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.

Papel dos receptores do tipo 5-HT3 na área septal medial sobre o controle da pressão sanguínea, do apetite por sódio e da ingestão hídrica / Role of receptor type 5-HT3 into the MSA under the control of blood pressure, the sodium appetite and fluid intake

Diferentes áreas do sistema nervoso central que participam da regulação cardiovascular recebem projeções de núcleos da rafe produtores de 5-HT. Diversos estudos têm também demonstrado a participação dos receptores serotoninérgicos nas respostas neuroendócrinas e emocionais ao estresse e no equilíbrio hidrossalino, assim os objetivos do referido trabalho foram: a) estudar o papel dos receptores do tipo 5-HT3 presentes na ASM (área septal medial) sobre as respostas cardiovasculares ao estresse de contenção em ratos; b)verificar a possível interação entre os receptores colinérgicos muscarínicos e 5-HT3 presentes na ASM no controle cardiovascular; c) verificar o papel dos receptores do tipo 5-HT3 na ASM sobre o controle hidrossalino. Foram utilizados ratos Wistar (280-300g) submetidos ao implante de uma cânula guia na ASM. Os animais destinados aos estudos cardiovasculares receberam implante de catéter carotídeo para análise da PA. No momento do experimento referente ao estresse os animais receberam injeção de m-CPBG e ondansetrona na ASM e 15 min após a microinjeção foram submetidos ao estresse de contenção com registro da PA. Para análise da interação entre os receptores muscarínicos e os receptores serotoninérgicos do tipo -HT3 os animais receberam previamente atropina, antagonista colinérgico muscarínico, e após 10 min receberam ondansetrona com registro constante da PA por mais 110min. No protocolo experimental para depleção de sódio os animais receberam microinjeções de furosemida 24h antes do experimento tendo disponíveis bebedouros de água destilada. No momento do experimento os animais receberam microinjeções de m-CPBG e ondansetrona e após 15 min os volumes de água e salina 1,5% foram registrados por 2h. Para análise do efeito do bloqueio dos receptores 5-HT3 sobre o comportamento de ingestão de água os animais foram submetidos a privação hídrica por 24h. No momento do experimento microinjeções de alina, m-CPBG ondansetrona foram feitas na ASM com medida dos volumes ingeridos ao longo de 2h. Verificamos que os receptores serotoninérgicos do tipo 5-HT3 presentes na ASM inibem o aumento da PA em animais submetidos ao estresse, além disso, verificamos também que a resposta hipertensiva decorrente do bloqueio dos receptores serotoninérgicos do tipo 5-HT3 depende da integridade funcional dos receptores colinérgicos muscarínicos. Por outro lado, tanto a ativação, quanto o bloqueio dos receptores serotoninérgicos do tipo 5-HT3 presentes na ASM parecem não mediar a ingestão de sódio em animais sódio-depletados nem ingestão de água em animais sob privação hídrica.

Expression and function of G-protein-coupled receptorsin the male reproductive tract

This review focuses on the expression and function of muscarinic acetylcholine receptors (mAChRs), α1-adrenoceptors and relaxin receptors in the male reproductive tract. The localization and differential expression of mAChR and α1-adrenoceptor subtypes in specific compartments of the efferent ductules, epididymis, vas deferens, seminal vesicle and prostate of various species indicate a role for these receptors in the modulation of luminal fluid composition and smooth muscle contraction, including effects on male fertility. Furthermore, the activation of mAChRs induces transactivation of the epidermal growth factor receptor (EGFR) and the Sertoli cell proliferation. The relaxin receptors are present in the testis, RXFP1 in elongated spermatids and Sertoli cells from rat, and RXFP2 in Leydig and germ cells from rat and human, suggesting a role for these receptors in the spermatogenic process. The localization of both receptors in the apical portion of epithelial cells and smooth muscle layers of the vas deferens suggests an involvement of these receptors in the contraction and regulation of secretion.

Esta revisão enfatiza a expressão e a função dos receptores muscarínicos, adrenoceptores α1 e receptores para relaxina no sistema reprodutor masculino. A expressão dos receptores muscarínicos e adrenoceptores α1 em compartimentos específicos de dúctulos eferentes, epidídimo, ductos deferentes, vesícula seminal e próstata de várias espécies indica o envolvimento destes receptores na modulação da composição do fluido luminal e na contração do músculo liso, incluindo efeitos na fertilidade masculina. Além disso, a ativação dos receptores muscarínicos leva à transativação do receptor para o fator crescimento epidermal e proliferação das células de Sertoli. Os receptores para relaxina estão presentes no testículo, RXFP1 nas espermátides alongadas e células de Sertoli de rato e RXFP2 nas células de Leydig e germinativas de ratos e humano, sugerindo o envolvimento destes receptores no processo espermatogênico. A localização de ambos os receptores na porção apical das células epiteliais e no músculo liso dos ductos deferentes de rato sugere um papel na contração e na regulação da secreção.

Trypanosoma cruzi infection induces up-regulation of cardiac muscarinic acetylcholine receptors in vivo and in vitro

The pathogenesis of chagasic cardiomyopathy is not completely understood, but it has been correlated with parasympathetic denervation (neurogenic theory) and inflammatory activity (immunogenic theory) that could affect heart muscarinic acetylcholine receptor (mAChR) expression. In order to further understand whether neurogenic and/or immunogenic alterations are related to changes in mAChR expression, we studied two models of Trypanosoma cruzi infection: 1) in 3-week-old male Sprague Dawley rats chronically infected with T. cruzi and 2) isolated primary cardiomyocytes co-cultured with T. cruzi and peripheral blood mononuclear cells (PBMC). Using [³H]-quinuclidinylbenzilate ([³H]-QNB) binding assays, we evaluated mAChR expression in homogenates from selected cardiac regions, PBMC, and cultured cardiomyocytes. We also determined in vitro protein expression and pro-inflammatory cytokine expression in serum and cell culture medium by ELISA. Our results showed that: 1) mAChR were significantly (P < 0.05) up-regulated in right ventricular myocardium (means ± SEM; control: 58.69 ± 5.54, N = 29; Chagas: 72.29 ± 5.79 fmol/mg, N = 34) and PBMC (control: 12.88 ± 2.45, N = 18; Chagas: 20.22 ± 1.82 fmol/mg, N = 19), as well as in cardiomyocyte transmembranes cultured with either PBMC/T. cruzi co-cultures (control: 24.33 ± 3.83; Chagas: 43.62 ± 5.08 fmol/mg, N = 7 for both) or their conditioned medium (control: 37.84 ± 3.84, N = 4; Chagas: 54.38 ± 6.28 fmol/mg, N = 20); 2) [³H]-leucine uptake was increased in cardiomyocytes co-cultured with PBMC/T. cruzi-conditioned medium (Chagas: 21,030 ± 2321; control 10,940 ± 2385 dpm, N = 7 for both; P < 0.05); 3) plasma IL-6 was increased in chagasic rats, IL-1â, was increased in both plasma of chagasic rats and in the culture medium, and TNF-á level was decreased in the culture medium. In conclusion, our results suggest that cytokines are involved in the up-regulation of mAChR in chronic Chagas disease.

Bradycardia induced by Mesobuthus tamulus scorpion venom involves muscarinic receptor-G-protein-coupled cell signaling pathways.

Indian red scorpion (Mesobuthus tamulus; MBT) envenomation produces various cardio-respiratory abnormalities including cardiac dysrhythmias. The underlying cell signaling pathways for the cardiac dysrhythmias produced by MBT venom are not known. The present study was therefore conducted to delineate the second messenger signaling pathways involved in MBT venom-induced atrial rhythm changes. The effects of venom and various antagonists were examined on spontaneously beating rat right atrial preparations in vitro. The MBT-venom produced an increase (35%), a decrease (45%) and again an increase (50%) in rate at 0.03, 0.3 and 3.0 microg/ml of venom, respectively. On the other hand, force of contraction exhibited a concentration-dependent rise (up to 40%) at all concentrations of venom. Pretreatment with atropine (0.3 microM) blocked the decrease in atrial rate at 0.3 microg/ml concentration of venom while no such blockade was seen in force of contraction. Submaximal concentration of ACh (0.1 nM) decreased the atrial rate by 25%. In the presence of MBT venom (0.3 microg/ml), ACh-induced fall in atrial rate was enhanced. The venom-induced fall in atrial rate and augmentation of ACh response were blocked by pertussis toxin (PTx; a Gi-inhibitor) or methylene blue (a G-cyclase inhibitor). The results indicate that the decrease in atrial rate produced by venom is mediated muscarinic by receptors via Gi-guanylyl cyclase mediated cell signaling pathways.

Development of fetal nicotine and muscarinic receptors in utero

The role of acetylcholine in the central and peripheral nervous systems is well established in adults. Cholinergic modulation of vascular functions and body fluid balance has been extensively studied. In the embryo-fetus, cholinergic receptors are widespread in the peripheral and central systems, including smooth muscle and the epithelial lining of the cardiovascular, digestive, and urinary systems, as well as in the brain. Fetal nicotine and muscarinic receptors develop in a pattern (e.g., amount and distribution) related to gestational periods. Cholinergic mechanisms have been found to be relatively intact and functional in the control of vascular homeostasis during fetal life in utero at least during the last third of gestation. This review focuses on the development of fetal nicotine and muscarinic receptors, and provides information indicating that central cholinergic systems are well developed in the control of fetal blood pressure and body fluid balance before birth. Therefore, the development of cholinergic systems in utero plays an important role in fetal vascular regulation, gastrointestinal motility, and urinary control.

Supersensitivity of the cholinergic muscarinic system in the rat's brain is induced by high concentrations of Cu+2

Transition metals have been described as regulators of receptor's function. here, we studied the effects of chronic administration of Cu2+ or the Cu2+ chelator penicillamine (PA) on the functional and binding properties of the muscarinic receptors (MR) on selected areas of rat's brain. Groups of 10 Sprague-Dawley rats were treated daily, for 45 days with either 1) 1 mg/Kg CuSO4 (Cu2+), 2) 100 mg/Kg PA, or 3) saline solution. Double T-maze and motility cages were used for behavioral testing and the binding assays were performed using [3H]-QNB or [3H]-N-MSCP as MR's ligands. Cu2+ brain levels were measured in the cerebral cortex by atomic absorption spectrophotometer. Results showed that PA treated rats displayed a significant decrease of locomotor's activity (LA) and rearing behavior (RB), but a significant increases in memory efficiency (ME). Cu2+ treated rats displayed diminished RB with no significant changes in LA. Cu2+ treated rats displayed higher MR's density (Bmax) in cortex (C), striatum (S), and hippocampus (H). An increase in Bmax was also observed in PA treated rats, but only in C and S. Finally, Cu2+ tissue concentration was significantly higher in C of both Cu2+ and with PA treated animals. In conclusion, 45 days of Cu2+ or PA treatment induced brain hypercuprosis, which was associated with MR binding supersensitivity; however, change in ME was only observed in PA treated rats suggesting that might be still another factor in these experiments besides Cu2+ (i.e., Zn2+ or PA itself) involved in memory modulation.

Phenyldiguanide activates cardiac receptors to produce responses by involving three different efferent pathways in anaesthetized rats.

The present study was undertaken to determine the afferent and efferent pathways involved in the phenyldiguanide (PDG)-induced reflex response in rats. Intravenous (iv) injection of PDG (10 microg/kg), produced hypotension, bradycardia and apnea over a period of time. Bilateral vagotomy abolished the PDG-induced reflex changes. Atropine (2 mg/kg; iv) blocked only the bradycardiac response produced by PDG, while prazosin (0.5 mg/kg; iv) blocked the hypotensive response, and bilateral vagotomy in these animals abolished the apneic response. In separate series of experiments, intrapericardial injection of lignocaine abolished the hypotensive and bradycardiac responses evoked by PDG in artificially ventilated rats. The results reveal that the PDG-induced reflex is mediated through vagal afferents originating from the heart and efferents involve three different pathways. The bradycardiac response was through the muscarinic receptors, the hypotension is mediated through alpha1 adrenoceptors and the apnea presumably through the spinal motoneurones supplying the respiratory muscles.

Atropine and ODQ antagonize tetanic fade induced by L-arginine in cats

Although it has been demonstrated that nitric oxide (NO) released from sodium nitrite induces tetanic fade in the cat neuromuscular preparations, the effect of L-arginine on tetanic fade and its origin induced by NO have not been studied in these preparations. Furthermore, atropine reduces tetanic fade induced by several cholinergic and anticholinergic drugs in these preparations, whose mechanism is suggested to be mediated by the interaction of acetylcholine with inhibitory presynaptic muscarinic receptors. The present study was conducted in cats to determine the effects of L-arginine alone or after pretreatment with atropine or 1H-[1,2,4]oxadiazole [4,3-a]quinoxalin-1-one (ODQ) on neuromuscular preparations indirectly stimulated at high frequency. Drugs were injected into the middle genicular artery. L-arginine (2 mg/kg) and S-nitroso-N-acetylpenicillamine (SNAP; 16 µg/kg) induced tetanic fade. The Nw-nitro-L-arginine (L-NOARG; 2 mg/kg) alone did not produce any effect, but reduced the tetanic fade induced by L-arginine. D-arginine (2 mg/kg) did not induce changes in tetanic fade. The tetanic fade induced by L-arginine or SNAP was reduced by previous injection of atropine (1.0 µg/kg) or ODQ (15 µg/kg). ODQ alone did not change tetanic fade. The data suggest that the NO-synthase-GC pathway participates in the L-arginine-induced tetanic fade in cat neuromuscular preparations. The tetanic fade induced by L-arginine probably depends on the action of NO at the presynaptic level. NO may stimulate guanylate cyclase increasing acetylcholine release and thereby stimulating presynaptic muscarinic receptors

Inhibition of carbachol-induced inositol phosphate accumulation in the embryonic retina promoted by kainate and veratridine

In the present study, we report that low concentrations of the glutamate ionotropic agonist kainate decreased the turnover of [3H]-phosphoinositides ([3H]-InsPs) induced by muscarinic receptors in the chick embryonic retina. When 100 muM carbachol was used, the estimated IC50 value for kainate was 0.2 muM and the maximal inhibition of ~50 percent was obtained with 1 muM or higher concentrations of the glutamatergic agonist. Our data also show that veratridine, a neurotoxin that increases the permeability of voltage-sensitive sodium channels, had no effect on [3H]-InsPs levels of the embryonic retina. However, 50 muM veratridine, but not 50 mM KCl, inhibited ~65 percent of the retinal response to carbachol. While carbachol increased [3H]-InsPs levels from 241.2 + 38.0 to 2044.5 + 299.9 cpm/mg protein, retinal response decreased to 861.6 + 113.9 cpm/mg protein when tissues were incubated with carbachol plus veratridine. These results suggest that the accumulation of phosphoinositides induced by activation of muscarinic receptors can be inhibited by the influx of Na+ ions triggered by activation of kainate receptors or opening of voltage-sensitive sodium channels in the chick embryonic retina.

Muscarinic M1 and M3 receptor in rat striatum: A binding study

In this study, the authors set out to determine the presence of M3 muscarinic receptors in rat striatum by examining the binding of [3H]N-methyl-scopolamine([3H]NMS) to striatal membranes and its displacement by antagonists with different affinity for M1 and M3 muscarinic receptors (pirenzepine; 4-diphenylacetoxy-N-methylpiperidine methiodide, 4-DAMP; and the p-fluoro analog of hexahydro-sila-difenidol, pFHHSiD). The specific binding of [3H]NMS to membranes from rat striatum (551 ñ 40 fmol.mg prot.-1, KD 0.11 ñ 0.01 nM) was displaced in a concentration-dependent manner by all three antagonists tested. Inhibition curves best fit to a single-site model for 4-DAMP(pKi 9.1 ñ 0.1), whereas for both pirenzepine and pFHHSiD, the best fit was to the two-site model. The pKi values for the high-affinity (8.0 ñ 0.2) and low-affinity (6.7 ñ 0.2) components for pirenzepine-mediated inhibition of [3H]NMS binding correspondend to those reported for M1 and M3 receptors, respectively. The pKi values for the high-affinity (7.7 ñ 0.1) and low-affinity (7.1 ñ 0.2) components for pFHHSiD inhibition were in good agreement with those reported for M3 and M1 receptors, respectively. Altogether, these results indicate the presence in rat striatum of both M1 and M3 muscarinic receptors. These findings might be relevant to the design and use of mucarinic antagonists in the treatment of neurological disorders such as Parkinson's disease

Subcellular distribution and pharmacological characterization of muscarinic receptors in tracheal smooth muscle

: Subcellular fractions isolated from tracheal smooth muscle have been identified using biochemical markers and measuring the [3H]QNB muscarinic receptor binding activity in these fractions. This muscarinic receptor (mAchR) activity was slightly enriched 1.6 times in the crude mitochondrial fraction (M), 2.6 times in the crude microsomal fraction (P), and greatly enriched in the highly purified plasma membranes fractions, being 5.3 times in a heavy plasma membrane fraction designed as P2 and 9.1 times in a light plasma membrane fraction named P1 fraction. The muscarinic receptor subtypes present in the subcellular fractions were identified using competition experiments. The binding of five selective antagonists, pirenzepine, AF-DX 116, hexahydrodifenidol, methoctramine and 4-DAMP were examined. In this sense, the M1 antagonist pirenzepine showed pKi's values between 6.44-7.45 and the M2 antagonist AF-DX 116 showed pKi's values ranging from 6.75 to 7.45 being the lowest pKi's values here described. The antagonist hexahydrodifenidol showed higher affinities than pirenzepine-derivated compounds with pKi's values from 7.25 to 7.65. The antagonist 4-DAMP exhibited pKi's values from 8.18-8.41. Finally, methoctramine showed similar affinities as 4-DAMP, with pKi's ranging from 8.09 to 8.22 suggesting the existence of M2 receptors in these fractions. These data suggest that M2 mAchR are present in all articulate fractions here studied. It is important to emphasize that the M2 muscarinic receptor presents in the light plasma membrane fraction (P1) shows poor selectivity towards the muscarinic antagonists being different from the M2 mAchRs associated with other subcellular fractions isolated from bovine tracheal smooth muscle

Lack of interaction of angiotensin converting enzyme inhibitors with muscarinic and neuropeptide Y receptors

In studies conducted in patients undergoing cardiac catheterizations, some hemodynamic changes were observed after the acute sublingual administration of the angiotensin converting enzyme inhibitors (ACEI) captopril, enalapril, and lisinopril. These changes consisted of an increase in pulmonary artery pressure, pulmonary vascular resistance (PVR) and induction of hypoxia. The pressure changes were transitory and disappeared after 25 min. The possible mechanisms involved in these changes may relate to interactions of the ACEI with peripheral receptor systems for hormones and neurotransmitters. We have thus undertaken the task of evaluating the potential effect of ACEI on biological receptor molecules. We have begun with studies on muscarinic receptors, and the recently characterized neuropeptide Y (NPY) receptors of endothelial cells. Equilibrium binding assays with 3H-QNB have been conducted for muscarinic receptors using rat brain synaptosomes, due to its expression of multiple muscarinic receptors subtypes. In addition 125BH-NPY binding assays were conducted on intact adrenal medullary endothelial cells. Enalapril and captopril, 10(-7) to 10(-3) M, were not able to produce significant inhibition of either muscarinic or NPY receptor probes. The paradoxical changes elicited by sublingual ACEI seems not to involve interaction with muscarinic or NPY receptors

NaCl increases the dissociation constant (Koff) of 3H-QNB muscarinic receptor complex in an airway smooth muscle plasma membrane fraction

Muscarinic receptors associated to plasma membrane fractions isolated from bovine airway smooth muscle were previously characterized by using 3H Quinuclidinyl-benzilate (3H-QNB) binding (Bécemberg, et al. Arch. Venez. Farm Terap. (1986) 5: 244-256). 3H-QNB binding to P1 plasma membrane fraction was time dependent. In order to study the effect of NaCl on the ligand binding properties of muscarinic receptor, a kinetic approach for the evaluation of koff of the 3H-QNB muscarinic receptor complex (QNB-mR) perfomed in the presence or absence of NaCl is done. The 3H-QNB bound to the receptor is released by atropine through an exchange reaction at the antagonist binding site. NaCl decreases the t1/2 of complexes and increases the velocity of dissociation of 3H-QNB-mR producing a significant change in Koff values, whichever conditions the 3H-QNB-mR complexes are formed. Pretreatment of P1 plasma membranes fraction with N-ethyl maleimide (1mM NEM) does not alter Koff value. In addition, in these NEM-treated P1 membranes, NaCl is unable to further increase the Koff value. The effect of NEM blocking this "activator effect" of NaCl suggests thar sulphydryl groups in the receptor structure may be involved in this NaCl effect on the 3H-QNB binding to muscarinic receptors

Effects of undernutrition on the ontogeny of muscarinic receptors and acetylcholinesterase activity in the rat motor cortex

The effect of undernutrition on the ontogeny of muscarinic receptor density and acetylcholineasterase (AChE) activity was studied in the motor cerebral cortex homogenates of female Wistar rats aged 12-19 days, 2-3 months and 5-8 months. Experimental animals were fed a protein-calorie deficient diet. The dinding assays, using (3H)-N-methylscopolamine as a ligand, indicated a significant increase in muscarinic receptor density in the motor cortex of 5-8- monthes old control rats (36%) compared to neonatal controls. No increase was observed for undernourished rats at any of the ages studied. No significant difference was observbed in AChE activity at any age of either group studied

Biochemical and pharmacological characterization of octylglucoside-solubilized muscarinic receptors derived from bovine tracheal smooth muscle

La actividad de receptor colinérgico muscarínico fue investigada en las fracciones subcelulares del músculo liso traqueal de bovino. Esta actividad de receptor muscarínico fue determinada mediante un ensayo basado en la unión específica del H-QNB (antagonista muscarínico Bencilato de Quinuclidinilo tritiado) utilizando un método simple y reproducible que permite separar el ligando radioactivo que se encuentra libre del unido, mediante un procedimiento de centrifugación en columnas; lo cual permite obtener valores muy bajos como "blanco". Esta actividad de receptor muscarínico fue encontrada enriquecida 4 veces en la fracción microsomal (P) con respecto al Extracto original y después de una etapa adicional de purificación utilizando un gradiente discontinuo de sacarosa se concentró 7 veces dicha actividad en la fracción de membranas plasmáticas (P-1). A partir de esta fracción de membranas plasmáticas enriquecida en receptor muscarínico y utilizando un método original que emplea un detergente no iónico como es el Octilglucósido al 1% se obtuvo una fracción de receptor muscarínico "solubilizado", después de centrifugar a 200.000 x g x 30 min. El material solubilizado fue filtrado a través de una columna de Sephadex G-50, con la finalidad de remover el detergente y el eluente obtenido de esta columna se denominó receptor muscarínico "solubilizado". El procedimiento descrito inicialmente para evaluar la interacción entre el receptor muscarínico y el H-QNB, también permite determinar bajo las mismas condiciones experimentales, la unión del H-QNB, tanto a los receptores muscarínicos unidos a las membranas plasmáticas como a los receptores presentes en el "solubilizado". Se encontró que el receptor muscarínico "solubilizado" y el asociado a las membranas plasmáticas mostraron idénticas propiedades bioquímicas y farmacológicas

Bases iónicas de la acción colinérgica en las células cardíacas / Ionic basis of the cholinergic action in the cardiac cells

La acetilcolina (Ach) se une en la membrana de las células del corazón a receptores colinérgicos muscarínicos, produciendo un aumento en la conductancia al potasio y una disminución de la conductancia al calcio. Estas alteraciones son las responsables de los efectos asociados a la acción parasimpática sobre el corazón. Así, el aumento de la conductancia al potasio se relaciona con el cronotropismo negativo e indirectamente con el inotropismo negativo inducidos por la estimulación vagal. La disminución de la conductancia al calcio se relaciona con el inotropismo negativo y con la disminución de la velocidad de conducción A-V. Existe evidencia que indica que los cambios de conductancia al potasio se producen por un aumento, bien sea del número de canales funcionales o de la conductancia por canal, de los canales iónicos responsables de la corriente de fondo (IK1). En el caso del calcio, una disminución en el número de canales funcionales es el mecanismo responsable de la disminución de la corriente lenta hacia adentro (Isi). Es probable que entre la interacción Ach- receptor y las alteraciones de la conductancia existan reacciones intermedias, en donde los nucleótidos cíclicos AMPc y GMPc parecen estar involucrados, al menos para el caso de la conductancia para el calcio

Effecto of the methylxantine derivative pentoxifylline on benzodiazepine and muscarinic binding sites in rats cerebral cortex

La inyección subcutánea del derivado metilxantínico pentoxifilina (3, 7, dimetil-1-(5-oxohexil)-xantina) redujo 3 h más tarde la concentración de sitios receptores para benzodiazepinas en la corteza cerebral de la rata. In vitro la pentoxifilina compitió efectivamente por la unión 3H-flunitrazepam a membranas cerebrales, demostrando poseer una potencia 10 veces mayor que la cafeína. Inyectada 2 veces por día durante 5 días, la pentoxifilina indujo un aumento de la concentración cerebral de receptores para benzodiazepinas. Estos resultados indican que la pentoxifilina modifica en forma distinta la concentración de receptores cerebrales para benzodiazepinas dependiendo del tiempo de tratamiento