cAMP induction by ouabain promotes endothelin-1 secretion via MAPK/ERK signaling in beating rabbit atria

Adenosine 3',5'-cyclic monophosphate (cAMP) participates in the regulation of numerous cellular functions, including the Na(+)-K(+)-ATPase (sodium pump). Ouabain, used in the treatment of several heart diseases, is known to increase cAMP levels but its effects on the atrium are not understood. The aim of the present study was to examine the effect of ouabain on the regulation of atrial cAMP production and its roles in atrial endothelin-1 (ET-1) secretion in isolated perfused beating rabbit atria. Our results showed that ouabain (3.0 micromol/L) significantly increased atrial dynamics and cAMP levels during recovery period. The ouabain-increased atrial dynamics was blocked by KB-R7943 (3.0 micromol/L), an inhibitor for reverse mode of Na(+)-Ca(2+) exchangers (NCX), but did not by L-type Ca2+ channel blocker nifedipine (1.0 micromol/L) or protein kinase A (PKA) selective inhibitor H-89 (3.0 micromol/L). Ouabain also enhanced atrial intracellular cAMP production in response to forskolin and theophyline (100.0 micromol/L), an inhibitor of phosphodiesterase, potentiated the ouabain-induced increase in cAMP. Ouabain and 8-Bromo-cAMP (0.5 micromol/L) markedly increased atrial ET-1 secretion, which was blocked by H-89 and by PD98059 (30 micromol/L), an inhibitor of extracellular-signal-regulated kinase (ERK) without changing ouabain-induced atrial dynamics. Our results demonstrated that ouabain increases atrial cAMP levels and promotes atrial ET-1 secretion via the mitogen-activated protein kinase (MAPK)/ERK signaling pathway. These findings may explain the development of cardiac hypertrophy in response to digitalis-like compounds.

Effect of ouabain on intracellular Ca(2+) concentration in rat vascular smooth muscle cells in vitro / 南方医科大学学报

<p><b>OBJECTIVE</b>To explore the effect of ouabain on intracellular Ca(2+) concentration ([Ca(2+)]i) in thoracic aorta vascular smooth muscle cells (VSMCs) in vitro.</p><p><b>METHODS</b>Primary SD rat thoracic aorta VSMCs were cultured by tissue adherent method and identified by immunochemistry. The binding ability between ouabain and VSMCs was detected by autoradiography, and fluo 3-AM (a Ca(2+) fluorescent probe) was employed to investigate whether ouabain affected VSMCs within a short period of time. The effect of a truncated fragment of the sodium pump α2 subunit was assayed in antagonizing the effect of ouabain on [Ca(2+)]i in the VSMCs.</p><p><b>RESULTS</b>Within the concentration range of 0.1-100 nmol/L, ouabain was found to dose-dependently bind to the VSMCs. Different concentrations of ouabain (0-3200 nmol/L) caused a transient, dose-dependent increase in [Ca(2+)]i in the VSMCs, which was antagonized by the application of the truncated fragment of sodium pump α2 subunit.</p><p><b>CONCLUSIONS</b>Elevations in [Ca(2+)]i in the VSMCs can be the cytological basis of high ouabain-induced hypertension. The truncated fragment of the sodium pump α2 subunit can antagonize ouabain-induced increase of [Ca(2+)]i in the VSMCs, which provides a clue for understanding the pathogenesis of and devising a therapeutic strategy for high ouabain-induced hypertension.</p>

Ouabain and asthenospermia / 中华男科学杂志

Asthenospermia accounts for about 30% of the causes of male infertility. Currently, most drugs for asthenospermia lack specificity and desirable therapeutic efficiency. An insight into the pathogenesis of asthenospermia is important for the development of specific therapies for this disease. The protein Na+/K(+)- ATPase α4 isoform (NKA4) presents in both mature testis tissue and the sperm tail, the absence or reduced activity of which may significantly decrease sperm motility. Ouabain is a natural inhibitor of NKA4, suppressing its activity by specifically binding the ouabain site in it. The hypothalamus and adrenal cortex excrete an ouabain-like steroid hormone called endogenous ouabain (EO), which may be associated with the pathogenesis of asthenospermia by inhibiting the activity of NKA4, affecting Na+/H+ exchange, Na+/Ca2+ exchange and sperm cell membrane potential, and eventually reducing sperm motility.

Damage of Spiral Ganglion Cell Induced by Ouabain Application in Cat / 대한이비인후과학회지

BACKGROUND AND OBJECTIVES: Auditory neuropathy is a hearing disorder characterized by the absence or the marked impairment of the auditory brainstem responses with the preservation of the cochlear microphonics (CMs) and otoacoustic emissions. This suggests that outer hair cell (OHC) function is normal but proximal auditory function to OHCs is impaired. It is assumed that the lesion is localized at the level of the inner hair cells (IHCs), auditory nerve fibers, or the synapse between them. This study was aimed to observe the change of hearing threshold and pathology of spiral ganglion cell induced by ouabain application, and present basic data to explain the auditory neuropathy. MATERIALS AND METHOD: Twenty ears of twenty normal hearing cats were used in this study. Cats were treated with 100 microL ouabain (1 mM) applied on the round window. After three days, compound action potential (CAP) and CM were measured and the cochlea was obtained. Pathologic change of spiral ganglion cell was evaluated under light microscope after H&E stain. Normal saline was injected for the control group. RESULTS: In the ouabain group, CAP threshold was increased in all tested frequencies (p0.05). There was significant difference between CAP and CM threshold shift (p<0.001). In the control group, there was no significant difference in CAP and CM thresholds. Light microscopic findings show that the condensed chromatin and nuclear fragments of spiral ganglion cells of an ear was exposed to ouabain, and outer hair cell and inner hair cell were not damaged. CONCLUSION: This study shows that the CAP threshold was significantly increased but the CM threshold was not changed in the ouabain group. Ouabain induced damage of spiral ganglion cells. This study is not sufficient to explain auditory neuropathy because threshold shift of CAP is not obvious, but it would be helpful to explain that selective damage of spiral ganglion cell would be the mechanism of auditory neuropathy.

Proliferating cell nuclear antigen involved in the repair process of ouabain-induced brain damage independent of hypertension in rats / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Ouabain is a mammalian adrenocortical hormone that is involved in the pathogenesis of hypertension by inhibiting Na-K ATPase activity. It also participates in a variety of kinase-mediated signaling pathways associated with Na-K ATPase. Previous studies have shown that ouabain can cause cardiac remodeling independent of elevated blood pressure and that proliferating cell nuclear antigen (PCNA) plays a coordinating role for numerous proteins involved in multiple processes associated with DNA synthesis. Therefore, we hypothesized that ouabain might play a role in the cerebral cortex through signaling pathways independent of hypertension. And PCNA might be involved in this process.</p><p><b>METHODS</b>Male Sprague-Dawley rats were treated with ouabain or with 0.9% nitric sodium as the control group. Systolic blood pressure was recorded weekly. After four weeks of treatment, morphological changes in the cerebral cortex were analyzed using light and transmission electron microscopy. The expression of PCNA in the cerebral cortex was evaluated by immunohistochemistry, real time quantitative PCR, and Western blotting.</p><p><b>RESULTS</b>After 4-week treatment, there was no significant difference in systolic blood pressure compared with the control group, but both structural deterioration and up-regulated expression of PCNA in the brain was induced by ouabain treatment.</p><p><b>CONCLUSIONS</b>These results suggest that ouabain induces alterations in the brain structure, and this effect is independent of blood pressure. PCNA might be involved in the repair process of ouabain-induced brain damage.</p>

Effect of ouabain on the pathogenesis of hypertension in rats / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Ouabain and digoxin are important cardiac glycoside and related to many cardiovascular diseases. The purpose of this study was to investigate the changes of sodium pump α-subunit expression in rats and compare the effects of ouabain (OUA) and digoxin (DIG) on the development of hypertension.</p><p><b>METHODS</b>In situ hybridization was performed. Specific sequence oligonucleotide probe tailing with a Dig-dUTP hybrid to target nucleic acids of the sodium pump α-subunit. According to counting positive particles sodium pump subunit expression was analyzed with statistical methods.</p><p><b>RESULTS</b>On day 16 of drug administration, the blood pressure of rats increased significantly in the OUA group. In the DIG group, the blood pressure revealed no significant difference when compared to the control group. In addition, the effects of OUA and DIG on sodium pump α-subunit RNA expression in tissues differed.</p><p><b>CONCLUSIONS</b>OUA and DIG can not only change the configuration of the sodium pump to depress their activity, but also influence their gene expression which is important in the mechanism of hypertension. This may be a key point in the pathogenesis of hypertension in the manner in which OUA differs from DIG and changes the sodium pump gene expression in the arteries and kidneys of rats.</p>

Relationship between ouabain and asthenozoospermia / 华中科技大学学报（医学）（英德文版）

A growing number of researches have shown that ouabain can regulate mammalian sperm function and male reproduction by modulating the sperm motility, capacitation and acrosome reaction in vitro. This study further examined the relationship between ouabain and asthenozoospermia. In this study, the rat was intraperitoneally injected with ouabain at different concentrations (low-dose ouabain group: 12.5 μg/kg body weight per day, and high-dose ouabain group: 25 μg/kg body weight per day) for 30 days to establish the asthenozoospermia model. The sperms from 60 males with normal fertility were incubated with ouabain of gradient concentrations (10(-7)-10(-2) mol/L) for 4 h. The sperm motility was evaluated under a microscope. Moreover, the endogenous ouabain (EO) level was determined in seminal plasma of mild or severe asthenozoospermia patients and males with normal fertility by competitive inhibition ELISA. The results showed that the sperm motility was significantly diminished in the rats treated with different concentrations of ouabain. The number of motile sperms (grades a and b) was decreased greatly in a time- and dose-dependent manner in 10(-5)-10(-2) mol/L ouabain groups (P<0.01), while no obvious change in sperm motility was observed in 10(-7)-10(-6)mol/L groups even for 4-h incubation (P>0.05). Furthermore, the EO level was significantly increased in asthenozoospermia patients as compared with that in males with normal fertility (25.27±1.71 μg/L in mild asthenozoospermia patients, 26.52±1.82 μg/L in severe asthenozoospermia patients, 19.31±1.45 μg/L in normal fertility men) (P<0.01). In conclusion, rat asthenozoospermia was successfully established by intraperitoneal injection of ouabain, and 10(-5) mol/L ouabain was sufficient enough to inhibit sperm motility in vitro. Moreover, EO, a normal constituent of seminal plasma, was highly expressed in asthenozoospermia males as compared with normal fertility ones.

Antiarrhythmic effect of TJ0711 / 药学学报

To study the antiarrhythmic effect of the newly developed alpha/beta-blocker TJ0711, a variety of animal models of arrhythmia were induced by CaCl2, ouabain and ischemia/reperfusion. Glass microelectrode technique was used to observe action potentials of right ventricular papillary muscle of guinea pig. The onset time of arrhythmia induced by CaCl2 was significantly prolonged by TJ0711 at 0.75, 1.5 and 3 mg x kg(-1) doses. TJ0711 (1.5 and 3 mg x kg(-1)) can significantly shorten the ventricular tachycardia (VT) and ventricular fibrillation (VF) duration, the incidence of VF and mortality were significantly reduced. On ischemia-reperfusion-induced arrhythmic model, TJ0711 (0.25, 0.5, 1 and 2 mg x kg(-1)) can significantly reduce the ventricular premature contraction (PVC), VT, VF incidence, mortality, arrhythmia score with a dose-dependent manner. At the same time, rats serum lactate dehydrogenase (LDH) and creatine kinase (CK) activities decreased significantly by TJ0711 (1 and 2 mg x kg(-1)). Ouabain could cause arrhythmia in guinea pigs, when TJ0711 (0.375, 0.75, 1.5 and 3 mg x kg(-1)) was given, the doses of ouabain inducing a variety of arrhythmia PVC, VT, VF, cardiac arrest (CA) were significantly increased with a dose-dependent manner. In the TJ0711 0.1-30 micromol x L(-1) concentration range, guinea pig right ventricular papillary muscle action potential RP (rest potential), APA (action potential amplitude) and V(max) (maximum velocity of depolarization) were not significantly affected. APD20, APD50 and APD90 had a shortening trend but no statistical difference with the increase of TJ0711 concentration. TJ0711 has antiarrhythmic effect on the sympathetic nerve excitement and myocardial cell high calcium animal arrhythmia model. Myocardial action potential zero phase conduction velocity and resting membrane potential were not inhibited by TJ0711. APD20, APD50 and APD90 were shortened by TJ0711 at high concentration. Its antiarrhythmic action mechanism may be besides the action of blocking beta1 receptor, may also have a strong selective blocking action on alpha1 receptor and reducing intracellular calcium concentration.

Effect of m-calpain in PKCα-mediated proliferation of pulmonary artery smooth muscle cells by low dose of ouabain.

There is growing evidence that ouabain, a cardiotonic steroid may promote growth of cardiac and vascular myocytes, indicating its novel role in cell growth and proliferation, without appreciable inhibition of the sodium pump. The mechanism(s) by which low dose of ouabain produces pulmonary artery smooth muscle cell proliferation, a prerequisite for right ventricular hypertrophy, is currently unknown. Here, we analyzed the effects of low dose of ouabain (10 nM) on increase in [Ca2+]i, m-calpain and protein kinase C (PKC) activities on pulmonary artery smooth muscle cell proliferation and determined their sequential involvement in this scenario. We treated bovine pulmonary artery smooth muscle cells with a low dose of ouabain (10 nM) and determined [Ca2+]i in the cells by fluorometric assay using fura2-AM, m-calpain activity by fluorometric assay using SLLVY-AMC as the substrate, PKC activity using an assay kit and assay of Na+/K+ATPase activity spectrophotometrically. We purified m-calpain and PKCα by standard chromatographic procedure by HPLC and then studied cleavage of the purified PKCα by m-calpain using Western immunoblot method. Subsequently, we performed cell proliferation assay utilizing the redox dye resazunin. We used selective inhibitors of [Ca2+]i (BAPTA-AM), m-calpain (MDL28170), PKCα (Go6976) and determined their involvement in ouabain (10 nM)-mediated smooth muscle cell proliferation. Our results suggested that treatment of bovine pulmonary artery smooth muscle cells with a low dose of ouabain (10 nM) increased [Ca2+]i and subsequently stimulated m-calpain activity and proteolytically activated PKCα in caveolae (signaling microdomain also known as signalosomes) of the cells. Upon activation, PKCα increased the smooth muscle cell proliferation via Go/G1 to S/G2-M phase transition. Thus, [Ca2+]i-mCalpain-PKCα signaling axis plays a crucial role during low dose of ouabain-mediated pulmonary artery smooth muscle cell proliferation.

Decreased BDNF levels in amygdala and hippocampus after intracerebroventricular administration of ouabain / Diminuição dos níveis de BDNF em amígdala e hipocampo após a administração intracerebroventricular de ouabaína

OBJECTIVE: The present study aims to investigate the effects of ouabain intracerebroventricular injection on BDNF levels in the amygdala and hippocampus of Wistar rats.METHODS: Animals received a single intracerebroventricular injection of ouabain (10-3 and 10-2 M) or artificial cerebrospinal fluid and immediately, 1h, 24h, or seven days after injection, BDNF levels were measured in the rat's amygdala and hippocampus by sandwich-ELISA (n = 8 animals per group).RESULTS: When evaluated immediately, 3h, or 24h after injection, ouabain in doses of 10-2 and 10-3 M does not alter BDNF levels in the amygdala and hippocampus. However, when evaluated seven days after injection, ouabain in 10-2 and 10-3 M, showed a significant reduction in BDNF levels in both brain regions evaluated.DISCUSSION: In conclusion, we propose that the ouabain decreased BDNF levels in the hippocampus and amygdala when assessed seven days after administration, supporting the Na/K ATPase hypothesis for bipolar illness.

OBJETIVO: O presente estudo tem como objetivo investigar os efeitos da injeção intracerebroventricular de ouabaína sobre os níveis de BDNF na amígdala e no hipocampo de ratos Wistar.MÉTODOS: Os animais receberam uma única injeção intracerebroventricular de ouabaína (10-3 and 10-2 M) ou fluido cerebroespinhal artificial e, imediatamente, 3h, 24h ou sete dias após a injeção, os níveis de BDNF foram mensurados na amígdala e hipocampo dos ratos por ELISA sandwich (n = 8 animais por grupo).RESULTADOS: Quando avaliados imediatamente após a injeção, 3h ou 24h, ouabaína nas doses 10-2 e 10-3 M não alterou os níveis de BDNF em ambas as estruturas avaliadas. Entretanto, quando avaliados sete dias após a injeção, ouabaína nas doses 10-2 e 10-3 M mostrou uma significante redução nos níveis de BDNF em amígdala e hipocampo.CONCLUSÃO: Em conclusão, propõe-se que a administração de ouabaína diminuiu os níveis de BDNF em amígdala e hipocampo quando avaliados sete dias após a injeção, suportando a hipótese da participação da Na/K ATPase no transtorno bipolar.

Detrimental effects of ouabain on cochlear spiral ganglion cells in rats / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To investigate the detrimental effects of ouabain on cochlear spiral ganglion neurons (SGCs) in vivo and in vitro.</p><p><b>METHODS</b>Seventy-five male SD rats were randomly divided into 5 groups. In addition to the normal control group, rats in other 4 groups received 0.01, 0.02, 0.05 mmol/L of Ouabain or saline through cochlear scala tympani drilling. Seven days after surgery, the hearing threshold was measured by distortion product otoacoustic emissions (DPOAE) and auditory brainstem response (ABR) in rats. In the in vitro study, SGNs were isolated from SD rats (E14) and treated with 1 × 10(-8) mmol/L of Ouabain. The damaged of SGCs were detected after ouabain treatment using immunohistochemistry, transmission electron microscope and scanning electron microscope in vitro.</p><p><b>RESULTS</b>After administration of Ouabain, DPOAE did not change significantly. No significant difference in the amplitude of DPOAE could be observed among all the groups (P > 0.05). Compared with saline and normal control, ABR threshold was significantly increased in the Ouabain treated groups (P < 0.05), which correlated with the concentration of Ouabain. Electron microscopy showed that after treated with Ouabain, SGCs presented degenerative changes, including collapse of organelle structures, the karyotheca dissolved, myelin sheath disintegrating. Ouabain could damage type I SGCs but not type II SGNs.</p><p><b>CONCLUSIONS</b>Ouabain can damage SGCs, either in the in vivo or in vitro conditions.</p>

Hypertensive effects of the iv administration of picomoles of ouabain

Ouabain, an endogenous digitalis compound, has been detected in nanomolar concentrations in the plasma of several mammals and is associated with the development of hypertension. In addition, plasma ouabain is increased in several hypertension models, and the acute or chronic administration of ouabain increases blood pressure in rodents. These results suggest a possible association between ouabain and the genesis or development and maintenance of arterial hypertension. One explanation for this association is that ouabain binds to the α-subunit of the Na+ pump, inhibiting its activity. Inhibition of this pump increases intracellular Na+, which reduces the activity of the sarcolemmal Na+/Ca2+ exchanger and thereby reduces Ca2+ extrusion. Consequently, intracellular Ca2+ increases and is taken up by the sarcoplasmic reticulum, which, upon activation, releases more calcium and increases the vascular smooth muscle tone. In fact, acute treatment with ouabain enhances the vascular reactivity to vasopressor agents, increases the release of norepinephrine from the perivascular adrenergic nerve endings and promotes increases in the activity of endothelial angiotensin-converting enzyme and the local synthesis of angiotensin II in the tail vascular bed. Additionally, the hypertension induced by ouabain has been associated with central mechanisms that increase sympathetic tone, subsequent to the activation of the cerebral renin-angiotensin system. Thus, the association with peripheral mechanisms and central mechanisms, mainly involving the renin-angiotensin system, may contribute to the acute effects of ouabain-induced elevation of arterial blood pressure.

effect of paroxetine on ouabain-induced toxicity in isolated guinea pig atria

It has been reported that selective serotonin reuptake inhibitors [SSRIs] possess some cardiac effects. In the present study we have investigated the effect of paroxetine [PX], a potent SSRI agent, on spontaneously as well as ouabain-induced arrhythmia beating isolated guinea-pig atria. The Guinea-pig heart was rapidly removed; the auricles were dissected out in oxygenated modified Krebs solution. The rate and force of spontaneous contractions were recorded isometrically with a photosensitive transducer. PX [1-16 micro g/ml] caused a dose-dependent decrease in the rate of contractions [14-70%] and contractile force [8-16%]. Ouabain alone [1.2 micro g/ml] produced arrhythmia at 7.2 +/- 1.5 min and asystole at 20.1 +/- 3.1 min. Pretreatment with PX [4 micro g/ml] significantly increased the time of arrhythmia onset to 19.8 min. In addition, PX prolonged the duration of action beating from 20.1 +/- 3.1 min to 43.1 +/- 2.6 and delayed the occurrence of asystole. The pattern of contractile force by PX + ouabain treatment was more regular than that observed after administration of ouabain alone. The above findings may the probably be due to the inhibition of cardiac Na[+] and Ca[2+] channels or autonomic nervous system. Results also suggest that PX may reduce the membrane conductance through inhibition of ionic channels to prevent ouabain-induced arrhythmia

Transplantation of Neural Differentiated Human Mesenchymal Stem Cells into the Cochlea of an Auditory-neuropathy Guinea Pig Model

The aim of this study was to determine the effects of transplanted neural differentiated human mesenchymal stem cells (hMSCs) in a guinea pig model of auditory neuropathy. In this study, hMSCs were pretreated with a neural-induction protocol and transplanted into the scala tympani of the guinea pig cochlea 7 days after ouabain injury. A control model was made by injection of Hanks balanced salt solution alone into the scala tympani of the guinea pig cochlea 7 days after ouabain injury. We established the auditory neuropathy guinea pig model using 1 mM ouabain application to the round window niche. After application of ouabain to the round window niche, degeneration of most spiral ganglion neurons (SGNs) without the loss of hair cells within the organ of Corti and increasing the auditory brain responses (ABR) threshold were found. After transplantation of neural differentiated hMSCs, the number of SGNs was increased, and some of the SGNs expressed immunoreactivity with human nuclear antibody under confocal laser scanning microscopy. ABR results showed mild hearing recovery after transplantation. Based on an auditory neuropathy animal model, these findings suggest that it may be possible to replace degenerated SGNs by grafting stem cells into the scala tympani.

A low concentration of ouabain (0.18 µg/kg) enhances hypertension in spontaneously hypertensive rats by inhibiting the Na+ pump and activating the renin-angiotensin system

We investigated the effects of low ouabain concentrations on systolic (SAP) and diastolic (DAP) arterial pressures and on pressor reactivity in 3-month-old male spontaneously hypertensive rats (SHR). Arterial blood pressure (BP) and pressor reactivity to phenylephrine (PHE) were investigated before and after 0.18 ìg/kg ouabain administration (N = 6). The influence of hexamethonium (N = 6), canrenone (N = 6), enalapril (N = 6), and losartan (N = 6) on ouabain actions was evaluated. Ouabain increased BP (SAP: 137 ± 5.1 to 150 ± 4.7; DAP: 93.7 ± 7.7 to 116 ± 3.5 mmHg; P<0.05) but did not change PHE pressor reactivity. Hexamethonium reduced basal BP in control but not in ouabain-treated rats. However, hexamethonium + ouabain increased DAP sensitivity to PHE. Canrenone did not affect basal BP but blocked ouabain effects on SAP. However, after canrenone + ouabain administration, DAP pressor reactivity to PHE still increased. Enalapril and losartan reduced BP and abolished SAP and DAP responses to ouabain. Enalapril + ouabain reduced DAP reactivity to PHE, while losartan + ouabain reduced SAP and DAP reactivity to PHE. In conclusion, a small dose of ouabain administered to SHR increased BP without altering PHE pressor reactivity. Although the renin-angiotensin system (RAS), Na+ pump and autonomic reflexes are involved in the effects of ouabain on PHE reactivity, central mechanisms might blunt the actions of ouabain on PHE pressor reactivity. The effect of ouabain on SAP seems to depend on the inhibition of both Na+ pump and RAS, whereas the effect on DAP seems to depend only on RAS.

Altered levels of nodal excitability by rate-dependent inhibitory effects of essential oil of Citrus aurantium on the electrophysiological properties of isolated perfused rabbit AV-Node: protective role in the prevention of ouabain toxicity / Alteración de los niveles de excitabilidad nodal por efectos inhibitorios velocidad-dependiente del aceite esencial de Citrus aurantium sobre las propiedades electrofisiológicas del nodo AV Aislado y perfundido de conejo: papel protector en la prevención de toxicidad por ouabaína

Treatment of supraventricular arrhythmia includes a wide range of medical interventions. Herbal remedies are suitable alternatives to synthetic drugs due to their availability, minimal side effects and lower price. Pharmacological studies and traditional medical literature point to the cardiovascular effects of Citrus aurantium L. (Rutaceae) in many instances. In the present study we used isolated perfused AV-node of rabbit as an experimental model to determine the effect of various concentrations of essential oil of C. aurantium (0.1-0.3 v/v) on the nodal conduction time and refractoriness of an isolated rabbit AV-nodal preparations. Selective stimulation protocols were used to independently quantify AV nodal recovery, facilitation and fatigue in 18 rabbits. Our results showed concentration-dependent and rate-independent suppressive effects of essence of C. aurantium on the Wenchebach cycle length (WBCL), AV Conduction Time (AVCT) and effective and functional refractory periods (ERP & FRP). Functional properties such as facilitation and fatigue were significantly increased by this plant. Citrus aurantium plays a protective role against the toxic effects of ouabaine by increasing AV nodal conduction time and refractoriness. The above results indicated differential effects of C. aurantium on slow and fast pathways, with a dominant role on fast pathways. This research has explained the protective role of C. aurantium on ouabaine toxicity. All results indicated the potential anti-arrhythmic effects of C. aurantium in treating supraventricular tachyarrhythmia.

El tratamiento de la arritmia supraventricular incluye una amplia gama de intervenciones médicas. Los remedios herbarios son alternativas adecuadas a las drogas sintéticas debido a su disponibilidad, con escasos efectos secundarios y bajo precio. Estudios farmacológicos y la literatura médica tradicional señalan los efectos cardiovasculares de Citrus aurantium L. (Rutaceae) en muchos casos. En el presente estudio se usaron aislados perfundidos del nodo AV de conejo como modelo experimental para determinar el efecto de diferentes concentraciones de aceite esencial de C. aurantium (0,1-0,3 v/v) sobre en el tiempo de conducción nodal y refractariedad. Un protocolo de estimulación selectiva se utilizó para cuantificar de forma independiente la recuperación, la facilitación y la fatiga del nodo AV en 18 conejos. Nuestros resultados muestran efectos supresores dependientes de la concentración e independiente de la velocidad de la esencia de C. aurantium sobre la duración del ciclo Wenchebach (WBCL), tiempo de conducción AV (AVTC) y períodos refractarios eficaz y funcional (PRE y PRF). Propiedades funcionales tales como la facilitación y la fatiga se incrementaron de manera significativa por esta planta. La Citrus aurantium juega un papel protector contra los efectos tóxicos de ouabaína al incrementar el tiempo de conducción AV nodal y la refractariedad. Los resultados indican efectos diferenciales de C. aurantium sobre las vías lentas y rápidas, con un papel dominante en las vías rápidas. Esta investigación ha explicado el papel protector de C. aurantium sobre la toxicidad ouabaine. Todos los resultados indican los posibles efectos anti-arrítmicos de C. aurantium en el tratamiento de taquiarritmias supraventriculares.

Gene expression profiling of the proliferative effect of periplocin on mouse cardiac microvascular endothelial cells / 中国结合医学杂志

<p><b>OBJECTIVE</b>Periplocin is an active digitalis-like component from Cortex Periplocae, which has been widely used in the treatment of heart diseases in China for many years. According to the recommendations on the cardiovascular effect of periplocin from in vivo experiments, subsequent in vitro experiments are greatly needed for the global assessment of periplocin. The objective of this study is to investigate the cell proliferation effect and the mechanism of periplocin on endothelial cells.</p><p><b>METHODS</b>The proliferative activity of periplocin (0.4, 2, 10, 50, 250 micromol/L; 6, 12, 24, 48, 72 h) was investigated by a comparison with the well-reported cardiac glycoside, ouabain, on mouse cardiac microvascular endothelial cells (CMEC). 3-(4,5-dimethylthiazolyl)-2,5-diphenyltetrazolium bromide (MTT), lactate dehydrogenase (LDH) and 5-bromo-2-deoxyuridine (BrdU) assays were used to evaluate cell proliferation and viability. Subsequently, cDNA microarray experiments were performed on periplocin- (50 micromol/L) and ouabain- (50 micromol/L) treated cells, and data was analyzed by ArrayTrack software.</p><p><b>RESULTS</b>Periplocin could increase cell viability to a level lower than ouabain in the MTT analysis, but decrease LDH release simultaneously. The BrdU incorporation assay showed an increase in cell proliferation with 2-50 micromol/L periplocin. Genes related to protein serine/threonine kinase were the most significantly enriched in the 160 genes identified in periplocin versus the control. In the 165 genes regulated by periplocin versus ouabain, GTP-binding was the most altered term.</p><p><b>CONCLUSIONS</b>The results demonstrated the proliferation action of periplocin on CMEC. Meanwhile, its lower cytotoxicity compared to ouabain provides a new insight into the treatment of heart failure.</p>

Role of renal sympathetic nerves in renal sodium transport in ouabain-hypertensive rats / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the role of renal sympathetic nerves in renal sodium transport in ouabain-hypertensive rats (OHR).</p><p><b>METHODS</b>Sixteen male SD rats with sham renal denervation (Sham-RDNX) and 16 with renal denervation (RDNX) were randomly into normal control group and ouabain group to receive intraperitoneal injection of normal saline (1 ml/kg) and ouabain (27.8 µg/kg) once a day, respectively. Systolic blood pressure (SBP), heart rate and body weight were recorded weekly. Food consumption of the rats was determined twice a week. After a 4-week treatment, blood and 24 h urine samples were collected to measure the serum and urinary concentration of sodium, trace lithium and creatinine. Endogenous creatinine clearance rate (Ccr), fractional excretions of sodium (FENa), fractional excretions of lithium (FELi) and fractional reabsorption of sodium in the postproximal tubules (FDRNa) were calculated. Plasma renin activity was determined by radioimmunoassay. Norepinephrine was extracted from the renal tissue and assayed for norepinephrine content by HPLC.</p><p><b>RESULTS</b>The body weight, food intake and heart rate showed no significant difference among the 4 groups (P > 0.05). After 4 weeks, the SBP of control RDNX group (CDNX) was significantly lower than that of the control Sham-DNX group (Csham)(P < 0.05); the SBP of ouabain RDNX group (ODNX) was also significantly lower than that of ouabain Sham-DNX group (Osham) (P < 0.05); RNDX lowered SBP by about 10 mmHg in both ouabain groups and control groups. The SBP was significantly higher in Osham and ODNX groups than in the corresponding control groups (P < 0.01), also significantly higher in ODNX group than in Csham group (P < 0.01). Ccr showed no significant difference among the 4 groups(P > 0.05). FENa, FELi and FDRNa were significantly lower in ouabain groups than in the corresponding control groups (P < 0.05, P < 0.01, P < 0.05), but FENa, FELi and FDRNa of ODNX group were similar with those of Osham group (P > 0.05); FENa , FELi and FDRNa were similar between CDNX and Csham groups (P > 0.05). The plasma renin activity was comparable between the 4 groups (P > 0.05). Renal norepinephrine level was markedly reduced in RDNX group compared with that in Sham-RDNX group in both ouabain and control groups (P < 0.01).</p><p><b>CONCLUSION</b>The increase of proximal tubule sodium reabsorption in OHR is not dependent on the renal sympathetic nerve.</p>

A key role for Na+/K+-ATPase in the endothelium-dependent oscillatory activity of mouse small mesenteric arteries

Oscillatory contractile activity is an inherent property of blood vessels. Various cellular mechanisms have been proposed to contribute to oscillatory activity. Mouse small mesenteric arteries display a unique low frequency contractile oscillatory activity (1 cycle every 10-12 min) upon phenylephrine stimulation. Our objective was to identify mechanisms involved in this peculiar oscillatory activity. First-order mesenteric arteries were mounted in tissue baths for isometric force measurement. The oscillatory activity was observed only in vessels with endothelium, but it was not blocked by L-NAME (100 µM) or indomethacin (10 µM), ruling out the participation of nitric oxide and prostacyclin, respectively, in this phenomenon. Oscillatory activity was not observed in vessels contracted with K+ (90 mM) or after stimulation with phenylephrine plus 10 mM K+. Ouabain (1 to 10 µM, an Na+/K+-ATPase inhibitor), but not K+ channel antagonists [tetraethylammonium (100 µM, a nonselective K+ channel blocker), Tram-34 (10 µM, blocker of intermediate conductance K+ channels) or UCL-1684 (0.1 µM, a small conductance K+ channel blocker)], inhibited the oscillatory activity. The contractile activity was also abolished when experiments were performed at 20°C or in K+-free medium. Taken together, these results demonstrate that Na+/K+-ATPase is a potential source of these oscillations. The presence of α-1 and α-2 Na+/K+-ATPase isoforms was confirmed in murine mesenteric arteries by Western blot. Chronic infusion of mice with ouabain did not abolish oscillatory contraction, but up-regulated vascular Na+/K+-ATPase expression and increased blood pressure. Together, these observations suggest that the Na+/K+ pump plays a major role in the oscillatory activity of murine small mesenteric arteries.

Ventricular performance and Na+-K+ ATPase activity are reduced early and late after myocardial infarction in rats

Myocardial infarction leads to compensatory ventricular remodeling. Disturbances in myocardial contractility depend on the active transport of Ca2+ and Na+, which are regulated by Na+-K+ ATPase. Inappropriate regulation of Na+-K+ ATPase activity leads to excessive loss of K+ and gain of Na+ by the cell. We determined the participation of Na+-K+ ATPase in ventricular performance early and late after myocardial infarction. Wistar rats (8-10 per group) underwent left coronary artery ligation (infarcted, Inf) or sham-operation (Sham). Ventricular performance was measured at 3 and 30 days after surgery using the Langendorff technique. Left ventricular systolic pressure was obtained under different ventricular diastolic pressures and increased extracellular Ca2+ concentrations (Ca2+e) and after low and high ouabain concentrations. The baseline coronary perfusion pressure increased 3 days after myocardial infarction and normalized by 30 days (Sham 3 = 88 ± 6; Inf 3 = 130 ± 9; Inf 30 = 92 ± 7 mmHg; P < 0.05). The inotropic response to Ca2+e and ouabain was reduced at 3 and 30 days after myocardial infarction (Ca2+ = 1.25 mM; Sham 3 = 70 ± 3; Inf 3 = 45 ± 2; Inf 30 = 29 ± 3 mmHg; P < 0.05), while the Frank-Starling mechanism was preserved. At 3 and 30 days after myocardial infarction, ventricular Na+-K+ ATPase activity and contractility were reduced. This Na+-K+ ATPase hypoactivity may modify the Na+, K+ and Ca2+ transport across the sarcolemma resulting in ventricular dysfunction.