RESUMEN
Prague hereditary hypertriglyceridemic (HTG) rats constitute a genetic model of hypertension associated with hyperlipidemia and insulin resistance. Various cell alterations, including changes in membrane dynamics, ion transport, and decreased platelet responses to thrombin have been observed in this strain. As hypertriglyceridemia appears to be associated with reduced endothelium-dependent vasodilation and platelet aggregation, we examined whether triglycerides could modulate cell responsiveness through changes in cyclic nucleotides in platelets of HTG rats. From the age of 6 weeks, these hypertensive animals were subjected for 10 weeks to interventions that modified circulating triglycerides levels (2.17+/-0.09 mmol/l), leading to their reduction (gemfibrozil treatment, 0.87+/-0.05 mmol/l) or elevation (high fructose intake, 3.23+/-0.07 mmol/l). Basal cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) contents were 15% and 48% lower in isolated platelets of HTG rats than in those of Lewis controls. cAMP level was further reduced in HTG rats subjected to high fructose intake. Irrespective of their plasma triglyceride levels, the thrombin-induced increase in platelet cGMP levels present in Lewis rats was absent in platelets of HTG rats. In contrast, no strain- or treatment-related differences were observed in the magnitude or kinetics of cGMP response to exogenous nitric oxide (NO). NO-induced cGMP and cAMP changes were associated in an opposite manner with trimethylamino-diphenylhexatriene (TMA-DPH) anisotropy, a biophysical parameter that reflects the microviscosity of the outer part of the cell membrane. Our results indicate that the attenuation of platelet responsiveness to thrombin in HTG rats represents a strain difference that cannot merely be due to a difference in plasma triglyceride levels. Platelet hyporesponsiveness to agonists such as thrombin in HTG rats cannot be explained by a change in levels of inhibitory cyclic nucleotides, since they were actually found to be low and not high.
Asunto(s)
Plaquetas/metabolismo , Hipertensión/etiología , Hipertrigliceridemia/sangre , Nucleótidos Cíclicos/metabolismo , Animales , Plaquetas/efectos de los fármacos , AMP Cíclico/metabolismo , AMP Cíclico/farmacología , GMP Cíclico/metabolismo , GMP Cíclico/farmacología , Hipertensión/sangre , Hipertensión/genética , Hipertrigliceridemia/complicaciones , Hipertrigliceridemia/genética , Masculino , Modelos Animales , Óxido Nítrico/farmacología , Nucleótidos Cíclicos/farmacología , Ratas , Ratas Endogámicas Lew , Ratas Mutantes , Trombina/farmacología , Triglicéridos/sangre , Triglicéridos/farmacologíaRESUMEN
OBJECTIVE: To search for alterations of cytosolic pH and cell calcium handling in platelets and erythrocytes of Dahl rats susceptible and resistant to salt-induced hypertension. DESIGN AND METHODS: Blood pressure, plasma lipids, platelet cytosolic calcium concentration ([Ca2+]i) and pH (pHi) together with thrombin-induced changes in these parameters as well as erythrocyte [Ca2+]i and 45Ca influx were determined in Dahl salt-sensitive (SS/Jr) and salt-resistant (SR/Jr) rats aged 9, 15 and 24 weeks, which were fed a low-salt diet (0.3% NaCl), and in animals fed high-salt diet (4% NaCl) for 5-10 weeks since weaning. RESULTS: With a low salt intake platelet pHi was lower in SS/Jr than it was in SR/Jr rats, whereas basal platelet [Ca2+]i was similar in rats of both strains. The difference in basal pHi between SS/Jr and SR/Jr rats increased progressively with age of animals. A high salt intake from youth did not influence platelet [Ca2+]i in rats of either strain but it caused an earlier decrease in pHi in SR/Jr than it did in SS/Jr rats. Thrombin stimulation induced similar elevations of pHi and [Ca2+]i in rats of both strains, irrespective of age, salt intake and response of blood pressure to salt intake. Erythrocyte 45Ca influx and [Ca2+]i were greater for SS/Jr rats but only the latter parameter was correlated positively to blood pressure. Both regulation of platelet pHi and erythrocyte Ca2+ handling were significantly related to plasma lipid levels. CONCLUSIONS: Platelets of SS/Jr rats fed a low-salt diet were characterized by a lower basal cytosolic pHi but unchanged [Ca2+]i relative to those of SR/Jr rats. Hypertension induced by high salt intake was associated with increased erythrocyte [Ca2+]i but not with elevation of platelet [Ca2+]i or alteration of response to stimulation with thrombin.
Asunto(s)
Plaquetas/metabolismo , Calcio/metabolismo , Eritrocitos/metabolismo , Hipertensión/sangre , Lípidos/sangre , Animales , Plaquetas/efectos de los fármacos , Presión Sanguínea , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Citosol/metabolismo , Eritrocitos/efectos de los fármacos , Concentración de Iones de Hidrógeno , Hipertensión/etiología , Hipertensión/fisiopatología , Masculino , Ratas , Ratas Endogámicas , Cloruro de Sodio DietéticoRESUMEN
The effects of the imidazole compound SK&F 96365 on Ca2+ movements and production of nitric oxide (NO) and von Willebrand factor (vWF) have been investigated in human endothelial cells. Changes in cytosolic Ca2+ concentration ([Ca2+]i) were measured with Fura-2. Real-time production of NO was monitored with a porphyrinic microsensor and the release of vWF with an enzyme-linked immunosorbent assay. Irrespective of the transmembrane Ca2+ gradient, 30 microM SK&F 96365 doubled [Ca2+]i suggesting a Ca2+ release from intracellular stores. The SK&F 96365-induced [Ca2+]i rise was not accompanied by detectable NO and vWF production, while 1 microM thapsigargin enhanced [Ca2+]i 2.5 times, doubled the secretion of vWF and increased the NO production to 10 +/- 4 nM (n = 5). Pretreatment with SK&F 96365 prevented thapsigargin from increasing [Ca2+]i, NO production and vWF secretion. To investigate the mechanism by which SK&F 96365 released Ca2+ from internal pools, its effect and that of thapsigargin on the ATP-dependent 45Ca2+ uptake into platelet membrane vesicles were compared. SK&F 96365 as thapsigargin, dose-dependently reduced the initial rate of 45Ca2+ uptake. In conclusion, we demonstrate that, in the absence of Ca2+ entry from the extracellular space, the [Ca2+]i increase elicited by SK&F 96365 or thapsigargin is not sufficient to initiate NO synthesis and vWF secretion. This confirms the important role of Ca2+ influx in endothelial secretion processes.
Asunto(s)
Bloqueadores de los Canales de Calcio/farmacología , ATPasas Transportadoras de Calcio/antagonistas & inhibidores , Calcio/metabolismo , Imidazoles/farmacología , Óxido Nítrico/biosíntesis , Factor de von Willebrand/biosíntesis , Adenosina Trifosfato/farmacología , Transporte Biológico/efectos de los fármacos , Transporte Biológico/fisiología , Plaquetas/enzimología , Plaquetas/ultraestructura , Calcio/farmacocinética , Radioisótopos de Calcio/farmacocinética , ATPasas Transportadoras de Calcio/metabolismo , Células Cultivadas/efectos de los fármacos , Células Cultivadas/enzimología , Células Cultivadas/metabolismo , Citoplasma/metabolismo , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Células Epiteliales , Epitelio/efectos de los fármacos , Epitelio/enzimología , Humanos , Membranas Intracelulares/metabolismo , Tapsigargina/farmacología , Venas Umbilicales/citología , Factor de von Willebrand/metabolismoRESUMEN
As previously described for endothelin-3, platelet exposure to cyclic GMP-elevating agents such as sodium nitroprusside and M&B-22948 (2-o-propoxyphenyl-8-azapurin-6-one), a cGMP phosphodiesterase inhibitor, lowered Ca2+ mobilization in response to thrombin. Interestingly, when cGMP phosphodiesterases were blocked, endothelin-3 produced a dose-dependent cGMP accumulation (P < 0.001). Since endothelin-3 has been proposed to decrease the activity of Ca2+ accumulating pumps, we examined whether this latter effect could be mediated by a rise in cGMP content. Cyclic GMP decreased in a dose-dependent manner the initial rate and plateau value of the ATP-dependent 45Ca2+ uptake in platelet membrane vesicles (P = 0.006 for each). Furthermore, combined treatment with endothelin-3 and M&B-22948 or a moderate concentration of Na(+)-nitroprusside further reduced the thrombin-evoked Ca2+ discharge (P = 0.004 and 0.01, respectively), suggesting that endothelin-3 pre-exposure had reduced the amount of mobilizable Ca2+. We propose that the depletion of platelet Ca2+ stores and the reduction of Ca2+ release evoked by endothelin-3 could be due, at least in part, to the elevation of cGMP content and to a decrease in Ca2+ accumulating pump activity.
Asunto(s)
Plaquetas/metabolismo , Calcio/metabolismo , GMP Cíclico/metabolismo , Endotelina-3/farmacología , Análisis de Varianza , Transporte Biológico Activo/efectos de los fármacos , Calcio/antagonistas & inhibidores , GMP Cíclico/fisiología , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Humanos , Indicadores y Reactivos/farmacología , Nitroprusiato/farmacología , Inhibidores de Fosfodiesterasa/farmacología , Purinonas/farmacología , Tapsigargina/farmacología , Trombina/farmacologíaRESUMEN
The reduction by nitric oxide donors of Ca2+ mobilization in stimulated platelets lead us to investigate the direct effect of authentic NO on ATP-dependent Ca2+ uptake into platelet membrane vesicles. The effects of NO were compared to those of thapsigargin and 2,5-di-(t-butyl)-1,4-benzohydroquinone, two specific inhibitors of the sarco/endoplasmic reticulum Ca2+-ATPases. All three compounds modulated the initial rate of ATP-dependent Ca2+ uptake. NO effects on the initial rate of active Ca2+ uptake were biphasic, with an inhibition above 10 nmol/L and a stimulation below this concentration. These effects could not be attributed to cGMP, its usual effector molecule, or to nitrite ions, its metabolic product. NO inhibitory effects were decreased after a five min incubation, indicating that they were due to a short-lived compound and reversible. These results suggest that NO is functionally coupled to SERCA pumps of the dense tubular system through a cGMP-independent mechanism.
Asunto(s)
Plaquetas/metabolismo , Calcio/metabolismo , Óxido Nítrico/farmacología , Transporte Biológico Activo/efectos de los fármacos , ATPasas Transportadoras de Calcio/antagonistas & inhibidores , Membrana Celular/metabolismo , Sistema Libre de Células , GMP Cíclico/farmacología , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Humanos , Nitritos/farmacología , Terpenos/farmacología , Tapsigargina , Factores de TiempoRESUMEN
Multiple cell membrane alterations have been described in humans and animals with various genetic forms of hypertension and/or dyslipidemia. The aim of our study was to characterize some properties of platelets and/or erythrocytes (cytosolic calcium handling, intracellular pH regulation and thrombin responsiveness) in a new model of genetic hypertension associated with hyperlipidemia-Prague hereditary hypertriglyceridemic (HTG) rats. There were no differences in basal cytosolic Ca2+ values in platelets or erythrocytes of HTG rats and control Wistar rats. Ca2+ influx into erythrocytes was also similar in HTG and control rats. In both strains Ca2+ influx correlated positively with plasma triglycerides. The slope of this relationship was less steep in HTG than in Wistar rats. Cytosolic Ca2+ response to thrombin stimulation was smaller in HTG platelets, which were also characterized by a major reduction of thrombin-induced Mn2+ entry through receptor-operated Ca2+ channels. Platelets of HTG rats had the same basal intracellular pHi values and similar buffering capacity as control rats but their pHi response to thrombin stimulation was substantially reduced. It can be concluded that reduced responsiveness to thrombin stimulation is a major alteration found in platelets of hypertensive hereditary hypertriglyceridemic rats.
Asunto(s)
Plaquetas/efectos de los fármacos , Calcio/metabolismo , Hipertrigliceridemia/metabolismo , Trombina/farmacología , Animales , Plaquetas/citología , Membrana Celular/metabolismo , Concentración de Iones de Hidrógeno , Hipertensión/metabolismo , Hipertrigliceridemia/genética , Canales Iónicos/metabolismo , Transporte Iónico , Manganeso/metabolismo , Ratas , Ratas Endogámicas , Ratas Wistar , Intercambiadores de Sodio-Hidrógeno/metabolismoRESUMEN
Platelet cytosolic free calcium concentration ([Ca2+]i) and intracellular pH (pHi) (including their responses to thrombin), as well as erythrocyte [Ca2+]i and 45Ca2+ influx, were studied in Lyon hypertensive (LH) and normotensive (LN) rats aged 3 months. Platelets of LH rats were characterized by substantially elevated basal [Ca2+]i values, higher [Ca2+]i levels after thrombin stimulation, and enhanced initial rate of thrombin-induced Mn2+ entry through receptor-operated Ca2+ channels. Basal platelet pHi values were not significantly different in LH and LN animals but thrombin elicited a significant alkalinization only in LH platelets. Erythrocytes of LH rats had an enhanced initial rate of 45Ca2+ and tended to elevated [Ca2+]i levels. Our data indicate profound alterations in cell Ca2+ handling in platelets and erythrocytes of LH rats, similar to those previously described in spontaneously hypertensive rats of the Okamoto-Aoki strain. The analysis of the relations between blood pressure, plasma lipids, and cell Ca2+ handling suggested that triglycerides, but not cholesterol, might be involved in altered platelet Ca2+ handling in LH rats.
Asunto(s)
Plaquetas/metabolismo , Calcio/sangre , Citosol/metabolismo , Eritrocitos/metabolismo , Hipertensión/sangre , Líquido Intracelular/metabolismo , Lípidos/sangre , Animales , Plaquetas/efectos de los fármacos , Presión Sanguínea , Citosol/efectos de los fármacos , Eritrocitos/efectos de los fármacos , Concentración de Iones de Hidrógeno , Hipertensión/fisiopatología , Líquido Intracelular/efectos de los fármacos , Masculino , Ratas , Trombina/farmacologíaRESUMEN
In stimulated platelets, endothelin-3 (ET-3) has been previously shown to attenuate Ca2+ mobilization. Using the calcium indicator chlortetracycline, the present study demonstrates that 0.5 microM ET-3 produced a 24% reduction in the Ca2+ pool mobilized by A23187. ET-3 up to 1 microM dose-dependently decreased the initial velocity and steady state value of 45Ca(2+)-uptake into platelet membrane vesicules (p < 0.001). In addition, ET-3 partially reversed the inhibitory effects of half maximally effective concentrations of thapsigargin and 2,5-di-(t-butyl)-1,4-benzohydroquinone, two specific inhibitors of the sarco/endoplasmic reticulum Ca(2+)-ATPases. These results suggest that ET-3 is functionally coupled to Ca(2+)-pumps of the dense tubular system. Based on these findings, we propose that ET-3 decreases the activity of Ca(2+)-pumps in the dense tubular system which accumulates less Ca2+, leading to lowered Ca2+ release in response to agonists.
Asunto(s)
Plaquetas/metabolismo , Calcio/sangre , Endotelinas/farmacología , Análisis de Varianza , Transporte Biológico Activo/efectos de los fármacos , Plaquetas/efectos de los fármacos , Calcimicina/farmacología , Membrana Celular/metabolismo , Clortetraciclina , Citosol/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Técnicas In Vitro , Cinética , Peróxidos/farmacología , Especies Reactivas de Oxígeno/farmacología , Trombina/farmacología , terc-ButilhidroperóxidoRESUMEN
To analyze the mechanisms by which endothelin-3 (ET-3) attenuates agonist-mediated Ca2+ mobilization from an internal pool, we investigated ET-3 effects on 45Ca2+ uptake in platelet membrane vesicles. They were compared to those of thapsigargin (Tg), a specific inhibitor of the dense tubule Ca2+ pumps. In the absence of ATP, ET-3 up to 1 microM did not affect the amount of Ca2+ bound to membrane sites. In the presence of ATP, ET-3 dose-dependently reduced the initial rate and the extent of Ca2+ uptake (p < 0.001). In comparison, Tg dose-dependently inhibited both the ATP-independent Ca2+ binding (p < 0.001) and the ATP-dependent Ca2+ accumulation (p < 0.001), with half-maximal effects at 7 nM. Pretreatment with 1 microM ET-3 decreased the inhibitory effect of 10 nM Tg, but only on the initial rate of ATP-dependent Ca2+ uptake (p = 0.04; n = 6). These results indicate that ET-3 is functionally coupled to Ca(2+)-ATPases of the dense tubules. Its inhibitory effects are probably due to inhibition of the catalytic cycle of the Ca2+ pumps. Such inhibition could lead to a depletion of Ca2+ stores and therefore to reduced Ca2+ release in response to agonists.
Asunto(s)
Plaquetas/efectos de los fármacos , Calcio/metabolismo , Endotelinas/farmacología , Adenosina Trifosfato/farmacología , Plaquetas/metabolismo , Humanos , Transporte Iónico/efectos de los fármacos , Terpenos/farmacología , TapsigarginaRESUMEN
Cytosolic Ca2+ concentration ([Ca2+]i) and 45Ca2+ influx were investigated in erythrocytes from conscious spontaneously hypertensive rats (SHR) and their normotensive controls Wistar-Kyoto (WKY). [Ca2+]i was evaluated with fura-2 and intra- and extra-cellular calibration parameters were compared. Irrespective of the calibration parameters used, erythrocyte [Ca2+]i was always significantly higher in SHR than in WKY and Wistar rats (by 25 and 40%, p < 0.01 and 0.001). A rise of the external Ca2+ concentration from 1 to 2 mmol/l increased less [Ca2+]i in SHR than in WKY erythrocytes (17 vs 37%, p < 0.01). SHR erythrocytes incorporated more 45Ca2+ than those from WKY, with an initial rate of 45Ca2+ uptake higher by 57% than that of WKY erythrocytes (p < 0.05). Vanadate ions, after corrections of their quenching effect on red cell and fura-2 fluorescence signals, increased [Ca2+]i by 19% in WKY erythrocytes (p = 0.05), but did not modify the SHR values. They also increased 45Ca2+ accumulation and the initial rate of 45Ca2+ influx in WKY erythrocytes only (p < 0.01). This study indicates that, when compared to WKY rats, erythrocytes from SHR are characterized by higher [Ca2+]i values, higher initial rate of Ca2+ influx and low sensitivity to vanadate ions.
Asunto(s)
Calcio/sangre , Eritrocitos/efectos de los fármacos , Eritrocitos/metabolismo , Hipertensión/sangre , Vanadatos/farmacología , Animales , Viscosidad Sanguínea , Calcio/farmacocinética , Radioisótopos de Calcio , Proteínas de Unión al Calcio/metabolismo , Estado de Conciencia , Masculino , Ratas , Ratas Endogámicas SHR , Ratas Endogámicas WKY , Ratas WistarRESUMEN
This study was designed to investigate the effects of a hypertensive stimulus, high salt intake, in hypertension-prone (SBH) and -resistant (SBN) Sabra rats on erythrocyte Na+ content (Na+i), Ca2+ influx and cytosolic Ca2+ concentration ([Ca2+]i). The relationships of these parameters to plasma lipids, circulating digoxin-like immunoreactivity and membrane microviscosity, determined by the fluorescence anisotropy of trimethylamino-diphenylhexatriene (TMA-DPH) and diphenylhexatriene (DPH), were also evaluated. Erythrocytes of SBH rats were characterized by increased [Ca2+]i, unchanged Ca2+ influx and reduced Na+i. There were no significant differences in the plasma digoxin-like immunoreactivity between the two strains. High-salt intake decreased membrane microviscosity (DPH anisotropy) in SBH rats but did not alter the above parameters. Erythrocyte [Ca2+]i correlated positively with diastolic blood pressure and negatively with erythrocyte Na+i. Membrane dynamics evaluated by the two fluorescent probes did not correlate with [Ca2+]i, Ca2+ influx or Na+i whereas DPH anisotropy was inversely related to blood pressure. These relationships were independent of plasma cholesterol or triglycerides. It can be concluded that 1) similarly to earlier observations in essential hypertension and spontaneously hypertensive rats, erythrocyte [Ca2+]i correlates positively with blood pressure in salt-dependent hypertension, and 2) increased erythrocyte Na+ content need not be a hallmark of hypertension.
Asunto(s)
Calcio/sangre , Digoxina , Membrana Eritrocítica/efectos de los fármacos , Hipertensión/sangre , Saponinas , Cloruro de Sodio Dietético/administración & dosificación , Sodio/sangre , Animales , Presión Sanguínea , Proteínas Sanguíneas/análisis , Cardenólidos , Citosol/metabolismo , Susceptibilidad a Enfermedades , Polarización de Fluorescencia , Corazón/efectos de los fármacos , Hipertensión/fisiopatología , Lípidos/sangre , Masculino , Tamaño de los Órganos , Ratas , ViscosidadRESUMEN
Since Ca2+ ions seem to directly participate in the control of erythrocyte membrane structure and deformability and because cell Ca2+ metabolism has been repeatedly proposed to be modified in hypertension, the intracellular calcium ion concentration ([Ca2+]i) was investigated in red blood cells from hypertensive and normotensive subjects. [Ca2+]i was measured by using the fluorescent Ca2+ chelator fura-2. Red blood cell [Ca2+]i was increased in hypertensive compared with normotensive subjects in the whole population and further increased when hypertensive were compared with age-matched normotensive subjects. An inverse relation between age and [Ca2+]i was observed when calculated with blood pressure adjusted. In hypertensive patients, high [Ca2+]i values were associated with a reduced erythrocyte deformability. The initial rate of 45Ca2+ uptake did not differ between the two blood pressure groups. Similarly, when the extracellular Ca2+ concentration was elevated from 1 to 2 mmol/l, [Ca2+]i increased by 16 +/- 4% (p less than 0.03) in red blood cells from both groups, thus maintaining a significant difference between hypertensive and normotensive subjects. Under these conditions, the addition of 10(-7) mol/l nicardipine, a dihydropyridine Ca2+ antagonist, decreased [Ca2+]i by 15 +/- 4% (p less than 0.05) and 7 +/- 5% in erythrocytes from hypertensive and normotensive subjects, respectively, thereby reducing the difference in [Ca2+]i observed between these two groups. This nicardipine effect was positively correlated to the initial [Ca2+]i. In the presence of 5 mumol/l W7, a calmodulin antagonist, [Ca2+]i increased significantly only in erythrocytes from hypertensive patients (26 +/- 6%, p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)
Asunto(s)
Calcio/sangre , Eritrocitos/metabolismo , Hipertensión/sangre , Calmodulina/antagonistas & inhibidores , Calmodulina/sangre , Femenino , Humanos , Membranas Intracelulares/metabolismo , Masculino , Persona de Mediana Edad , Concentración Osmolar , Valores de Referencia , Reología , Sulfonamidas/farmacologíaRESUMEN
1. Despite the fact that numerous studies provide evidence for the existence of an endogenous inhibitor of the cell membrane Na(+)-K+ pump in plasma, little is known about the relationships between this factor and the main haemodynamic parameters. 2. In order to shed some light on this, we attempted to correlate haemodynamic parameters, measured during heart catheterization in a group of 22 cardiopathic subjects, with plasma digitalis-like activity levels, determined by two different procedures. 3. The ability of plasma to inhibit a human renal Na(+)-K(+)-ATPase showed an inverse correlation with cardiac index and a direct correlation with peripheral resistance. Plasma cross-reactivity with digoxin antibodies correlated directly with left atrial pressure. 4. These results furnish confirmation of a number of theoretical assumptions which attribute to the digitalis-like factor the ability to modify the contractility of the cardiovascular system.
Asunto(s)
Proteínas Sanguíneas/análisis , Cardiopatías/sangre , Hemodinámica/fisiología , Potasio/metabolismo , Saponinas , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Sodio/metabolismo , Adulto , Anciano , Anticuerpos/metabolismo , Transporte Biológico Activo/efectos de los fármacos , Transporte Biológico Activo/fisiología , Presión Sanguínea/fisiología , Proteínas Sanguíneas/metabolismo , Cardenólidos , Cateterismo Cardíaco , Digoxina/inmunología , Femenino , Humanos , Masculino , Persona de Mediana Edad , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Resistencia Vascular/fisiologíaRESUMEN
Na+ ions, which can play a pathogenic role in the development of high blood pressure, have been reported to regulate membrane enzymatic activities, receptor-ligand interaction and coupling of G-protein receptors to their effectors. This study was designed to investigate the in vitro effects of Na+ ions on membrane dynamic properties. The fluorescence anisotropy values of TMA-DPH (trimethylamino-diphenylhexatriene, probe selectively incorporated into the outer leaflet of the plasma membrane) was evaluated in platelets and erythrocytes of sodium-dependent hypertension-prone and -resistant rats of the Sabra Strain. Whereas no difference was observed between the 2 strains, TMA-DPH anisotropy was found to be strongly influenced in platelets by external Na+ ions. In the absence of external Na+, TMA-DPH anisotropy increased in human and rat platelets. In contrast, Na+ ions did not affect the anisotropy when the probe was inserted into erythrocyte ghosts. This indicates that Na+ ions can acutely regulate order parameter and microviscosity of platelet plasma membrane in the regions explored by the probe.
Asunto(s)
Plaquetas/metabolismo , Membrana Eritrocítica/metabolismo , Fluidez de la Membrana/fisiología , Sodio/metabolismo , Animales , Membrana Celular/metabolismo , Difenilhexatrieno/análogos & derivados , Difenilhexatrieno/metabolismo , Polarización de Fluorescencia , Masculino , Ratas , Ratas Endogámicas , Sodio/farmacología , ViscosidadRESUMEN
Variations of Ca2+ influx (evaluated by the initial rate of 45Ca2+ uptake) and cytosolic free Ca2+ concentration ([Ca2+]i, measured with fura-2) were investigated in human erythrocytes. When external Ca2+ concentration ([Ca2+]o) rose from 1 to 2 mM, the initial rate of Ca2+ influx nearly doubled whereas [Ca2+]i increased only by 15%. Nicardipine dose-dependently decreased both initial rate of Ca2+ influx and [Ca2+]i (up to 53 and 18%. respectively at 10(-6) M). The less marked changes in [Ca2+]i than in Ca2+ influx indicate a partial adjustment of the Ca2+ extruding-pump activity to of Ca2+ influx. In vivo administration of nicardipine reduced [Ca2+]i only when its initial value exceeded 80 nM and prevented the rise in [Ca2+]i induced by the increase in [Ca2+]o. Our results indicate that nicardipine may reduce Ca2+ influx in human erythrocytes and participate in the control of [Ca2+]i when elevated.
Asunto(s)
Canales de Calcio/metabolismo , Calcio/sangre , Eritrocitos/metabolismo , Nicardipino/farmacología , Canales de Calcio/efectos de los fármacos , Citosol/efectos de los fármacos , Citosol/metabolismo , Relación Dosis-Respuesta a Droga , Eritrocitos/efectos de los fármacos , Humanos , Nicardipino/administración & dosificación , Vanadatos/farmacologíaRESUMEN
Digitalis compounds are known to increase Ca2+ influx in various cells. As platelet cytosolic free [Ca2+] and plasma digitalis-like activity have been reported independently to be higher in some untreated hypertensive patients than in normotensive subjects, we have investigated a possible relationship between these two parameters. Platelet cytosolic free [Ca2+], determined using Quin-2, the capacity of plasma extracts to inhibit renal Na+, K(+)-ATPase activity and ouabain binding on human erythrocytes were measured in parallel in 25 untreated hypertensive patients and 11 normotensive subjects. Enhanced values for all 3 parameters were observed in the same hypertensive patient. Platelet cytosolic free [Ca2+] was positively correlated to the plasma digitalis-like activity, which was evaluated by the inhibition of Na+, K(+)-ATPase activity and ouabain binding (r = 0.430, P = .010 and r = 0.448, P = .006, respectively). These relationships were independent of age and blood pressure. These results indicate that endogenous digitalis-like compounds may participate in the control of cytosolic free [Ca2+], in agreement with their proposed hypertensive role.
Asunto(s)
Proteínas Sanguíneas/metabolismo , Calcio/sangre , Digoxina , Hipertensión/sangre , Saponinas , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Adolescente , Adulto , Anciano , Plaquetas/metabolismo , Cardenólidos , Citosol , Eritrocitos/metabolismo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Ouabaína/sangre , ATPasa Intercambiadora de Sodio-Potasio/sangreRESUMEN
Inhibition of the Na(+)-K+ pump by digitalis compounds has been reported to increase intracellular Na+ and Ca2+ concentrations and to stimulate Na(+)-H+ exchange. The activity of endogenous digitalis-like compounds, proposed to promote natriuresis and to raise blood pressure, has been found to be increased in volume expansion and hypertension. The enhanced cytosolic [Ca2+] present in platelets from hypertensive patients may thus originate from inhibition of the Na(+)-K+ pump by endogenous inhibitors, enhanced mobilization of internal Ca2+ stores due to phospholipase C activation and/or structural membrane defects. In unstimulated platelets from essential hypertensives, the increase in [Ca2+]i depends on external Ca2+, thereby underlining the importance of Ca2+ influx. The observation that [Ca2+]i was also enhanced in erythrocytes (p = 0.03) demonstrates that intracellular stores are not required for this rise. Plasma digitalis-like activity was positively correlated with platelet [Ca2+]i (inhibition of renal Na+,K(+)-ATPase, competition with ouabain binding, p less than 0.01). Platelet [Ca2+]i also rose during chronic digoxin administration (p less than 0.02) but not after acute in vitro ouabain treatment. The alkalinisation of platelet cytosol (p = 0.005) also agrees with the stimulation of the Na(+)-H(+)-exchange. In conclusion, these results are compatible with a participation of endogenous Na(+)-K+ pump inhibitors in the control of cytoplasmic [Ca2+] and cell excitability.
Asunto(s)
Plaquetas/metabolismo , Proteínas Sanguíneas/metabolismo , Calcio/sangre , Digoxina , Hipertensión/sangre , Saponinas , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Adulto , Cardenólidos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Potasio/sangre , Potasio/metabolismo , Serotonina/sangre , Sodio/sangre , Sodio/metabolismoRESUMEN
A plasma Na+-K+ ATPase inhibitor, which is estimated by a technique in which it competes with ouabain for binding on red cells, was measured in three groups of individuals: (a) normotensive subjects without a family history of hypertension, (b) normotensive subjects with a family history of hypertension, (c) untreated essential hypertensive subjects. The mean value of the inhibitor in group (b) subjects was significantly higher than the mean value in group (a). The mean value in group (c) subjects was also significantly higher than in group (a) subjects. However, the means of the values in groups (b) and (c) were not significantly different. There was a significant positive correlation between the levels of the inhibitor and the urinary Na+ excretion in all subjects. However, there was no correlation between the inhibitor levels and mean arterial pressure. The relevance of these results to the pathophysiology of hypertension in the black African subject is discussed.
Asunto(s)
Población Negra , Hipertensión/sangre , Natriuréticos/sangre , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Adulto , Anciano , Femenino , Humanos , Hipertensión/genética , Hipertensión/fisiopatología , Masculino , Persona de Mediana Edad , Nigeria , Volumen Plasmático , Sodio/metabolismo , ATPasa Intercambiadora de Sodio-Potasio/sangreRESUMEN
It has been postulated that one or more plasma digitalis-like compounds may play an important role in body fluid regulation and in essential hypertension, although very little is known about their possible role in general haemodynamics. We therefore measured plasma inhibition of human kidney Na+,K+-ATPase and plasma cross-reactivity with digoxin antibodies in 11 normotensive cardiopathic subjects admitted to our clinic for heart catheterization. Possible correlations with haemodynamic parameters were studied. Plasma digoxin-like activity correlated directly with left atrial pressure and with pulmonary circulation data. The ability of the plasma to inhibit Na+,K+-ATPase showed an inverse correlation with cardiac output and cardiac index. No correlations were found with any of the other parameters measured, notably systemic resistance, blood pressure and natriuresis. These findings suggest the presence of more than one substance sharing chemical properties with digitalis: (1) a substance cross-reacting with digoxin antibodies and dependent on pulmonary vascular congestion; and (2) a substance capable of inhibiting the Na+-K+ pump and present in large amounts in heart diseases with a reduced cardiac index.
Asunto(s)
Proteínas Sanguíneas/metabolismo , Cateterismo Cardíaco , Digoxina , Saponinas , Adulto , Anciano , Cardenólidos , Enfermedad Coronaria/fisiopatología , Cardiopatías Congénitas/fisiopatología , Hemodinámica , Humanos , Persona de Mediana Edad , Radioinmunoensayo , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidoresRESUMEN
A decrease in platelet 5-HT content linked to partial inhibition of 5-HT uptake has been described in essential hypertension. Transport of 5-HT through platelet membrane is dependent upon transmembranal Na+ and K+ gradients. It is inhibited by Na+, K+-ATPase inhibitors such as ouabain and endogenous digitalis-like compounds isolated from hemodiafiltrate. The activity of such compounds in plasma extracts, measured by inhibition of Na+,K+-ATPase or ouabain binding to human erythrocytes, and platelet 5-HT content were determined in parallel in essential hypertensive patients. Significant negative correlations were observed between these parameters in men, suggesting that high levels of digitalis-like compounds can affect platelet 5-HT content. In addition, in essential hypertensive patients, total plasma cholesterol was inversely related to both platelet 5-HT content (n = 15, r = -0.594, P less than 0.02) and maximal velocity of 5-HT uptake (n = 15, r = -0.717, P less than 0.003). In normotensive control subjects, no variation of platelet 5-HT content with cholesterol was observed. This suggests that the platelet membranes of essential hypertensive patients are more sensitive to increases in plasma cholesterol than those of normotensive subjects.