Effect of constant light and immobilization stress on rat submandibular saliva secretory response induced by cholinergic and peptidergic agonists

The aim of this work was to analyse the parasympathetic control of submandibular saliva secretory response to cholinergic and peptidergic agonists in rats chronically exposed to constant light or repeated immobilization. Thirty two adult male Wistar rats were used: LL (8 rats exposed to constant light for 20 days), IMO (8 rats submitted to 14:10 h light: dark cycle and immobilized 2 hours daily for 7 days), and control (16 rats not exposed to stress and submitted to 14:10 hours light:dark cycle). Saliva was collected under anesthesia from the salivary ducts of submandibular glands under increasing doses of methacholine and substance P. Secretory responses (µg/saliva/mg dry weight gland) to methacholine were significantly higher in LL and IMO groups compared to control for the following doses (µg/kg body weight): 3 (153±9 versus 46±3, p<0.001 and 76±3 versus 40±3, p<0.001), 10 (379±23 versus 277±8, p<0.001 and 275±19 versus 250±10, p<0.01) and 30 (729±25 versus 695±19, p<0.05 and 1008±39 versus 640±20, p<0.001). Also, responses to substance P were significantly increased in LL and IMO groups compared to control for the following doses: 0.2 (80±3 versus 30±3, p<0.01 and 94±16 versus 31±3, p<0.001), 0.5 (328±20 versus 231±16, p<0.01 and 531±31 versus 219±25,p<0.001), 1 (681±35 versus 547±30, p<0.01 and 1031±63 versus 563±53, p<0.001), and 5 (2222±88 versus 1868±59, p<0.01 and 3230±145 versus 1921±218, p<0.001). In conclusion, supersensitivity of secretory response to both agonists suggests that chronic exposure of rats to stressors capable of activating the sympathetic adrenal system promotes inhibition of the parasympathetic control of salivary secretion.

Vascular function in arteries of intertidal fish Girella laevifrons (Kyphosidae) / Função vascular nas artérias do peixe marinho Girella laevifrons (Kyphosidae)

Preliminary studies showed that dorsal artery contraction mediated by acetylcholine (ACh) is blocked with indomethacin in intertidal fish (Girella laevifrons). Our objective was to characterise the cholinergic pathway in several artery vessels of the G. laevifrons. Afferent and efferent branchial, dorsal and mesenteric arteries were dissected of 6 juvenile specimens, isometric tension studies were done using dose response curves (DRC) for Ach (10–13 to 10–3 M), and cholinergic pathways were obtained by blocking with atropine or indomethacin. CRC to ACh showed a pattern of high and low sensitivity. Furthermore, these contractions were blocked in the presence of atropine and indomethacin in all vessels. Our results suggest that contraction observed with acetylcholine is mediated by receptors that activate a cyclooxygenase contraction pathway.

Estudos preliminares mostraram que a contração da artéria dorsal mediada por acetilcolina (ACh) é bloqueada com indometacina em peixes marinhos Girella laevifrons. Nosso objetivo foi caracterizar a via colinérgica em várias artérias de G. laevifrons. Artérias aferentes e eferentes branquiais, dorsais e mesentéricas foram dissecadas de 6 espécimes juvenis. Os estudos de tensão isométrica foram feitos utilizando-se a curva dose - resposta (CDR) para Ach (10–13 a 10–3M), e identificaram-se as vias colinérgicas, bloqueando com atropina e indometacina. CRC para ACh mostrou um padrão de alta e baixa sensibilidade. Essas contrações foram bloqueadas na presença de atropina e indometacina em todas as artérias avaliadas. Nossos resultados sugerem que a contração observada com acetilcolina é mediada por receptores muscarínicos que ativam uma ciclo-oxigenase.

Resistance Exercise Restores Endothelial Function and Reduces Blood Pressure in Type 1 Diabetic Rats / Exercício Resistido Restaura a Função Endotelial e Reduz a Pressão Arterial de Ratos Diabéticos Tipo 1

Background: Resistance exercise effects on cardiovascular parameters are not consistent. Objectives: The effects of resistance exercise on changes in blood glucose, blood pressure and vascular reactivity were evaluated in diabetic rats. Methods: Wistar rats were divided into three groups: control group (n = 8); sedentary diabetic (n = 8); and trained diabetic (n = 8). Resistance exercise was carried out in a squat device for rats and consisted of three sets of ten repetitions with an intensity of 50%, three times per week, for eight weeks. Changes in vascular reactivity were evaluated in superior mesenteric artery rings. Results: A significant reduction in the maximum response of acetylcholine-induced relaxation was observed in the sedentary diabetic group (78.1 ± 2%) and an increase in the trained diabetic group (95 ± 3%) without changing potency. In the presence of NG-nitro-L-arginine methyl ester, the acetylcholine-induced relaxation was significantly reduced in the control and trained diabetic groups, but not in the sedentary diabetic group. Furthermore, a significant increase (p < 0.05) in mean arterial blood pressure was observed in the sedentary diabetic group (104.9 ± 5 to 126.7 ± 5 mmHg) as compared to that in the control group. However, the trained diabetic group showed a significant decrease (p < 0.05) in the mean arterial blood pressure levels (126.7 ± 5 to 105.1 ± 4 mmHg) as compared to the sedentary diabetic group. Conclusions: Resistance exercise could restore endothelial function and prevent an increase in arterial blood pressure in type 1 diabetic rats. .

Fundamento: Os efeitos do exercício resistido sobre os parâmetros cardiovasculares não são consistentes. Objetivos: Foram avaliados os efeitos do exercício resistido sobre as alterações na glicemia, reatividade vascular e pressão arterial de ratos diabéticos. Métodos: Ratos Wistar foram divididos em três grupos: grupo controle (n = 8), diabético sedentário (n = 8) e diabético treinado (n = 8). O exercício resistido foi realizado no aparelho de agachamento para ratos e consistiu em três séries de dez repetições com uma intensidade de 50%, três vezes por semana, durante 8 semanas. As alterações na reatividade vascular foram avaliadas em anéis de artéria mesentérica superior. Resultados: Foi observada uma redução significativa da resposta máxima dos relaxamentos induzidos por acetilcolina no grupo diabético sedentário (78,1% ± 2) e um aumento do grupo diabético treinado (95 ± 3%), sem alterar a potência. Na presença de NG-nitro-L-arginina metil éster, os relaxamentos induzidos por acetilcolina foram significativamente reduzidos nos grupos controle e diabético treinado, mas não no grupo diabético sedentário. Além disso, foi observado um aumento significativo (p < 0,05) da pressão arterial média no grupo diabético sedentário de 104,9 ± 5 para 126,7 ± 5 mmHg, quando comparado ao grupo controle. Por outro lado, o grupo diabético treinado apresentou redução significativa (p < 0,05) nos níveis da pressão arterial média de 126,7 ± 5 mmHg para 105,1 ± 4 mmHg, quando comparado ao diabético sedentário. Conclusões: O exercício resistido foi capaz de restaurar a funcionalidade endotelial e impedir o aumento da pressão arterial em ratos com ...

Tratamento da síndrome da bexiga hiperativa idiopática refratária / Treatment of idiopathic refractory overactive bladder syndrome

A síndrome da bexiga hiperativa é um distúrbio caracterizado pela presença de urgência miccional, podendo ou não haver incontinência urinária associada. Afeta homens e mulheres em iguais proporções, mas tem maior impacto sobre a população feminina, já que a incidência de incontinência é maior. O tratamento de primeira linha consiste em medidas comportamentais, treinamento vesical e fisioterapia, podendo-se associar tratamento farmacológico em casos persistentes. As drogas mais frequentemente utilizadas são os anticolinérgicos, medicações eficazes, porém associadas a efeitos adversos incômodos que frequentemente limitam a aderência. Uma proporção considerável de mulheres não obtém êxito com a combinação de medidas conservadoras e medicações anticolinérgicas, seja por eficácia limitada, seja por efeitos colaterais intoleráveis. Para essa população, modalidades de tratamento de segunda e terceira linha podem trazer alívio sintomático e melhora da qualidade de vida. Além disso, foi recentemente aprovada uma nova classe de drogas para o tratamento da bexiga hiperativa, os agonistas de receptores ?3-adrenérgicos, que prometem eficácia equivalente aos anticolinérgicos sem os efeitos adversos que os limitam...

Overactive bladder syndrome is a condition characterized by urgency, with or without urinary incontinence. It affects men and women equally, but has a greater impact on the female population, given the higher prevalence of urgency-incontinence. First line treatment consists on lifestyle interventions, bladder drills and physical therapy. Pharmacological treatment may be associated in persistent cases. The most commonly used medications are anticholinergics, which are efficacious, but limited by a variety of bothersome adverse effects that impair treatment compliance. A significant proportion of women don?t experience a successful outcome with the combination of conservative measures and treatment with anticholinergics, owing both to limited efficacy and to intolerable adverse effects that lead to treatment discontinuation. For this population, second and third line therapies may provide symptomatic relief, with great improvement in health related quality of life. Also, a new class of drugs, the ?3-adrenergic receptor agonists, has recently been approved for the treatment of overactive bladder, and may provide similar efficacy to currently used anticholinergic drugs without the associated adverse effects...

Intervalos de referencia para agregación plaquetaria en individuos sanos / Platelet aggregation reference interval for healthy individuals / Intervalos de referência para agregação plaquetária

La agregación por transmisión de luz (ATL) es el método más usado por laboratorios clínicos y de investigación para evaluar la función plaquetaria y es considerado actualmente el estándar de oro; no obstante, la ATL aún no es un método estandarizado, pese a los esfuerzos de organismos internacionales como la International Society of Thrombosis and Hemostasis (ISTH). Organismos regulatorios recomiendan que cada laboratorio clínico determine e informe sus intervalos de referencia (IR) para cada agonista que utiliza. Se presenta los IR de este laboratorio en la determinación de agregación plaquetaria mediante ALT utilizando adenosin difosfato (ADP), colágeno y adrenalina como agonistas. Para ello se diseñó un estudio de corte transversal sobre una muestra a conveniencia de voluntarios(as) aparentemente sanos; se usó un agregómetro óptico y se emplearon los siguientes agonistas y concentraciones finales: ADP 5 µM, colágeno 2 µM y adrenalina 10 µM. Se definió IR como los percentiles 2,5 y 97,5 (P2,5 y P97,5) del Porcentaje de Agregación Plaquetaria Máxima APM%. Participaron 63 individuos, rango de edad 18 a 66 años, 79,4% sexo femenino. Los valores de APM% fueron: ADP P2,5=49% y P97,5=87%; colágeno P2,5=43% y P97,5=86%; adrenalina P2,5=42% y P97,5=85%. Atendiendo a recomendaciones internacionalmente aceptadas, se presentan los IR de APM (%) por el método ATL en este laboratorio (ASCARDIO, Barquisimeto, Venezuela), lo que permite al clínico basar sus decisiones en evidencia válida y pertinente.

Platelet aggregation tests by means of light transmission (LTA), the current gold standard, are the most commonly used methods used to evaluate platelet function at clinical and research laboratories. However, LTA has not been determinastandardized despite the work from international organizations such as the International Society of Thrombosis and Homeostasis (ISTH). Regulatory agencies recommend that each clinical laboratory establishes and informs its own Reference Internal (RI) for all agonists they use. RI are presented for our laboratory using the following agonists: diphosphate (ADP), collagen y adrenaline and the pertaining methodology. To assess our RI for platelet aggregation tests by LTA, a cross-sectional study was designed with a convenience sample of healthy volunteer men and women using an optical aggregometer with the following agonist and final concentration: adenosine diphosphate (ADP) 5 µM, collagen 2 µM and adrenaline 10 µM. The RIs were defined as Percentiles 2.5 (P2,5) and 97.5 (P97.5) of the percentage of maximal aggregation (%MA). 63 subjects participate, age range 18 to 66,79.4% were female. The IR for %MA were: ADP P2,5=49% and P97.5=87%; collagen P2,5=43% and P97.5=86%; adrenaline P2,5=42% and P97.5=85%. In agreement with international accepted recommendation guidelines, RIs for the %MA values were presented by LTA done in our clinical laboratory (ASCARDIO, Barquisimeto, Lara State, Venezuela), that allows physicians to base their clinical decision process on valid and pertinent information.

A agregometria por transmissão de luz (ATL) é o método mais utilizado pelos laboratórios clínicos e de pesquisa para avaliar a função plaquetária e é atualmente considerada o padrão-ouro; no entanto, a ATL ainda não é um método padronizado, apesar dos esforços das agências internacionais como a International Society of Thrombosis and Hemostasis (ISTH). Agências reguladoras recomendam que cada laboratório clínico determine e comunique seus intervalos de referência (IR) para cada agonista utilizado. São apresentados o IR de nosso laboratório para determinar a agregação plaquetária através de ALT usando Difosfato de Adenosina (ADP), Colágeno e Adrenalina como agonistas. Para isso foi elaborado um estudo de corte transversal em uma amostra de conveniência de voluntários(as) aparentemente saudáveis , foi usado agregômetro ótico e foram utilizados os seguintes agonistas e concentrações finais: 5µ M ADP, Colágeno 2µM e Adrenalina 10µM. Definiu-se o IR com os percentis 2,5 e 97.5 (P2,5 e P97.5) do Percentual de Agregação Plaquetária Máxima APM%. Participaram 63 indivíduos, na faixa etária 18-66 anos, 79,4% do sexo feminino. Os valores de APM% foram: ADPP2,5=49% e P97.5=87%; Colágeno P2,5=43% e P99,5=86%, Adrenalina P2,5=42% e P97.5=85%. Atendendo às recomendações aceitas internacionalmente, apresentam-se os IR de APM% pelo método ATL em nosso laboratório (ASCARDIO, Barquisimeto, Venezuela), o que permite ao médico basear as suas decisões em evidências válidas e pertinentes.

Role of rennin-angiotensin system in cholinergic agonist carbachol-induced cardiovascular responses in ovine fetus / 生理学报

To investigate the mechanisms underlying the cholinergic agonist carbachol-induced cardiovascular responses, changes of renin-angiotensin system were examined in fetal hormonal systems. In the ovine fetal model under stressless condition, the cardiovascular function was recorded. Blood samples were collected before (during baseline period) and after the intravenous administration of carbachol. Simultaneously, the levels of angiotensin I (Ang I), angiotensin II (Ang II) and vasopressin in the fetal plasma were detected by immunoradiological method. Also, blood gas, plasma osmolality and electrolyte concentrations were analyzed in blood samples. Results showed that in chronically prepared ovine fetus, intravenous infusion of carbachol led to a significant decrease of heart rate (P < 0.05), and a transient decrease followed by an increase of blood pressure (P < 0.05) within 30 min. After the intravenous infusion of carbachol, blood concentrations of Ang I and Ang II in near-term ovine fetus were both significantly increased (P < 0.05); however, blood concentration of vasopressin, values of blood gas, electrolytes and plasma osmolality in near-term ovine fetus were not significantly changed (P > 0.05). Blood levels of Ang I and Ang II in the atropine (M receptor antagonist) + carbachol intravenous administration group was lower than those in the carbachol group without atropine administration (P < 0.05). In conclusion, this study indicates that the near-term changes of cardiovascular system induced by intravenous administration of carbachol in ovine fetus, such as blood pressure and heart rate, are associated with the changes of hormones of circulatory renin-angiotensin system.

Histological, biochemical and pharmacologycal characterization of the gastric muscular layer in Chagas disease / Caracterização histológica, bioquímica e farmacológica da musculatura gástrica na doença de Chagas

PURPOSE: To assess in vitro the correlation between the number of neurons and the sensitivity to cholinergic drugs and acetylcholinesterase activity in chagasic patients. METHODS: A 3x1 cm strip of the muscle layer of the anterior part of the stomach, always close to the angular incisure, was removed from 10 chronic chagasic patients (6 men) submitted to megaesophagus or megacolon surgery and from 10 non-chagasic patients (4 men) submitted to other types of surgery (control group), aged on average 52.3 and 50.1 years, respectively, for histological and pharmacological studies. The action of cholinergic drugs was investigated in isolated preparations according to the superfusion method of Ferreira and Costa, and acetylcholinesterase activity was determined by the method of Ellman. For neuron count, the strips were cut into 8 µm sections according to the method standardized by Alcântara. RESULTS: There was a difference in number of neurons between the chagasic (5,6) and control (7,3) groups. Acetylcholinesterase activity, in moles of hydrolyzed substrate per minute per gram tissue, was reduced in chagasic patients (4,32) compared to the controls (7,30). No hypersensitivity of the gastric musculature to cholinergic drugs was detected, with a reduced maximum response to carbachol and betanechol in the chagasic group. CONCLUSIONS: The reduction of neurons in the myenteric plexus of the stomach of chronic chagasic patients can be demonstrated even in the absence of clinical chagasic gastropathy. The hypersensitivity of the gastric musculature to cholinergic drugs probably depends on intense denervation. The reduced acetylcholinesterase activity demonstrates the involvement of the cholinergic innervation in the stomach of chronic chagasic patients. There was no correlation between number of neurons, sensitivity to cholinergic drugs and acetylcholinesterase activity in the gastric musculature of chagasic and non-chagasic patients.

OBJETIVO: Avaliar in vitro a correlação entre o número de neurônios e a sensibilidade a drogas colinérgicas e a atividade da acetilcolinesterase em pacientes chagásicos. MÉTODOS: Em 10 pacientes chagásicos crônicos (6 homens) submetidos à cirurgia de megaesôfago ou de megacólon e em 10 pacientes não chagásicos (4 homens) submetidos a outros tipos de cirurgia (grupo controle), respectivamente com idade média de 52,3 e 50,1 anos, retirou-se uma tira de 3x1 cm da camada muscular da parede anterior do estômago, sempre junto á cisura angular, que serviu para os estudos histológicos e farmacológicos. A ação de drogas colinérgicas foi feita em preparação isolada de acordo com o método de superfusão de Ferreira e Costa, e a determinação da atividade da acetilcolinesterase pelo método de Ellman. Para a contagem de neurônios a tira muscular foi submetida a cortes de 8 micra segundo método padronizado por Alcântara. RESULTADOS: Houve diferença do número de neurônios entre os grupos chagásico (5,6) e controle (7,3). A atividade da acetilcolinesterase mostrou-se diminuída nos chagásicos (4,32) expressa como número de moles do substrato hidrolisado por minuto por grama de tecido, em relação aos controles (7,30). Não se encontrou hipersensibilidade da musculatura gástrica a drogas colinérgicas, encontrando-se inclusive efeito máximo reduzido ao carbacol e betanecol no grupo chagásico. CONCLUSÕES: A redução de neurônios no plexo mioentérico do estômago de pacientes chagásicos crônicos pode ser demonstrada mesmo na ausência de gastropatia chagásica clínica. A hipersensibilidade da musculatura gástrica a drogas colinérgicas provavelmente depende de desnervação intensa. A redução da atividade da acetilcolinesterase demonstra o comprometimento da inervação colinérgica no estômago de pacientes chagásicos crônicos. Não houve correlação entre número de neurônios, sensibilidade a drogas colinérgicas e atividade da acetilcolinesterase na musculatura gástrica de pacientes chagásicos ou não chagásicos.

Avaliação morfológica e dos mecanismos de mobilização de Ca2+ pela glicose e acetilcolina em células pancreáticas humanas / Morphologic evaluation and Ca2+ mobilization by glicose and acetylcholine in human pancreatic cells

OBJETIVOS: Avaliar a morfologia das organelas e do citoesqueleto em células pancreáticas humanas cultivadas e a mobilização de Ca2+ em resposta à glicose e ACh por medidas fluorimétricas. MATERIAL E MÉTODOS: As células foram semeadas em lamínulas, fixadas e marcadas com uma combinação de fluoróforos: o núcleo foi corado com DAPI e as mitocôndrias, com Mytotracker Red. Foram utilizados faloidina e anticorpos secundários conjugados com Alexa Fluor verde e vermelho fluorescentes (488 e 594) para identificar proteína actina F e receptor muscarínico tipo M3, respectivamente. Para estudar a mobilização de Ca2+, as células foram incubadas com fura-2/AM. RESULTADOS: As células pancreáticas humanas apresentaram morfologia preservada com grande quantidade de mitocôndrias. Na região de maior densidade celular, evidenciou-se as pseudo-ilhotas e os receptores muscarínicos M3. Por meio da elevação da [Ca2+]c, devido à ação da glicose e ACh, mostrou-se preservação da capacidade responsiva a esses estímulos e foi dependente de concentração desses agonistas. A glicose promoveu uma resposta sustentada e a ACh induziu uma resposta bifásica. CONCLUSÃO: As células pancreáticas humanas cultivadas conservaram sua morfologia. A mobilização de Ca2+ em resposta à glicose e a ACh confirma a sua funcionalidade. Os receptores muscarínicos M3 estão presentes nessas células.

AIMS: The proposal of this study was to analyze morphology of the organelles and cytoskeleton in human pancreatic cells cultured and the mobilization of the cytosolic calcium ([Ca2+]c) in response to glucose and ACh by fluorimetry method. MATERIAL AND METHODS: The cells were plated on glass coverslips, fixed and stained with a combination of fluorophores: the nuclei were stained with DAPI and mitochondria with Mytotracker Red. It was used phalloidin and the secondary antibodies Alexa Fluor conjugated green and red-fluorescent (488 and 594) to identify the protein cell actin F and type M3 muscarinic receptor respectively. The cells also were loaded with fura-2/AM to study Ca2+ mobilization. RESULTS: The human pancreatic cells show characteristics morphologically preserved with great amount of mitochondria. In region major cell density was evidenced pseudo-islets and type M3 muscarinic receptors. Through increase of [Ca2+]c due to action of glucose and ACh were shown that the cellsÆ capacity to respond to these stimuli were conserved. The elevation of the [Ca2+]c depended on concentration by glucose-induced promoting sustained phase and ACh-induced a biphasic response. CONCLUSION: The morphologic characteristics of human pancreatic cells cultured were preserved. The Ca2+ mobilization in response to glucose and ACh confirmed its functionality. The expression of the M3 muscarinic receptors in human pancreatic cell cultured was demonstrated.

Effects of arecoline and nicotine on the expression of hTERT in oral keratinocytes / 中华口腔医学杂志

<p><b>OBJECTIVE</b>To investigate the effects of arecoline and nicotine on the expression of human telomerase reverse transcriptase (hTERT) mRNA and protein in cultured normal human oral keratinocytes (KC).</p><p><b>METHODS</b>The experiments were divided into arecoline group, arecoline/nicotine group and control group. The hTERT mRNA and protein expression of KC was examined by reverse transcription polymerase chain reaction (RT-PCR) and Western blot.</p><p><b>RESULTS</b>Arecoline could induce the hTERT mRNA and protein expression of KC in a dose dependent manner, the hTERT mRNA and protein expression of KC was higher in 0.030, 0.060, 0.090 g/L arecoline group than control group (P < 0.001). Nicotine (0.025 g/L) increased hTERT mRNA and protein expression of KC induced by arecoline.</p><p><b>CONCLUSIONS</b>Arecoline could increase the expression of hTERT mRNA and protein in oral keratinocytes. Nicotine had a synergistic effect on arecoline. hTERT over-expression induced by arecoline and nicotine may play an important role in the malignant transformation of oral submucous fibrosis.</p>

Prevención de los efectos adversos anticolinérgicos inducidos por los antipsicóticos en la 3ª edad: descripción de un caso en tratamiento con clozapina / Prevention of anticholinergic adverse effect induced by antipsychotics in aged: report of a case in treatment with clozapine

La seguridad y eficacia del uso de Clozapina en ancianos es actualmente objeto de estudio. Los pacientes geriátricos parecen presentar una mayor predisposición a sufrir hipotensión ortoestática y una mayor reactividad a los efectos anticolinérgicos, como también son más sensibles a los efectos adversos sobre el SNC. Se describe en el presente estudio el caso de una paciente portadora de un cuadro esquizofrénico catatónico en tratamiento con Clozapina, que presentó marcados síntomas anticolinérgicos y que mejoró de sus síntomas tras la suspensión del fármaco y el cambio a otro antipsicótico atípico (Risperidona). El estudio de niveles plasmáticos de Clozapina mostró que aún con 50 mg/día alcanzaba niveles plasmáticos de 200 ng/dl. Se analiza en el presente reporte la necesidad de una adecuada evaluación clínica y la importancia del control de Niveles Plasmáticos.

Relationship between carbachol hyperstimulation-induced pancreatic acinar cellular injury and trypsinogen or NF-kappaB activation in rats in vitro / 华中科技大学学报（医学）（英德文版）

The relationship between M3 cholinergic receptor agonist (carbachol) hyperstimulation-induced pancreatic acinar cellular injury and trypsinogen activation or NF-kappaB activation in rats was studied in vitro. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, the active protease inhibitor (pefabloc), and NF-kappaB inhibitor (PDTC) in vitro. Intracellular trypsin activity was measured by using a fluorogenic substrate. The cellular injury was evaluated by measuring the leakage of LDH from pancreatic acinar cells. The results showed that as compared with control group, 10(-3) mol/L carbachol induced a significant increase of the intracellular trypsin activity and the leakage of LDH from pancreatic acinar cells. Pretreatment with 2 mmol/L pefabloc could significantly decrease the activity of trypsin and the leakage of LDH from pancreatic acinar cells (P 0.05). It was concluded that intracellular trypsinogen activation is likely involved in pancreatic acinar cellular injury induced by carbachol hyperstimulation in vitro. NF-kappaB activation may not be involved in pancreatic acinar cellular injury induced by carbachol hyperstimulation in vitro.

Protein kinase C isoform specificity of cholinergic potentiation of glucose-induced pulsatile 5-HT/ insulin release from mouse pancreatic islets

Thymeleatoxin (TMX), an activator of Ca2+-sensitive protein kinase C (cPKC) isoforms, was used to assess the PKC isoform specificity of cholinergic potentiation of glucose (11 mM)-induced pulsatile 5-HT/insulin release (PIR) from single mouse pancreatic islets. TMX (100 nM) and carbachol (Cch, 50 mM) enhanced PIR ~ 3-fold while reducing the underlying [Ca2+]i oscillations (duration and amplitude) by ~ 40-50 percent. Both effects were ablated by the specific PKC inhibitor bisindolylmaleimide and chronic TMX pretreatment. Cch also evoked an initial transient [Ca2+]i rise and surge of 5-HT release, which remained unaffected by chronic TMX pretreatment. It is concluded that the immediate cholinergic responses are insensitive to cPKC. In contrast, specific activation of a cPKC isoform mediates sustained cholinergic potentiation of glucose-induced insulin secretion.

Lack of effect of Helicobacter pylori on symptom improvement with a prokinetic medication, cisapride, in patients with non-ulcer dyspepsia.

OBJECTIVE: This study was undertaken to determine whether H. pylori infection has an effect on the improvement of dyspeptic symptoms in response to a prokinetic agent, cisapride, in patients with non-ulcer dyspepsia (NUD). MATERIAL AND METHOD: 35 NUD patients (16 M, 19 F) who had no underlying medical condition and negative upper endoscopy were included in the present study. Each patient received a 2-wk treatment of cisapride (Prepulsid, 10 mg, tid ac). H. pylori infection was determined using a rapid urease test (CLO test). Gastric emptying (GE) scintigraphy and dyspeptic symptom scores were evaluated before and at the end of the treatment. GE was evaluated in 22 healthy volunteers as normal controls. RESULTS: Half time (T1/2) GE of NUD patients was 90.9 +/- 28 min which was significantly longer than controls (77.6 +/- 14 min; p < 0.05) and was shortened to 73.6 +/- 22 min (p < 0.0001) at the end of the treatment. Cisapride significantly improved total dyspeptic symptom scores [7 (2-18) to 3 (0-11), p < 0.0001]. The symptom score improvement was not affected by H. pylori infection [H. pylori positive: 6 (2-18) to 2.5 (0-9), p < 0.0001; H. pylori negative: 9 (4-16) to 3 (0-11), p < 0.0001] or GE status [delayed GE: 10 (5-16) to 3 (15), p < 0.05; non delayed GE: 6 (2-18) to 2 (0-11); p < 0.0001]. CONCLUSIONS: Cisapride improves dyspeptic symptoms regardless of H. pylori and GE status. These results suggest that gastric emptying and H. pylori infection are not essential to determine prior to prescribing a prokinetic agent, cisapride, in patients with NUD.

Preliminary studies of the mechanism of carbachol increase contraction in rat ventricular myocytes / 中国应用生理学杂志

<p><b>AIM</b>To study whether the mAchR agonist Carbachol(Cch, nonselective) causes increased contractions and L-type Ca2+ current [L(Ca(L))] in ventricular myocytes and the mechanism of these effects.</p><p><b>METHODS</b>The effect of Cch on the I(Ca(L)) and Na/Ca exchange current (I(Na/Ca) was studied in patch-clamped ventricular myocytes isolated from rat hearts and superfused with Tyrode's solution (35 degrees C, 1.8 mmol/L [Ca2+]o) and stimulated at 0.2 Hz and 1.0 Hz evoke contractions of single myocyte.</p><p><b>RESULTS</b>(1) An increase of stimulation frequency decreased the contractions of myocytes(negative inotropic effects). (2) 100 micromol/L Cch increased contraction in 6 cells by 28% (delta L0.2 Hz/ delta L1.0 Hz > or = 1.25) at 1.0 Hz stimulus frequency. (3) The mAchR antagonist Atropine prevented the Cch effect. The mAchR agonist McN-A-343 (M1-selective) did not change the contractions in most of the cells. (4) 100 micromol/L Cch had no significant effect on basal I(Ca(L)), but increased the tail current density on repolarization from +10 mV to -40 mV, suggested that Cch increased I(Na/Ca).</p><p><b>CONCLUSION</b>The increase of cell contractions by Cch is apparently mediated by M2 mAchR and eventually increased I(Na/Ca). The increase Ca2+ content of the SR is reflected by the greater magnitude of I(Na/Ca). These results provide an explanation for the increased contraction of the rat ventricular myocytes by Cch and without changes in I(Ca(L)).</p>

Electroacupuncture ameliorates experimental colitis induced by acetic acid in rat

The effect of electroacupuncture (EA) on experimental colitis was investigated in Sprague-Dawley rats. Colitis was induced by intracolonic instillation of 4% acetic acid. EA (2 Hz, 0.05 ms, 2 V for 20min) was applied to bilateral Hoku (LI-4) and Zusanli (ST-36) on 12 hrs and 36 hrs after induction of colitis. EA-treatment significantly reduced the macroscopic damage and the myeloperoxidase activity of colonic samples at 3 days post-induction of colitis. Colitic colon showed a decreased in vitro motility. However, colonic motility of EAtreated group was not significantly different from that of normal group. The anti-inflammatory effect of EA was not inhibited by a glucocorticoid receptor antagonist, RU-486, but suppressed by a beta-adrenoceptor antagonist, propranonol. These results suggest that EA-treatment has a beneficial effect on colitis, and its anti-inflammatory effect is mediated by beta-adrenoceptor activation but not by endogenous glucocorticoiddependent mechanism.

Five subtypes of muscarinic receptors are expressed in gastric smooth muscles of guinea pig

Muscarinic receptors play key roles in the control of gastrointestinal smooth muscle activity. However, specific physiological functions of each subtype remain to be determined. In this study, the nonselective cation channel activated by carbachol (ICCh) was examined in circular smooth muscle cells of the guinea pig gastric antrum using patch-clamp technique. 4-DAMP inhibited ICCh dose- dependently with IC50 of 1.1 +/- 0.1 nM (n = 6). GTPgS- induced current, however, was not inhibited by 10 nM 4-DAMP. ICCh was not recorded in pertussis- toxin (PTX)-pretreated smooth muscle cells of gastric antrum. ICCh values in response to 10 mM CCh at a holding potential of 60 mV were -330 32 pA (n=4) and -15 +/- 3 pA (n = 6) in the control and PTX-treated cells, respectively (P<0.01). Sensitivities to nanomolar 4-DAMP and PTX suggest the possible involvement of m4 subtype. Using sequence information obtained from cloned guinea pig muscarinic receptor genes, it is possible to amplify the cDNAs encoding m1-m5 from guinea pig brain tissue. Single cell RT-PCR experiments showed that all five subtypes of muscarinic receptor were present in circular smooth muscle cells of the guinea pig gastric antrum. Together with our previous results showing that Go protein is important for activation of ACh-activated NSC channels, our results suggest that ICCh might be activated by acetylcholine through m4 subtype as well as m2 and m3 subtypes in guinea-pig stomach.

Effects of arecoline on calcium channel currents and caffeine-induced calcium release in isolated single ventricular myocyte of guinea pig / 华中科技大学学报（医学）（英德文版）

The effects of Arecoline (Are) on calcium mobilization were investigated. In isolated single ventricular myocyte of guinea pig, patch clamp whole cell recording techniques were used to record the current of L-type calcium channel and cytosolic Ca2+ level ([Ca2+]i) labeled with fluorescence probe Fluo-3/AM was measured under a laser scanning confocal microscope. Results revealed that Are (3-100 mumol/L) could inhibit L-type calcium current in a concentration-dependent manner and the value of IC50 was 33.73 mumol/L (n = 5). In the absence of extracellular calcium, the resting levels of [Ca2+]i was not affected by Are (n = 6, P > 0.05), but pretreatment with Are (30 mumol/L) could significantly inhibit the [Ca2+]i elevation induced by caffeine (10 mmol/L, n = 6, P < 0.01). It was concluded that Are could inhibit not only calcium influx through L-type calcium channel but also calcium release from sarcoplasmic reticulum.

comparative study of the effect of some salivary stimulants on the ultrastructure of parotid acini in albino rats

Parotid gland has a dual nerve supply: sympathetic and parasympathetic. They both activate salivary secretion. Parasympathetic stimulation results in copious watery secretion whereas sympathetic stimulation results in small amount of enzyme-rich secretion. The aim of the present study was to compare between the effect of three different drugs [alpha and beta adrenergic and cholinergic] on the ultrastructure of parotid acini in albino rats. Twenty eight adult male albino rats were used in this study. They were divided into four groups: group I [control group], group II [pilocarpine group], group III [noradrenaline group] and group IV [isoproterenol group]. Group II, III and IV were further subdivided according to the duration of treatment into subgroup A that received one single injection and subgroup B that received a daily injection for seven days. 1. Pilocarpine group showed wide intercellular canaliculi of parotid acini denoting the release of profuse watery secretion. 2. Isoproterenol group revealed degranulation of serous acinar cells denoting the release of enzyme-rich secretion. 3. Noreadrenaline group showed degenerative changes in the cytoplasmic organelles as well as the nuclei of serous acinar cells. Thus, noreadrenaline is not recommended to be used clinically as a salivary stimulant

Carbachol potentiates cholera toxin-induced secretion in a colonic epithelial cell line (HT29-19A) and rat ileal mucosa in vitro.

Recent studies show that enteric nerves are involved in the action of cholera toxin, both in vivo and in vitro. The aim of this study was to investigate in vitro the influence of carbachol, a cholinergic agonist, on the action of cholera toxin. Cultured HT29-19A cell lines and rat ileal mucosa were used in an Ussing chamber for the measurement of short-circuit current induced by cholera toxin. Cyclic AMP was measured from HT29-19A cell lines by standard radio-immunoassay. Pre-treatment of the HT29-19A cell lines with carbachol potentiated cholera toxin-induced secretory response, and enhanced accumulation of cAMP. Carbachol also potentiated the cholera toxin-secretory response in the rat ileal mucosa, but only following pretreatment with the prostaglandin synthesis inhibitor, indomethacin. There was synergistic interaction between cholera toxin and cholinergic neurotransmitter carbachol on the intestinal epithelium. Cholinergic agonists may play a role in regulating the secretory response to the toxin. Such interaction is masked in the intact tissues in vitro due to the release of prostaglandins during isolation.