Electromechanical uncoupling in a molluscan muscle examined by the sucrose gap technique

Summary Membrane potential and tension ofBusycon radular protractor muscles were studied by sucrose gap methods.Excitation-contraction (EC) coupling was examined in response to acetylcholine (ACh) and high K which depolarized the fibres and induced tension, but without action potential firing. Potassium depolarization did not follow predictions expected from the Nernst equation at low and very high K levels, and maximum tension was found at about 100 mM K. EC coupling was very sensitive to [Ca]_o. Ca-free media eliminated K- and ACh-induced tension but with normal depolarization, showing full electromechanical uncoupling.Ionophore A23187 enhanced K- and ACh-induced responses and X-537A enhanced ACh responses, demonstrating acute dependence of activation on [Ca]_o in this muscle. The calcium antagonists nifedipine and nisoldipine reduced tension in the muscle only at very high concentrations, and both agents slightly reduced K- and ACh-induced depolarization.Verapamil reduced K- and ACh-induced tension but paradoxically it enhanced the depolarizing actions of these agents leading to electromechanical uncoupling. Abscisic acid (ABA) enhanced ACh- and K-induced tension and simultaneously enhanced their depolarizing actions. Ionophores and ABA appear to enhance calcium influx which may secondarily influence sodium influx.Calcium antagonists have no consistent actions on this muscle, suggesting that calcium channel activity of the radular protractor may be different from that seen in mammalian visceral muscles.Abbreviations ABRM Anterior byssus retractor muscle - ACh acetylcholine - ABA abscisic acid - EC excitation-contraction - SR sarcoplasmic reticulum - EGTA ethylene-diamine-tetraacetic acid     

The electrical response of Phaseolus vulgaris roots to abrupt exposure to hydroquinone

Previous reports have suggested the primary mode of action of the allelochemical hydroquinone involves disruption of root cell membrane transport. Here we report the effects of hydroquinone on common bean (Phaseolus vulgaris) plants. Growth of leaves, roots and stems were all inhibited by 14 day exposure to 0.01 mM or 0.25 mM hydroquinone. Chlorophyll fluorescence (Fv/Fm) was inhibited by 0.25 mM hydroquinone. The membrane potential of P. vulgaris root cortex cells briefly hyperpolarized and subsequently slowly transiently depolarized upon abrupt exposure to a range of hydroquinone concentrations. Both the hyperpolarization and depolarization were concentration dependent but appeared saturable. Root cells exposed to 0.03 mM hydroquinone hyperpolarized 3.4 mV (+/− 0.6 s.e.) 3 minutes after the start of exposure then depolarized 36.7 mV (+/− 3.9) with no effect evident after 24 hours. Individual recordings showed a response to as little as 0.001 mM hydroquinone. Exposure of P. vulgaris root cells to arbutin, a nontoxic monoglucoside of hydroquinone, produced a similar but much smaller (approximately 25%) electrical response. Exposure of root cells of Antennaria microphylla, a known allelopathic source (donor plant) of hydroquinone, also produced a much smaller hyperpolarization and depolarization response. It is concluded that the electrical response to hydroquinone by P. vulgaris root cells and the changes in membrane transport they represent are not sufficiently large or long lasting enough to disrupt mineral and water uptake leading to plant injury. The possibility, however, that these events are related to initiation of signal transduction events leading to cell death is discussed.Key words: allelopathy, hydroquinone, membrane potential, depolarization, hyperpolarization, Phaseolus vulgaris, Antennaria microphylla     

Effect of various ionic compositions upon the membrane potentials during activation of sea urchin eggs

The membrane potentials of sea urchin (Hemicentrotus pulcherrimus) eggs before and after fertilization and their changes during the membrane elevation induced by intracellular electrical stimulation were recorded in solutions of various ionic compositions. Upon fertilization, the membrane potential (?10 mV) depolarized and reversed polarity by a few mV, then gradually returned to a new steady level ranging between ?50 and ?60 mV. The activation potential is closely associated with a transient increase in the membrane permeability. The potential of the unfertilized egg is hyperpolarized by monovalent anions (Br^?, Cl^? and NO₃^?) and depolarized slightly by K⁺. In contrast, the membrane of the fertilized egg is markedly depolarized by K⁺. Suppression of depolarization associated with an increase of the membrane permeability was recorded in Na-free medium (Tris-HCl). The selective increase in permeability to monovalent anions is thought to alternate with the selective increase in permeability to K⁺through the mediation of a transient increase of Na⁺-permeability at the time of fertilization. No causal relationship between the membrane elevation and the depolarization was established because the breakdown of the cortical granules occurs without depolarization or an increase in membrane permeability.     

Muscle Contraction during Hyperpolarizing Currents in the Crab

Isolated muscle fibers from the motor legs of the crab Trichodactilus dilocarcinus were submitted to strong hyperpolarizing currents of varied intensities which produced tension during the current pulse. Threshold for tension was obtained with intensities of about 0.2 x 10^–5 A, changing E_m to ca. –150 mV (starting from a resting potential ofca. –80 mV). At the closure of the anodic square pulse, a second phase of tension usually appeared superimposed upon the one obtained during hyperpolarization. The first phase of tension increased with the increase of Ca⁺⁺ concentration in the bath. Sr⁺⁺ produced the same type of mechanical output as Ca⁺⁺. When added to the normal Ca⁺⁺ concentration, Ba⁺⁺ and Mn⁺⁺ in low concentrations (up to 21.5 mM) also increased the tension of this phase, but at higher concentrations they blocked both phases while Mg⁺⁺ did not alter the tension. Of all the divalent cations employed, only Sr⁺⁺ is capable of developing tension as a substitute for Ca⁺⁺ in the external media. Procaine administered in a dosage (5 x 10^–3 W/V)which would suppress the contracture due to caffeine (10 mM), did not modify the tension developed during the hyperpolarization. The preceding data indicate that the Ca⁺⁺ required for tension during hyperpolarization comes from sites which would differ from those usually postulated for tension due to depolarization in the muscle fibers of other crustaceans (American crayfish). Furthermore, the external source of Ca⁺⁺ appears to be one mainly implicated in the induction of tension due to inward current pulses.     

High K(+)-induced membrane depolarization attenuates endothelium-dependent pulmonary vasodilation

Impairment of endothelium-dependent pulmonary vasodilation has been implicated in the development of pulmonary hypertension. Pulmonary vascular smooth muscle cells and endothelial cells communicate electrically through gap junctions; thus, membrane depolarization in smooth muscle cells would depolarize endothelial cells. In this study, we examined the effect of prolonged membrane depolarization induced by high K(+) on the endothelium-dependent pulmonary vasodilation. Isometric tension was measured in isolated pulmonary arteries (PA) from Sprague-Dawley rats, and membrane potential was measured in single PA smooth muscle cells. Increase in extracellular K(+) concentration from 4.7 to 25 mM significantly depolarized PA smooth muscle cells. The 25 mM K(+)-mediated depolarization was characterized by an initial transient depolarization (5-15 s) followed by a sustained depolarization that could last for up to 3 h. In endothelium-intact PA rings, ACh (2 microM), levcromakalim (10 microM), and nitroprusside (10 microM) reversibly inhibited the 25 mM K(+)-mediated contraction. Functional removal of endothelium abolished the ACh-mediated relaxation but had no effect on the levcromakalim- or the nitroprusside-mediated pulmonary vasodilation. Prolonged ( approximately 3 h) membrane depolarization by 25 mM K(+) significantly inhibited the ACh-mediated PA relaxation (-55 +/- 4 vs. -29 +/- 2%, P < 0.001), negligibly affected the levcromakalim-mediated pulmonary vasodilation (-92 +/- 4 vs. -95 +/- 5%), and slightly but significantly increased the nitroprusside-mediated PA relaxation (-80 +/- 2 vs. 90 +/- 3%, P < 0. 05). These data indicate that membrane depolarization by prolonged exposure to high K(+) concentration selectively inhibited endothelium-dependent pulmonary vasodilation, suggesting that membrane depolarization plays a role in the impairment of pulmonary endothelial function in pulmonary hypertension.     

Mechanical and electrical effects of high-frequency and high-intensity stimulation of muscle

A unique tension response can be obtained by stimulating an isometrically held skeletal muscle or a single muscle fiber by a train of high-frequency pulses (2,000 pps) at higher-than-normal intensity, or by a long DC pulse. It is called the tetanoid response, and it is composed of three well-defined stages. Initially, tension develops rapidly, and mechanical output (Po) reaches about 0.35. Subsequently, this tension is maintained at a nearly steady level for the remainder of stimulation. After stimulation, a final increase of tension takes place. Intracellular electrical recordings show that the initial development of tension is elicited by two or three action potentials generated at the beginning of the stimulation, and that no additional action potentials are generated for the remainder of stimulation. During stimulation, part of the fiber membrane (regarded in cross-section) is depolarized, which generates tension, and part of the membrane is hyperpolarized. With termination of stimulation, a single action potential is elicited via anode-break excitation (ABE) on the hyperpolarized portion of the membrane, which gives rise to the final increase of tension.     

Noninactivating tension in rat skeletal muscle. Effects of thyroid hormone

Inactivation of excitation-contraction coupling was examined in extensor digitorum longus (EDL) and soleus muscle fibers from rats injected daily with tri-iodothyronine (T3, 150 micrograms/kg) for 10-14 d. Steady-state activation and inactivation curves for contraction were obtained from measurements of peak potassium contracture tension at different surface membrane potentials. The experiments tested the hypothesis that noninactivating tension is a "window" tension caused by the overlap of the activation and inactivation curves. Changes in the amplitude and voltage dependence of noninactivating tension should be predicted by the changes in the activation and inactivation curves, if noninactivating tension arises from their overlap. After T3 treatment, the area of overlap increased in EDL fibers and decreased in soleus fibers and the overlap region was shifted to more negative potentials in both muscles. Noninactivating tension also appeared at more negative membrane potentials after T3 treatment in both EDL and soleus fibers. The effects of T3 treatment were confirmed with a two microelectrode voltage-clamp technique: at the resting membrane potential (-80 mV) contraction in response to a brief test pulse required less than normal depolarization in EDL, but more than normal depolarization in soleus fibers. After T3 treatment, the increase in contraction threshold at depolarized holding potentials (attributed to inactivation) occurred at more depolarized holding potentials in EDL, or less depolarized holding potentials in soleus. The changes in contraction threshold could be accounted for by the effects of T3 on the activation and inactivation curves. In conclusion, (a) T3 appeared to affect the expression of both activation and inactivation characteristics, but the activation effects could not be cleanly distinguished from T3 effects on the sarcoplasmic reticulum and contractile proteins, and (b) the experiments provided evidence for the hypothesis that the noninactivating tension is a steady-state "window" tension.     

Endothelium inhibits the palytoxin-induced depolarization and Ca2+ mobilization in porcine coronary artery through endothelium-derived hyperpolarizing factor and nitric oxide released by palytoxin

Palytoxin induced increases in cytosolic Ca²⁺ and tension, which were dependent on external Ca²⁺, and depolarized the membrane in endothelium-denuded porcine coronary arteries. When the endothelium was present, however, these effects were greatly inhibited, suggesting that some factors from endothelium inhibited the palytoxin-actions. Pretreatment with 100 μM N^ω-nitro-L-arginine partially reversed the inhibitory effect of endothelium on the Ca²⁺ movement and the contraction but not that on the depolarization. Pretreatment with 10 μM indomethacin did not affect the inhibition. These results suggest that palytoxin released both nitric oxide and endothelium-derived hyperpolarizing factor (EDHF) from the endothelium, both of which counteracted the actions of palytoxin on smooth muscle cells. It is thought that the palytoxin-induced depolarization was attenuated by hyperpolarization due to EDHF.     

Spontaneous action potentials and resting potential shifts in fertilized eggs of the tunicate Clavelina

Electrical activity in the fertilized egg of the tunicate Clavelina was studied with microelectrode recording and voltage clamp techniques. The resting potential could assume either of two stable values (approximately ?70 or ?30 mV) and could be shifted between these values by direct current stimulation. Spontaneous shifts between two stable resting potentials were also seen. Egg cells produced action potentials spontaneously and in response to depolarizing stimuli. Inward currents were carried by both Na and Ca ions and a prominent outward potassium current was seen with depolarization to voltages above ?15 mV. The steady-state current-voltage relationship (I–V curve) of the membrane showed two voltages where the net membrane current equaled zero: approximately ?35 and ?70 mV. Between these two voltages, membrane current was inward and carried by noninactivating Na and Ca currents. Inward rectification, which was blocked by external Rb, occurred at voltages below ?70 mV. The voltage dependence of inward rectification is thought by the authors to be important for establishing the more negative resting potential; it is also thought the presence of inward current which does not inactivate completely at voltages more negative than about ?20 mV is an important determinant of the more depolarized resting potential.     

Prolonged membrane potential depolarization in cingulate pyramidal cells after digit amputation in adult rats

The anterior cingulate cortex (ACC) plays an important role in higher brain functions including learning, memory, and persistent pain. Long-term potentiation of excitatory synaptic transmission has been observed in the ACC after digit amputation, which might contribute to plastic changes associated with the phantom pain. Here we report a long-lasting membrane potential depolarization in ACC neurons of adult rats after digit amputation in vivo. Shortly after digit amputation of the hind paw, the membrane potential of intracellularly recorded ACC neurons quickly depolarized from ~-70 mV to ~-15 mV and then slowly repolarized. The duration of this amputation-induced depolarization was about 40 min. Intracellular staining revealed that these neurons were pyramidal neurons in the ACC. The depolarization is activity-dependent, since peripheral application of lidocaine significantly reduced it. Furthermore, the depolarization was significantly reduced by a NMDA receptor antagonist MK-801. Our results provide direct in vivo electrophysiological evidence that ACC pyramidal cells undergo rapid and prolonged depolarization after digit amputation, and the amputation-induced depolarization in ACC neurons might be associated with the synaptic mechanisms for phantom pain.     

The Mode of Transverse Spread of Contraction Initiated by Local Activation in Single Crayfish Muscle Fibers

Isolated single crayfish muscle fibers were locally activated by applying negative current pulses to a pipette whose tip was in contact with the fiber surface. The contraction initiated by a moderate depolarization spread inwards in a graded manner according to the magnitude and duration of depolarization. Increase of the depolarized area increased the distance of the inward spread for a given amount of depolarization. If a large area of the surface membrane was depolarized with a large pipette for a sufficiently long time, the contraction spread not only inwards, but further transversely passing through the center of the fiber. Successive brief depolarizations given at an appropriate interval could produce contraction more effectively for a given amount of total current than did a prolonged depolarization. On the other hand, the contraction initiated by a strong negative current was observed to spread around the whole perimeter but not through the center of the fiber. Each type of local contraction always spread along the striation pattern and not longitudinally. Possible mechanisms of these responses are discussed in connection with the transverse tubular system of the muscle fibers.     

Effects of conditioning polarization on the membrane ionic currents in rat myometrium

Summary Membrane ionic currents were measured in pregnant rat uterine smooth muscle under voltage clamp conditions by utilizing the double sucrose gap method, and the effects of conditioning pre-pulses on these currents were investigated. With depolarizing pulses, the early inward current was followed by a late outward current. Cobalt (1mm) abolished the inward current and did not affect the late outward currentper se, but produced changes in the current pattern, suggesting that the inward current overlaps with the initial part of the late outward current. After correction for this overlap, the inward current reached its maximum at about +10 mV and its reversal potential was estimated to be +62 mV. Tetraethylammonium (TEA) suppressed the outward currents and increased the apparent inward current. The increase in the inward current by TEA thus could be due to a suppression of the outward current. The reversal potential for the outward current was estimated to be –87 mV. Conditioning depolarization and hyperpolarization both produced a decrease in the inward current. Complete depolarization block occurred at a membrane potential of –20 mV. Conditioning hyperpolarization experiments in the presence of cobalt and/or TEA revealed that the decrease in the inward current caused by conditioning hyperpolarization was a result of an increase in the outward current overlapping with the inward current. It appears that a part of the potassium channel population is inactivated at the resting membrane potential and that this inactivation is removed by hyperpolarization.     

Membrane potentials in respiring and respiration-deficient yeasts monitored by a fluorescent dye

Changes in fluorescence of 3,3′-dipropylthiodicarbocyanine iodide which had been equilibrated with suspensions of the wild-type yeast Saccharomyces cerevisiae and of respiration-deficient mutants were followed. The changes have been attributed to changes of yeast membrane potentials, since the fluorescence with wild-type yeast could be affected in a predictable manner by uncouplers and the pore-forming agent nystatin. As in other systems, a rise of steady-state fluorescence was ascribed to depolarization and a drop of the fluorescence to hyperpolarization. (1) A considerable rise in steady-state fluorescence was brought about by addition of antimycin A or some other mitochondrial inhibitors to respiring cells. A major part of the composite membrane potential monitored in intact yeast cells appeared to be represented by the membrane potential of mitochondria. (2) Addition of D-glucose and of other substrates of hexokinase, including non-metabolizable 2-deoxy-D-glucose, induced a two-phase response of fluorescence, indicating transient depolarization followed by repolarization. Such a response was not elicited by other sugars which had been reported to be transported into the cells by a glucose carrier or by D-galactose in galactose-adapted cells. The depolarization was explained by electrogenic ATP exit from mitochondria to replenish the ATP consumed in the hexokinase reaction and the repolarization by subsequent activation of respiration. (3) In non-respiring cells only a drop in fluorescence was induced by glucose and this was ascribed to an ATP-dependent polarization of the plasma membrane. (4) Steady-state fluorescence in suspensions of respiration-deficient mutants, lacking cytochrome a, cytochrome b, or both, was high and remained unaffected by uncouplers and nystatin. This indicates that membranes of the mutants may have been entirely depolarized. A partial polarization, apparently restricted to the plasma membrane, could be achieved by glucose addition.     

Influence of Phenolic Acids on Ion Uptake: IV. Depolarization of Membrane Potentials

The membrane potentials of aged, excised barley (Hordeum vulgare L.) root cells were rapidly depolarized by the addition of salicylic acid (o-hydroxybenzoic acid) to the buffered medium bathing root segments. Initial values for membrane potentials were restored very slowly (within 100 minutes) by replacing the phenolic solution by phenolic-free buffer. Several other naturally occurring benzoic and cinnamic acids depolarized cell membrane potentials. The cinnamic acids consistently caused a greater depolarization than the correspondingly substituted benzoic acids. A strong positive correlation was found between the depolarization values (ΔE) for the benzoic acids and their lipid solubilities. This study supports the hypothesis that the inhibition of ion uptake brought about by naturally occurring phenolic acids is caused by a generalized increase in membrane permeability to inorganic ions.     

Effects of External Calcium Deprivation on Single Muscle Fibers

Deprivation of external calcium causes sudden potentiation of the twitch response of single muscle fibers. The potentiation was 64 ± 8%. Potentiation is simultaneous with membrane depolarization occurring after Ca⁺⁺ removal. This depolarization amounted to 9 ± 2 mv. Ca⁺⁺ removal also alters the action potential. 3 min after calcium withdrawal, action potential amplitude fell by 36 ± 3 mv; maximum rates of rise and fall of the spike decreased by 55 ± 5 and 63 ± 5% respectively. Changes in shape of the A. P. differ from those seen with other potentiators of the twitch response, such as Zn⁺⁺. After short exposure to calcium-free media, potassium-induced contractures show potentiation of peak tension. The S-shaped curve relating potassium contracture tension to log [K]_o shifts to the left after such treatment. Calcium deprivation also increased the rate of relaxation of the contractures. This effect depends on the duration of calcium deprivation, and is probably related to the effect of calcium lack on the membrane. The change in relaxation occurred immediately after calcium deprivation, and was reversed by sudden readmission of calcium. Relaxation of twitch and tetanus responses also were affected by Ca lack, but not as rapidly as potassium contractures. The results suggest that external calcium is not directly involved in the process responsible for tension development, supporting the view that this process is mediated by translocation of intracellular calcium. The relaxation process, however, appears to be rapidly affected by deprivation of external calcium.     

Effects of Inhibitors on the Light-Induced Changes in Electric Potential in Pulvinar Motor Cells of Phaseolus vulgaris L.

To analyze the mechanism of the light-induced changes in electricpotential in motor cells of the pulvinus of Phaseolus vulgarisL., inhibitors were applied to the pulvinus by the xylem perfusionmethod. The membrane potential was –60 to –80 mV,which indicated that the polarization was less than that ofcells of a pulvinus in air. A pulse (30 s) of blue light (BL)induced transient depolarization of the membrane in the motorcells. Red light (RL) caused hyperpolarization of the membrane.The magnitude of BL pulse-induced transient depolarization wasgreater under the hyperpolarized state caused by the RL. The membrane was depolarized to –30 to –40 mV onperfusion with the respiratory inhibitor NaN₃ (1 mM) and a pulseof BL or irradiation with RL did not cause any change in thepotential in the depolarized state. Hyperpolarization of themembrane by RL was inhibited by perfusion with DCMU (50 µM),an inhibitor of electron transport in photosynthesis. However,the magnitude of the depolarization induced by the pulse ofBL was not affected. Perfusion with a proton ionophore CCCP(100µM) depolarized the membrane and no change in thepotential was induced by a pulse of BL or by RL in the depolarizedstate. The extent of the BL pulse-induced depolarization of the membranewas proportional to the magnitude of the membrane potentialat the time of which the pulse of BL was applied. It is suggestedthat the active component of the membrane potential was inhibitedby the pulse of BL. The experimental results further supportthe hypothesis that BL inhibits the activity of the proton ATPaseand, thus, causes loss of the electrogenic component of themembrane potential of the pulvinar motor cells. (Received June 22, 1992; Accepted August 24, 1992)     

The Role of Sodium Current in the Radial Spread of Contraction in Frog Muscle Fibers

The membrane potential of isolated muscle fibers was controlled with a two-electrode voltage clamp, and the radial extent of contraction elicited by depolarizing pulses of increasing magnitude was observed microscopically. Depolarizations of the fiber surface only 1–2 mv greater than the contraction threshold produced shortening throughout the entire cross-section of the muscle fiber. The radial spread of contraction was less effective in fibers exposed to tetrodotoxin or to a bathing medium with a greatly reduced sodium concentration. The results provide evidence that depolarization of a muscle fiber produces an increase in sodium conductance in the T tubule membrane and that the resultant sodium current contributes to the spread of depolarization along the T system.     

Ionic interconversion of pacemaker and nonpacemaker cultured chick heart cells

Trypsin-dispersed cells from hearts (ventricles) of 7 to 8 day chick embryos were cultured 3 to 21 days. The cells became attached to the culture dish and assembled into monolayer communities. By means of a bridge circuit, one microelectrode was used for simultaneously passing current and recording membrane potentials (V_m). The input resistance, calculated by the measured ΔV_m for a known step of current, averaged 10 MΩ. Electrotonic depolarization of nonpacemaker cells had no effect on frequency of firing. Within 2 min after addition of Ba⁺⁺ (5 to 10 mM) to the Tyrode bath, the cells became partially depolarized and quiescent nonpacemaker cells developed oscillations in V_m which led to action potentials. With time, the depolarization became nearly complete and the input resistance increased 2 to 10 times. During such sustained depolarizations, action potentials were no longer produced and often tiny oscillations were observed; however, large action potentials developed during hyperpolarizing pulses. Thus, the automaticity of the depolarized cell became apparent during artificial repolarization. Sr⁺⁺ (5 to 10 mM) initially produced hyperpolarization and induced automaticity in quiescent nonpacemaker cells. Elevated [K⁺]_o (20 to 30 mM) suppressed automaticity of pacemaker cells and decreased R_m concomitantly. Thus, Ba⁺⁺ probably converts nonpacemaker cells into pacemaker cells independently of its depolarizing action. Ba⁺⁺ may induce automaticity and depolarization by decreasing g _K, and elevated [K⁺]_o may depress automaticity by increasing g _K. The data support the hypothesis that the level of g _K determines whether a cell shall function as a pacemaker.     

The effect of ions upon the response of smooth muscle to cooling

The slow tonic responses of the anterior byssus retractor of Mytilus edulis to rapid cooling were investigated by simultaneously recording tension and resting potential changes after soaking the muscle in banthine, a powerful neuromuscular blocking agent. The quantitative relations between the amount of cooling and the amount of associated depolarization necessary for contraction at various concentrations of potentiating potassium can be expressed in a family of curves. The plateaus of the curves for sea water and for potassium-free sea water were beneath the depolarization value necessary for contraction, so that it is clear that no amount of cooling with sea water alone or with potassium-free sea water would ever be effective. When the muscle is treated with subthreshold amounts of potassium and rapidly cooled in various concentrations of sodium ion and calcium ion, respectively, the sodium and calcium do not affect the amount of depolarization. Acetylcholine, in subthreshold amounts, has a potentiating effect, but, unlike potassium and cooling, acts through the nervous apparatus. Mytilus muscle will respond to cooling with tonic contraction whenever a critical threshold amount of depolarization is achieved. Cooling alone cannot trigger the contraction since it cannot bring about sufficient depolarization. Cooling can result in contraction, however, if used in conjunction with some other subthreshold depolarizing agent. Cooling affects the contractile mechanism by first causing membrane breakdown and depolarization.     

Early changes in membrane potential of Saccharomyces cerevisiae induced by varying extracellular K+, Na+ or H+ concentrations

Recently we introduced a fluorescent probe technique that makes possible to convert changes of equilibrium fluorescence spectra of 3,3’-dipropylthiadicarbocyanine, diS-C₃(3), measured in yeast cell suspensions under defined conditions into underlying membrane potential differences, scaled in millivolts (Plasek et al. in J Bioenerg Biomembr 44: 559–569, 2012). The results presented in this paper disclose measurements of real early changes of plasma membrane potential induced by the increase of extracellular K⁺, Na⁺ and H⁺ concentration in S. cerevisiae with and without added glucose as energy source. Whereas the wild type and the ?tok1 mutant cells exhibited similar depolarization curves, mutant cells lacking the two Trk1,2 potassium transporters revealed a significantly decreased membrane depolarization by K⁺, particularly at lower extracellular potassium concentration [K⁺]_out. In the absence of external energy source plasma membrane depolarization by K⁺ was almost linear. In the presence of glucose the depolarization curves exhibited an exponential character with increasing [K⁺]_out. The plasma membrane depolarization by Na⁺ was independent from the presence of Trk1,2 transporters. Contrary to K⁺, Na⁺ depolarized the plasma membrane stronger in the presence of glucose than in its absence. The pH induced depolarization exhibited a fairly linear relationship between the membrane potential and the pH_o of cell suspensions, both in the wild type and the Δtrk1,2 mutant strains, when cells were energized by glucose. In the absence of glucose the depolarization curves showed a biphasic character with enhanced depolarization at lower pH_o values.