Curcuma Longa Abolishes Phenylephrine-Induced Contractions in Isolated Aortic Artery of Rats

Curcuma longa has biological effects. Its cardiovascular activities are yet to be scientifically studied. Objectives: To investigate the vasorelaxant effects of the aqueous extract of Curcuma longa (AECL). Methods: Aortic annuli of normotensive rats, with or without endothelium, were set up in a data storage system with nutrient solution in recipients, with scientifically recommended temperature, aeration and tension. Over contraction by Phenylephrine, the AECL (1, 3, 10, 30, 100, 300 and 1000 µg/mL) was incubated before and after incubation with atropine or L-name or indomethacin. An AECL concentration-response curve was also built over contractions caused by elevation of extracellular K+. Data were significant when p < 0.05, with GraphPad Prism 6.0 software resolutions. Results: The AECL induced 100% vasorelaxation also in the endothelium-free annuli. The part of the endothelium-dependent effect had EC50 = 4.32 ± 0.05 µg/mL. With inhibition of NO production, the EC50 increased to 126.50 ± 2.35 µg/mL; after inhibition of prostacyclin production, to 124.6 ± 0.05 µg/mL; and after muscarinic blockade, to 437.10 ± 0.2 µg/mL. Opening of K+ channels (relaxation of 56.98%) and VOCC blockade (relaxation of 31.56%) were evident. Conclusion: AECL induced significant vasorelaxation, being more significant in the presence of endothelium. The muscarinic pathway seems to be the main one involved in this effect, followed by the NO production and prostacyclin pathways. The activity in K+ channels by AECL was more significant than its VOCC blockade. The use of other models and tools to study action mechanisms will be important and elucidating

Expression of Muscarinic Receptors and the Effect of Tiotropium Bromide in Aged Mouse Model of Chronic Asthma / 결핵및호흡기질환

BACKGROUND: Efficacy and safety of tiotropium bromide, a muscarinic receptor antagonist, in treatment of asthma have been reported. However, its effect on airway remodeling in chronic asthma of the elderly has not been clearly verified. The objective of this study was to investigate the effect of tiotropium and expression of muscarinic receptors as its related mechanism in an aged mouse model of chronic asthma with airway remodeling. METHODS: BALB/c female mice age 6 weeks, 9 and 15 months were sensitized and challenged with ovalbumin (OVA) for three months. Tiotropium bromide was administered during the challenge period. Airway hyperresponsiveness (AHR) and pulmonary inflammation were measured. Parameters of airway remodeling, and expression levels of M2 and M3 receptors were examined. RESULTS: Total cell with eosinophils, increased in the OVA groups by age, was decreased significantly after treatment with tiotropium bromide, particularly in the age group of 15 months. AHR and levels of interleukin (IL)-4, IL-5, and IL-13 were decreased, after tiotropium administration. In old aged group of 9- and 15-months-treated groups, hydroxyproline contents and levels of α-smooth muscle actin were attenuated. Tiotropium enhanced the expression of M2 but decreased expression of M3 in all aged groups of OVA. CONCLUSION: Tiotropium bromide had anti-inflammatory and anti-remodeling effects in an aged mouse model of chronic asthma. Its effects seemed to be partly mediated by modulating expression M3 and M2 muscarinic receptors. Tiotropium may be a beneficial treatment option for the elderly with airway remodeling of chronic asthma.

Functional and biochemical characteristics of urinary bladder muscarinic receptors in long-term alloxan diabetic rats / Características funcionais e bioquímicas dos receptores muscarínicos da bexiga de ratos com diabetes crônico induzido por aloxana

Objective To re-examine the function of the urinary bladder in vivoas well as to determine the functional and biochemical characteristics of bladder muscarinic receptors in long-term alloxan-induced diabetes rats.Methods Two-month-old male Wistar rats were injected with alloxan and the animals showing blood glucose levels >300mg/dL together with age-paired untreated animals were kept for 11 months. Body weight, bladder weight, blood glucose, and urinary volume over a period of 24 hours were determined in both groups of animals. A voiding cystometry in conscious control and diabetic rats was performed to determine maximal micturition pressure, micturition contraction interval and duration as well as voided and post-voiding residual volume. In addition, concentration-response curves for bethanechol in isolated bladder strips, as well as [3H]-N methyl-scopolamine binding site characteristics in bladder homogenates were determined.Results Mean bladder weight was 162.5±21.2mg versus 290±37.9mg in control and treated animals, respectively (p<0.05). Micturition contraction amplitude (34.6±4.7mmHg versus 49.6±2.5mmHg), duration (14.5±1.7 seconds versus 23.33±4.6 seconds) and interval (87.5±17.02 seconds versus 281.11±20.24 seconds) were significantly greater in alloxan diabetic rats. Voided urine volume per micturition contraction was also significantly higher in diabetic animals. However the post-voiding residual volume was not statistically different. Bethanechol potency (EC50 3µM versus 5µM) and maximal effect (31.2±5.9g/g versus 36.1±6.8g/g) in isolated bladder strips as well as number (169±4fmol/mg versus 176±3fmol/mg protein) and affinity (0.69±0.1nM versus 0.57±0.1nM) of bladder muscarinic receptors were also not statistically different.Conclusion Bladder function in vivo is altered in chronic alloxan-induced diabetes rats without changes in functional and biochemical characteristics of bladder muscarinic receptors.

Objetivo Reestudar o funcionamento da bexiga in vivo e determinar as características funcionais e bioquímicas dos receptores muscarínicos vesicais de ratos com diabetes crônico induzido por aloxana.Métodos Ratos Wistar de dois meses de idade receberam injeção de aloxana, e os animais que apresentaram glicemia >300mg/dL foram mantidos por 11 meses junto de outros não tratados e pareados por idade. Nos dois grupos de animais, peso corpóreo, peso da bexiga, glicemia e volume urinário de 24 horas foram medidos. Em ambos os grupos, realizou-se a cistometria miccional em animais não anestesiados. Foram determinados os seguintes parâmetros: pressão máxima de micção, intervalo e contração de micção, bem como o volume de esvaziamento e o volume residual pós-miccional. Além disso, foram determinadas as curvas de concentração-resposta a betanecol em preparações isoladas de bexiga e também as características dos sítios de ligação da [3H]-N-metil-escopolamina em homogenatos de bexiga.Resultados O peso médio da bexiga foi de 162,5±21,2mg versus290±37,9mg nos animais controles e tratados, respectivamente (p<0,05). A amplitude de contração (34,6±4,7mmHg versus 49,6±2,5mmHg), a duração (14,5±1,7 segundos versus 23,33±4,6 segundos) e o intervalo (87,5±17,02 segundos versus 281,11±20,24 segundos) de micção foram significantemente maiores nos ratos tratados com aloxana. O volume de urina eliminada durante a contração miccional também foi maior nos animais diabéticos. Contudo, o volume residual pós-miccional não foi estatisticamente diferente. Não foram observadas diferenças na resposta ao betanecol (EC50 3µM versus 5µM) e no seu efeito máximo (31,2±5,9g/g versus 36,1±6,8g/g) em preparações isoladas de bexiga, bem como no número total (169±43fmol/mgversus 176±3fmol/mg) e na afinidade (0,69±0,1nMversus 0,57±0,1nM) dos receptores muscarínicos da bexiga.Conclusão O funcionamento da bexiga in vivo está alterado no diabetes crônico induzido por aloxana, porém sem alterações funcionais e bioquímicas nos receptores muscarínicos da bexiga.

Role of vagus nerve-muscarinic cholinergic receptor pathway in mitigation of myocardial ischemia-reperfusion injury by intrathecal morphine postconditioning in rats / 中华麻醉学杂志

Objective evaluate the role of vagus nerve?muscarinic cholinergic receptor ( M recep?tor) pathway in mitigation of myocardial ischemia?reperfusion (I∕R) injury by intrathecal morphine postcon?ditioning in rats. Methods Seventy adult male Sprague?Dawley rats in which intrathecal catheters were suc?cessfully placed without complications, weighing 250-350 g, were randomly assigned into 7 groups ( n=10 each) using a random number table:sham operation group (Sham group), I∕R group, intrathecal morphine postconditioning group ( MP group) , vagal transection ( VT) group, VT+ intrathecal morphine postcondi?tioning group (VT+MP group), atropine (ATP, M receptor antagonist) + morphine postconditioning group ( ATP+MP group) , and ATP group. Myocardial I∕R was produced by occlusion of the anterior descending branch of left coronary artery for 30 min followed by 2 h of reperfusion. Morphine ( 3μg∕kg, 10μl) was in?trathecally infused over 5 min starting from onset of reperfusion in MP group. Normal saline 10 μl was in?trathecally infused over 5 min starting from onset of reperfusion in NS group. The bilateral vagus nerves were transected at 10 min before reperfusion in VT+MP group. Atropine ( 0?1 mg∕kg, 0?5 ml) was intravenously infused over 5 min starting from 10 min before reperfusion in ATP+MP group. The occurrence of cardiac ar? rhythmia ( premature ventricular contractions ( PVCs) and ventricular tachycardia ( VT)∕ventricular fibrilla?tion ( VF) ) within the first 30 min of reperfusion was recorded. The rats were sacrificed at 120 min of reper?fusion, and myocardial specimens were obtained for determination of myocardial infarct size ( IS) as a per?centage of area at risk (AAR). IS∕AAR ratio was calculated. Results Compared with Sham group, the number of PVCs and VT∕VF and IS∕AAR ratio were significantly increased in the other groups. Compared with I∕R group, the number of PVCs and VT∕VF and IS∕AAR ratio were significantly decreased in MP group. Compared with MP group, the number of PVCs and VT∕VF and IS∕AAR ratio were significantly in?creased in VT+MP and ATP+MP groups. Conclusion Vagus nerve?M receptor pathway is involved in miti?gation of myocardial I∕R injury by intrathecal morphine postconditioning in rats.

Effects of Halothane, Enflurane or Isoflurane on the m1 or m3 Muscarinic Receptor Expressed in Xenopus Oocytes / 대한마취과학회지

BACKGROUND: Muscarinic receptors are distributed abundantly in the central nervous system and peripheral visceral organs, and have a close relationship with anesthesia. We investigated the effects of halothane, enflurane or isoflurane on m1 or m3 muscarinic signaling. METHODS: Using two electrode voltage clamps, Ca2+ -activated Cl- currents (ICl(Ca)) were measured in Xenopus oocytes injected with an m1 or m3 receptor mRNA. ICl(Ca) was induced with the application of acetyl beta-methylcholine with or without exposure to volatile anesthetics. RESULTS: Halothane depressed the m1 and m3 receptor function significantly (m1; 0.49 +/- 0.17 microampere -> 0.1 +/- 0.05 microampere, m3; 0.74 +/- 0.2 -> 0.23 +/- 0.06 microampere, P 0.15 +/- 0.04 microampere, m3; 0.95 +/- 0.34 -> 0.19 +/- 0.05 microampere, P 0.3 +/- 0.09 microampere, P 0.5). From a concentration-response curve, enflurane decreased m1 and m3 signaling dose-dependently. CONCLUSIONS: Our data suggests that m1 and m3 muscarinic receptors were depressed by halothane, enflurane or isoglurane except for the fact that the m1 receptor was not affected by isoflurane.

Preparation of m_1AChR-G_(11)fusion protein and m_4AChR-G_(16)fusion protein and effects of various ligands on them / 中国病理生理杂志

AIM: To prepare m 1AChR-G 11 and m 4AChR-G 16 fusion protein in Baculovirus-Sf9 cell system and detect the effects of various muscarinic ligands on the interaction between m 1AChR and G 11 and m 4AChR and G 16 ,and screen different kinds of ligands specific for m 1 and m 4. METHODS: To prepare fused DNA of m 1AChR-G 11 and m 4AChR-G 16 in two PCR, then expressed in Sf9 cells and detect the pharmacological function of m 1AChR-G 11 fusion protein and m 4AChR-G 16 fusion protein by [ 3H]QNB and [ 35 S]GTP?S binding experiments; To expore the way of the activation of m 1AChR-G 11 and m 4AChR-G 16 fusion protein by various ligands includingcetylcholine (ACh), Pilocarpine (Pilo), 4-hydroxy-2-butynyl-1-trimethylammonium-m-chloro-carbanilatechloride (McN-A-343),tetrandrine, pirenzepine (PZ), alcuronium, atropine,R-(+)-hyoscyamine and gallamine by displacement by GDP on [ 35 S]GTP?S binding experiments. RESULTS: The expression levels of m 1AChR-G 11 and m 4AChR-G 16 fusion protein were (45 39?2 62) nmol?g -1 protein, (47 04?1 58) nmol?g -1 protein. The affinity of GDP to G 11 and G 16 partner changed in the presence of different muscarinic ligands. CONCLUSION: The m 1AChR-G 11 and m 4AChR-G 16 showed the pharmacological specificity to m 1 and m 4 receptor and the efficient signaling of the two partners. Ligands of m 1AChR and m 4AchR mediated different signal transduction by changing the affinity of G 11 /G 16 and GDP. So m1AChR-G 11 fusion protein and m 4AChR-G 16 fusion protein can be taken as a tool to screen ligands specific for m 1AChR and m 4AChR.

The Effects of Vecuronium and Pancuronium on the Tension of the Isolated Rat Tracheal Smooth Muscle / 대한마취과학회지

BACKGROUND: Non-depolarizing muscle relaxants have their muscle relaxing effect by competing with acetylcholine (ACh) at the nicotinic receptor level. What are the effects of such muscle relaxants on the tracheal smooth muscle? This present study was set up to address the question as to how vecuronium and pancuronium influence the tracheal smooth muscle. METHODS: Sixty male Sprague-Dawley rat tracheal smooth muscles were isolated at optimal length for isometric force. The preparations were set up in an organ bath containing Tyrode's solution. And isometric force displacement transducer and physiograph were used to record the change in force. After the equilibration period the preparations were contracted with ACh 10(-5) M and carbachol 3x10(-7)M seperately. The preparations were washed with fresh tyrode's solution and allowed to return passively to resting tone. Then the cumulartive effect of ACh (from 3 10(-7) M through 10(-5) M) and carbachol (CCh, from 10(-8) M through 3 10(-6) M) were produced before and after pretreating the preparation with vecuronium (10(-5) M and 10(-6) M) and pancuronium (10(-5) M and 10(-6) M) respectively. Also, we studied the changes of contraction produced by neostigmine before and after pretreatment with vecuronium (10(-5) M and 3 10(-5) M) and pancuronium (3 10(-6) M and 3 10(-5) M). RESULTS: Vecuronium shifted the ACh dose-response curve of the tracheal contraction to the left (p0.05). CONCLUSIONS: Vecuronium inhibits the ACh hydrolyzing enzyme, especially acetylcholinesterase. Therefore it potentiates ACh contraction in the tracheal smooth muscle, but not the CCh contraction, while pancuronium has a different effect in comparison with vecuronium. That is, at a low concentration it reveals an antagonistic effect on the muscarinic M2 receptor and at a higher concentration it has an antagonistic effect on the muscarinic M3 receptor in the tracheal smooth muscle.

The Antiallodynic Effect of Edrophonium and Neostigmine in a Neuropathic Pain Model / 대한마취과학회지

BACKGROUND: Peripheral nerve injury may produce a syndrome consisting of spontaneous pain, allodynia and hyperpathia. Cholinesterase inhibitors are known to have an antinociceptive effect in hot plate and tail flick tests and to be mediated by spinal muscarinic system. The purpose of the current study was to determine the effect of intrathecally (i.t.) administered edrophonium and neostigmine on the touch-evoked allodynia and to identify the antagonism of antiallodynia in a rat model of neuropathic pain. METHODS: Sprague Dawley rats were prepared with tight ligation of left L5/L6 spinal nerves with 6~0 black silk and chronic lumbar intrathecal catheters. After obtaining the baseline hindpaw withdrawal scores, edrophonium (3~100ug) or neostigmine (0.3~10ug) was administered intrathecally. Tactile allodynia was measured using von Frey filaments and allodynic threshold was calculated by updown method. Motor dysfunction was assessed by observing righting/stepping reflex responses and abnormal weight bearing. To examine the reversal of antiallod ynia, muscarinic receptor antagonist atropine (10ug) or nicotinic receptor antagonist mecamylamine (10ug) was injected intrathecally 5 min. prior to injection of edrophonium or neostigmine. RESULTS: I.t. edrophonium and i.t. neostigmine produced a dose dependent antagonism of allodynic state but had moderate to severe effect on motor weakness at doses of 3 and 10 g of neostigmine. Pretreatment with i.t. atropine yielded a complete antagonism of antiallodynia in both drugs, but i.t. mecamylamine did not significantly reverse incresed allodynic threshold. CONCLUSIONS: These experiments suggest that i.t. edrophonium or i.t. neostigmine produces a dose dependent antagonism on touch-evoked allodynia at the spinal level and this antagonism is likely due to spinal muscarinic system.

Effect of desflurane pretreatment on the density of muscarinic receptors on hypoxic/reoxygenated cardiac myocytes / 中华麻醉学杂志

Objective To investigate the effect of desflurane pretreatment on the density of muscarinic (M)-receptors on anoxic/reoxygenated rat cardiac myocytes. Methods Myocytes were obtained from ventricle of 2-3 day SD rats and incubated for 3 days. Then they were randomly allocated to one of 5 groups: group I received no treatment and served as control ( n = 5); group Ⅱ underwent 2 h hypoxia (liquid culture medium was insufflated with a gas mixture of 1 % CO2 , 1 % O2 and 98% N2) followed by 1h reoxygenation ( n = 6) ; group Ⅲ was pre-treated with 1.5 MAC desflurane (9% ) for 20 min, followed by 10 min washout before the same duration of hypoxia/reoxygenation as in group II ( n = 6) ; group IV underwent longer duration of hypoxia (48 h) followed by 24 reoxygenation ( n = 5) ; group V was pre-treated with 1.5 MAC desflurane (9% ) for 20 min followed by 10 min washout, then underwent longer duration of hypoxia/reoxygenation as in group IV ( n = 5) . The density of M-receptors was measured by the radio-ligand binding assay (RLBA) .Results The density of M-receptors was significantly lower in group Ⅱthan that in group I ( P

RELATIONSHIP BETWEEN DENSITY OF MUSCARINIC RECEPTOR SUBTYPE AND BLADDER INSTABILITY IN RATS / 解放军医学杂志

To study the relationship between the density of muscarinic receptor (MR) subtype and the occurrence of bladder instability. The rat model of unstable bladder was established, and the density of MR subtype was determined in normal and unstable bladders. The results showed that there were only m 2 RNA and m 3 RNA present in the detrusor of normal and unstable bladders, and m 1, m 2 or m 3 mRNA subtypes were not detected. In the normal bladder, the amount of m 2 RNA (1 67?0 42) was obviously greater than m 3 RNA (0 64?0 19), and the ratio of them was 2 59∶1. In the unstable bladder, the density of both m 2 and m 3 increased to (2 03?0 65) and (1 53?0 46) respectively, and the increase of m 3 was more significant ( P