Guinea-pig Taenia Coli Research Articles

P2X3 receptors participate in purinergic inhibition of gastrointestinal smooth muscle.

The ATP analogue α,β-meATP is a potent relaxant of gastrointestinal smooth muscle, but its molecular target is uncertain inside the gut. α,β-meATP relaxed the carbachol-precontracted guinea-pig taenia coli in a concentration-dependent manner (EC50, 2.0±0.1μM). A luciferase-based assay confirmed that α,β-meATP solutions were minimally contaminated with ATP. α,β-meATP-evoked relaxations were inhibited by the competitive P2Y1 antagonist MRS2179 (pA2=5.36), but also by the competitive P2X3 antagonist, A-317491 (pA2=5.51). When MRS2179 and A-317491 were applied together, residual α,β-meATP responses converted from brief to prolonged relaxations. Sodium nitroprusside (a nitric oxide donor) also caused prolonged relaxations. Immunohistochemistry revealed that P2X3 receptors were present in myenteric ganglion cells and their varicose nerve terminals. The amplitude of α,β-meATP responses was not inhibited by TTX (NaV channel blocker) and ωCgTx (N-type CaV channel blocker). However, responses to α,β-meATP were inhibited by TEA (non-selective K+-channel blocker), indicating that relaxations involved opening K+-channels. The findings of this study are consistent with the conclusion that α,β-meATP stimulates Ca2+-permeable P2X3 receptors on varicose nerve terminals to release inhibitory nucleotides: 1) ATP and β-NAD release results in P2Y1-mediated brief relaxations; 2) another released transmitter (possibly NO) results in prolonged relaxations. Prejunctional P2X3 receptors represent a purinergic feed-forward mechanism to augment the action of inhibitory nerves on gut motility. This positive feed-forward mechanism may counter-balance the known negative feedback mechanism caused by adenosine and prejunctional A1 receptors on inhibitory motor nerves.

Introduction to Purinergic Signaling.

Purinergic signaling was proposed in 1972, after it was demonstrated that adenosine 5'-triphosphate (ATP) was a transmitter in nonadrenergic, noncholinergic inhibitory nerves supplying the guinea-pig taenia coli. Later, ATP was identified as an excitatory cotransmitter in sympathetic and parasympathetic nerves, and it is now apparent that ATP acts as a cotransmitter in most, if not all, nerves in both the peripheral nervous system and central nervous system (CNS). ATP acts as a short-term signaling molecule in neurotransmission, neuromodulation, and neurosecretion. It also has potent, long-term (trophic) roles in cell proliferation, differentiation, and death in development and regeneration. Receptors to purines and pyrimidines have been cloned and characterized: P1 adenosine receptors (with four subtypes), P2X ionotropic nucleotide receptors (seven subtypes) and P2Y metabotropic nucleotide receptors (eight subtypes). ATP is released from different cell types by mechanical deformation, and after release, it is rapidly broken down by ectonucleotidases. Purinergic receptors were expressed early in evolution and are widely distributed on many different nonneuronal cell types as well as neurons. Purinergic signaling is involved in embryonic development and in the activities of stem cells. There is a growing understanding about the pathophysiology of purinergic signaling and there are therapeutic developments for a variety of diseases, including stroke and thrombosis, osteoporosis, pain, chronic cough, kidney failure, bladder incontinence, cystic fibrosis, dry eye, cancer, and disorders of the CNS, including Alzheimer's, Parkinson's. and Huntington's disease, multiple sclerosis, epilepsy, migraine, and neuropsychiatric and mood disorders.

ROLE OF Na+, K+, 2Cl– -COTRANSPORTER AND POTASSIUM CONDUCTIVITY IN THE REALIZATION OF THE EFFECT OF CARBON MONOXIDE, IN THE SMOOTH MUSCLE OF VISCERAL ORGANS

The aim of this study was to investigate the role of the Na + , K + , 2Cl – -transport and potassium conductance, in the implementation of the effects of carbon monoxide in the SMC taenia coli and ureter of guinea pig. Materials and methods. The study was conducted by the method of double saccharose bridge. Was studied the effects of carbon monoxide donor CORM2 in ureter and smooth muscle of guinea pig of taenia coli, in normal Krebs solution, against inhibitor-bumetanide and potassium channel blocker tetraethylammonium chloride (TEA). Studied the effects of bumetanide in normal Krebs solution and the background of TEA. Main results. CORM2 causes inhibition of contractile activity of SMC taenia coli and guinea pig ureter. Its effect is weakened by bumetanide and TEA. Bumetanide causes inhibition of contractile activity of SMC taenia coli, and its effect is weakened by TEA. Conclusions. In this way, it shows that there are tissue-specific mechanisms of interrelation effects CO and ion conductivity.

Bitter taste receptor agonists mediate relaxation of human and rodent vascular smooth muscle

Taste-sensing type 2 receptors (TAS2Rs) have been implicated in extraoral functions. Airway smooth muscle expresses TAS2Rs and is strongly relaxed by TAS2R agonists. We hypothesised that TAS2R agonists might affect vascular smooth muscle as well. Moreover, the general pharmacological profile of TAS2R agonists, which are used to investigate the functions of TAS2R׳s, are undefined. The aim of this study was to pharmacologically characterise the effects of five prototype TAS2R agonists in vascular smooth muscle. Responses to the TAS2R agonists were investigated in guinea-pig aorta and taenia coli, mouse aorta (wild-type and caveolin-1−/− mice) and human pulmonary arteries. Chloroquine, denatonium, dextromethorphan, noscapine and quinine, agonists for TAS2R3, TAS2R4, TAS2R10 and TAS2R14, induced strong endothelium-independent relaxations (responses between 82–96% of maximal relaxations) in phenylephrine pre-contracted guinea-pig aorta that persisted in the presence of L-type Ca2+ and KCa1.1-channel blockers. Experiments in guinea-pig taenia coli revealed that denatonium and quinine also inhibited relaxations to phenylephrine, indicating antagonism of α-adrenoceptors. Only chloroquine and noscapine mediated relaxations when the guinea pig aorta was pre-contracted by U-46619 or PGF2α. Relaxations to chloroquine and noscapine after U-46619 pre-contractions were however markedly impaired in aortae from caveolin-1−/− mice. Chloroquine and noscapine mediated relaxations of human pulmonary arteries that expressed also mRNA for TAS2R3, TAS2R4, TAS2R10 and TAS2R14, at levels similar to that of the α1A adrenoceptor. Notwithstanding whether TAS2Rs are involved or not, TAS2R agonists have profound effects on vascular smooth muscle. Chloroquine and noscapine are of special interest as their effects cannot be accounted for by conventional pathways.

Purinergic signalling: from discovery to current developments.

New FindingsWhat is the topic of this review?This is a personal historical review about the discovery and the main conceptual advances leading to our current understanding of purinergic signalling. The contributions of leading figures in the field are acknowledged. It includes the discovery of purinergic neuromuscular and synaptic transmission, cotransmission, the identification of P1 (adenosine), P2X nucleotide ion channel and P2Y nucleotide G protein-coupled receptors, the identity of ectonucleotidases and release of ATP from cells by mechanical stimulation and mechanosensory transduction.What advances does it highlight?It highlights the pathophysiology of purinergic signalling and recent therapeutic developments.This lecture is about the history of the purinergic signalling concept. It begins with reference to the paper by Paton & Vane published in 1963, which identified non-cholinergic relaxation in response to vagal nerve stimulation in several species, although they suggested that it might be due to sympathetic adrenergic nerves in the vagal nerve trunk. Using the sucrose gap technique for simultaneous mechanical and electrical recordings in smooth muscle (developed while in Feldberg’s department in the National Institute for Medical Research) of the guinea-pig taenia coli preparation (learned when working in Edith Bülbring’s smooth muscle laboratory in Oxford Pharmacology), we showed that the hyperpolarizations recorded in the presence of antagonists to the classical autonomic neurotransmitters, acetylcholine and noradrenaline, were inhibitory junction potentials in response to non-adrenergic, non-cholinergic neurotransmission, mediated by intrinsic enteric nerves controlled by vagal and sacral parasympathetic nerves. We then showed that ATP satisfied the criteria needed to identify a neurotransmitter released by these nerves. Subsequently, it was shown that ATP is a cotransmitter in all nerves in the peripheral and central nervous systems. The receptors for purines and pyrimidines were cloned and characterized in the early 1990s, and immunostaining showed that most non-neuronal cells as well as nerve cells expressed these receptors. The physiology and pathophysiology of purinergic signalling is discussed.

Convergent Synthesis of the Potent P2Y Receptor Antagonist MG 50-3-1 Based on a Regioselective Ullmann Coupling Reaction

MG 50-3-1 (3, trisodium 1-amino-4-{4-[4-chloro-6-(2-sulfophenylamino)-1,3,5-triazin-2-ylamino]-2-sulfophenylamino}-9,10-dioxo-9,10-dihydroanthracene 2-sulfonate) is the most potent and selective antagonist (IC50 4.6 nM) for “P2Y1-like” nucleotide-activated membrane receptors in guinea-pig taenia coli responsible for smooth muscle relaxation. Full characterization of the compound, however, e.g., at the human P2Y1 receptor, which is a novel potential target for antithrombotic drugs, as well as other P2 receptor subtypes, has been hampered due to difficulties in synthesizing the compound in sufficient quantity. MG 50-3-1 would be highly useful as a biological tool for detailed investigation of signal transduction in the gut. We have now developed a convenient, fast, mild, and efficient convergent synthesis of 3 based on retrosynthetic analysis. A new, regioselective Ullmann coupling reaction under microwave irradiation was successfully developed to obtain 1-amino-4-(4-amino-2-sulfophenylamino)-9,10-dioxo-9,10-dihydro­anthracene 2-sulfonate (8). Four different copper catalysts (Cu, CuCl, CuCl2, and CuSO4) were investigated at different pH values of sodium phosphate buffer, and in water in the absence or presence of base. Results showed that CuSO4 in water in the presence of triethylamine provided the best conditions for the regioselective Ullmann coupling reaction yielding the key intermediate compound 8. A new synthon (sodium 2-(4,6-dichloro-1,3,5-triazin-2-ylamino)benzenesulfonate, 13) which can easily be obtained on a gram scale was prepared, and 13 was successfully coupled with 8 yielding the target compound 3.

Purinergic signaling

AbstractThe concept of purinergic neurotransmission was proposed in 1972, after it was shown that adenosine 5′‐triphosphate (ATP) was a transmitter in non‐adrenergic, non‐cholinergic inhibitory nerves in the guinea pig taenia coli. Subsequently, ATP was identified as a cotransmitter in sympathetic and parasympathetic nerves, and it is now recognized that ATP acts as a cotransmitter in most nerves in both the peripheral nervous system and central nervous system (CNS). ATP acts as a fast excitatory neurotransmitter or neuromodulator and has potent long‐term (trophic) roles in cell proliferation, differentiation, and death in development and regeneration, as well as in disease. Three subclasses of receptors to purines and pyrimidines have been identified, P1 adenosine receptors (with four subtypes), P2X ionotropic nucleotide receptors (seven subtypes), and P2Y metabotropic nucleotide receptors (eight subtypes). ATP is released physiologically by many different cell types by mechanical deformation, and after release ATP undergoes rapid enzymatic degradation by ectonucleotidases. Purinergic receptors appeared early in evolution and have a widespread distribution on many different non‐neuronal cell types as well as neurons. There is evidence for the involvement of purinergic signaling in embryonic development and in the activities of stem cells. There is a rapidly growing literature about the pathophysiology of purinergic signaling, and there are therapeutic developments for a variety of diseases, including stroke and thrombosis, osteoporosis, kidney failure, bladder incontinence, cystic fibrosis, dry eye, cancer, and disorders of the CNS, such as Alzheimer's, Parkinson's, and Huntington's disease, multiple sclerosis, epilepsy, migraine, and neuropsychiatric disorders. WIREs Membr Transp Signal 2012, 1:116–125. doi: 10.1002/wmts.14For further resources related to this article, please visit the WIREs website.

Effect of vanadate on cooling-induced tension changes in K-depolarized smooth muscles from guinea-pigs

Sodium vanadate reversed cooling-induced relaxation of K-depolarized taenia coli of guinea-pigs but failed to reverse it in the portal vein and uterus. It potentiated cooling-induced contraction of K-depolarized vas deferens and ureter. These effects were not mediated by the inhibition of Na,K-ATPase, but by the inhibition of the Ca-pump.

Introductory overview of purinergic signalling

Purinergic neurotransmission was proposed in 1972 following identification of adenosine 5'-triphosphate (ATP) as the transmitter in non-adrenergic, non-cholinergic inhibitory nerves in guinea-pig taenia coli. Subsequently ATP was identified as a co-transmitter in sympathetic, parasympathetic and most nerves in the peripheral and central nervous systems. ATP acts as a short-term signalling molecule in neurotransmission, neuromodulation and secretion and has long-term (trophic) roles in cell proliferation, differentiation and death in development and regeneration. Three subclasses of purine and pyrimidine receptors have been identified, P1 adenosine receptors (4 subtypes), P2X ionotropic nucleotide receptors (7 subtypes) and P2Y metabotropic receptors (8 subtypes). ATP is released physiologically by many cell types by mechanical deformation and, after release, ATP undergoes ectonucleotidase degradation. Purinergic receptors appeared early in evolution and have a widespread distribution on many non-neuronal cells and neurons. Purinergic signalling is involved in embryonic and stem cell development. There is a rapidly growing literature about the pathophysiology of purinergic signalling including therapeutic developments for diseases, including stroke, thrombosis, osteoporosis, kidney failure, bladder incontinence, cystic fibrosis, dry eye, cancer and brain disorders.

Involvement of Secondary Intracellular Messengers in the Mechanisms of Purinergic Inhibition of Intestinal Smooth Muscles

We studied the role of adenylate cyclase and phospholipase C in the control of ATP-induced relaxation of carbachol-evoked contraction of smooth muscles of the guinea-pig taenia coli. We showed that ATP-induced relaxation of carbachol-caused contraction is completely realized under control conditions via activation of inositol trisphosphate-sensitive (InsP3) receptors of the sarcoplasmic reticulum of smooth muscle cells (SMCs). In the case where phospholipase C was blocked, the relaxing action of ATP on smooth muscles continues to be mediated mostly by activation of InsP3 receptors, but other mechanisms begin to participate in this process. Intracellular processes are also involved in ATP-induced relaxation where the signal is transferred from purine receptors via activation of phospholipase C under conditions of parallel activation of adenylate cyclase by forskolin; these processes also include activation of InsP3 receptors of the sarcoplasmic reticulum of SMCs and some other events. After U73122-induced blocking of phospholipase C and forskolin-induced activation of adenylate cyclase, ATP-caused relaxation can completely be removed by an inhibitor of InsP3 receptors, 2-APB. This indicates that, under the above conditions, such relaxation is realized exclusively via InsP3 receptors of the sarcoplasmic reticulum of SMCs. At the same time, ATP-induced relaxation caused by activation of phospholipase C and inactivation of adenylate cyclase is also nearly completely realized with involvement of InsP3 receptors of the sarcoplasmic reticulum. However, removing the activity of phospholipase C under conditions of blocking of adenylate cyclase and InsP3 receptors of the sarcoplasmic reticulum in SMCs leads to the recovery of ATP-induced relaxation with the participation of the other intracellular processes. Therefore, two intracellular messengers, phospholipase C and adenylate cyclase, are involved in purinergic inhibition of smooth muscles. Upon their action, multiple intracellular signal pathways are triggered. The level of their participation can be influenced by the initial functional state of intestinal SMCs. These changes are always directed toward the maintenance of normal functioning of the respective organs.

Kinase-related protein/telokin inhibits Ca2+-independent contraction in Triton-skinned guinea pig taenia coli

KRP (kinase-related protein), also known as telokin, has been proposed to inhibit smooth muscle contractility by inhibiting the phosphorylation of the rMLC (regulatory myosin light chain) by the Ca2+-activated MLCK (myosin light chain kinase). Using the phosphatase inhibitor microcystin, we show in the present study that KRP also inhibits Ca2+-independent rMLC phosphorylation and smooth muscle contraction mediated by novel Ca2+-independent rMLC kinases. Incubating KRP-depleted Triton-skinned taenia coli with microcystin at pCa>8 induced a slow contraction reaching 90% of maximal force (Fmax) at pCa 4.5 after approximately 25 min. Loading the fibres with KRP significantly slowed down the force development, i.e. the time to reach 50% of Fmax was increased from 8 min to 35 min. KRP similarly inhibited rMLC phosphorylation of HMM (heavy meromyosin) in vitro by MLCK or by the constitutively active MLCK fragment (61K-MLCK) lacking the myosin-docking KRP domain. A C-terminally truncated KRP defective in myosin binding inhibited neither force nor HMM phosphorylation. Phosphorylated KRP inhibited the rMLC phosphorylation of HMM in vitro and Ca2+-insensitive contractions in fibres similar to unphosphorylated KRP, whereby the phosphorylation state of KRP was not altered in the fibres. We conclude that (i) KRP inhibits not only MLCK-induced contractions, but also those elicited by Ca2+-independent rMLC kinases; (ii) phosphorylation of KRP does not modulate this effect; (iii) binding of KRP to myosin is essential for this inhibition; and (iv) KRP inhibition of rMLC phosphorylation is most probably due to the shielding of the phosphorylation site on the rMLC.

Multidrug Resistance Protein Transporter and Ins(1,4,5)P3-Sensitive Ca2+-Signaling Involved in Adenosine Triphosphate Export via Gq Protein–Coupled NK2-Receptor Stimulation With Neurokinin A

The purpose of this study is to identify the membrane transport machinery and cell signaling involved in the neurokinin A–inducible release of adenosine triphosphate (ATP) as an autocrine/paracrine signal from cultured guinea-pig taenia coli (T. coli) smooth muscle cells (SMCs). ATP release evoked by neurokinin A was inhibited by L-659877, a NK2-receptor antagonist; by modulators for Ins(1,4,5)P3-sensitive Ca2+-signaling, U-73122, thapsigargin, and 2-APB; and by W-7, a calmodulin inhibitor, and staurosporine, a protein kinase C (PKC) inhibitor, but not by wortmannin, a phosphoinositide 3-kinase inhibitor. The evoked release was suppressed by a multidrug resistance protein (MRP)-transporter inhibitors, MK-571, indomethacin, and benzbromarone, but not by CFTR-inh 172, a CFTR-Cl− channel blocker, and α-glycyrrhetinic acid, a gap junction hemichannel blocker. Neurokinin A caused a marked accumulation of Ins(1,4,5)P3 and an increase in [Ca2+]i in the cultured cells. These findings suggest that stimulation of Gq/11 protein– coupled NK2 receptor with neurokinin A caused a substantial release of ATP from cultured T. coli SMCs and that the evoked release may be mediated by Ins(1,4,5)P3-sensitive Ca2+-signaling, further by PKC and Ca2+/calmodulin signals, and finally by an activation of MRP transporters as the membrane device.

The intracellular action of Mn2+ in the Mn2+-induced contraction in the guinea-pig taenia coli

The intracellular action of Mn2+ in the Mn2+-induced contraction in the guinea-pig taenia coli

A constitutive model for smooth muscle including active tone and passive viscoelastic behaviour

A new constitutive model for the biomechanical behaviour of smooth muscle tissue is proposed. The active muscle contraction is accomplished by the relative sliding between actin and myosin filaments, comprising contractile units in the smooth muscle cells. The model includes a chemical part, governing the cross-bridge (myosin head) cycling, that is responsible for the filament sliding. The number of activated cross-bridges govern the contractile force generated and also the contraction speed. A strain-energy function is used to describe the mechanical behaviour of the smooth muscle tissue. Besides the active contractile apparatus, the mechanical model also incorporates a passive viscoelastic part. The constitutive model was calibrated with respect to experiments on smooth muscle tissue from swine carotid artery and guinea pig taenia coli, in terms of isometric and isotonic tensile test results. The model was fully able to reproduce the experimental results.

Purinergic cotransmission

Adenosine 5'-triphosphate (ATP) is a cotransmitter with classical transmitters in most nerves in the peripheral and central nervous systems, although the proportions vary between tissues and species and in different developmental and pathophysiological circumstances. There was early evidence that ATP was released together with acetylcholine (ACh) from motor nerves supplying skeletal muscle, although it was considered at the time as a molecule involved in the vesicular uptake and storage of ACh. Later it was shown that in the developing neuromuscular junction, released ATP acted on P2X receptor ion channels as a genuine cotransmitter with ACh. Adenosine triphosphate was shown to be released from sympathetic nerves supplying the guinea-pig taenia coli in 1971. Soon after, the possibility was raised that ATP was coreleased with noradrenaline from sympathetic nerves to guinea-pig seminal vesicle, cat nictitating membrane and guinea-pig vas deferens. Sympathetic purinergic cotransmission has also been demonstrated in many blood vessels. Parasympathetic nerves supplying the urinary bladder use ACh and ATP as cotransmitters; ATP acts through P2X ionotropic receptors, whereas the slower component of the response is mediated by the metabotropic muscarinic receptor. Adenosine triphosphate and glutamate appear to be cotransmitters in primary afferent sensory neurons. Adenosine triphosphate, calcitonin gene-related peptide and substance P coexist in some sensory-motor nerves. A subpopulation of intramural enteric nerves provides non-adrenergic, non-cholinergic inhibitory innervation of gut smooth muscle. Three cotransmitters are involved, namely ATP, nitric oxide and vasoactive intestinal polypeptide. In recent years, studies have shown that ATP is released with ACh, noradrenaline, glutamate, gamma-aminobutyric acid, 5-hyroxytryptamine and dopamine in different subpopulations of neurons in the central nervous system.

Spasmogenic effect of the aqueous extract of <i>Tamarindus indica</i> l. (caesalpiniaceae) on the contractile activity of guinea-pig <i>Taenia coli</i>

The effect of aqueous extract of Tamarindus indica (AETI) was studied on the guinea pig taenia coli, due to its use for treatment of constipation in traditional medicines. AETI, at concentrations ranging from 10(-8) mg/ml to 10(-2) mg/ml, increased the spontaneous contractile activity of guinea pig taenia coli in a dose-dependent manner (EC50 = 4x10(-6) mg/ml). This activity was unaffected by atropine. In high K(+), Ca(2+)-free solution containing EDTA, AETI as well as acetylcholine, used as a control, induced tonic contraction. These results suggest that the plant extract exert a spasmogenic effect that would not involve cholinergic mechanism of action. However, these active principles could mobilize both extra cellular calcium and intracellular calcium from internal stores.

Cupric ions at high concentrations induce rigor-like contraction in a Ca2+-free medium in the guinea pig taenia coli

Cupric ions at high concentrations induce rigor-like contraction in a Ca2+-free medium in the guinea pig taenia coli

The sources of Ca2+ and the intracellular action involved in cupric ions-induced contraction in the guinea pig taenia coli

The sources of Ca2+ and the intracellular action involved in cupric ions-induced contraction in the guinea pig taenia coli

Effects of Tetrapleura Tetraptera (Taub) Fruit Extract On Some Isolated Tissues: Possible Mechanism(S) of Antihypertensive Action

Hypotensive activities of the alcoholic extract of T. tetraptera fruits have been earlier demonstrated in cats, rats and rabbits. In the present study, we demonstrate myorelaxant actions of the extract in isolated smooth muscle preparations. Relaxations were dose-dependent: EC50s were 0.3 ± 0.01, 2.4 ± 0.1 and 4.3 ± 0.01 mg/ml for rat anococcygeus, rat uterus and guinea-pig taenia coli respectively. Also, spasmogenic actions of noradrenaline, carbachol, Ca2+ and K+ in the rat anococcygeus and uterus were all suppressed by the extract. The effect of the extract on Ca2+-induced contractions were investigated in guinea-pig isolated taenia coli and atria preparations and compared to nifedipine, a calcium- channel blocker . Pre-treatment with T. tetraptera (1-10 mg/ml) and nifedipine (0.01-0.1 μg/ml) for 15 minutes in Ca2+-free K+ (100 mM) medium caused a right ward shift of the concentrationresponse curves for CaCl2-induced contractions in atria and taenia coli with depressed maximal response, suggestive of a non-competitive interaction. In all tissues investigated, the extract nonspecifically reduced both the natural tone and agonist-induced contractions. Thus smooth muscle relaxant actions of the extract may partly account for the hypotensive actions of T. tetraptera fruits. Journal of Science and Technology Vol. 28 (1) 2008 pp. 23-35

Alteration of phosphocreatine level of guinea-pig taenia coli upon exposure to imidazole buffer

Alteration of phosphocreatine level of guinea-pig taenia coli upon exposure to imidazole buffer