RESUMEN
When frog sartorius muscles were exposed to diisopropylfluorophosphate, the amplitude and half-decay time of the end-plate current decreased; the half-decay time became almost potential-independent and the equilibrium potential for the end-plate current was more negative than during control conditions. When the excess reagent was removed by washing so that only the phosphorylated acetylcholinesterase remained, the amplitude of the end-plate current was restored, while its half-decay time was markedly increased. These findings reveal that this organophosphate significantly affects the receptor-ionic conductance modulator complex in addition to its well-known anticholinesterase activity.
Asunto(s)
Isoflurofato/farmacología , Unión Neuromuscular/efectos de los fármacos , Receptores de Droga/efectos de los fármacos , Animales , Anuros , Transporte Biológico/efectos de los fármacos , Estimulación Eléctrica , Potenciales de la Membrana/efectos de los fármacos , Contracción Muscular , Unión Neuromuscular/metabolismo , Potasio/metabolismo , Receptores Colinérgicos/efectos de los fármacos , Sodio/metabolismoRESUMEN
Although Ca(2+)-induced Ca2+ release (CICR) via ryanodine receptors has been found to occur in intact neurons, little is known about the physiological processes that regulate it. We studied the effects of cyclic ADP-ribose (cADPR) on CICR in cultured bullfrog sympathetic neurons by fura-2 fluorescence recording and patch-clamp techniques. cADPR applied through a patch pipette augmented action potential- or depolarizing pulse-induced rises in intracellular Ca2+ without a change in Ca2+ entry initiating the responses, but not in the presence of ryanodine. Likewise, cADPR enhanced a single or oscillatory rise(s) in intracellular Ca2+ induced by caffeine. These results strongly suggest that cADPR can be an endogenous modulator of ryanodine receptors in neurons.
Asunto(s)
Adenosina Difosfato Ribosa/análogos & derivados , Cafeína/farmacología , Canales de Calcio/fisiología , Calcio/metabolismo , Ganglios Simpáticos/fisiología , Neuronas/fisiología , Adenosina Difosfato Ribosa/farmacología , Animales , Canales de Calcio/efectos de los fármacos , Células Cultivadas , ADP-Ribosa Cíclica , Activación del Canal Iónico/efectos de los fármacos , Activación del Canal Iónico/fisiología , Cinética , Potenciales de la Membrana/efectos de los fármacos , Neuronas/efectos de los fármacos , Rana catesbeiana , Factores de TiempoRESUMEN
Ca2+-induced Ca2+ release (CICR) occurs via activation of ryanodine receptors (RyRs) in frog motor nerve terminals after RyRs are primed for activation by repetitive Ca2+ entries, thereby contributing to synaptic plasticity. To clarify how the mechanism of CICR becomes activable by repetitive Ca2+ entries, we studied effects of a RyR modulator, cyclic ADP-ribose (cADPr), on CICR by Ca2+ imaging techniques. Use-dependent binding of fluorescent ryanodine and its blockade by ryanodine revealed the existence of RyRs in the terminals. Repetition of tetani applied to the nerve produced repetitive rises in intracellular Ca2+ ([Ca2+]i) in the terminals. The amplitude of each rise slowly waxed and waned during the course of the stimulation. These slow rises and decays were blocked by ryanodine, indicating the priming, activation and inactivation of CICR. Uncaging of caged-cADPr loaded in the terminals increased the amplitude of short tetanus-induced rises in [Ca2+]i and the amplitude, time to peak and half decay time of the slow waxing and waning rises in [Ca2+]i evoked by repetitive tetani. A cADPr blocker, 8-amino-cADPr, loaded in the terminals decreased the slow waxing and waning component of rises and blocked all the actions of exogenous cADPr. It is concluded that cADPr enhances the priming and activation of CICR. The four-state model for RyRs suggests that cADPr inhibits the inactivation of CICR and increases the activation efficacy of RyR.
Asunto(s)
Calcio/metabolismo , ADP-Ribosa Cíclica/análogos & derivados , Neuronas Motoras/citología , Terminales Presinápticos/efectos de los fármacos , Terminales Presinápticos/metabolismo , Animales , ADP-Ribosa Cíclica/farmacología , Técnicas In Vitro , Neuronas Motoras/efectos de los fármacos , Unión Neuromuscular/citología , Ranidae , Rianodina/metabolismo , Factores de TiempoRESUMEN
In this prospective multicenter study, we investigated the course of depression and anxiety during hematopoietic stem cell transplantation (HSCT) until 5 years after transplantation adjusting for medical information. Patients were consulted before HSCT (n=239), at 3 months (n=150), 12 months (n=102) and 5 years (n=45) after HSCT. Depression and anxiety were assessed with the Hospital Anxiety and Depression Scale (HADS). Detailed medical and demographic information was collected. Prevalence rates were compared with an age- and gender-matched control group drawn from a large representative sample (n=4110). The risk of depression before HSCT was lower for patients than for the control group (risk ratio (RR), 0.56; 95% confidence interval (CI), 0.39/0.81). Prevalence rates of depression increased from 12 to 30% until 5 years post HSCT. Anxiety rates were most frequently increased before HSCT (29%, RR, 1.31; 95% CI, 1.02/1.68) and then reached a stable level comparable to the background population (RR 0.83, 95% CI, 0.56/1.22). This study confirms the low levels of depression in the short term after HSCT and identifies depression as a long-term effect. Furthermore, it confirms previous results of heightened anxiety before HSCT. Surveillance of symptoms of anxiety during the short-term phase of HSCT and of depression during the following years is crucial.
Asunto(s)
Ansiedad/etiología , Depresión/etiología , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Adulto , Estudios de Casos y Controles , Femenino , Alemania/epidemiología , Trasplante de Células Madre Hematopoyéticas/psicología , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Factores de TiempoRESUMEN
Although allogeneic hematopoietic stem cell transplantation (HSCT) features severe physical and psychological strain, no previous study has prospectively investigated fatigue beyond 3 years after transplantation. We investigated the temporal course of fatigue over 5 years, compared patients with the general population (GP) and tested for treatment- and complication-related risk factors. Patients were assessed before conditioning (T0, N=239) and at 100-day (T1, N=150), 1-year (T2, N=102) and 5-year (T3, N=45) follow-up. We measured fatigue with the Multidimensional Fatigue Inventory-20. Patients were compared with the GP at T0 and at T3. Global fatigue increased from T0 to T1 (t=3.85, P<0.001), decreased from T1 to T2 (t=-2. 92, P=0.004) and then remained stable (t=0.45, P=0.656). No difference in global fatigue was found between T0 and T3 (t=0.68, P=0.497). Compared with the GP, patients showed higher global fatigue at T0 (t=-6.02, P<0.001) and T3 (t=-2.50, P=0.014). These differences reached meaningful effect sizes (d⩾0.5). Acute and chronic GvHD predicted global fatigue at T1 (γ=0.34, P=0.006) and T2 (γ=0.38, P=0.010), respectively. To conclude, fatigue among allogeneic HSCT patients improves with time, finally returning to pretransplantation levels. However, even after 5 years, the difference from the GP remains relevant. Patients with GvHD are at risk for increased fatigue.
Asunto(s)
Fatiga/etiología , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Adulto , Fatiga/diagnóstico , Femenino , Estudios de Seguimiento , Enfermedad Injerto contra Huésped/patología , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Factores de Riesgo , Factores de Tiempo , Acondicionamiento Pretrasplante/efectos adversos , Acondicionamiento Pretrasplante/métodos , Trasplante Homólogo/efectos adversosRESUMEN
We reported that NK4, composed of the N-terminal hairpin and subsequent four kringle domains of hepatocyte growth factor (HGF), acts as the competitive antagonist for HGF. We now provide the first evidence that NK4 inhibits tumor growth and metastasis as an angiogenesis inhibitor as well as an HGF antagonist. Administration of NK4 suppressed primary tumor growth and lung metastasis of Lewis lung carcinoma and Jyg-MC(A) mammary carcinoma s.c. implanted into mice, although neither HGF nor NK4 affected proliferation and survival of these tumor cells in vitro. NK4 treatment resulted in a remarkable decrease in microvessel density and an increase of apoptotic tumor cells in primary tumors, which suggests that the inhibition of primary tumor growth by NK4 may be achieved by suppression of tumor angiogenesis. In vivo, NK4 inhibited angiogenesis in chick chorioallantoic membranes and in rabbit corneal neovascularization induced by basic fibroblast growth factor (bFGF). In vitro, NK4 inhibited growth and migration of human microvascular endothelial cells induced by bFGF and vascular endothelial growth factor (VEGF) as well as by HGF. HGF and VEGF activated the Met/HGF receptor and the KDR/VEGF receptor, respectively, whereas NK4 inhibited HGF-induced Met tyrosine phosphorylation but not VEGF-induced KDR phosphorylation. NK4 inhibited HGF-induced ERK1/2 (p44/42 mitogen-activated protein kinase) activation, but allowed for bFGF- and VEGF-induced ERK1/2 activation. These results indicate that NK4 is an angiogenesis inhibitor as well as an HGF antagonist, and that the antiangiogenic action of NK4 is independent of its activity as HGF antagonist. The bifunctional properties of NK4 to act as an angiogenesis inhibitor and as an HGF antagonist raises the possibility that NK4 may prove therapeutic for cancer patients.
Asunto(s)
Inhibidores de la Angiogénesis/farmacología , Inhibidores de Crecimiento/farmacología , Factor de Crecimiento de Hepatocito/antagonistas & inhibidores , Factor de Crecimiento de Hepatocito/farmacología , Neovascularización Patológica/tratamiento farmacológico , Alantoides/irrigación sanguínea , Alantoides/efectos de los fármacos , Animales , Anticuerpos/farmacología , Células CHO , Carcinoma Pulmonar de Lewis/irrigación sanguínea , Carcinoma Pulmonar de Lewis/tratamiento farmacológico , Carcinoma Pulmonar de Lewis/secundario , Movimiento Celular/efectos de los fármacos , Embrión de Pollo , Corion/irrigación sanguínea , Corion/efectos de los fármacos , Cricetinae , Endotelio Vascular/citología , Endotelio Vascular/efectos de los fármacos , Activación Enzimática , Humanos , Neoplasias Pulmonares/irrigación sanguínea , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/secundario , Masculino , Neoplasias Mamarias Experimentales/irrigación sanguínea , Neoplasias Mamarias Experimentales/tratamiento farmacológico , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Fosforilación/efectos de los fármacos , Proteínas Proto-Oncogénicas c-met/metabolismo , Conejos , Proteínas Recombinantes/farmacología , Tirosina/metabolismoRESUMEN
Hepatocyte growth factor (HGF) is involved in malignant behavior of cancers as a mediator in tumor-stromal interactions through enhancing tumor invasion and metastasis. We found recently that NK4, a four-kringle fragment of HGF, functions as both an HGF-antagonist and an angiogenesis inhibitor. We have now determined whether blockade of the HGF-c-Met/HGF receptor pathway and tumor angiogenesis by administration of recombinant NK4 would inhibit growth, invasion, and metastasis of human pancreatic carcinoma implanted into the pancreas of nude mice. When treatment with NK4 or anti-HGF neutralizing antibody was initiated from the third day after orthotopic injection of SUIT-2 human pancreatic cancer cells, both NK4 and anti-HGF antibody suppressed the conversion of orthotopic pancreatic tumors from carcinoma in situ to aberrantly invading cancers during days 3-14. On the other hand, when the treatment was begun on day 10, a time when cancer cells were already invading surrounding tissues, NK4 but not anti-HGF antibody inhibited tumor growth, peritoneal dissemination, and ascites accumulation at 4 weeks after the inoculation. Antitumor effects of NK4 correlated with decreased microvessel density in pancreatic tumors thereby indicating that the antiangiogenic activity of NK4 may have mainly contributed to its antitumor effects. Moreover, although NK4-treatment was initiated from the end stage (day 24 after tumor inoculation), NK4 prolonged survival time of mice, and the suppression of peritoneal dissemination, ascites accumulation, and invasion of metastasized cancer cells into the peritoneal wall were remarkable. We propose that simultaneous targeting of both tumor angiogenesis and the HGF-mediated invasion-metastasis may prove to be a new approach to treating patients with pancreatic cancer.
Asunto(s)
Antineoplásicos/farmacología , Factor de Crecimiento de Hepatocito/farmacología , Mitógenos , Neoplasias Pancreáticas/tratamiento farmacológico , Animales , División Celular/efectos de los fármacos , Progresión de la Enfermedad , Inhibidores de Crecimiento/farmacología , Factor de Crecimiento de Hepatocito/metabolismo , Humanos , Masculino , Ratones , Invasividad Neoplásica , Neovascularización Patológica/tratamiento farmacológico , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Neoplasias Peritoneales/prevención & control , Neoplasias Peritoneales/secundario , Proteínas Proto-Oncogénicas c-met/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Invasion of various carcinoma cells follows their interaction with stromal cells. Hepatocyte growth factor (HGF), four-kringle-containing growth factor, is a mesenchymal or stromal-derived mediator which affects the growth and the invasiveness of carcinoma cells. We now have evidence that a four-kringle-containing antagonist for HGF, HGF/NK4 inhibits invasion of tumors in vivo, as well as in vitro. HGF/NK4 competitively inhibited the binding of HGF to Met/ HGF receptors on GB-d1 human gallbladder carcinoma cells. HGF induced invasion of the cells through Matrigel basement membrane components and into collagen gels, but HGF-induced invasion was inhibited by HGF/NK4. Invasion of GB-d1 cells was induced by co-cultivation with stromal fibroblasts, which mimics tumor-stromal interaction, but it was almost completely suppressed by HGF/NK4. Likewise, invasive growth induced by HGF in collagen gels in GB-dl cells, HuCC-T1 human cholangiocarcinoma cells, and ME-180 human uterus cervical carcinoma cells was also strongly inhibited by HGF/NK4. When GB-d1 cells were implanted subcutaneously into nude mouse, tumor cells invaded muscular tissue, but the infusion of HGF/NK4 inhibited this invasion. Furthermore, HGF/NK4 increased apoptotic cell death of GB-d1 cells and inhibited tumor growth in vivo. These results indicate that HGF/ NK4 may inhibit growth and invasion of carcinoma cells, as mediated by HGF during tumor-stromal interactions. We propose that there is a unique therapeutic potential for HGF/NK4 to prevent tumor invasion and perhaps even metastasis.
Asunto(s)
Neoplasias de la Vesícula Biliar/fisiopatología , Factor de Crecimiento de Hepatocito/antagonistas & inhibidores , Kringles , Invasividad Neoplásica , Anciano , Animales , Femenino , Humanos , Masculino , Metaloproteinasa 9 de la Matriz , Inhibidores de la Metaloproteinasa de la Matriz , Ratones , Ratones Desnudos , Persona de Mediana Edad , Trasplante de Neoplasias , Fosforilación , Proteínas Proto-Oncogénicas c-met/metabolismo , Células Tumorales Cultivadas , Tirosina , Activador de Plasminógeno de Tipo Uroquinasa/antagonistas & inhibidoresRESUMEN
A mechanism of the long-term potentiation of transmitter release induced by adrenaline (ALTP) was studied by recording intracellularly the fast excitatory postsynaptic potentials (fast EPSPs). The ALTP was produced during the blockade of K+ channels at the presynaptic terminals by tetraethylammonium (TEA). The synaptic delay, possibly reflecting a relative change in the duration of an action potential at the presynaptic terminal, was not changed during the course of the ALTP. By contrast, it was significantly lengthened by TEA and other K+ channel inhibitors (4-aminopyridine and Cs+) that markedly enhanced the evoked release of transmitter. The magnitude of facilitation of the fast EPSP, induced by a conditional stimulus to the preganglionic nerve, was decreased during the generation of the ALTP, but was unchanged during the potentiation of transmitter release caused by TEA. These results, together with theoretical considerations applying the residual Ca2+ hypothesis to the facilitation, suggest that the enhancement of transmitter release during the ALTP is not caused by an increased Ca2+ influx during a presynaptic impulse owing to the blockade of K+ channel or the modulation of Ca2+ channel, but presumably is induced by a rise in the basal level of free Ca2+ in the presynaptic terminal.
Asunto(s)
Epinefrina/farmacología , Ganglios Simpáticos/metabolismo , Neurotransmisores/metabolismo , Animales , Fenómenos Biomecánicos , Calcio/metabolismo , Conductividad Eléctrica , Espacio Extracelular/metabolismo , Modelos Biológicos , Concentración Osmolar , Potasio/metabolismo , Rana catesbeiana , Tetraetilamonio , Compuestos de Tetraetilamonio/farmacología , Factores de TiempoRESUMEN
Fluorescent ryanodine revealed the distribution of ryanodine receptors in the submembrane cytoplasm (less than a few micrometers) of cultured bullfrog sympathetic ganglion cells. Rises in cytosolic Ca(2+) ([Ca(2+)](i)) elicited by single or repetitive action potentials (APs) propagated at a high speed (150 microm/s) in constant amplitude and rate of rise in the cytoplasm bearing ryanodine receptors, and then in the slower, waning manner in the deeper region. Ryanodine (10 microM), a ryanodine receptor blocker (and/or a half opener), or thapsigargin (1-2 microM), a Ca(2+)-pump blocker, or omega-conotoxin GVIA (omega-CgTx, 1 microM), a N-type Ca(2+) channel blocker, blocked the fast propagation, but did not affect the slower spread. Ca(2+) entry thus triggered the regenerative activation of Ca(2+)-induced Ca(2+) release (CICR) in the submembrane region, followed by buffered Ca(2+) diffusion in the deeper cytoplasm. Computer simulation assuming Ca(2+) release in the submembrane region reproduced the Ca(2+) dynamics. Ryanodine or thapsigargin decreased the rate of spike repolarization of an AP to 80%, but not in the presence of iberiotoxin (IbTx, 100 nM), a BK-type Ca(2+)-activated K(+) channel blocker, or omega-CgTx, both of which decreased the rate to 50%. The spike repolarization rate and the amplitude of a single AP-induced rise in [Ca(2+)](i) gradually decreased to a plateau during repetition of APs at 50 Hz, but reduced less in the presence of ryanodine or thapsigargin. The amplitude of each of the [Ca(2+)](i) rise correlated well with the reduction in the IbTx-sensitive component of spike repolarization. The apamin-sensitive SK-type Ca(2+)-activated K(+) current, underlying the afterhyperpolarization of APs, increased during repetitive APs, decayed faster than the accompanying rise in [Ca(2+)](i), and was suppressed by CICR blockers. Thus, ryanodine receptors form a functional triad with N-type Ca(2+) channels and BK channels, and a loose coupling with SK channels in bullfrog sympathetic neurons, plastically modulating AP.
Asunto(s)
Potenciales de Acción/fisiología , Canales de Calcio/fisiología , Calcio/farmacología , Ganglios Simpáticos/fisiología , Canal Liberador de Calcio Receptor de Rianodina/fisiología , Animales , Calcio/metabolismo , Electrofisiología , Transporte Iónico/fisiología , Rana catesbeianaRESUMEN
Ca(2+)-induced Ca(2+) release (CICR) enhances a variety of cellular Ca(2+) signaling and functions. How CICR affects impulse-evoked transmitter release is unknown. At frog motor nerve terminals, repetitive Ca(2+) entries slowly prime and subsequently activate the mechanism of CICR via ryanodine receptors and asynchronous exocytosis of transmitters. Further Ca(2+) entry inactivates the CICR mechanism and the absence of Ca(2+) entry for >1 min results in its slow depriming. We now report here that the activation of this unique CICR markedly enhances impulse-evoked exocytosis of transmitter. The conditioning nerve stimulation (10-20 Hz, 2-10 min) that primes the CICR mechanism produced the marked enhancement of the amplitude and quantal content of end-plate potentials (EPPs) that decayed double exponentially with time constants of 1.85 and 10 min. The enhancement was blocked by inhibitors of ryanodine receptors and was accompanied by a slight prolongation of the peak times of EPP and the end-plate currents estimated from deconvolution of EPP. The conditioning nerve stimulation also enhanced single impulse- and tetanus-induced rises in intracellular Ca(2+) in the terminals with little change in time course. There was no change in the rate of growth of the amplitudes of EPPs in a short train after the conditioning stimulation. On the other hand, the augmentation and potentiation of EPP were enhanced, and then decreased in parallel with changes in intraterminal Ca(2+) during repetition of tetani. The results suggest that ryanodine receptors exist close to voltage-gated Ca(2+) channels in the presynaptic terminals and amplify the impulse-evoked exocytosis and its plasticity via CICR after Ca(2+)-dependent priming.
Asunto(s)
Canales de Calcio/metabolismo , Exocitosis/fisiología , Plasticidad Neuronal/fisiología , Terminales Presinápticos/metabolismo , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Animales , Calcio/metabolismo , Estimulación Eléctrica , Electrofisiología , Técnicas In Vitro , Placa Motora/metabolismo , Neuronas Motoras/metabolismo , Músculo Esquelético/inervación , Músculo Esquelético/metabolismo , Neurotransmisores/metabolismo , RanidaeRESUMEN
The extent to which Ca2+-induced Ca2+ release (CICR) affects transmitter release is unknown. Continuous nerve stimulation (20-50 Hz) caused slow transient increases in miniature end-plate potential (MEPP) frequency (MEPP-hump) and intracellular free Ca2+ ([Ca2+]i) in presynaptic terminals (Ca2+-hump) in frog skeletal muscles over a period of minutes in a low Ca2+, high Mg2+ solution. Mn2+ quenched Indo-1 and Fura-2 fluorescence, thus indicating that stimulation was accompanied by opening of voltage-dependent Ca2+ channels. MEPP-hump depended on extracellular Ca2+ (0.05-0.2 mM) and stimulation frequency. Both the Ca2+- and MEPP-humps were blocked by 8-(N, N-diethylamino)octyl3,4,5-trimethoxybenzoate hydrochloride (TMB-8), ryanodine, and thapsigargin, but enhanced by CN-. Thus, Ca2+-hump is generated by the activation of CICR via ryanodine receptors by Ca2+ entry, producing MEPP-hump. A short interruption of tetanus (<1 min) during MEPP-hump quickly reduced MEPP frequency to a level attained under the effect of TMB-8 or thapsigargin, while resuming tetanus swiftly raised MEPP frequency to the previous or higher level. Thus, the steady/equilibrium condition balancing CICR and Ca2+ clearance occurs in nerve terminals with slow changes toward a greater activation of CICR (priming) during the rising phase of MEPP-hump and toward a smaller activation during the decay phase. A short pause applied after the end of MEPP- or Ca2+-hump affected little MEPP frequency or [Ca2+]i, but caused a quick increase (faster than MEPP- or Ca2+-hump) after the pause, whose magnitude increased with an increase in pause duration (<1 min), suggesting that Ca2+ entry-dependent inactivation, but not depriming process, explains the decay of the humps. The depriming process was seen by giving a much longer pause (>1 min). Thus, ryanodine receptors in frog motor nerve terminals are endowed with Ca2+ entry-dependent slow priming and fast inactivation mechanisms, as well as Ca2+ entry-dependent activation, and involved in asynchronous exocytosis. Physiological significance of CICR in presynaptic terminals was discussed.
Asunto(s)
Calcio/metabolismo , Calcio/farmacología , Exocitosis/fisiología , Neuronas Motoras/citología , Terminales Presinápticos/metabolismo , Potenciales de Acción/fisiología , Animales , Bloqueadores de los Canales de Calcio/farmacología , Quelantes/farmacología , Ácido Egtácico/análogos & derivados , Ácido Egtácico/farmacología , Potenciales Evocados Motores/efectos de los fármacos , Potenciales Evocados Motores/fisiología , Exocitosis/efectos de los fármacos , Colorantes Fluorescentes , Ácido Gálico/análogos & derivados , Ácido Gálico/farmacología , Activación del Canal Iónico/efectos de los fármacos , Activación del Canal Iónico/fisiología , Magnesio/farmacología , Neuronas Motoras/química , Neuronas Motoras/metabolismo , Neurotransmisores/metabolismo , Terminales Presinápticos/química , Terminales Presinápticos/efectos de los fármacos , Ranidae , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Factores de TiempoRESUMEN
Periodic, synchronized Ca2+ signals appeared 30-120 min after the application of tetrodotoxin, 4-aminopyridine and Cs+, and became stable in interval (6-47s) for hours. The Ca2+ signals were accompanied by excitatory or inhibitory postsynaptic potentials (excitatory postsynaptic currents (EPSCs) for the former) and blocked by the simultaneous application of 6-cyano-7-nitroquinoxaline-2,3-dione and 3-((RS)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid or treatment with Ca2+ -free solution, nicardipine, or omega-conotoxin MVIIC (omegaCTX), but not with ryanodine, caffeine, thapsigargin or CPP alone. Nicardipine largely, but omegaCTX less, blocked Ca2+ action potentials or voltage pulse-induced Ca2+ currents at the cell soma, while omegaCTX completely blocked autaptic EPSCs. Ca2+ signals within a neuron occurred almost simultaneously in the cell soma and all the processes (> 200 microm), while the latency between Ca2+ signals of neighbouring neurons varied over hundreds of ms like that of Ca2 action potential induction from EPSPs. Ca2+ signals propagated in random directions throughout neural circuits. Thus, when Na+ and K+ channels are blocked, Ca2+ action potentials spontaneously occur somewhere in a neuron, eventually propagate via the cell soma to the presynaptic terminals and activate excitatory synaptic transmission, causing synchronized Ca2+ signals. The results further suggest that the axon of hippocampal neurones have the potential ability to convey coded information via Ca2+ action potentials.
Asunto(s)
Potenciales de Acción/fisiología , Relojes Biológicos/fisiología , Señalización del Calcio/fisiología , Calcio/metabolismo , Red Nerviosa/fisiología , 4-Aminopiridina/farmacología , Animales , Canales de Calcio/fisiología , Señalización del Calcio/efectos de los fármacos , Células Cultivadas , Cesio/farmacología , Hipocampo/citología , Hipocampo/metabolismo , Neuronas/citología , Neuronas/fisiología , Técnicas de Placa-Clamp , Ratas , Ratas Wistar , Transmisión Sináptica/fisiología , Tetrodotoxina/farmacologíaRESUMEN
Modifications to the optics of a conventional confocal laser-scanning microscope were made to allow imaging intracellular Ca(2+)-dependent fluorescence with a UV laser (351 or 364 nm). Modifications included: (1) a chromatic compensation lens in the laser path; (2) the design of a practically achromatic relay lens; (3) a longer tube length for the objective; and (4) highly reflective mirrors maximizing fluorescence measurement. This UV laser-scanning confocal microscope (UV-CLSM) yielded a lateral resolution of < 0.3 micron and an axial resolution of < 1.5 microns and a relevant field size of 100 microns in diameter for a 40X objective). The effects of varying the focal length of a compensation lens, the degree of the correction for the coverglass thickness of objective and the detector aperture size on the quality of image formation were examined. Finally, UV-CLSM revealed optical sections of fine and complex structures of bullfrog sympathetic neurones loaded with a Ca(2+)-sensitive fluorescent probe. Changes in intracellular free Ca2+ distribution in response to high [K+] or caffeine were demonstrated. In addition, an increase in the intracellular concentration of caffeine applied externally was clearly imaged in space and time and distinguished from a resultant rise in [Ca2+]i. Thus, the UV-CLSM developed is suitable for ratiometric intracellular Ca2+ measurements and other biological studies.
Asunto(s)
Calcio/análisis , Microscopía Confocal/instrumentación , Microscopía Confocal/métodos , Microscopía Ultravioleta/instrumentación , Microscopía Ultravioleta/métodos , Animales , Neuronas/metabolismo , Rana catesbeiana , Sistema Nervioso Simpático/metabolismoRESUMEN
How depolarization-induced Ca2+ entry or caffeine activates Ca(2+)-induced Ca2+ release (CICR) in the cytoplasm and nucleoplasm was studied by recording intracellular Ca2+ ([Ca2+]i) with a confocal microscope in cultured bullfrog sympathetic ganglion cells. The amplitude and propagation speed of voltage pulse-induced rises in [Ca2+]i were greater in the submembrane (< 5 microns depth) region than in the core region, and delayed and smaller, but significant, in the nucleus. Ryanodine and dantrolene reduced the rises in [Ca2+]i in both the cytoplasm and nucleus. A rapid application of high K+ solution induced global rises in [Ca2+]i in both the cytoplasm and nucleoplasm, which were decreased by dantrolene. Caffeine produced a slow, small rise in [Ca2+]i which grew into a global, regenerative rise both in the cytoplasm and nucleoplasm with some inward gradient in the cytoplasm. Each of the high [Ca2+]i phases during caffeine-induced [Ca2+]i oscillation began in the submembrane region, while low [Ca2+]i phases started in the core region. These results suggest that CICR activated by Ca2+ entry or caffeine occurs predominantly in the submembrane region causing an inwardly spreading Ca2+ wave or [Ca2+]i oscillations, and that the nuclear envelope can cause CICR in the nucleoplasm, which is delayed due to Ca2+ diffusion barrier at the nuclear pores.
Asunto(s)
Calcio/metabolismo , Ganglios Simpáticos/metabolismo , Animales , Cafeína/farmacología , Señalización del Calcio , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Polaridad Celular/efectos de los fármacos , Células Cultivadas , Citoplasma/efectos de los fármacos , Citoplasma/metabolismo , Dantroleno/farmacología , Ganglios Simpáticos/citología , Ganglios Simpáticos/efectos de los fármacos , Membranas Intracelulares/efectos de los fármacos , Membranas Intracelulares/metabolismo , Microscopía Confocal , Modelos Biológicos , Potasio/metabolismo , Potasio/farmacología , Rana catesbeiana , Rianodina/farmacología , Canal Liberador de Calcio Receptor de Rianodina/efectos de los fármacos , Canal Liberador de Calcio Receptor de Rianodina/metabolismoRESUMEN
Seventeen adults and 11 children, a group of 18 familial hypercholesterolemic (FHC) and 10 normal subjects, were fed products with and without locust bean gum (LBG) (8 to 30 g/day) to assess the hypolipidemic effect of LBG. Identical food products with and without LBG were consumed by two groups (A and B) of arbitrarily assigned patients using a cross-over design. Plasma cholesterol, low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol, very low-density lipoprotein cholesterol and triglycerides were measured at 2-wk intervals and compared to control feeding periods. In group A, FHC C decreased 10% and LDL-C 11%, normal subjects decreased C 6% and LDL-C 10% (p less than 0.001). In group B, FHC C decreased 17% and LDL-C 19%, normal subjects decreased cholesterol 11%, and LDL-C 6% (p less than 0.001). Cholesterol and LDL-C were lowered in FHC children in both groups. High-density lipoprotein/LDL ratios increased in both groups. The use of food products with LBG in children and adults is a unique approach to treating FHC. LBG food acceptance was good, and there were no significant side effects. LBG in food products appears to be an effective, safe approach to controlling hyperlipidemia.
Asunto(s)
Hiperlipoproteinemia Tipo II/dietoterapia , Lípidos/sangre , Polisacáridos/uso terapéutico , Adolescente , Adulto , Niño , Colesterol/sangre , HDL-Colesterol , LDL-Colesterol , VLDL-Colesterol , Fibras de la Dieta/análisis , Femenino , Galactanos , Humanos , Hiperlipoproteinemia Tipo II/sangre , Lipoproteínas HDL/sangre , Lipoproteínas LDL/sangre , Lipoproteínas VLDL/sangre , Masculino , Mananos , Persona de Mediana Edad , Gomas de PlantasRESUMEN
Fully refined, bleached, deodorized corn oil and soy oil, and lightly hydrogenated, winterized soy oil were compared for effectiveness in lowering plasma cholesterol. Twenty-four, healthy, young college students were the subjects for the 10-wk studies. At the 300 cal level, the corn oil and unhydrogenated soy oil diets contained approximately 53 g of polyunsaturated and 26 g of saturated fat. The hydrogenated soy oil diet contained 42 and 25 g, respectively. All diets contained approximately 700 mg of cholesterol. Corn oil and unhydrogenated soy oil were equally effective in lowering both total and low density lipoprotein cholesterol. Lightly hydrogenated soy oil was also quite effective, but less so that the more unsaturated oils. Triglycerides were also lowered, but very low density and high density lipoprotein cholesterol concentrations, as well as total high density lipoproteins, were scarcely affected. All of the polyunsaturated fat diets produced small but statistically significant reductions in the cholesterol to protein ratio of all three lipoproteins.
Asunto(s)
Grasas de la Dieta/farmacología , Glycine max , Lípidos/sangre , Adulto , Colesterol/sangre , Femenino , Humanos , Hidrogenación , Lipoproteínas/sangre , Masculino , Aceites/farmacología , Relación Estructura-Actividad , Triglicéridos/sangre , Zea maysRESUMEN
Twenty children ages 3 to 17 yr, eight with normal lipids and 12 with familial hypercholesterolemia were studied on a metabolic unit for 14 days to evaluate fecal bile acid and fecal neutral sterol excretion. The diet contained a moderately low cholesterol content, 180 to 200 mg/day. Stools were collected in three separate, 3-day pools. Fecal bile acids and fecal neutral sterols were measured using two stool markers and thin-layer, and gas-liquid chromatography techniques. Fecal neutral sterol and fecal bile acid excretion were the same for normal and familial hypercholesterolemic children on a mg/kg basis. Fecal neutral sterols in familial hypercholesterolemic children decreased with age, p less than 0.001; fecal bile acid excretion also appeared to decrease with age, but less significantly, p less than 0.07. Although the familial hypercholesterolemic children have significantly increased plasma and potentially elevated tissue or total body cholesterol, the excretion of fecal bile acids and fecal neutral sterols did not differ between familial hypercholesterolemic and normal children.
Asunto(s)
Ácidos y Sales Biliares/metabolismo , Colesterol/metabolismo , Heces/análisis , Hiperlipoproteinemia Tipo II/metabolismo , Esteroles/metabolismo , Adolescente , Envejecimiento , Niño , Preescolar , Colesterol en la Dieta/administración & dosificación , Femenino , Humanos , Hiperlipoproteinemia Tipo II/genética , MasculinoRESUMEN
The efficacy of clofibrate (CPIB) and nicotinic acid (NA) in the treatment of type III hyperlipoproteinemia was evaluated in 5 male subjects in a randomized cross-over study with clofibrate 1 g b.i.d. and NA 3 g/day (given either b.i.d. or t.i.d.). Following a baseline period of 6 weeks, each drug was given for 12 weeks with samples for lipid and lipoprotein determinations obtained at 6, 9, and 12 weeks. Both clofibrate and NA resulted in a significant reduction from baseline of total cholesterol (23% and 28%), VLDL cholesterol (49% and 56%), total triglycerides (40% and 43%), and VLDL triglycerides (46% and 48%), as well as a significant increase in HDL cholesterol (22% and 28%) and HDL/LDL ratio (31% and 62%). The HDL/LDL ratio was higher on NA than clofibrate (0.47 +/- 0.19 vs. 0.38 +/- 0.09, P less than 0.05). Four subjects were continued in the study and treated sequentially with NA 3.0 g/day (alternate to the previous schedule) and gemfibrozil 1.2 g/d in divided doses. Each of the 4 regimens resulted in a significant change from baseline of each of the measured lipid and lipoprotein determinations except LDL cholesterol. Comparison among the treatment regimens revealed no differences except for significantly higher HDL cholesterol and HDL/LDL ratio with NA given t.i.d.
Asunto(s)
Clofibrato/administración & dosificación , Hiperlipoproteinemia Tipo III/tratamiento farmacológico , Hipolipemiantes/administración & dosificación , Niacina/administración & dosificación , Ácidos Pentanoicos/administración & dosificación , Valeratos/administración & dosificación , Adulto , Colesterol/sangre , HDL-Colesterol , LDL-Colesterol , VLDL-Colesterol , Clofibrato/uso terapéutico , Esquema de Medicación , Quimioterapia Combinada , Gemfibrozilo , Humanos , Hiperlipoproteinemia Tipo III/sangre , Hipolipemiantes/uso terapéutico , Lipoproteínas HDL/sangre , Lipoproteínas LDL/sangre , Lipoproteínas VLDL/sangre , Masculino , Persona de Mediana Edad , Niacina/uso terapéutico , Ácidos Pentanoicos/uso terapéutico , Distribución Aleatoria , Triglicéridos/sangreRESUMEN
In an attempt to compare the cholesterol-lowering effects of equivalent doses of D- and L-thyroxine, 10 euthyroid, hypercholesterolemic subjects were treated with graded doses of each medication in a cross-over design using thyrotropin suppression following thyrotropin-releasing hormone administration as the end-point. The mean thyrotropin-suppressive dose of D-thyroxine was 2.4 +/- 0.66 mg per day, which resulted in mean reductions of 10 percent in total plasma cholesterol, 10 percent in plasma low-density lipoprotein cholesterol, and 11 percent in plasma high-density lipoprotein cholesterol. The mean thyrotropin-suppressive dose of L-thyroxine was 135 +/- 46 micrograms per day, which resulted in mean reductions of 7 percent in total plasma cholesterol, 6 percent in plasma low-density lipoprotein cholesterol, and 14 percent in plasma high-density lipoprotein cholesterol. The reductions in total, low-density, and high-density cholesterol achieved with D-thyroxine were not significantly different from those achieved with L-thyroxine. Neither medication produced a significant increase in heart rate or ventricular ectopy as determined by Holter monitoring. These data do not support the belief that D-thyroxine has a preferential cholesterol-lowering effect in humans when compared with equivalent doses of L-thyroxine. In addition, both D- and L-thyroxine reduced plasma high-density lipoprotein cholesterol.