RESUMEN

BACKGROUND: Intestinal ischemia and reperfusion (I/R) injury leads to abnormalities in motility, namely delay of transit, caused by damage to myenteric neurons. Alterations of the nitrergic transmission may occur in these conditions. This study investigated whether an in vitro I/R injury may affect nitric oxide (NO) production from the myenteric plexus of the guinea pig ileum and which NO synthase (NOS) isoform is involved. METHODS: The distribution of the neuronal (n) and inducible (i) NOS was determined by immunohistochemistry during 60 min of glucose/oxygen deprivation (in vitro ischemia) followed by 60 min of reperfusion. The protein and mRNA levels of nNOS and iNOS were investigated by Western-immunoblotting and real time RT-PCR, respectively. NO levels were quantified as nitrite/nitrate. KEY RESULTS: After in vitro I/R the proportion of nNOS-expressing neurons and protein levels remained unchanged. nNOS mRNA levels increased 60 min after inducing ischemia and in the following 5 min of reperfusion. iNOS-immunoreactive neurons, protein and mRNA levels were up-regulated during the whole I/R period. A significant increase of nitrite/nitrate levels was observed in the first 5 min after inducing I/R and was significantly reduced by N(ω) -propyl-l-arginine and 1400 W, selective inhibitors of nNOS and iNOS, respectively. CONCLUSIONS & INFERENCES: Our data demonstrate that both iNOS and nNOS represent sources for NO overproduction in ileal myenteric plexus during I/R, although iNOS undergoes more consistent changes suggesting a more relevant role for this isoform in the alterations occurring in myenteric neurons following I/R.

Asunto(s)

Íleon/enzimología , Plexo Mientérico/enzimología , Óxido Nítrico Sintasa de Tipo II/metabolismo , Óxido Nítrico Sintasa de Tipo I/metabolismo , Daño por Reperfusión/enzimología , Animales , Western Blotting , Cobayas , Inmunohistoquímica , Masculino , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo I/análisis , Óxido Nítrico Sintasa de Tipo II/análisis , ARN Mensajero/análisis , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa

RESUMEN

BACKGROUND: Ischemic episodes lead to profound functional and structural alterations of the gastrointestinal tract which may contribute to disorders of intestinal motility. Enhancement of glutamate overflow and the consequent activation of NMDA (N-methyl-D-aspartate) receptors may participate to such changes by modulating different enteric neurotransmitter systems, including cholinergic motor pathways. METHODS: The molecular mechanism/s underlying activation of NMDA receptors in the guinea pig ileum were investigated after glucose/oxygen deprivation (in vitro ischemia) and during reperfusion. KEY RESULTS: The number of ileal myenteric neurons positive for NR1, the functional subunit of NMDA receptors, and its mRNA levels were unchanged after in vitro ischemia/reperfusion. In these conditions, the protein levels of NR1, and of its phosphorylated form by protein kinase C (PKC), significantly increased in myenteric neurons, whereas, the levels of NR1 phosphorylated by protein kinase A (PKA) did not change, with respect to control values. Spontaneous glutamate overflow increased during in vitro ischemia/reperfusion. In these conditions, the NMDA receptor antagonists, D(-)-2-amino-5-phosphonopentanoic acid [(D)-AP5] (10 µmol L(-1)) and 5,7-dichlorokynurenic acid (5,7-diClKyn acid) (10 µmol L(-1)) and the PKC antagonist, chelerythrine (1 µmol L(-1)), but not the PKA antagonist, H-89 (1 µmol L(-1)), were able to significantly depress the increased glutamate efflux. CONCLUSIONS & INFERENCES: The present data suggest that in the guinea pig ileum during in vitro ischemia/reperfusion, NR1 protein levels increase. Such event may rely upon posttranscriptional events involving NR1 phosphorylation by PKC. Increased NR1 levels may, at least in part, explain the ability of NMDA receptors to modulate a positive feedback on ischemia/reperfusion-induced glutamate overflow.

Asunto(s)

Íleon/inervación , Plexo Mientérico/fisiopatología , Proteína Quinasa C/fisiología , Receptores de N-Metil-D-Aspartato/fisiología , Daño por Reperfusión/fisiopatología , 2-Amino-5-fosfonovalerato/farmacología , Animales , Benzofenantridinas/farmacología , Cobayas , Íleon/irrigación sanguínea , Técnicas In Vitro , Isoquinolinas/farmacología , Ácido Quinurénico/análogos & derivados , Ácido Quinurénico/farmacología , Masculino , Modelos Animales , Fosforilación , Proteína Quinasa C/antagonistas & inhibidores , Proteína Quinasa C/efectos de los fármacos , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidores , Receptores de N-Metil-D-Aspartato/efectos de los fármacos , Sulfonamidas/farmacología

RESUMEN

In the guinea pig colon, chronic sympathetic denervation entails supersensitivity to inhibitory mu-opioid agents modulating cholinergic neurons. The mechanism underlying such adaptive change has not yet been unravelled, although protein kinase C (PKC) may be involved. A previous study indirectly demonstrated that activation of mu-opioid receptors on myenteric neurons facilitates PKC activity. Such coupling may counteract the inhibitory action of mu-opioid agents on acetylcholine overflow, since PKC, per se, increases this parameter. After chronic sympathetic denervation such restraint abates, representing a possible mechanism for development of supersensitivity to mu-opioid agents. In the present study, this hypothesis was further investigated. After chronic sympathetic denervation, Ca(2+)-dependent PKC activity was reduced in colonic myenteric plexus synaptosomes. The mu-opioid agent, DAMGO, increased Ca(2+)-dependent PKC activity in synaptosomes obtained from normal, but not from denervated animals. In myenteric synaptosomes obtained from this experimental group, protein levels of Ca(2+)-dependent PKC isoforms betaI, betaII and gamma decreased, whereas alpha levels increased. In whole-mount preparations, the four Ca(2+)-dependent PKC isoforms co-localized with mu-opioid receptors on subpopulations of colonic myenteric neurons. The percentage of neurons staining for PKCbetaII, as well as the number of mu-opioid receptor-positive neurons staining for PKCbetaII, decreased in denervated preparations. The same parameters related to PKCalpha, betaI or gamma remained unchanged. Overall, the present data strengthen the concept that mu-opioid receptors located on myenteric neurons are coupled to Ca(2+)-dependent PKCs. After chronic sympathetic denervation, a reduced efficiency of this coupling may predominantly involve PKCbetaII, although also PKCbetaI and gamma, but not PKCalpha, may be implicated.

Asunto(s)

Calcio/metabolismo , Colon/inervación , Proteína Quinasa C/metabolismo , Receptores Opioides mu/fisiología , Sistema Nervioso Simpático/fisiología , Animales , Western Blotting , Desnervación , Encefalina Ala(2)-MeFe(4)-Gli(5)/farmacología , Cobayas , Inmunohistoquímica , Masculino

RESUMEN

BACKGROUND: The effects of thyroid deprivation on the autonomic modulation to the heart remain controversial. METHODS: In this study in patients followed for thyroid carcinoma, we investigated (1) heart rate variability parameters and the baroreflex gain and (2) intracellular catecholamine levels in circulating lymphocytes during short-term hypothyroidism (phase 1) and after reinstitution of TSH-suppressive thyroid hormone replacement (phase 2). RESULTS: The RR interval value (p < 0.01) and systolic blood pressure (p < 0.05) were higher in phase 1 than in phase 2. The low-frequency/high-frequency (LF/HF) ratio was significantly lower in the hypothyroid state (p < 0.05), with a higher HF component (p < 0.05). After adjusting for mean RR interval in the regression model, the difference between the power of RR interval oscillations calculated in the two states was greater for the LF band (p = 0.005) and it was borderline significant for the HF band (p = 0.052). The baroreflex gain alpha(LF) index was similar in the two phases. The stimulus-induced cellular production of norepinephrine and epinephrine in peripheral blood mononuclear cells was significantly higher in phase 2. CONCLUSION: The neurally-mediated influences on the sinus node and the study of intracellular catecholamine production suggest a reduced sympathoexcitation in hypothyroidism compared with the treatment phase. The early increase in blood pressure observed after thyroid hormone withdrawal is not due to impaired sensitivity of the baroreflex arc.

Asunto(s)

Catecolaminas/orina , Frecuencia Cardíaca/fisiología , Hipotiroidismo/tratamiento farmacológico , Hipotiroidismo/fisiopatología , Hormonas Tiroideas/uso terapéutico , Sistema Nervioso Autónomo/efectos de los fármacos , Sistema Nervioso Autónomo/fisiología , Barorreflejo/efectos de los fármacos , Barorreflejo/fisiología , Presión Sanguínea/efectos de los fármacos , Presión Sanguínea/fisiología , Carcinoma Papilar/diagnóstico por imagen , Carcinoma Papilar/radioterapia , Carcinoma Papilar/cirugía , Células Cultivadas , Dopamina/orina , Epinefrina/orina , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Linfocitos/citología , Linfocitos/metabolismo , Norepinefrina/orina , Cintigrafía , Nodo Sinoatrial/efectos de los fármacos , Nodo Sinoatrial/fisiología , Hormonas Tiroideas/sangre , Neoplasias de la Tiroides/diagnóstico por imagen , Neoplasias de la Tiroides/radioterapia , Neoplasias de la Tiroides/cirugía , Tiroidectomía , Imagen de Cuerpo Entero

RESUMEN

Intracellular free calcium concentrations (Ca++i) were studied in polymorphonuclear leukocytes (PMNs) from 13 athyreotic patients who had been previously treated by total thyroidectomy and radioiodine therapy for differentiated thyroid carcinoma, and from age- and sex-matched euthyroid healthy controls. Patients were studied twice, when hypothyroid (visit 1) and after restoration of euthyroidism by L-T4 TSH-suppressive therapy (visit 2). PMNs from patients at visit 1 had significantly lower resting (Ca++)i levels compared to both visit 2 and controls. Values at visit 2 did not differ from those of the controls. Stimulus-induced (Ca++)i rise was also significantly blunted at visit 1 and normalized at visit 2, possibly through a differential contribution of distinct intracellular Ca++ stores, as suggested by the response pattern to the chemotactic agent, N-formyl-Met-Leu-Phe (fMLP), to the selective SERCA pump inhibitor, thapsigargine, and to the mitochondrial uncoupler, carbonyl cyanide p-trifluoromethoxyphenyl-hydrazone (FCCP). In vitro treatment of PMNs from healthy subjects with high TSH concentrations impaired intracellular Ca++ store function. Both resting (Ca++)i levels and fMLP-induced (Ca++)i rise increased in the presence either of low-concentration TSH or of T4, but effects of TSH and T4 were not additive. T3, rT3, and TRIAC had no effect. In conclusion, this study provides evidence for a direct relationship between thyroid status and (Ca++)i homeostasis in human PMNs, mainly related to direct actions of TSH and T4 on these cells.

Asunto(s)

Calcio/metabolismo , Neutrófilos/metabolismo , Hormonas Tiroideas/farmacología , Tirotropina/farmacología , Adulto , Anciano , Antitiroideos/uso terapéutico , ATPasas Transportadoras de Calcio/antagonistas & inhibidores , Carbonil Cianuro p-Trifluorometoxifenil Hidrazona/farmacología , Femenino , Humanos , Técnicas In Vitro , Radioisótopos de Yodo , Masculino , Persona de Mediana Edad , N-Formilmetionina Leucil-Fenilalanina/farmacología , Neutrófilos/efectos de los fármacos , ATPasas Transportadoras de Calcio del Retículo Sarcoplásmico , Tapsigargina/farmacología , Hormonas Tiroideas/sangre , Neoplasias de la Tiroides/radioterapia , Neoplasias de la Tiroides/cirugía , Tiroidectomía , Tirotropina/sangre , Tiroxina/farmacología , Desacopladores/farmacología

RESUMEN

The authors report two cases of catechol-O-methyltransferase (COMT) inhibitor-induced asymptomatic hepatic dysfunction in women with Parkinson disease. The patients were genotyped for the UDP-glucuronosyltransferase (UGT) 1A9 gene (which encodes the main COMT inhibitor-metabolizing enzyme), and found to carry mutations leading to defective glucuronidation activity. This suggests that UGT1A9 poor metabolizer genotype(s) may be a predisposing factor for COMT inhibitor-induced hepatotoxicity.

Asunto(s)

Antiparkinsonianos/efectos adversos , Inhibidores de Catecol O-Metiltransferasa , Enfermedad Hepática Inducida por Sustancias y Drogas , Inhibidores Enzimáticos/efectos adversos , Glucuronosiltransferasa/genética , Hepatopatías/genética , Adulto , Anciano , Benzofenonas/efectos adversos , Catecol O-Metiltransferasa/metabolismo , Catecoles/efectos adversos , Análisis Mutacional de ADN , Inhibidores Enzimáticos/metabolismo , Femenino , Genotipo , Glucuronatos/metabolismo , Humanos , Hígado/efectos de los fármacos , Hígado/enzimología , Hígado/fisiopatología , Hepatopatías/enzimología , Persona de Mediana Edad , Mutación/genética , Nitrilos , Nitrofenoles/efectos adversos , Enfermedad de Parkinson/tratamiento farmacológico , Polimorfismo Genético/genética , Tolcapona , UDP Glucuronosiltransferasa 1A9

RESUMEN

In this study, we investigated whether changes in the regulatory mechanisms of apoptosis and oxidative stress may be detected, peripherally, in patients with Parkinson's disease (PD). For this purpose, we measured caspase-3 activity, Bcl-2 concentrations, peripheral benzodiazepine receptor (PBR) expression and Cu/Zn superoxide dismutase (SOD) concentrations in lymphocytes of untreated PD patients, patients treated only with L-Dopa or with L-Dopa and dopamine agonists and healthy volunteers. Caspase-3 activity was significantly increased in all PD patient groups. Patients treated with L-Dopa and dopamine agonists showed the lowest values of Bcl-2, coupled with the highest density of PBRs, while increased levels of Cu/Zn SOD were found in the group under monotherapy with L-Dopa. We also found, in PD patients, clear, negative correlations between Bcl-2 levels and both duration and severity of the disease. Our findings point to the existence of changes in the regulatory mechanisms of apoptosis in PD patients -- observable outside the central nervous system -- which seem to be modulated by the pharmacological treatment with dopaminergic agents.

Asunto(s)

Antiparkinsonianos/uso terapéutico , Apoptosis/fisiología , Caspasas/metabolismo , Dopaminérgicos/uso terapéutico , Linfocitos/metabolismo , Enfermedad de Parkinson/tratamiento farmacológico , Enfermedad de Parkinson/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/biosíntesis , Anciano , Apoptosis/efectos de los fármacos , Caspasa 3 , Supervivencia Celular/efectos de los fármacos , Progresión de la Enfermedad , Agonistas de Dopamina/farmacología , Femenino , Humanos , Levodopa/farmacología , Linfocitos/efectos de los fármacos , Linfocitos/enzimología , Masculino , Persona de Mediana Edad , Sistema Nervioso Periférico/efectos de los fármacos , Sistema Nervioso Periférico/metabolismo , Receptores de GABA-A/efectos de los fármacos , Receptores de GABA-A/metabolismo , Superóxido Dismutasa/metabolismo

RESUMEN

The actions of adenosine, adenosine 5'-triphosphate (ATP), 2-methylthio adenosine diphosphate ADP (2-MeSADP), 2-methylthio ATP (2-MeSATP), alpha,beta-methylene ATP (alpha,beta-meATP) and uridine triphosphate (UTP) on isolated segments of mouse stomach (fundus), duodenum, ileum and colon were investigated. The localization of P2Y(1), P2Y(2), P2Y(4), P2X(1) and P2X(2) receptors and neuronal nitric oxide synthase (NOS) were examined immunohistochemically, and P2Y(1) mRNA was examined with in situ hybridization. The order of potency for relaxation of longitudinal muscle of all regions was: 2-MeSADP>/=2-MeSATP>alpha,beta-meATP>ATP=UTP=adenosine. This is suggestive of P2Y(1)-mediated relaxation and perhaps a further P2Y receptor subtype sensitive to alpha,beta-meATP. As ATP and UTP are equipotent, the presence of a P2Y(2) receptor is indicated. ATP responses were inhibited by the P2Y(1)-selective antagonist MRS 2179, and suramin. P2Y(1) receptors were visualized immunohistochemically in the smooth muscle of the ileum and in a subpopulation for myenteric neurones, which also stained for NOS. P2Y(1) mRNA was localized in neurones in both myenteric and submucosal ganglia in the ileum. Taken together, these results suggest that ATP was acting on non-adrenergic, non-cholinergic inhibitory neurons, which release both nitric oxide (NO) and ATP. Reduced relaxations to 2-MeSADP by tetrodotoxin and N(omega)-nitro-L-arginine methyl ester, are consistent with this possibility. Adenosine acts via P1 receptors to relax smooth muscle of the mouse gut. Segments of mouse colon (in contrast to the stomach and small intestine) were contracted by nucleotides with the potency order: 2-MeSATP>alpha,betameATP>ATP; the contractions showed no desensitization and were antagonized by suramin and PPADS, consistent with responses mediated by P2X(2) receptors. Immunoreactivity to P2X(2) receptors was demonstrated on both longitudinal and circular muscle of the colon, but not in the other regions of the gut, except for a small subpopulation of myenteric neurones. In summary, neuronal P2Y(1) receptors appear to mediate relaxation, largely through NO in all regions of the mouse gut, and to a lesser extent by P2Y(1), P2Y(2) and a novel P2Y receptor subtype responsive to alpha,beta-meATP in smooth muscle, while P2X(2) receptors mediate contraction of colonic smooth muscle.

Asunto(s)

Fenómenos Fisiológicos del Sistema Digestivo , Sistema Digestivo/química , Receptores Purinérgicos P2/análisis , Receptores Purinérgicos P2/fisiología , Animales , Sistema Digestivo/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Técnicas In Vitro , Masculino , Ratones , Ratones Endogámicos C57BL , Relajación Muscular/efectos de los fármacos , Relajación Muscular/fisiología , Neuronas/química , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Óxido Nítrico Sintasa/análisis , Óxido Nítrico Sintasa de Tipo I , Agonistas del Receptor Purinérgico P2 , Antagonistas del Receptor Purinérgico P2 , Receptores Purinérgicos P2Y1

RESUMEN

BACKGROUND AND AIM: From February 1998 to September 1999, the Italian Ministry of Health limited angiotensin-II type-1 (AT1) receptor antagonist reimbursement only to patients who necessitated discontinuation of angiotensin converting enzyme (ACE) inhibitor treatment due to cough or angioedema. We assessed the consequences of this decision on the reporting of ACE inhibitor-associated adverse drug reactions (ADRs). METHODS: ACE inhibitor-associated ADRs reported to the national pharmacovigilance system in 1997-2000 in the district of Varese (northern Italy, more than 820,000 inhabitants) were retrieved and analysed. The dispensation of ACE inhibitors and AT, receptor antagonists reimbursed by the National Health System was also examined. RESULTS: There were 228 reports of ACE inhibitor-associated ADRs, and cough was the ADR reported in 93.4% of cases. There were no reports of cough in 1997, 50 in 1998, 156 in 1999 and 7 in 2000. In 1998-1999, the dispensation of ACE inhibitors showed little variation, while that of AT1 receptor antagonists grew about twofold. CONCLUSIONS: There was a clear correlation between ACE inhibitor-associated ADR reporting and limitation to AT1 receptor antagonist reimbursement status. Drug reimbursement policies should thus be added to the list of factors that may bias ADR reporting, and health authorities should be aware of this potential bias when defining specific measures to regulate prescription and use of new drugs.

Asunto(s)

Sistemas de Registro de Reacción Adversa a Medicamentos/estadística & datos numéricos , Antagonistas de Receptores de Angiotensina , Inhibidores de la Enzima Convertidora de Angiotensina/efectos adversos , Prescripciones de Medicamentos/estadística & datos numéricos , Mecanismo de Reembolso/tendencias , Adulto , Sistemas de Registro de Reacción Adversa a Medicamentos/tendencias , Anciano , Anciano de 80 o más Años , Tos/tratamiento farmacológico , Femenino , Humanos , Masculino , Persona de Mediana Edad , Mecanismo de Reembolso/estadística & datos numéricos

RESUMEN

In isolated human neutrophils, diazepam (10 nmol/l to 10 micromol/l) concentration-dependently increased migration and phagocytosis. Diazepam-induced migration and phagocytosis were inhibited by the peripheral benzodiazepine receptor (PBR) antagonist PK11195 (10 micromol/l). The PBR agonist Ro5-4864 (10 nmol/l to 10 micromol/l) did not affect migration but slightly enhanced phagocytosis, while clonazepam, which binds to the central-type benzodiazepine receptors but has no affinity for PBRs, was ineffective on both parameters up to 10 micromol/l. Phagocytosis induced by diazepam or Ro5-4864 was inhibited by the Ca2+ channel blocker L-verapamil (10 micromol/l), which however did not affect the action of diazepam on migration. Competition binding experiments performed by fluorescent staining of PBRs showed that diazepam directly interacts with PBRs on human neutrophils. Both diazepam and Ro5-4864 (10 nmol/l to 10 micromol/l) induced a rise of intracellular free Ca2+ concentrations ([Ca2+]i), which was inhibited by PK11195 (10 micromol/l) and L-verapamil (10 micromol/l) and prevented by extracellular Ca2+ chelation with EGTA (5 mmol/l). In conclusion, experimental evidence indicates that in human neutrophils diazepam stimulates both migration and phagocytosis through activation of PBRs. Diazepam-induced [Ca2+]i changes depend on a PBR-operated, L-verapamil-sensitive increase in the plasma membrane permeability and subsequent extracellular Ca2+ entry, and contribute to diazepam-induced phagocytosis. On the contrary, the effect of diazepam on migration seems to occur through Ca2+ -independent mechanisms.

Asunto(s)

Calcio/metabolismo , Quimiotaxis de Leucocito/efectos de los fármacos , Diazepam/farmacología , Neutrófilos/efectos de los fármacos , Fagocitosis/efectos de los fármacos , Receptores de GABA-A/metabolismo , Bloqueadores de los Canales de Calcio/farmacología , Células Cultivadas , Citometría de Flujo , Agonistas de Receptores de GABA-A , Antagonistas de Receptores de GABA-A , Humanos , Técnicas In Vitro , Isoquinolinas/farmacología , Neutrófilos/fisiología , Verapamilo/farmacología

RESUMEN

OBJECTIVE: To test the hypothesis that comparison of defined daily dose (DDD) and drug user data may help to estimate drug exposure in a defined population and provide information about drug prescribing patterns. METHODS AND RESULTS: First, comparison of DDD figures with the number of apparent drug users (ADU, i.e., individuals for whom at least one prescription of the drug had been dispensed during a given time period) is demonstrated to correspond to the product of the prescribed daily dose (PDD) and the proportion of days in which the drug had been taken (days of treatment/days in a time period, D). The resulting equation (DDD/day)/ ADU in a time period = PDD x D is then applied to the analysis of different sets of drug dispensation data. Examples show that this approach may be helpful to monitor drug prescribing patterns over time. Moreover, in definite situations, it may provide reliable estimates of either PDD or D. CONCLUSIONS: Comparison of DDD and drug user data is suggested to be a cost-effective strategy to monitor drug prescribing patterns from an epidemiological perspective, which may be useful to researchers involved in drug utilisation studies as well as to health authorities for monitoring and regulatory purposes.

Asunto(s)

Prescripciones de Medicamentos/estadística & datos numéricos , Utilización de Medicamentos/estadística & datos numéricos , Preparaciones Farmacéuticas/administración & dosificación , Algoritmos , Difosfonatos , Métodos Epidemiológicos , Italia/epidemiología , Inhibidores de Agregación Plaquetaria , Población , Ticlopidina

RESUMEN

To assess the role of AMPA and kainate receptors in modulating neurotransmitter release from the myenteric plexus, the effect of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) and kainic acid on endogenous acetylcholine (ACh) and noradrenaline (NA) overflow from the guinea-pig isolated colon was studied. AMPA inhibited spontaneous ACh overflow and increased electrically-evoked NA overflow. Kainic acid did not influence both ACh and NA overflow. AMPA-mediated effects on ACh and NA overflow were significantly reduced by the AMPA/kainate antagonist, 6-cyano-7-nitroquinoxaline-2,3-dione, CNQX. The inhibitory effect of AMPA on ACh overflow could be due, at least in part, to the AMPA-induced NA overflow as it was greatly reduced after adrenoceptor blockade and virtually abolished in sympathetically-denervated animals. The possible functional significance of these findings was studied by measuring the efficiency of the peristaltic reflex in the presence of the different agonists. The efficiency of peristalsis was enhanced by AMPA, whereas it was not modified by kainic acid. In conclusion, AMPA receptors, but not kainate receptors, may play a role in the modulation of ACh and NA release and of peristalsis in the guinea-pig colon.

Asunto(s)

Acetilcolina/metabolismo , Colon/fisiología , Norepinefrina/metabolismo , Receptores AMPA/fisiología , Antagonistas Adrenérgicos alfa/farmacología , Antagonistas Adrenérgicos beta/farmacología , Animales , Colon/efectos de los fármacos , Colon/inervación , Relación Dosis-Respuesta a Droga , Estimulación Eléctrica , Agonistas de Aminoácidos Excitadores/farmacología , Cobayas , Técnicas In Vitro , Ácido Kaínico/farmacología , Músculo Liso/efectos de los fármacos , Músculo Liso/inervación , Músculo Liso/fisiología , Plexo Mientérico/efectos de los fármacos , Plexo Mientérico/metabolismo , Peristaltismo/efectos de los fármacos , Peristaltismo/fisiología , Propranolol/farmacología , Receptores AMPA/agonistas , Simpatectomía , Yohimbina/farmacología , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiónico/farmacología

RESUMEN

We studied the effects of two diazepam-binding inhibitor (DBI)-derived peptides, triakontatetraneuropeptide (DBI 17-50, TTN) and eiksoneuropeptide (DBI 51-70, ENP), on cytosolic free Ca2+ concentrations ([Ca2+]i), chemotaxis, superoxide anion (O2-) generation, and phagocytosis in human neutrophils. Both TTN and ENP induced a rapid and transient rise of [Ca2+]i. The effect of TTN depended on the presence of extracellular Ca2+, whereas the effect of ENP also persisted after extracellular Ca2+ chelation. TTN induced neutrophil chemotaxis, stimulated O2- generation, and enhanced phagocytosis. ENP did not affect cell migration and oxidative metabolism but enhanced phagocytosis. Both peptides modulated N-formyl-methionyl-leucyl-phenylalanine- and phorbol myristate acetate-induced O2- generation. Because neutrophils express benzodiazepine receptors of the peripheral type (pBRs) and DBI-derived peptides may interact with such receptors, we investigated the possible role of pBRs in TTN- or ENP-induced effects. The synthetic pBR ligand RO 5-4864 increased [Ca2+]i through extracellular Ca2+ influx and this effect was prevented by the pBR antagonist PK-11195. RO 5-4864, however, was ineffective on neutrophil migration and O2- generation and only slightly affected phagocytosis. Moreover, PK-11195 delayed the [Ca2+]i rise induced by TTN but did not significantly affect its extent, and had no effect on the [Ca2+]i rise induced by ENP. We conclude that DBI-derived peptides induce [Ca2+]i changes and modulate neutrophil function mainly through pBR-independent pathways. In view of the wide cell and tissue distribution of DBI in the brain and in peripheral organs, modulation of neutrophil function by DBI-derived peptides may be relevant for both the neuroimmune network and the development and regulation of the inflammatory processes.

Asunto(s)

Calcio/sangre , Quimiotaxis de Leucocito/fisiología , Neuropéptidos/farmacología , Neutrófilos/fisiología , Fragmentos de Péptidos/farmacología , Fagocitosis/efectos de los fármacos , Superóxidos/sangre , Antineoplásicos/farmacología , Benzodiazepinonas/farmacología , Citosol/metabolismo , Humanos , Hipolipemiantes/farmacología , Isoquinolinas/farmacología , Cinética , N-Formilmetionina Leucil-Fenilalanina/farmacología , Neutrófilos/efectos de los fármacos , Acetato de Tetradecanoilforbol/farmacología

RESUMEN

A rapid and simple HPLC-ED method is described to identify and measure catecholamines (CTs) and their major metabolites in immune cells. Using this method, intracellular CTs were quantified in human peripheral blood mononuclear cells (PBMCs), T and B lymphocytes, monocytes and granulocytes. Immune cell subsets were separated by density gradient centrifugation and immunomagnetic cell sorting. CTs were also found in the human hematopoietic cell lines NALM-6 (pre-B) and (in smaller amounts) in Jurkat (T lymphoblastoid) and U937 (promonocytic). In cultured PBMCs, intracellular CTs were reduced by both the tyrosine hydroxylase inhibitor alpha-methyl-p-tyrosine and the chromaffin granule depletant reserpine. In NALM-6 cells, both alpha-methyl-p-tyrosine and the dopamine-beta-hydroxylase inhibitor disulfiram reduced intracellular CTs, supporting the presence of active synthetic pathways in these cells. Since sympathoadrenergic mechanisms play a key role in the interactions between the immune system and the nervous system, these findings may be relevant for a better understanding of the neuro-immune network.

Asunto(s)

Catecolaminas/análisis , Cromatografía Líquida de Alta Presión/métodos , Electroquímica/métodos , Células Madre Hematopoyéticas/química , Leucocitos Mononucleares/química , Linfocitos B/química , Separación Celular , Disulfiram/farmacología , Granulocitos/química , Humanos , Células Jurkat , Monocitos/química , Reserpina/farmacología , Linfocitos T/química , Factores de Tiempo , Células U937 , alfa-Metiltirosina/farmacología

RESUMEN

Enteric ganglia can maintain integrated functions, such as the peristaltic reflex, in the absence of input from the central nervous system, which has a modulatory role. Several clinical and experimental observations suggest that homeostatic control of gut function in a changing environment may be achieved through adaptive changes occurring in the enteric ganglia. A distinctive feature of enteric ganglia, which may be crucial during the development of adaptive responses, is the vicinity of the final effector cells, which are an important source of mediators regulating cell growth. The aim of this review is to focus on the possible mechanisms underlying neuronal plasticity in the enteric nervous system and to consider approaches to the study of plasticity in this model. These include investigations of neuronal connectivity during development, adaptive mechanisms that maintain function after suppression of a specific neural input, and the possible occurrence of activity-dependent modifications of synaptic efficacy, which are thought to be important in storage of information in the brain. One of the applied aspects of the study of plasticity in the enteric nervous system is that knowledge of the underlying mechanisms may eventually enable us to develop strategies to correct neuronal alterations described in several diseases.

Asunto(s)

Sistema Nervioso Entérico/fisiología , Plasticidad Neuronal/fisiología , Adaptación Biológica , Envejecimiento/fisiología , Fenómenos Fisiológicos del Sistema Digestivo , Sistema Nervioso Entérico/crecimiento & desarrollo , Ambiente , Humanos

RESUMEN

In the guinea-pig colon, acetylcholine (ACh) release from intrinsic cholinergic motor neurons is inhibited by adrenoceptors, opioid and muscarinic receptors. Chronic sympathetic denervation resulted in supersensitivity to the inhibitory effect of DAMGO (mu-opioid agonist) on ACh release and on the peristaltic reflex. After chronic treatment with naltrexone (NTX) supersensitivity to DAMGO and subsensitivity to UK14,304 (alpha2-adrenoceptor agonist) developed for both functional parameters. The facilitatory effect of scopolamine on ACh release remained unchanged after chronic NTX treatment, whereas it was potentiated after chronic sympathetic denervation. These data suggest the existence of a functional interaction between different inhibitory pathways modulating cholinergic motor neurons in the guinea-pig colon. Namely, chronic manipulation of an inhibitory pathway may entail adaptive sensitivity changes in another inhibitory pathway so that homeostasis can be maintained.

Asunto(s)

Colon/inervación , Acetilcolina/metabolismo , Agonistas alfa-Adrenérgicos/farmacología , Animales , Tartrato de Brimonidina , Fibras Colinérgicas/fisiología , Desnervación , Cobayas , Neuronas Motoras/fisiología , Naltrexona/farmacología , Norepinefrina/metabolismo , Peristaltismo/fisiología , Quinoxalinas/farmacología , Receptores Adrenérgicos alfa 2/fisiología , Receptores Opioides/fisiología , Sistema Nervioso Simpático/fisiología

RESUMEN

We report that neutrophil function was impaired in former heroin addicts on chronic naltrexone maintenance. Of the subjects, 62.5% had elevated plasma ACTH, 25% had elevated plasma cortisol and one subject had increased urinary cortisol. All subjects showed enhanced expression of opioid receptors on monocytes, neutrophils and lymphocytes. In vitro, incubation with therapeutically relevant concentrations of naltrexone induced a slow increase of neutrophil cytoplasmatic free Ca(2+)concentrations ([Ca(2+)]()E2>i) and slowed down the [Ca(2+)]()E2>i rise induced by N-formyl-methionyl-leucyl-phenylalanine. Neither naltrexone nor its metabolite beta-naltrexol affected human neutrophil function in vitro. We conclude that impairment of neutrophil function during chronic naltrexone may be related to opioid receptor overexpression. With this regard, the possible role of naltrexone-induced [Ca(2+)]()E2>i changes deserves further investigation. 1999 Academic Press.

Asunto(s)

Leucocitos/efectos de los fármacos , Naltrexona/uso terapéutico , Antagonistas de Narcóticos/uso terapéutico , Neutrófilos/efectos de los fármacos , Receptores Opioides/efectos de los fármacos , Adolescente , Hormona Adrenocorticotrópica/sangre , Hormona Adrenocorticotrópica/efectos de los fármacos , Adulto , Calcio/metabolismo , Femenino , Dependencia de Heroína/tratamiento farmacológico , Humanos , Hidrocortisona/sangre , Hidrocortisona/orina , Leucocitos/citología , Leucocitos/metabolismo , Masculino , Naltrexona/análogos & derivados , Naltrexona/farmacología , Neutrófilos/metabolismo , Neutrófilos/fisiología , Receptores Opioides/biosíntesis

RESUMEN

Evidence is presented that human neutrophils contain catecholamines and several of their metabolites. In vitro, incubation with alpha-methyl-p-tyrosine or pargyline affects intracellular dopamine, norepinephrine and their metabolites, suggesting catecholamine synthesis and degradation by these cells. Reserpine reduces intracellular dopamine and norepinephrine and desipramine reduces intracellular norepinephrine, suggesting the presence of storage and uptake mechanism. In view of the ability of catecholamines to affect neutrophil function, the present results support the hypothesis that autoregulatory adrenergic mechanisms may exist in these cells.

Asunto(s)

Catecolaminas/metabolismo , Neutrófilos/metabolismo , Adulto , Femenino , Humanos , Masculino , alfa-Metiltirosina/farmacología

RESUMEN

Evidence has been obtained that peripheral blood mononuclear cells contain dopamine, norepinephrine, epinephrine, and their metabolites. Pharmacologic inhibition of tyrosine hydroxylase or monoamine oxidase profoundly affected intracellular catecholamines (CTs) and their metabolites, indicating that these cells are able to synthesize and breakdown CTs. The sensitivity of intracellular CTs to reserpine and the presence of CTs in the extracellular medium suggest that CTs are stored and released. Moreover, the increase of extracellular CTs in the presence of monoamine uptake blockers point to the presence of functional uptake mechanisms. Altogether, these results indicate the existence of a CT lifecycle in human mononuclear cells and warrant further studies to investigate the role of adrenergic autoregulatory mechanisms in modulation of the immune response and in the pathogenesis of diseases involving the immune system.

Asunto(s)

Catecolaminas/metabolismo , Leucocitos Mononucleares/metabolismo , Neuroinmunomodulación/fisiología , Adulto , Calcio/sangre , Inhibidores Enzimáticos/farmacología , Humanos , Líquido Intracelular/metabolismo , Monoaminooxidasa/sangre , Inhibidores de la Monoaminooxidasa/farmacología , Pargilina/farmacología , Tirosina 3-Monooxigenasa/antagonistas & inhibidores , Tirosina 3-Monooxigenasa/sangre , alfa-Metiltirosina/farmacología

RESUMEN

Doctors' attitudes to adverse drug reactions (ADRs) and perception of drug-related risk for ADR occurrence were investigated in four hospitals in Northern Italy using an interviewer-administered questionnaire. ADRs were a relevant concern in medical practice for 80% of the respondents and had been observed by 87%. ADRs were perceived to occur in no more than 5% of hospitalized patients and serious ADRs in less than 1%. The response patterns, however, differed according to the ward of work and the year of graduation of the doctors. Antibacterials, NSAIDs and antiarrhythmics were rated as higher risk drugs, while diuretics, lipid lowering agents, antihistamines, antiemetics and antispasmodics were rated as lower risk drugs. Risk perception was dishomogeneous mainly with respect to the ward of work. The estimated frequency of ADR occurrence, the perception of drug-related risk and previous ADR reporting behaviour were clearly correlated. The present results suggest that personal factors affect doctors' attitudes to ADRs, perception of drug-related risk and ADR reporting behaviour and may thus be relevant in developing and targeting educational strategies aimed at increasing awareness of ADRs and at encouraging ADR reporting.