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Effects of noradrenaline on potassium reflux, membrane potential and electrolyte levels in tissue slices prepared from guinea-pig liver.

1. Some effects of noradrenaline on potassium efflux, electrolyte levels, membrane potential and current distribution in guinea-pig liver slices have been examined.2. The slices (thickness ca. 300 mum) were prepared from the median lobe of the liver and incubated at 38 degrees C in a mammalian Ringer fluid containing 2 mM pyruvate. After an initial recovery period, the ionic composition of the tissue remained stable for several hours.3. The steady-state contents of sodium, potassium and chloride were 296, 266 and 272 m-equiv/kg dry tissue respectively. The inulin space was 29 ml./100 g wet tissue.4. Most if not all of the tissue potassium was exchangeable. The rate constant for (42)K efflux was 0.019 min(-1).5. Noradrenaline (1 muM) markedly increased the efflux of (42)K and within 2 min caused tissue potassium to fall by 8%. At the same time the sodium content rose.6. Traverses of the slices with micro-electrodes showed many negative-going deflexions of 30-40 mV in amplitude. The evidence suggests that these correspond to the membrane potentials of the parenchymal cells.7. Noradrenaline (1 muM) caused a reversible hyperpolarization of about 10 mV. The response became larger on replacing external chloride by isethionate or methylsulphate, but was little affected by a reduction in external potassium.8. After slices had been bathed in potassium and chloride-free solutions for several min, restoration of external potassium caused the membrane potential to increase by up to 10 mV. This hyperpolarization, but not that caused by noradrenaline, was abolished by ouabain.9. Noradrenaline reduced the amplitude and quickened the time course of electrotonic potentials set up by current pulses from another microelectrode, suggesting that the membrane conductance had risen.10. Although certain mechanisms based on electrogenic active transport processes with unusual properties have not been excluded, the present findings are more simply explained by supposing that noradrenaline increases the potassium permeability of the parenchymal cell membrane.

Animals↗

Regulation of hepatic Na(+)-HCO3- cotransport and pH by membrane potential difference.

Hepatocytes possess several mechanisms for membrane acid-base transport, which work in concert to maintain intracellular pH (pHi) in a narrow physiological range, despite metabolic processes that produce and consume substantial quantities of H+ and HCO3-.Na(+)-H+ and Cl(-)-HCO3- exchangers contribute to recovery from intracellular acidosis and alkalosis, respectively, but are largely inoperative at physiological values of pHi. Recent studies indicate that hepatocytes also possess a mechanism for coupled transport of Na+ and HCO3- across the basolateral membrane. This appears to be the dominant pathway for membrane acid-base transport operative under basal conditions, mediates influx of Na+ and HCO3-, and is an important contributor to recovery from intracellular acidosis. In this review, the properties of hepatic Na(+)-HCO3- cotransport are described with emphasis on its effects on pHi and Na+ homeostasis and on the possible role of membrane potential difference as a signal modulating the rate of HCO3- influx and pHi of hepatocytes through effects on this transporter.

Animals↗

The timing of phasic transmitter release is Ca2+-dependent and lacks a direct influence of presynaptic membrane potential.

Ca2+ influx through voltage-gated Ca2+ channels and the resulting elevation of intracellular Ca2+ concentration, [Ca2+]i, triggers transmitter release in nerve terminals. However, it is controversial whether in addition to the opening of Ca2+ channels, membrane potential directly affects transmitter release. Here, we assayed the influence of membrane potential on transmitter release at the calyx of Held nerve terminals. Transmitter release was evoked by presynaptic Ca2+ uncaging, or by presynaptic Ca2+ uncaging paired with presynaptic voltage-clamp depolarizations to +80 mV, under pharmacological block of voltage-gated Ca2+ channels. Such a change in membrane potential did not alter the Ca2+ dependence of transmitter release rates or synaptic delays. We also found, by varying the amount of Ca2+ influx during Ca2+ tail-currents, that the time course of phasic transmitter release is not invariant to changes in release probability. Rather, the time difference between peak Ca2+ current and peak transmitter release became progressively shorter with increasing Ca2+ current amplitude. When this time difference was plotted as a function of the estimated local [Ca2+]i at the sites of vesicle fusion, a slope of approximately 100 micros per 10 microM [Ca2+]i was found, in reasonable agreement with a model of cooperative Ca2+ binding and vesicle fusion. Thus, the amplitude and time course of the [Ca2+]i signal at the sites of vesicle fusion controls the timing and the amount of transmitter release, both under conditions of brief periods of Ca2+ influx, as well as during step-like elevations of [Ca2+]i produced by Ca2+ uncaging.

Action Potentials↗

Measurements of membrane potentials in plant mitochondria with the safranine method.

The positively charged dye, safranine, has been used as an indicator of membrane potentials in mung bean (Phaseolus aureus) and Voodoo lily (Sauromatum guttatum) mitochondria under a variety of metabolic conditions. The spectral response of safranine has been calibrated with respect to a K(+) diffusion potential and was found to be linearly related to the developed potential within the range of 50 to 160 millivolts. Both respiration and ATP hydrolysis gave rise to a membrane potential of approximately 135 millivolts. Respiratory inhibitors such as cyanide and antimycin depolarized the potential, whereas rotenone has little effect. No potentials were developed during NADH supported cyanide insensitive respiration. It is concluded that safranine may be a useful spectrophotometric probe of the mitochondrial membrane potential.

Journal Article↗

Electrophysiological properties of Dictyostelium derived from membrane potential measurements with microelectrodes.

Electrical membrane properties of the cellular slime mold Dictyostelium discoideum were investigated with the use of intracellular microelectrodes. The rapid potential transients (1 msec) upon microelectrode penetration of normal cells had a negative-going peak-shaped time course. This indicates that penetration of a cell with a microelectrode causes a rapid depolarization, which can just be recorded by the microelectrode itself. Therefore, the initial (negative) peak potential transient value Ep (-19mV) should be used as an indicator of the resting membrane potential Em of D. discoideum before impalement, rather than the subsequent semistationary depolarized value En (-5 mV). Using enlarged cells such as giant mutant cells (Ep = -39 mV) and electrofused normal cells (Ep = -30 mV) improved the reliability of Ep as an indicator of Em. From the data we concluded that Em of D. discoideum cells bathed in (mM) 40 NaCl, 5 KCl and 1 CaCl2 is at least -50 mV. This potential was shown to be dependent on extracellular potassium. The average input resistance Ri of the impaled cells was 56 M omega for normal D. discoideum. However, our analysis indicates that the membrane resistance of these cells before impalement is greater than 1 G omega. Specific membrane capacitance was 1-3 pF/cm2. Long-term recording of the membrane potential showed the existence of a transient hyperpolarization following the rapid impalement transient. This hyperpolarization was associated with an increase in Ri of the impaled cell. It was followed by a depolarization, which was associated with a decrease in Ri. The depolarization time was dependent on the filling of the microelectrode. The present characterization of the electrical membrane properties of Dictyostelium cells is a first step in a membrane electrophysiological analysis of signal transduction in cellular slime molds.

Dictyostelium↗

Effect of hypoxia on membrane potential and resting conductance in rat hippocampal neurons.

The present patch-clamp study describes the effect of hypoxia at 30-31 degrees C on membrane potential and resting conductance in pyramidal cells from the hippocampal CA1 region in rat brain slices. The initial effect of hypoxia was a gradual hyperpolarization; the peak change in membrane potential measured over 15 min was -5.3 +/- 0.22 mV (P < 0.0001). After reoxygenation followed a transient hyperpolarization measuring -1.8 +/- 0.24 mV (P < 0.0001) and a subsequent normalization of the membrane potential, which after 5 min did not differ from its level prior to the hypoxic episode. Voltage-clamp analysis showed that the hypoxic hyperpolarization was related to an outward current at the holding potential (-60 mV) and an increase in resting conductance. The effect was not influenced by intracellular Cl- concentration, which indicated that it was not due to an inward flow of Cl- ions. The addition of tolbutamide, glibenclamide and dantrolene sodium did not affect the hypoxic hyperpolarization, neither did the presence of ATP in the pipette solution. The presence/absence of glucose in the perfusion medium did not influence the initial hyperpolarization during hypoxia; however, glucose seemed to prevent the subsequent depolarization under hypoxia. It was concluded that hypoxia caused an initial hyperpolarization of CA1 cells which was related to an increase in the resting conductance. The results did not suggest the involvement of ATP-sensitive K+ channels.

Adenosine Triphosphate↗

Neurotrophin-induced rapid enhancement of membrane potential oscillations in mesencephalic trigeminal neurons.

We have proposed that neurotrophins, in addition to their trophic actions, act as neuromodulators in the adult central nervous system. As a first step to test this hypothesis, we examined in the adult rat slice preparation whether nerve growth factor and neurotrophin-3 are capable of altering the excitability of neurons of the mesencencephalic trigeminal nucleus. In contrast to vehicle pressure microapplication, which did not evoke changes in the electrophysiological properties of these neurons, neurotrophin application produced a significant increase in amplitude of the membrane potential oscillatory activity that is observed in these cells and a significant decrease in their threshold current. The latency of these effects ranged from 2 to 80 s and the duration ranged from 2 to 11 min. Neurotrophin-3 induced a decrease in input resistance and resting membrane potential in 58% of the cells; nerve growth factor induced a decrease in input resistance and resting membrane potential in 35% of the neurons. The spike configuration and action potential afterhyperpolarization potential remained unchanged following neurotrophin application. Tetrodotoxin blocked the membrane potential oscillatory activity of trigeminal mesencephalic neurons. Neurotrophin-induced effects were not blocked by the tyrosine kinase inhibitor K-252a, whereas IgG-192, an antibody directed to the neurotrophin low-affinity receptor, enhanced excitability, as did neurotrophins. These results demonstrate that neurotrophins are capable of producing a rapid increase in the excitability of trigeminal mesencephalic neurons and suggest that their effects may be mediated by low-affinity neurotrophin receptors.

Action Potentials↗

The effect of beta-adrenergic agonists on the membrane potential of fat-cell mitochondria in situ.

1. The accumulation of [3H]methyltriphenylphosphonium by isolated fat-cells was used to estimate the membrane potential of mitochondria in situ. 2. Adrenaline caused a large decrease in the accumulation of [3H]methyltriphenylphosphonium. Mitochondria in fat-cells incubated in the presence of adrenaline had a very low calculated membrane potential. This effect was also given by isoprenaline (a beta-adrenergic agonist) and was blocked by propranolol (a beta-adrenergic antagonist). 3. The effect of isoprenaline could be partially antagonized by the use of media with high albumin concentrations. Addition of sodium oleate to saturate the fatty acid-binding sites on the albumin reversed this antagonism. 4. It is proposed that the decrease in the calculated mitochondrial membrane potential is due to the uncoupling effect of the non-esterified fatty acids released by the stimulation of lipolysis observed in the presence of beta-adrenergic agonists.

Adipose Tissue↗

Bcl-2 protects against FCCP-induced apoptosis and mitochondrial membrane potential depolarization in PC12 cells.

This report addresses the relation between Bcl-2 and mitochondrial membrane potential (DeltaPsi(m)) in apoptotic cell death. Rat pheochromocytoma (PC12) cells are differentiated into neuron-like cells with nerve growth factor (NGF). It is known that Bcl-2 can attenuate apoptosis induced by deprivation of neurotrophic factor. The protective effect of Bcl-2 has been correlated with preservation of DeltaPsi(m). Protonophores, such as carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP), collapse the proton gradient across the mitochondrial inner membrane, resulting in a complete abolition of the mitochondrial membrane potential. Based on the analysis of morphology, of phosphatidylserine exposure and of nuclear fragmentation we conclude that FCCP induces apoptosis in PC12 cells, which can be prevented by overexpression of Bcl-2. To determine whether the cytoprotective effect of Bcl-2 is due to stabilization of DeltaPsi(m), we investigated the effect of Bcl-2 on changes in DeltaPsi(m), induced by FCCP in PC12 cells. We showed that treatment with FCCP induced a reduction in DeltaPsi(m), as assessed with the lipophilic cationic membrane potential-sensitive dye JC-1, and that Bcl-2 protects against FCCP-induced changes in NGF differentiated PC12 cells. Our data indicate that Bcl-2 protects against FCCP-induced cell death by stabilizing DeltaPsi(m).

Animals↗

Fluorescent probe studies of sodium/sugar cotransport and membrane potential changes in isolated chicken enterocytes.

The fluoroprobe technique using a potential-sensitive dye, diS-C3(5), was applied to isolated enterocytes in order to correlate membrane potential changes with Na+/sugar cotransport. Cells were prepared from the chicken small intestine by enzymatic (Dispase) treatment combined with mechanical agitation. In the presence of Na+, addition of D-glucose (2 mM) to the suspension of cells doped with the dye (1.8 microM) gave rise to Na+-dependent increases in fluorescence indicative of depolarization of the cell membrane potential. The pH optimum for this response was ca. 7.3. Similar but smaller fluorescence increases were also evoked by D-galactose, alpha-methyl-D-glucoside, and 3-O-methyl-D-glucose, all known to be actively accumulated by enterocytes in a Na+-dependent fashion. These monosaccharide-evoked fluorescence changes were suppressed by lowering the extracellular Na+ concentration or by phlorizin. Monosaccharides with no reported Na+ requirement induced no such fluorescence responses. A disaccharide, maltose, elicited a smaller fluorescence increase which also was Na+ dependent and phlorizin inhibitable; this was interpreted as due to a possible supply of D-glucose by the membrane-bound disaccharidase systems. No change was observed with lactose, sucrose, or trehalose, however. The glucose-evoked fluorescence changes correlatable with depolarization were also confirmed using another carbocyanine, diO-C5(3). These results demonstrate that fluorescence from diS-C3(5) or diO-C5(3) can be a good measure for cotransport-associated membrane potential changes in isolated intestinal epithelial cells.

Animals↗

[Ion-exchanging ratio and membrane potential in the bacterium S. faecalis].

The membrane potentials (delta psi) in S. faecalis equal to -140 mV were measured by means of TPP+-electrodes. The values of delta psi remain constant under temperature variation from 37 degrees to 18 degrees C. The ion-exchanging ratio during glycolysis was 2H+: Lac-: K+ where 2H+ and Lac- are liberated from the cell. It is not excluded that an electrogenic pump exchanging 2H+ by K+ operates in the membrane of S. faecalis.

Cell Membrane↗

Ventral tegmental area afferents to the prefrontal cortex maintain membrane potential 'up' states in pyramidal neurons via D(1) dopamine receptors.

The electrophysiological nature of dopamine actions has been controversial for years, with data supporting both inhibitory and excitatory actions. In this study, we tested whether stimulation of the ventral tegmental area (VTA), the source of the dopamine innervation of the prefrontal cortex, would exert different responses depending on the membrane potential states that pyramidal neurons exhibit when recorded in vivo, and whether VTA stimulation would have a role in controlling transitions between these states. Prefrontal cortical neurons have a very negative resting membrane potential (down state) interrupted by plateau depolarizations (up state). Although the up state had been shown to be dependent on hippocampal afferents in nucleus accumbens neurons, our results indicate that neither hippocampal nor thalamic inputs are sufficient to drive up events in prefrontal cortical neurons. Electrical VTA stimulation resulted in a variety of actions, in many cases depending on the neuron membrane potential state. Trains of stimuli resembling burst firing evoked a long-lasting transition to the up state, an effect blocked by a D(1) antagonist and mimicked by chemical VTA stimulation. These results indicate that projections from the VTA to the prefrontal cortex may be involved in controlling membrane potential states that define assemblies of activable pyramidal neurons in this region.

Animals↗

Plasma membrane potential modulates chemotactic peptide-stimulated cytosolic free Ca2+ changes in human neutrophils.

The relationship between fMet-Leu-Phe-induced changes in the cytosolic free Ca2+ concentration [( Ca2+]i), plasma membrane potential depolarization, and metabolic responses was studied in human neutrophils. Receptor-activated depolarization occurred both at high and resting [Ca2+]i, but was inhibited at very low [Ca2+]i. Phorbol 12-myristate 13-acetate-induced plasma membrane depolarization, on the contrary, was independent of [Ca2+]i. The threshold fMet-Leu-Phe concentration for plasma membrane depolarization (10(-8) M) was at least 1 log unit higher than that for [Ca2+]i increases (5 X 10(-10) M) and coincident with that for NADPH oxidase activation. Nearly maximal [Ca2+]i increases were elicited by 3 X 10(-9) fMet-Leu-Phe in the absence of any significant plasma membrane potential change. This observation allowed us to investigate the effects of artificially induced plasma membrane depolarization and hyperpolarization at low fMet-Leu-Phe concentrations (10(-9) to 3 X 10(-9) M) which did not perturb plasma membrane potential. Depolarizing (gramicidin D at 10(-7) to 10(-6) M or KCl at 50 mM) and hyperpolarizing (valinomycin at 4 microM) treatments had little influence on unstimulated [Ca2+]i levels, whereas fMet-Leu-Phe-induced transients were significantly altered. Gramicidin D and KCl decreased the fMet-Leu-Phe-induced [Ca2+]i increases in Ca2+-containing or in Ca2+-free media. Valinomycin, on the contrary, increased receptor-stimulated [Ca2+]i increases, and the effect was larger in the presence of extracellular Ca2+. Valinomycin also strongly potentiated secretion. It is suggested that plasma membrane depolarization in human neutrophils is a physiological feedback mechanism inhibiting receptor-dependent [Ca2+]i changes.

Calcium↗

Membrane potential of mitochondrial measured with microelectrodes.

The membrane potentials of giant mitochondria from cuprizone-fed mice were found to be independent of metabolic state. Experiments are described in which the presence of the microelectrodes in the inner mitochondrial space, and the metabolic viability of the impaled mitochonidra, are validated.

Animals↗

Platelet membrane potential: simultaneous measurement of diSC3(5) fluorescence and optical density.

The role of membrane potential in the activation of human platelets by thrombin, ADP and PAF was assessed, using the fluorescent probe diSC3(5). Thrombin, ADP and PAF transiently depolarised the platelet membrane by 6-8 mV from its resting level (-70 mV). This depolarisation had a similar time course to that of shape change. The ionophores valinomycin and gramicidin hyperpolarised and depolarised the platelets respectively but did not activate them. In contrast, exposure of platelets to high K+ media both depolarised and caused them to change shape. Removal of Na+ from the suspension media abolished the depolarisation induced by thrombin, ADP and PAF but the platelets under these conditions were still capable of changing shape and aggregating. This result indicates that the observed depolarisation depends on Na+ fluxes. Amiloride or tetrodotoxin did not mimic the effect of Na+ removal suggesting that any Na+ movement involved does not go through the classic "Na+ channel". Thrombin, ADP and PAF still depolarised the platelet membrane in the absence of added Ca++. Under these conditions, however, the membrane did not repolarise. It is evident that all three agents, thrombin, ADP and PAF, change the membrane potential of human washed platelets through a similar mechanism and this change seems to be a consequence of stimulus-receptor interaction (and platelet activation?). A causal relationship however between these events cannot be clearly shown.

Adenosine Diphosphate↗

Effects of palmitoyl CoA and palmitoyl carnitine on the membrane potential and Mg2+ content of rat heart mitochondria.

Palmitoyl CoA and palmitoyl carnitine added to rat heart mitochondria in amounts above 20 and 50 nmoles/mg protein, respectively, induced a fall in transmembrane potential and loss of endogenous Mg2+. The dissipation of membrane potential by low concentrations of palmitoyl CoA in the presence of Ca2+, but not that of high concentrations of palmitoyl CoA alone, was prevented by either ruthenium red, Cyclosporin A or Mg2+, but reversed only by Mg2+. The fall of membrane potential induced by palmitoyl carnitine was not prevented by any of these factors. It is suggested that the action of both palmitoyl CoA and palmitoyl carnitine at high concentrations is due to a non specific disruption of membrane architecture, while that of low concentrations of palmitoyl CoA in the presence of Ca2+ is associated specifically with energy dissipation due to Ca2+ cycling.

Animals↗

[Phosphorylation of creatine and the membrane potential of heart mitochondria].

The method of penetrating ions was used to measure the membrane potential and to study the creatine phosphokinase reaction in rat heart mitochondria. The concentration of adenine nucleotides in the intermembrane space at different levels of the membrane potential was estimated. The role of the diffusional layer adjacent to the inner membrane of heart mitochondria for coupling of the creatine phosphokinase reaction with oxidative phosphorylation is discussed.

Adenine Nucleotides↗

Extracellular calcium, contractile activity and membrane potential in tail arteries from genetically hypertensive rats.

OBJECTIVE AND DESIGN: This study compares the effect of extracellular calcium on contractile responsiveness and membrane potential (E(m)) in arteries from stroke-prone spontaneously hypertensive rats (SHRSP) and Wistar-Kyoto normotensive (WKY) rats. METHODS: Isometric force and E(m) were measured in isolated tail artery strips using standard muscle bath and microelectrode techniques, respectively. RESULTS: The resting contractile force in SHRSP and WKY arteries was not influenced by the extracellular calcium concentration. However, the rate of force development in response to norepinephrine (3 x 10(-8) mol/l) was slowed when calcium was elevated and increased when calcium was reduced. Compared with WKY rats, this stabilizing action of calcium on contractions to norepinephrine was reduced in SHRSP. In 1.6 mmol/l calcium, resting E(m) in SHRSP did not differ from that in WKY rats. Calcium-free buffer caused depolarization in SHRSP and WKY rats. Reductions in calcium below physiological levels resulted in depolarization, whereas elevations in calcium caused hyperpolarization. Regardless of the calcium concentration, E(m) values in SHRSP did not differ from those in WKY rats. Norepinephrine (3 x 10(-8) mol/l) caused a depolarization in WKY rat and SHRSP arteries, and the magnitude of this depolarization was not influenced by calcium. Endothelium removal did not alter the stabilizing effects of calcium on the membrane potential or contractile activity in WKY rats or SHRSP. CONCLUSIONS: The reduced stabilizing effect of calcium on the contractile activity in SHRSP arteries is not due to an alteration in the general effect of the cation on the membrane potential.

Animals↗