrine. Furthermore, we found, that 7-MEOTA exerts better properties in most of the tests related to a possible AD treatment. Only the pharmacokinetics and a higher acetylcholinesterase and butyrylcholinesterase inhibitory potency would slightly give advantages to tacrine over 7-MEOTA, but concerning its lower toxicity, better antioxidant properties, interaction with muscarinic and nicotinic receptors and "safer" metabolization provide strong evidence for reconsider 7-MEOTA and its derivatives as candidate molecules for the treatment of AD.

• MeSH
• Cholinesterase Inhibitors pharmacology   MeSH
• Rats MeSH
• Oxidative Stress drug effects   MeSH
• Rats, Wistar MeSH
• Tacrine analogs & derivatives   pharmacology   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Comparative Study MeSH

The aim of the present study was to investigate the relaxatory function of adenosine receptor subtypes in rat urinary bladder, and if it is altered in the state of inflammation. The in vitro responses to the P1 receptor agonist adenosine were investigated in the presence of the general P2 receptor antagonist pyridoxal-phosphate-6-azophenyl-2',4'-disulfonic acid (PPADS; 1*10(-4)M). Experiments were performed on preparations from normal (healthy) rats and rats with cyclophosphamide (CYP; 100mg kg(-1) i. p.)-induced cystitis. The specific P1A(1) antagonist 1,3-dipropyl-8-cyclopentylxanthine (DPCPX; 1*10(-5)M) decreased the adenosine relaxatory response in normal bladders (-60%), but not in preparations from CYP pre-treated rats. Immunohistochemical findings support the hypothesis that the expression of P1A(1) receptors in the rat urinary bladder is decreased during cystitis. The adenosine-evoked relaxation was not affected by the specific P1A(2A) antagonist SCH 58261 (3*10(-7)M), neither in normal nor in CYP pre-treated rats. The relaxation to adenosine was, however, significantly increased by the specific P1A(3) antagonist MRS 1523 (1*10(-5)M) in preparations from both normal and CYP pre-treated rats, suggesting P1A(3) to be mediating bladder contraction. Thus, in the rat urinary bladder the relaxation to adenosine is mainly due to the P1A(1) receptor, while the P1A(3) receptor seems to be responsible for contractile responses. The DPCPX-resistant part of the relaxation is possibly due to the P1A(2B) receptor, the fourth subtype of the adenosine receptor family.

• MeSH
• Adenosine pharmacology   physiology   MeSH
• Purinergic P1 Receptor Agonists pharmacology   MeSH
• Purinergic P1 Receptor Antagonists pharmacology   MeSH
• Cystitis metabolism   physiopathology   MeSH
• Muscle, Smooth drug effects   metabolism   physiopathology   MeSH
• Rats MeSH
• Drug Resistance drug effects   physiology   MeSH
• Inflammation Mediators physiology   MeSH
• Urinary Bladder drug effects   metabolism   physiology   MeSH
• Disease Models, Animal MeSH
• Rats, Sprague-Dawley MeSH
• Purinergic P2 Receptor Antagonists pharmacology   MeSH
• Receptor, Adenosine A1 physiology   MeSH
• Muscle Contraction drug effects   physiology   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Obidoxime, a well-known bis-pyridinium reactivator, is often the preferred antidote of organophosphorus poisoning caused by pesticides and tabun. It is also considered to be an allosteric modulator of muscarinic receptors, preferably M2 sub-type. This study compared the effect of obidoxime and atropine in vivo and in vitro on the cholinergic stimulation of the rat heart (M2) and the urinary bladder (M3). The results showed that obidoxime exerts anti-muscarinic effects, that may play an important role in the treatment of organophosphourus poisoning, and that the muscarinic receptor inhibition profile shows M2 receptor selectivity. This anti-muscarinic effect is much smaller that the effect of atropine and might be due to the allosteric inhibition of the receptors. The results also indicate that the acetylcholinesterase inhibition and the muscarinic receptor antagonism occur at different concentrations and dose levels.

• MeSH
• Muscarinic Antagonists pharmacology   MeSH
• Atropine pharmacology   MeSH
• Cholinesterase Inhibitors pharmacology   MeSH
• Rats MeSH
• Urinary Bladder drug effects   MeSH
• Molecular Structure MeSH
• Obidoxime Chloride chemistry   pharmacology   MeSH
• Organophosphorus Compounds MeSH
• Organophosphate Poisoning MeSH
• Cholinesterase Reactivators chemistry   pharmacology   MeSH
• Heart Rate drug effects   MeSH
• Heart Atria drug effects   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Male MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

Trimedoxime is a bisquaternary oxime that is widely used in the treatment of organophosphorous poisoning caused by tabun and paraoxon. We tested its affinity to acetylcholinesterase (AChE), its mechanism of interaction and effect on the cholinergic system of the rat bladder. The half maximal inhibitory concentration (IC50) of trimedoxime to recombinant AChE was found to be 82.0 mM ± 30.1 mM. This represents a weak inhibition. Its interaction with AChE seems to be very similar to obidoxime - one aromatic nucleus interacts with the peripheral anionic site and the other with the residues TYR337 and TYR341 inside the cavity. Also the oxime moiety is moving towards the catalytic triade ready for the reactivation of the inhibited AChE. In the organ bath experiment no significant effect of trimedoxime was observed on the contraction of the detrusor caused by the muscarinic agonist metacholine.

• Keywords
• acetylcholinesterase, trimedoxime, antidote, muscaricnic receptors, reactivation,
• MeSH
• Acetylcholinesterase pharmacokinetics   pharmacology   drug effects   MeSH
• Antidotes pharmacology   therapeutic use   MeSH
• Cholinergic Antagonists pharmacokinetics   pharmacology   therapeutic use   MeSH
• Cholinergic Agents pharmacology   immunology   isolation & purification   MeSH
• Animal Experimentation MeSH
• Financing, Organized MeSH
• Urinary Bladder MeSH
• Organophosphorus Compounds pharmacokinetics   pharmacology   toxicity   MeSH
• Pesticides pharmacokinetics   pharmacology   toxicity   MeSH
• Rats, Sprague-Dawley MeSH
• Cholinesterase Reactivators pharmacokinetics   pharmacology   therapeutic use   MeSH
• Receptors, Muscarinic drug effects   MeSH
• Drug Synergism MeSH
• Trimedoxime pharmacokinetics   pharmacology   metabolism   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH