RESUMO
OBJECTIVE: We previously found that mirabegron exerts a relaxant effect in the presence of the ß3 -adrenoceptor antagonist SR58894A during carbachol-induced contraction in human and pig detrusor. The aim of this study was to explore the possible mechanism underlying the relaxant effects of mirabegron using detrusor smooth muscle. METHODS: Human tissue was obtained from urinary bladders of patients undergoing radical cystectomy at Kyushu University and Harasanshin Hospital. Pig tissue was obtained from an abattoir. Tension force (organ bath experiments) was measured in intact or permeabilised (α-toxin or ß-escin) detrusor smooth muscle strips. The contribution of cAMP-dependent signaling and the inhibition of Ca2+ sensitization to the relaxant effects of mirabegron were characterized using 1 µM SR58894A, 100 µM SQ22536 (an adenylyl cyclase inhibitor), 10 µM H-89 (a protein kinase [PK] A inhibitor), 10 µM Y-27632 (a selective Rho kinase inhibitor), and 10 µM GF-109203X (a selective PKC inhibitor). RESULTS: 30 µM Mirabegron impaired carbachol (0.03-1 µM)-induced contraction in human detrusor smooth muscle. SR58894A only partially attenuated the relaxant effects of mirabegron in human and pig detrusor strips precontracted with 1 µM carbachol. In α-toxin-permeabilized detrusor strips, tension force at 1 µM [Ca2+ ]i was decreased by mirabegron in a concentration-dependent manner. The relaxant effect of mirabegron was only slightly attenuated by H-89 and not significantly affected by SQ22536. Y-27632 potentiated the relaxation response to mirabegron, but attenuated responses to cAMP; GF-109203X had little effect. Mirabegron but not cAMP had a notable relaxant effect in the pig detrusor smooth muscle permeabilized with ß-escin. CONCLUSIONS: Mirabegron-induced relaxation of pig and human detrusor smooth muscle occurs via both a ß3 -adrenoceptor/cAMP-dependent and -independent pathway.
Assuntos
Acetanilidas/farmacologia , AMP Cíclico/metabolismo , Relaxamento Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Tiazóis/farmacologia , Bexiga Urinária/efeitos dos fármacos , Agentes Urológicos/farmacologia , Inibidores de Adenilil Ciclases/farmacologia , Idoso , Amidas/farmacologia , Animais , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Feminino , Humanos , Indóis/farmacologia , Isoquinolinas/farmacologia , Masculino , Maleimidas/farmacologia , Músculo Liso/metabolismo , Proteína Quinase C/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Piridinas/farmacologia , Sulfonamidas/farmacologia , Suínos , Bexiga Urinária/metabolismo , Quinases Associadas a rho/antagonistas & inibidoresRESUMO
Gastrointestinal tract motility may be demoted significantly after surgery operations at least in part due to anaesthetic agents, but there is no comprehensive explanation of the molecular mechanism(s) of such adverse effects. Anesthetics are known to interact with various receptors and ion channels including several subtypes of transient receptor potential (TRP) channels. Two members of the canonical subfamily of TRP channels (TRPC), TRPC4 and TRPC6 are Ca2+-permeable cation channels involved in visceral smooth muscle contractility induced by acetylcholine, the primary excitatory neurotransmitter in the gut. In the present study, we aimed to study the effect of anesthetics on muscarinic receptor-mediated excitation and contraction of intestinal smooth muscle. Here we show that muscarinic cation current (mICAT) mediated by TRPC4 and TRPC6 channels in mouse ileal myocytes was strongly inhibited by isoflurane (0.5mM), one of the most commonly used inhalation anesthetics. Carbachol-activated mICAT was reduced by 63 ± 11% (n = 5), while GTPγS-induced (to bypass muscarinic receptors) current was inhibited by 44 ± 9% (n = 6). Furthermore, carbachol-induced ileum and colon contractions were inhibited by isoflurane by about 30%. We discuss the main sites of isoflurane action, which appear to be G-proteins and muscarinic receptors, rather than TRPC4/6 channels. These results contribute to our better understanding of the signalling pathways affected by inhalation anesthetics, which may cause ileus, and thus may be important for the development of novel treatment strategies during postoperative recovery.
Assuntos
Carbacol/antagonistas & inibidores , Intestinos/efeitos dos fármacos , Isoflurano/farmacologia , Contração Muscular/efeitos dos fármacos , Músculo Liso/fisiologia , Canais de Cátion TRPC/metabolismo , Canal de Cátion TRPC6/metabolismo , Anestésicos Inalatórios/farmacologia , Animais , Cálcio/metabolismo , Carbacol/farmacologia , Fenômenos Eletrofisiológicos/efeitos dos fármacos , Intestinos/fisiologia , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Músculo Liso/citologia , Músculo Liso/efeitos dos fármacosRESUMO
Chlorogenic acid (CGA) is a polyphenol found in coffee and medicinal herbs such as Lonicera japonica. In this study, the effect of CGA-induced relaxation on carbachol (CCh)-induced contraction of mouse urinary bladder was investigated. CGA (30-300 µg/ml) inhibited CCh- or U46619-induced contraction in a concentration-dependent manner. SQ22536 (adenylyl cyclase inhibitor) recovered CGA-induced relaxation of CCh-induced contraction; however, ODQ (guanylyl cyclase inhibitor) did not have the same effect. In addition, 3-isobutyl-1-methylxanthine (IBMX) enhanced CGA-induced relaxation; however, forskolin or sodium nitroprusside did not have the same effect. Moreover, Ro 20-1724, a selective phosphodiesterase (PDE) 4 inhibitor, enhanced CGA-induced relaxation, but vardenafil, a selective PDE5 inhibitor, did not have the same effect. In the presence of CCh, CGA increased cyclic adenosine monophosphate (cAMP) level, whereas SQ22536 inhibited the increase of cAMP levels. Moreover, higher cAMP levels were obtained with CGA plus IBMX treatment than the total cAMP levels obtained with separate CGA and IBMX treatments. In conclusion, these results suggest that CGA inhibited CCh-induced contraction of mouse urinary bladder by partly increasing cAMP levels via adenylyl cyclase activation.
Assuntos
Carbacol/antagonistas & inibidores , Ácido Clorogênico/farmacologia , Contração Muscular/efeitos dos fármacos , Relaxamento Muscular/efeitos dos fármacos , Bexiga Urinária/efeitos dos fármacos , 1-Metil-3-Isobutilxantina/farmacologia , 4-(3-Butoxi-4-metoxibenzil)-2-imidazolidinona/farmacologia , Adenilil Ciclases/metabolismo , Animais , Carbacol/farmacologia , AMP Cíclico/metabolismo , Relação Dose-Resposta a Droga , Técnicas In Vitro , Masculino , Camundongos Endogâmicos , Inibidores da Fosfodiesterase 4/farmacologiaRESUMO
BACKGROUND: Propofol is a widely used intravenous general anesthetic. Acetylcholine (ACh) is critical in controlling epithelial ion transport. This study was to investigate the effects of propofol on ACh-evoked secretion in rat ileum epithelium. METHODS: The Ussing chamber technique was used to investigate the effects of propofol on carbachol (CCh)-evoked short-circuit currents (Isc). KEY RESULTS: Propofol (10-2 -10-6 mol/L) attenuated CCh-evoked Isc of rat ileum mucosa in a dose-dependent manner. The inhibitory effect of propofol was only evident after application to the serosal side. Pretreatment with tetrodotoxin (TTX, 0.3 µmol/L, n=5) had no effect on propofol-induced inhibitory effect, whereas serosal application of K+ channel inhibitor, glibenclamide, but not, an ATP-sensitive K+ channel inhibitor, largely reduced the inhibitory effect of propofol. In addition, pretreatment with either hexamethonium bromide (HB, nicotinic nACh receptor antagonist) or Cl- channel blockers niflumic acid and cystic fibrosis transmembrane conductance regulator (inh)-172 did not produce any effect on the propofol-induced inhibitory effect. CONCLUSIONS & INFERENCES: Propofol inhibits CCh-induced intestinal secretion by directly targeting basolateral K+ channels.
Assuntos
Carbacol/farmacologia , Cloretos/antagonistas & inibidores , Íleo/metabolismo , Mucosa Intestinal/metabolismo , Bloqueadores dos Canais de Potássio/farmacologia , Propofol/farmacologia , Anestésicos Intravenosos/farmacologia , Animais , Carbacol/antagonistas & inibidores , Cloretos/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Íleo/efeitos dos fármacos , Mucosa Intestinal/efeitos dos fármacos , Masculino , Canais de Potássio/fisiologia , Ratos , Ratos WistarRESUMO
Background: The hypomotility of colon observed in Hirschsprung's disease (HD) has been attributed to congenital aganglionosis only. So far, it is not clear whether the contractility of colonic smooth muscle in this condition is altered or not. Therefore, the present study attempted to understand the contractile status of colonic segments of HD patients by examining carbachol and endothelin (ET-1) evoked colonic smooth muscle contractions in vitro . Methods: Contractile responses were recorded from strips of colonic segments obtained from HD patients, using organ bath preparations. Cholinergic agonist carbachol and ET-1 along with their antagonists were used to evoke contractile responses. Thereafter, the samples were histopathologically confirmed for HD. Results: Colonic strips of HD did not show any spontaneous contractions but responded to carbachol and ET-1 to a lesser extent. In HD, response of carbachol was blocked by atropine and hexamethonium by nearly 73% and 50% respectively. ET-1 induced contractile responses were blocked by ET-A and ET-B antagonist up to 40%, signifying the possible role of ET-A and ET-B receptors in HD colon contractility. Conclusion: As evidenced by lack of spontaneous contractions and impaired carbachol and ET-1-induced contractile responses, it is concluded that, in addition to aganglionosis, decreased contractility of colonic smooth muscle may contribute to hypomotility observed in patients with HD.
Assuntos
Carbacol/farmacologia , Colo/efeitos dos fármacos , Endotelinas/farmacologia , Doença de Hirschsprung/fisiopatologia , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Atropina/administração & dosagem , Atropina/farmacologia , Carbacol/antagonistas & inibidores , Colo/fisiologia , Endotelinas/antagonistas & inibidores , Hexametônio/administração & dosagem , Hexametônio/farmacologia , Doença de Hirschsprung/metabolismo , Doença de Hirschsprung/patologia , Humanos , Músculo Liso/fisiologiaRESUMO
Flavonoid galetin 3,6-dimethyl ether (FGAL) has been isolated from the aerial parts of Piptadenia stipulaceae and has shown a spasmolytic effect in guinea pig ileum. Thus, we aimed to characterize its relaxant mechanism of action. FGAL exhibited a higher relaxant effect on ileum pre-contracted by histamine (EC50=1.9±0.4×10(-7) M) than by KCl (EC50=2.6±0.5×10(-6) M) or carbachol (EC50=1.8±0.4×10(-6) M). The flavonoid inhibited the cumulative contractions to histamine, as well as to CaCl2 in depolarizing medium nominally Ca(2+)-free. The flavonoid relaxed the ileum pre-contracted by S-(-)-Bay K8644 (EC50=9.5±1.9×10(-6) M) but less potently pre-contracted by KCl or histamine. CsCl attenuated the relaxant effect of FGAL (EC50=1.1±0.3×10(-6) M), but apamin or tetraethylammonium (1mM) had no effect (EC50=2.6±0.2×10(-7) and 1.6±0.3×10(-7) M, respectively), ruling out the involvement of small and big conductance Ca(2+)-activated K(+) channels (SKCa and BKCa, respectively). Either 4-aminopyridine or glibenclamide attenuated the relaxant effect of FGAL (EC50=1.8±0.2×10(-6) and 1.5±0.5×10(-6) M, respectively), indicating the involvement of voltage- and ATP-sensitive K(+) channels (KV and KATP, respectively). FGAL did not alter the viability of intestinal myocytes in the MTT assay and decreased (88%) Fluo-4 fluorescence, indicating a decrease in cytosolic Ca(2+) concentration. Therefore, the relaxant mechanism of FGAL involves pseudo-irreversible noncompetitive antagonism of histaminergic receptors, KV and KATP activation and blockade of CaV1, thus leading to a reduction in cytosolic Ca(2+) levels.
Assuntos
Cálcio/metabolismo , Flavonoides/farmacologia , Íleo/efeitos dos fármacos , Contração Muscular/efeitos dos fármacos , Canais de Potássio/agonistas , Éster Metílico do Ácido 3-Piridinacarboxílico, 1,4-Di-Hidro-2,6-Dimetil-5-Nitro-4-(2-(Trifluormetil)fenil)/farmacologia , 4-Aminopiridina/farmacologia , Animais , Apamina/farmacologia , Cloreto de Cálcio/antagonistas & inibidores , Cloreto de Cálcio/farmacologia , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Césio/farmacologia , Cloretos/farmacologia , Flavonoides/antagonistas & inibidores , Glibureto/farmacologia , Cobaias , Histamina/farmacologia , Antagonistas dos Receptores Histamínicos/farmacologia , Íleo/fisiologia , Células Musculares/efeitos dos fármacos , Bloqueadores dos Canais de Potássio/farmacologia , Cloreto de Potássio/antagonistas & inibidores , Cloreto de Potássio/farmacologia , TetraetilamônioRESUMO
Endocannabinoids (eCBs) are cannabis-like substances produced in the brain where their primary function is to regulate synaptic transmission by inhibiting neurotransmitter release in a retrograde fashion. We have recently demonstrated a novel mechanism regulating GABAergic transmission from neurons in the Substantia Nigra pars reticulata (SNr) to dopaminergic neurons in the Substantia Nigra pars compacta (SNc) mediated by eCBs. Production of eCBs was initiated by spillover of glutamate, yet the source of the glutamate was not determined (Freestone et al., 2014; Neuropharmacology 79 p467). The present study aimed at elucidating the potential role of glutamatergic terminals arising from neurons in the Subthalamic nucleus (STN) in driving the eCB-mediated modulation of this inhibitory transmission. GABAergic IPSCs or IPSPs evoked in SNc neurons by electrical stimuli delivered to the SNr region were transiently inhibited by electrical or pharmacological (U-tube application of muscarinic agonist carbachol [100 µM]) stimulation of the STN (to 74±5% and 69±4% respectively). In both stimulation protocols, the attenuation of GABAergic transmission was abolished by cannabinoid receptor 1 antagonist rimonabant (3 µM), and reduced by group 1 metabotropic glutamate receptor antagonist CPCCOEt (100 µM), consistent with a glutamate-initiated and eCB-mediated mechanism. The carbachol-induced attenuation of GABAergic transmission was abolished by M3 muscarinic receptor antagonist 4-DAMP (10 µM), confirming a specific activation of STN neurons. These results demonstrate that glutamatergic projection from the STN to dopaminergic SNc neurons underlies an eCB-mediated inhibition of GABAergic input to these neurons.
Assuntos
Endocanabinoides/fisiologia , Neurônios GABAérgicos/fisiologia , Parte Compacta da Substância Negra/fisiologia , Núcleo Subtalâmico/fisiologia , Transmissão Sináptica/fisiologia , Animais , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Cromonas/administração & dosagem , Cromonas/farmacologia , Neurônios Dopaminérgicos/fisiologia , Estimulação Elétrica , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Neurônios GABAérgicos/efeitos dos fármacos , Potenciais Pós-Sinápticos Inibidores/efeitos dos fármacos , Potenciais Pós-Sinápticos Inibidores/fisiologia , Parte Reticular da Substância Negra/efeitos dos fármacos , Parte Reticular da Substância Negra/fisiologia , Piperidinas/farmacologia , Pirazóis/farmacologia , Ratos , Rimonabanto , Núcleo Subtalâmico/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacosRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Pittosporum mannii Hook. f. (Pittosporaceae) is a plant widely used in traditional medicine in Cameroon for the treatment of many gastrointestinal disorders including diarrhea. To date, no pharmacological study on the antidiarrheal and the antispasmodic properties of this plant has been reported. The aim of the present study was to evaluate in vitro the relaxant activity of the aqueous extract of stem barks of P. mannii (PMAE) on rat duodenum. MATERIALS AND METHODS: Different concentrations of PMAE were tested separately (10-80 µg/mL) or cumulatively (5-80 µg/mL) on spontaneous and spasmogen (carbachol, histamine and KCl)-induced contractions of isolated rat duodenum strips. RESULTS: At concentrations ranging from 10 to 80 µg/mL, PMAE significantly decreased the tonus and the amplitude of spontaneous contractions. However, at high concentration (80 µg/mL), the extract elicited a transient relaxation was followed by a slight increase of tonus, while the amplitude remained lower compared to the normal spontaneous activity. The relaxant effect of the extract was not significantly affected in the presence of atropine (0.713 µg/mL) and promethazine (0.5 µg/mL). In addition, PMAE (20, 40, and 80 µg/mL) partially but significantly inhibited in a concentration related manner the contractions induced by carbachol (10(-9)-10(-4)M) and histamine (10(-9)-10(-4)M) on rat duodenum. PMAE (10-80 µg/mL) also significantly induced a concentration-dependent relaxation on KCl (20mM, 50mM, 10(-3)-6.10(-3)M)-induced contraction of rat duodenum. CONCLUSIONS: These results show that the aqueous extract of P. mannii stem barks possesses antispasmodic and spasmolytic effects at lower concentrations; therefore, supporting the use of the stem barks of this plant in the folk medicine for the treatment of diarrhea. However, caution should be paid while using higher concentrations that instead might produce spasmogenic effect and might worsen the diarrheal condition. The relaxant effect of PMAE appears to be non-specific of muscarinic or histaminic receptors, but may involve at least in part a mechanism of inhibition of the Ca(2+) influx into the smooth muscle cells through voltage-operated Ca(2+) channels.
Assuntos
Duodeno/efeitos dos fármacos , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Casca de Planta/química , Extratos Vegetais/farmacologia , Caules de Planta/química , Rosales/química , Animais , Atropina/farmacologia , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Relação Dose-Resposta a Droga , Duodeno/fisiologia , Histamina/farmacologia , Antagonistas dos Receptores Histamínicos/farmacologia , Técnicas In Vitro , Masculino , Medicina Tradicional , Contração Muscular/fisiologia , Músculo Liso/fisiologia , Parassimpatolíticos/farmacologia , Extratos Vegetais/química , Cloreto de Potássio/antagonistas & inibidores , Cloreto de Potássio/farmacologia , Prometazina/farmacologia , RatosRESUMO
Ciliary muscle is a smooth muscle characterized by a rapid response to muscarinic receptor stimulation and sustained contraction. Although it is evident that these contractions are Ca(2+)-dependent, detailed molecular mechanisms are still unknown. In order to elucidate the role of Ser/Thr protein phosphatase 2A (PP2A) in ciliary muscle contraction, we examined the effects of okadaic acid and other PP2A inhibitors on contractions induced by carbachol (CCh) and ionomycin in bovine ciliary muscle strips (BCM). Okadaic acid inhibited ionomycin-induced contraction, while it did not cause significant changes in CCh-induced contraction. Fostriecin showed similar inhibitory effects on the contraction of BCM. On the other hand, rubratoxin A inhibited both ionomycin- and CCh-induced contractions. These results indicated that PP2A was involved at least in ionomycin-induced Ca(2+)-dependent contraction, and that BCM had a unique regulatory mechanism in CCh-induced contraction.
Assuntos
Corpo Ciliar/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Ácido Okadáico/farmacologia , Fosfoproteínas Fosfatases/antagonistas & inibidores , Fosfoproteínas Fosfatases/fisiologia , Animais , Cálcio/fisiologia , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Bovinos , Técnicas In Vitro , Ionomicina/antagonistas & inibidores , Ionomicina/farmacologia , Micotoxinas/farmacologia , Polienos/farmacologia , Pironas/farmacologiaRESUMO
Suppressing anxiety and fear memory relies on bidirectional projections between the medial prefrontal cortex and the amygdala. Positive allosteric modulators of mGluR5 improve cognition in animal models of schizophrenia and retrieval of newly formed associations such as extinction of fear-conditioned behaviour. The increase in neuronal network activities of the medial prefrontal cortex is influenced by both mGluR1 and mGluR5; however, it is not well understood how they modulate network activities and downstream information processing. To map mGluR5-mediated network activity in relation to its emergence as a viable cognitive enhancer, we tested group I mGluR compounds on medial prefrontal cortex network activity via multi-electrode array neuronal spiking and whole-cell patch clamp recordings. Results indicate that mGluR5 activation promotes feed-forward inhibition that depends on recruitment of neuronal activity by carbachol-evoked up states. The rate of neuronal spiking activity under the influence of carbachol was reduced by the mGluR5 positive allosteric modulator, N-(1,3-Diphenyl-1H-pyrazolo-5-yl)-4-nitrobenzamide (VU-29), and enhanced by the mGluR5 negative allosteric modulator, 3-((2-methyl-1,3-thiazol-4-yl)ethynyl)pyridine hydrochloride (MTEP). Spontaneous inhibitory post-synaptic currents were increased upon application of carbachol and in combination with VU-29. These results emphasize a bias towards tonic mGluR5-mediated inhibition that might serve as a signal-to-noise enhancer of sensory inputs projected from associated limbic areas onto the medial prefrontal cortex neuronal microcircuit.
Assuntos
Vias Neurais/efeitos dos fármacos , Córtex Pré-Frontal/efeitos dos fármacos , Córtex Pré-Frontal/fisiologia , Receptor de Glutamato Metabotrópico 5/fisiologia , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Regulação Alostérica/efeitos dos fármacos , Animais , Benzamidas/farmacologia , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Sinergismo Farmacológico , Agonistas de Aminoácidos Excitatórios/farmacologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Potenciais Pós-Sinápticos Inibidores/efeitos dos fármacos , Potenciais Pós-Sinápticos Inibidores/fisiologia , Masculino , Metoxi-Hidroxifenilglicol/análogos & derivados , Metoxi-Hidroxifenilglicol/farmacologia , Vias Neurais/fisiologia , Córtex Pré-Frontal/citologia , Pirazóis/farmacologia , Piridinas/farmacologia , Ratos , Receptor de Glutamato Metabotrópico 5/agonistas , Receptores de Glutamato Metabotrópico/agonistas , Receptores de Glutamato Metabotrópico/fisiologia , Tiazóis/farmacologiaRESUMO
Tomopteris helgolandica Greeff 1879 (Tomopteridae) is a transparent holoplanktonic polychaete that can emit a bright light. In this study, we investigated the emission pattern and control of this deep-sea worm's luminescence. Potassium chloride depolarisation applied on anaesthetised specimens triggered a maximal yellow light emission from specific parapodial sites, suggesting that a nervous control pathway was involved. Pharmacological screening revealed a sensitivity to carbachol, which was confirmed by a dose-light response associated with a change in the light emission pattern, where physiological carbachol concentrations induced flashes and higher concentrations induced glows. The light response induced by its hydrolysable agonist, acetylcholine, was significantly weaker but was facilitated by eserine pretreatment. In addition, a specific inhibitory effect of tubocurarine was observed on carbachol-induced emission. Lastly, KCl- and carbachol-induced light responses were significantly reduced when preparations were pre-incubated in Ca(2+)-free artificial seawater or in different calcium channel blockers (verapamil, diltiazem) and calmodulin inhibitor (trifluoperazine) solutions. All of these results strongly suggest that T. helgolandica produces its light flashes via activation of nicotinic cholinergic receptors and a calcium-dependent intracellular mechanism involving L-type calcium channels.
Assuntos
Carbacol/farmacologia , Medições Luminescentes , Poliquetos/fisiologia , Cloreto de Potássio/metabolismo , Receptores Nicotínicos/metabolismo , Zooplâncton/fisiologia , Acetilcolina/farmacologia , Análise de Variância , Animais , Oceano Atlântico , Carbacol/agonistas , Carbacol/antagonistas & inibidores , Relação Dose-Resposta a Droga , Noruega , Água do Mar/química , Tubocurarina/farmacologiaRESUMO
Increased spontaneous firing (hyperactivity) is induced in fusiform cells of the dorsal cochlear nucleus (DCN) following intense sound exposure and is implicated as a possible neural correlate of noise-induced tinnitus. Previous studies have shown that in normal hearing animals, fusiform cell activity can be modulated by activation of parallel fibers, which represent the axons of granule cells. The modulation consists of a transient excitation followed by a more prolonged period of inhibition, presumably reflecting direct excitatory inputs to fusiform cells and an indirect inhibitory input to fusiform cells from the granule cell-cartwheel cell system. We hypothesized that since granule cells can be activated by cholinergic inputs, it might be possible to suppress tinnitus-related hyperactivity of fusiform cells using the cholinergic agonist, carbachol. To test this hypothesis, we recorded multiunit spontaneous activity in the fusiform soma layer (FSL) of the DCN in control and tone-exposed hamsters (10 kHz, 115 dB SPL, 4h) before and after application of carbachol to the DCN surface. In both exposed and control animals, 100 µM carbachol had a transient excitatory effect on spontaneous activity followed by a rapid weakening of activity to near or below normal levels. In exposed animals, the weakening of activity was powerful enough to completely abolish the hyperactivity induced by intense sound exposure. This suppressive effect was partially reversed by application of atropine and was usually not associated with significant changes in neural best frequencies (BF) or BF thresholds. These findings demonstrate that noise-induced hyperactivity can be pharmacologically controlled and raise the possibility that attenuation of tinnitus may be achievable by using an agonist of the cholinergic system.
Assuntos
Carbacol/farmacologia , Núcleo Coclear/fisiologia , Agonistas Muscarínicos/farmacologia , Ruído/efeitos adversos , Animais , Atropina/farmacologia , Limiar Auditivo/efeitos dos fármacos , Carbacol/antagonistas & inibidores , Núcleo Coclear/efeitos dos fármacos , Cricetinae , Interpretação Estatística de Dados , Fenômenos Eletrofisiológicos/efeitos dos fármacos , Potenciais Evocados Auditivos/fisiologia , Frequência Cardíaca/efeitos dos fármacos , Masculino , Mesocricetus , Antagonistas Muscarínicos/farmacologiaRESUMO
The aim of this study was to see if the crude extract of Lepidium sativum (Ls.Cr) exhibits species specificity in its antidiarrheal and antispasmodic activities along with insight into the underlying mechanisms using the in-vivo and in-vitro experiments. Ls.Cr inhibited castor oil-induced diarrhea in mice at doses (300 and 1000 mg/kg) three times higher dose than for rats. In isolated rat ileum and jejunum, Ls.Cr completely inhibited carbachol (CCh), low K⺠(25 mM) and high K⺠(80 mM)-induced contractions, while in guinea-pig tissues, Ls.Cr caused complete inhibition of only CCh-induced contraction. In rabbit tissues, Ls.Cr completely inhibited CCh and low Kâº-induced contractions sensitive to K⺠channel antagonists. Pretreatment of guinea-pig and rat tissues with Ls.Cr caused a rightward shift in CCh-induced contractions in a pattern similar to dicyclomine, while in rabbit and rat tissues, Ls.Cr shifted isoprenaline curves to the left similar to papaverine. These data indicate that the antidiarrheal and antispasmodic activities of L. sativum are species dependent, mediating its antispasmodic effect through combinations of multiple pathways including activation of K⺠channels, and inhibition of muscarinic receptors, Caâºâº channels and PDE enzyme. Rat tissues showed the highest potency. Based on the results, we recommend using multiple species to know the real pharmacological profile of medicinal products.
Assuntos
Antidiarreicos/farmacologia , Diarreia/tratamento farmacológico , Íleo/efeitos dos fármacos , Lepidium sativum/química , Parassimpatolíticos/farmacologia , Fitoterapia , Extratos Vegetais/farmacologia , Animais , Cálcio/agonistas , Carbacol/antagonistas & inibidores , Óleo de Rícino/efeitos adversos , Catárticos , Diarreia/induzido quimicamente , Feminino , Cobaias , Íleo/fisiologia , Jejuno/efeitos dos fármacos , Jejuno/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Especificidade de Órgãos , Extratos Vegetais/análise , Canais de Potássio/efeitos dos fármacos , Canais de Potássio/metabolismo , Coelhos , Ratos , Ratos Sprague-Dawley , Receptores Muscarínicos/efeitos dos fármacos , Receptores Muscarínicos/metabolismo , Especificidade da EspécieRESUMO
Functional studies have shown altered cholinergic mechanisms in the inflamed bladder, which partly depend on muscarinic receptor-induced release of nitric oxide (NO). The current study aimed to characterize which muscarinic receptor subtypes that are involved in the regulation of the nitrergic effects in the bladder cholinergic response during cystitis. For this purpose, in vitro examinations of carbachol-evoked contractions of inflamed and normal bladder preparations were performed. The effects of antagonists with different selectivity for the receptor subtypes were assessed on intact and urothelium-denuded bladder preparations. In preparations from cyclophosphamide (CYP; in order to induce cystitis) pre-treated rats, the response to carbachol was about 75% of that of normal preparations. Removal of the urothelium or administration of a nitric oxide synthase inhibitor re-established the responses in the inflamed preparations. Administration of 4-diphenylacetoxy-N-methylpiperidine (4-DAMP) inhibited the carbachol-induced contractile responses of preparations from CYP pre-treated rats less potently than controls. Pirenzepine and p-fluoro-hexahydro-sila-diphenidol (pFHHSiD) affected the carbachol-induced contractile responses to similar extents in preparations of CYP pre-treated and control rats. However, the Schild slopes for the three antagonists were all significantly different from unity in the preparations from CYP pre-treated rats. Again, L-NNA or removal of the urothelium eliminated any difference compared to normal preparations. This study confirms that muscarinic receptor stimulation in the inflamed rat urinary bladder induces urothelial release of NO, which counteracts detrusor contraction.
Assuntos
Cistite/fisiopatologia , Contração Muscular/fisiologia , Relaxamento Muscular/fisiologia , Receptores Muscarínicos/fisiologia , Bexiga Urinária/fisiopatologia , Urotélio/efeitos dos fármacos , Animais , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Ciclofosfamida , Cistite/induzido quimicamente , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Masculino , Antagonistas Muscarínicos/farmacologia , Contração Muscular/efeitos dos fármacos , Relaxamento Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Músculo Liso/fisiologia , Músculo Liso/fisiopatologia , Óxido Nítrico Sintase/antagonistas & inibidores , Nitroarginina/farmacologia , Piperidinas/farmacologia , Pirenzepina/farmacologia , Ratos , Ratos Sprague-Dawley , Bexiga Urinária/efeitos dos fármacos , Bexiga Urinária/fisiologia , Urotélio/cirurgiaRESUMO
Acetylcholine facilitates long-term potentiation (LTP) and long-term depression (LTD), substrates of learning, memory, and sensory processing, in which acetylcholine also plays a crucial role. Ca(2+) ions serve as a canonical regulator of LTP/LTD but little is known about the effect of acetylcholine on intracellular Ca(2+) dynamics. Here, we investigated dendritic Ca(2+) dynamics evoked by synaptic stimulation and the resulting LTP/LTD in layer 2/3 pyramidal neurons of the rat visual cortex. Under muscarinic stimulation, single-shock electrical stimulation (SES) inducing â¼20 mV EPSP, applied via a glass electrode located â¼10 µm from the basal dendrite, evoked NMDA receptor-dependent fast Ca(2+) transients and the subsequent Ca(2+) release from the inositol 1,4,5-trisphosphate (IP(3))-sensitive stores. These secondary dendritic Ca(2+) transients were highly localized within 10 µm from the center (SD = 5.0 µm). The dendritic release of Ca(2+) was a prerequisite for input-specific muscarinic LTP (LTPm). Without the secondary Ca(2+) release, only muscarinic LTD (LTDm) was induced. D(-)-2-amino-5-phosphopentanoic acid and intracellular heparin blocked LTPm as well as dendritic Ca(2+) release. A single burst consisting of 3 EPSPs with weak stimulus intensities instead of the SES also induced secondary Ca(2+) release and LTPm. LTPm and LTDm were protein synthesis-dependent. Furthermore, LTPm was confined to specific dendritic compartments and not inducible in distal apical dendrites. Thus, cholinergic activation facilitated selectively compartment-specific induction of late-phase LTP through IP(3)-dependent Ca(2+) release.
Assuntos
Cálcio/metabolismo , Neurônios Colinérgicos/fisiologia , Potenciação de Longa Duração/fisiologia , Agonistas Muscarínicos/farmacologia , Córtex Visual/metabolismo , Córtex Visual/fisiologia , Animais , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Dendritos/metabolismo , Dendritos/fisiologia , Interações Medicamentosas , Estimulação Elétrica/métodos , Antagonistas de Aminoácidos Excitatórios/farmacologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Feminino , Heparina/administração & dosagem , Heparina/farmacologia , Técnicas In Vitro , Potenciação de Longa Duração/efeitos dos fármacos , Depressão Sináptica de Longo Prazo/efeitos dos fármacos , Depressão Sináptica de Longo Prazo/fisiologia , Masculino , Microinjeções , Antagonistas Muscarínicos/farmacologia , Inibidores da Síntese de Proteínas/farmacologia , Ratos , Valina/administração & dosagem , Valina/análogos & derivados , Valina/farmacologia , Córtex Visual/efeitos dos fármacosRESUMO
Acute pancreatitis is a major health burden for which there are currently no targeted therapies. Premature activation of digestive proenzymes, or zymogens, within the pancreatic acinar cell is an early and critical event in this disease. A high-amplitude, sustained rise in acinar cell Ca(2+) is required for zymogen activation. We previously showed in a cholecystokinin-induced pancreatitis model that a potential target of this aberrant Ca(2+) signaling is the Ca(2+)-activated phosphatase calcineurin (Cn). However, in this study, we examined the role of Cn on both zymogen activation and injury, in the clinically relevant condition of neurogenic stimulation (by giving the acetylcholine analog carbachol) using three different Cn inhibitors or Cn-deficient acinar cells. In freshly isolated mouse acinar cells, pretreatment with FK506, calcineurin inhibitory peptide (CiP), or cyclosporine (CsA) blocked intra-acinar zymogen activation (n = 3; P < 0.05). The Cn inhibitors also reduced leakage of lactate dehydrogenase (LDH) by 79%, 62%, and 63%, respectively (n = 3; P < 0.05). Of the various Cn isoforms, the ß-isoform of the catalytic A subunit (CnAß) was strongly expressed in mouse acinar cells. For this reason, we obtained acinar cells from CnAß-deficient mice (CnAß-/-) and observed an 84% and 50% reduction in trypsin and chymotrypsin activation, respectively, compared with wild-type controls (n = 3; P < 0.05). LDH release in the CnAß-deficient cells was reduced by 50% (n = 2; P < 0.05). The CnAß-deficient cells were also protected against zymogen activation and cell injury induced by the cholecystokinin analog caerulein. Importantly, amylase secretion was generally not affected by either the Cn inhibitors or Cn deficiency. These data provide both pharmacological and genetic evidence that implicates Cn in intra-acinar zymogen activation and cell injury during pancreatitis.
Assuntos
Células Acinares/efeitos dos fármacos , Inibidores de Calcineurina , Calcineurina/genética , Carbacol/antagonistas & inibidores , Carbacol/toxicidade , Precursores Enzimáticos/metabolismo , Agonistas Nicotínicos/toxicidade , Células Acinares/enzimologia , Amilases/metabolismo , Animais , Calcineurina/fisiologia , Colecistocinina/farmacologia , Quimotripsina/metabolismo , DNA/genética , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/genética , Feminino , Genótipo , L-Lactato Desidrogenase/metabolismo , Masculino , Camundongos , Camundongos Knockout , Pâncreas/citologia , Pâncreas/efeitos dos fármacos , Pâncreas/enzimologia , Monoéster Fosfórico Hidrolases/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Tripsina/metabolismoRESUMO
BACKGROUND: Fibroblast to myofibroblast transition is believed to contribute to airway remodelling in lung diseases such as asthma and chronic obstructive pulmonary disease. This study examines the role of aclidinium, a new long-acting muscarinic antagonist, on human fibroblast to myofibroblast transition. METHODS: Human bronchial fibroblasts were stimulated with carbachol (10(-8) to 10(-5) M) or transforming growth factor-ß1 (TGF-ß1; 2 ng/ml) in the presence or absence of aclidinium (10(-9) to 10(-7) M) or different drug modulators for 48 h. Characterisation of myofibroblasts was performed by analysis of collagen type I and α-smooth muscle actin (α-SMA) mRNA and protein expression as well as α-SMA microfilament immunofluorescence. ERK1/2 phosphorylation, RhoA-GTP and muscarinic receptors (M) 1, 2 and 3 protein expression were determined by western blot analysis and adenosine 3'-5' cyclic monophosphate levels were determined by ELISA. Proliferation and migration of fibroblasts were also assessed. RESULTS: Collagen type I and α-SMA mRNA and protein expression, as well as percentage α-SMA microfilament-positive cells, were upregulated in a similar way by carbachol and TGF-ß1, and aclidinium reversed these effects. Carbachol-induced myofibroblast transition was mediated by an increase in ERK1/2 phosphorylation, RhoA-GTP activation and cyclic monophosphate downregulation as well as by the autocrine TGF-ß1 release, which were effectively reduced by aclidinium. TGF-ß1 activated the non-neuronal cholinergic system. Suppression of M1, M2 or M3 partially prevented carbachol- and TGF-ß1-induced myofibroblast transition. Aclidinium dose-dependently reduced fibroblast proliferation and migration. CONCLUSION: Aclidinium inhibits human lung fibroblast to myofibrobast transition.
Assuntos
Brônquios/citologia , Fibroblastos/efeitos dos fármacos , Antagonistas Muscarínicos/farmacologia , Miofibroblastos/efeitos dos fármacos , Tropanos/farmacologia , Actinas/biossíntese , Actinas/genética , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Diferenciação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Agonistas Colinérgicos/farmacologia , Colágeno Tipo I/biossíntese , Colágeno Tipo I/genética , Relação Dose-Resposta a Droga , Fibroblastos/metabolismo , Humanos , RNA Mensageiro/genética , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Fator de Crescimento Transformador beta1/farmacologia , Regulação para Cima/efeitos dos fármacosRESUMO
The present study deals with the pharmacological effects of the sesquiterpene alcohol (-)-α-bisabolol on various smooth-muscle preparations from rats. Under resting tonus, (-)-α-bisabolol (30-300 µmol/L) relaxed duodenal strips, whereas it showed biphasic effects in other preparations, contracting endothelium-intact aortic rings and urinary bladder strips, and relaxing these tissues at higher concentrations (600-1000 µmol/L). In preparations precontracted either electromechanically (by 60 mmol/L K(+)) or pharmacomechanically (by phenylephrine or carbachol), (-)-α-bisabolol showed only relaxing properties. The pharmacological potency of (-)-α-bisabolol was variable, being higher in mesenteric vessels, whereas it exerted relaxing activity with a lesser potency on tracheal or colonic tissues. In tissues possessing spontaneous activity, (-)-α-bisabolol completely decreased spontaneous contractions in duodenum, whereas it increased their amplitude in urinary bladder tissue. Administered in vivo, (-)-α-bisabolol attenuated the increased responses of carbachol in tracheal rings of ovalbumin-sensitized rats challenged with ovalbumin, but was without effect in the decreased responsiveness of urinary bladder strips in mice treated with ifosfamide. In summary, (-)-α-bisabolol is biologically active in smooth muscle. In some tissues, (-)-α-bisabolol preferentially relaxed contractions induced electromechanically, especially in tracheal smooth muscle. The findings from tracheal rings reveal that (-)-α-bisabolol may be an inhibitor of voltage-dependent Ca(2+) channels.
Assuntos
Contração Muscular/efeitos dos fármacos , Relaxamento Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Sesquiterpenos/farmacologia , Animais , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Cistite/induzido quimicamente , Cistite/tratamento farmacológico , Modelos Animais de Doenças , Duodeno/efeitos dos fármacos , Duodeno/fisiologia , Ifosfamida , Técnicas In Vitro , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Masculino , Sesquiterpenos Monocíclicos , Contração Muscular/fisiologia , Relaxamento Muscular/fisiologia , Músculo Liso/fisiologia , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/fisiologia , Ovalbumina , Fenilefrina/farmacologia , Ratos , Ratos Wistar , Traqueia/efeitos dos fármacos , Traqueia/fisiologia , Bexiga Urinária/efeitos dos fármacos , Bexiga Urinária/fisiologiaRESUMO
Angiotensin II (AngII) plays a key role in maintaining body fluid homeostasis. The physiological and behavioral effects of central AngII include increased blood pressure and fluid intake. In vitro experiments demonstrate that repeated exposure to AngII reduces the efficacy of subsequent AngII, and behavioral studies indicate that prior icv AngII administration reduces the dipsogenic response to AngII administered later. Specifically, rats given a treatment regimen of three icv injections of a large dose of AngII, each separated by 20 min, drink less water in response to a test injection of AngII than do vehicle-treated controls given the same test injection. The present studies were designed to test three potential explanations for the reduced dipsogenic potency of AngII after repeated administration. To this end, we tested for motor impairment caused by repeated injections of AngII, for a possible role of visceral distress or illness, and for differences in the pressor response to the final test injection of AngII. We found that repeated injections of AngII neither affected drinking stimulated by carbachol nor did they produce a conditioned flavor avoidance. Furthermore, we found no evidence that differences in the pressor response to the final test injection of AngII accounted for the difference in intake. In light of these findings, we are able to reject these three explanations for the observed behavioral desensitization, and, we suggest instead that the mechanism for this phenomenon may be at the level of the receptor.
Assuntos
Angiotensina II/farmacologia , Aprendizagem da Esquiva/efeitos dos fármacos , Comportamento de Escolha/efeitos dos fármacos , Ingestão de Líquidos/efeitos dos fármacos , Tolerância a Medicamentos , Vasoconstrição/efeitos dos fármacos , Angiotensina II/administração & dosagem , Animais , Carbacol/antagonistas & inibidores , Carbacol/farmacologia , Interações Medicamentosas , Injeções Intraventriculares , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Inhibition of type-5 phosphodiesterase by sildenafil decreases capacitative Ca2+ entry mediated by transient receptor potential proteins (TRPs) in the pulmonary artery. These families of channels, especially the canonical TRP (TRPC) subfamily, may be involved in the development of bronchial hyperresponsiveness, a hallmark of asthma. In the present study, we evaluated i) the effects of sildenafil on tracheal rings of rats subjected to antigen challenge, ii) whether the extent of TRPC gene expression may be modified by antigen challenge, and iii) whether inhibition of type-5 phosphodiesterase (PDE5) may alter TRPC gene expression after antigen challenge. Sildenafil (0.1 µM to 0.6 mM) fully relaxed carbachol-induced contractions in isolated tracheal rings prepared from naive male Wistar rats (250-300 g) by activating the NO-cGMP-K+ channel pathway. Rats sensitized to antigen by intraperitoneal injections of ovalbumin were subjected to antigen challenge by ovalbumin inhalation, and their tracheal rings were used to study the effects of sildenafil, which more effectively inhibited contractions induced by either carbachol (10 µM) or extracellular Ca2+ restoration after thapsigargin (1 µM) treatment. Antigen challenge increased the expression of the TRPC1 and TRPC4 genes but not the expression of the TRPC5 and TRPC6 genes. Applied before the antigen challenge, sildenafil increased the gene expression, which was evaluated by RT-PCR, of TRPC1 and TRPC6, decreased TRPC5 expression, and was inert against TRPC4. Thus, we conclude that PDE5 inhibition is involved in the development of an airway hyperresponsive phenotype in rats after antigen challenge by altering TRPC gene expression.