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Inhibitor Ouabain (inhibitor + ouabain)
Selected AbstractsLong-term modulation of glucose utilization by IL-1, and TNF-, in astrocytes: Na+ pump activity as a potential target via distinct signaling mechanismsGLIA, Issue 1 2002Céline Véga Abstract Interleukin-1, (IL-1,) and tumor necrosis factor-, (TNF-,) markedly stimulate glucose utilization in primary cultures of mouse cortical astrocytes. The mechanism that gives rise to this effect, which takes place several hours after application of cytokine, has remained unclear. Experiments were conducted to identify the major signaling cascades involved in the metabolic action of cytokine. First, the selective IL-1 receptor antagonist (IL-1ra) prevents the effect of IL-1, on glucose utilization in a concentration-dependent manner, whereas it has no effect on the action of TNF-,. Then, using inhibitors of three classical signaling cascades known to be activated by cytokines, it appears that the PI3 kinase is essential for the effect of both IL-1, and TNF-,, whereas the action of IL-1, also requires activation of the MAP kinase pathway. Participation of a phospholipase C-dependent pathway does not appear critical for both IL-1, and TNF-,. Inhibition of NO synthase by L-NAME did not prevent the metabolic response to both IL-1, and TNF-,, indicating that nitric oxide is probably not involved. In contrast, the Na+/K+ ATPase inhibitor ouabain prevents the IL-1,- and TNF-,-stimulated 2-deoxyglucose (2DG) uptake. When treatment of astrocytes with a cytokine was followed 24 h later by an acute application of glutamate, a synergistic enhancement in glucose utilization was observed. This effect was greatly reduced by ouabain. These data suggest that Na+ pump activity is a common target for both the long-term metabolic action of cytokines promoted by the activation of distinct signaling pathways and the enhanced metabolic response to glutamate. GLIA 39:10,18, 2002. © 2002 Wiley-Liss, Inc. [source] ,-Hydroxybutyrate binds to the synaptic site recognizing succinate monocarboxylate: A new hypothesis on astrocyte,neuron interaction via the protonation of succinateJOURNAL OF NEUROSCIENCE RESEARCH, Issue 7 2008Tünde Molnár Abstract Succinate (SUC), a citrate (CIT) cycle intermediate, and carbenoxolone (CBX), a gap junction inhibitor, were shown to displace [3H],-hydroxybutyrate ([3H]GHB), which is specifically bound to sites present in synaptic membrane subcellular fractions of the rat forebrain and the human nucleus accumbens. Elaboration on previous work revealed that acidic pH-induced specific binding of [3H]SUC occurs, and it has been shown to have a biphasic displacement profile distinguishing high-affinity (Ki,SUC = 9.1 ± 1.7 ,M) and low-affinity (Ki,SUC = 15 ± 7 mM) binding. Both high- and low- affinity sites were characterized by the binding of GHB (Ki,GHB = 3.9 ± 0.5 ,M and Ki,GHB = 5.0 ± 2.0 mM) and lactate (LAC; Ki,LAC = 3.9 ± 0.5 ,M and Ki,LAC = 7.7 ± 0.9 mM). Ligands, including the hemiester ethyl-hemi-SUC, and the gap junction inhibitors flufenamate, CBX, and the GHB binding site-selective NCS-382 interacted with the high-affinity site (in ,M: Ki,EHS = 17 ± 5, Ki,FFA = 24 ± 13, Ki,CBX = 28 ± 9, Ki,NCS-382 = 0.8 ± 0.1 ,M). Binding of the Na+,K+ -ATPase inhibitor ouabain, the proton-coupled monocarboxylate transporter (MCT)-specific ,-cyano-hydroxycinnamic acid (CHC), and CIT characterized the low-affinity SUC binding site (in mM: Ki,ouabain = 0.13 ± 0.05, Ki,CHC = 0.32 ± 0.07, Ki,CIT = 0.79 ± 0.20). All tested compounds inhibited [3H]SUC binding in the human nucleus accumbens and had Ki values similar to those observed in the rat forebrain. The binding process can clearly be recognized as different from synaptic and mitochondrial uptake or astrocytic release of SUC, GHB, and/or CIT by its unique GHB selectivity. The transient decrease of extracellular SUC observed during epileptiform activity suggested that the function of the synaptic target recognizing protonated succinate monocarboxylate may vary under different (patho)physiological conditions. Furthermore, we put forward a hypothesis on the synaptic activity-regulated signaling between astrocytes and neurons via SUC protonation. © 2008 Wiley-Liss, Inc. [source] The effect of intracellular acidification on the relationship between cell volume and membrane potential in amphibian skeletal muscleTHE JOURNAL OF PHYSIOLOGY, Issue 3 2005James A. Fraser The relationship between cell volume (Vc) and membrane potential (Em) in Rana temporaria striated muscle fibres was investigated under different conditions of intracellular acidification. Confocal microscope xz -scanning monitored the changes in Vc, whilst conventional KCl and pH-sensitive microelectrodes measured Em and intracellular pH (pHi), respectively. Applications of Ringer solutions with added NH4Cl induced rapid reductions in Vc that rapidly reversed upon their withdrawal. These could be directly attributed to the related alterations in extracellular tonicity. However: (1) a slower and persistent decrease in Vc followed the NH4Cl withdrawal, leaving Vc up to 10% below its resting value; (2) similar sustained decreases in resting Vc were produced by the addition and subsequent withdrawal of extracellular solutions in which NaCl was isosmotically replaced with NH4Cl; (3) the same manoeuvres also produced a marked intracellular acidification, that depended upon the duration of the preceding exposure to NH4Cl, of up to 0.53 ± 0.10 pH units; and (4) the corresponding reductions in Vc similarly increased with this exposure time. These reductions in Vc persisted and became more rapid with Cl, deprivation, thus excluding mechanisms involving either direct or indirect actions of pHi upon Cl, -dependent membrane transport. However they were abolished by the Na+,K+ -ATPase inhibitor ouabain. The Em changes that accompanied the addition and withdrawal of NH4+ conformed to a Nernst equation modified to include realistic NH4+ permeability terms, and thus the withdrawal of NH4+ restored Em to close to control values despite a persistent change in Vc. Finally these Em changes persisted and assumed faster kinetics with Cl, deprivation. The relative changes in Vc, Em and pHi were compared to predictions from the recent model of Fraser and Huang published in 2004 that related steady-state values of Vc and Em to the mean charge valency (zx) of intracellular membrane-impermeant anions, X,i. By assuming accepted values of intracellular buffering capacity (,i), intracellular acidification was shown to produce quantitatively predictable decreases in Vc. These findings thus provide experimental evidence that titration of the anionic zx by increased intracellular [H+] causes cellular volume decrease in the presence of normal Na+,K+ - ATPase activity, with Cl, -dependent membrane fluxes only influencing the kinetics of such changes. [source] Studies on the Prevention of Nigericin Action in Neuroblastoma X Glioma Hybrid (NG108,15) Cells,BASIC AND CLINICAL PHARMACOLOGY & TOXICOLOGY, Issue 2 2000Jeffrey A. Doebler Electrophysiological analysis of neuroblastoma X glioma hybrid (NG108,15) cells was used as an in vitro neuronal model system to evaluate antagonists of the K+ -selective carboxylic ionophore, nigericin. Changes in membrane electrical characteristics induced by nigericin with and without the simultaneous administration of antagonists were measured using intracellular microelectrode techniques. Bath application of nigericin (3 ,M) produced a severe hyperpolarization and blocked the generation of action potentials in response to electrical stimulation. Simultaneous administration of nigericin plus the Na+ -K+ pump inhibitor ouabain or drugs known to influence Ca++ signaling in cells, i.e., quinidine, compound R24571, verapamil or haloperidol, was able to significantly attenuate the hyperpolarization. All antagonists acted in a concentration-dependent manner. However, nigericin plus maximally effective concentrations of ouabain (1 ,M), verapamil (3 ,M) and haloperidol (3 and 10 ,M) resulted in moderate-to-severe depolarization by the end of 24 min. superfusions, suggesting that the concentrations of antagonists were excessive and that NG108,15 cell damage had occurred. In addition, none of the compounds studied was able to effectively prevent nigericin-induced blockade of action potentials. Thus, none of these antagonists appears suitable for transition to in vivo antidotal protection studies. [source] | ||||||||||