Intracellular Sodium Activity Research Articles

Electroneutral Na+/Cl- cotransport activity of zebrafish Slc12a10.1 expressed in Xenopus oocytes.

Na+/Cl- cotransporter 2 (Ncc2 or Slc12a10) is a membrane transport protein that belongs to the electroneutral cation-chloride cotransporter family. The Slc12a10 gene (slc12a10) is widely present in bony vertebrates but is deleted or pseudogenized in birds, some bony fishes, and most mammals. Slc12a10 is highly homologous to Ncc (Slc12a3 or Ncc1); however, there are only a few reports measuring the activity of Slc12a10. In this study, we focused on zebrafish Slc12a10.1 (zSlc12a10.1) and analyzed its activity using Xenopus oocyte electrophysiology. Analysis using Na+-selective microelectrodes showed that intracellular sodium activity (aNai) in zSlc12a10.1 oocytes was significantly decreased in Na+- or Cl--free medium and recovered when Na+ or Cl- was readded to the medium. Similar analysis using a Cl--selective microelectrode showed that intracellular chloride activity (aCli) in zSlc12a10.1 oocytes significantly decreased in Na+- or Cl--free medium and recovered when Na+ or Cl- was readded to the medium. When a similar experiment was performed with a voltage clamp, the membrane current did not change when aNai of zSlc12a10.1 oocytes was decreased in Na+-free medium. Molecular phylogenetic and synteny analyses suggest that gene duplication between slc12a10.2 and slc12a10.3 in zebrafish is a relatively recent event, whereas gene duplication between slc12a10.1 and the ancestral gene of slc12a10.2/slc12a10.3 occurred at least about 2 million years ago. slc12a10 deficiency was observed in species belonging to Ictaluridae, Salmoniformes, Osmeriformes, Batrachoididae, Syngnathiformes, Gobiesociformes, Labriformes, and Tetraodontiformes. These results indicate that zebrafish Slc12a10.1 is an electroneutral Na+/Cl-cotransporter and establish its evolutionary position among various teleost slc12a10 paralogs.NEW & NOTEWORTHY Na+/Cl- cotransporter 2 (Slc12a10; Ncc2) is a protein highly homologous to Ncc (Slc12a3; Ncc1); however, there are only a few reports measuring the activity of Slc12a10. Electrophysiological analysis of Xenopus oocytes expressing zebrafish Slc12a10.1 showed that Slc12a10.1 acts as an electroneutral Na+/Cl-cotransporter. This is the third report on the activity of Slc12a10, following previous reports on Slc12a10 in eels.

Augmentation and ionic mechanism of effect of β-N-methylamino- l-alanine in presence of bicarbonate on membrane potential of Retzius nerve cells of the leech Haemopis sanguisuga

The role of neurotoxic non-protein amino acid beta-N-methylamino- l-alanine (L-BMAA) as a putative causative agent of Western pacific amyotrophic lateral sclerosis/Parkinsonism dementia complex (ALS/PDC) has recently been reinvigorated. In view of this data we have investigated the strength and mechanism of effect of L-BMAA in presence of 20 mmol/L bicarbonate (a cofactor for BMAA) on membrane potential of the Leech Haemopis sanguisuga. Our results show that L-BMAA has excitatory effect in bicarbonate containing solution, which is more potent than in nominally bicarbonate free solution. This potentiation by bicarbonate is L-BMAA specific, as it was not exhibited by beta-N-oxalylamino- l-alanine. The effect of L-BMAA was partially blocked by non-NMDA receptor antagonist CNQX. Application of L-BMAA caused a decrease in input membrane resistance, an increase of intracellular sodium activity, and a decrease of intracellular potassium activity. Present findings indicate that BMAA could initiate excitotoxicity through activation of non-NMDA ionotropic glutamate receptors.

Cell Volume Regulation During Hyperosmotic Shrinkage Is Mediated by Na+/K+-ATPase and Na+–K+–2Cl− Cotransporter in Necturus Gastrics Surface Epithelial Cells

Cell volume regulation was investigated in gastric surface epithelial cells during hypertonic conditions. Isolated Necturus antral mucosa was perfused on the serosal side with Ringer's solution (pH 7.25, 95%O2/5%CO2) and on the mucosal side successively with 150-500 mM NaCl. Amiloride, ouabain, and bumetanide were used to experimentally inhibit Na+/H+, Na+/K+ ATPase or Na+-K+-2Cl- ion transporters. Intracellular sodium activity and cell volume changes were measured with liquid sensor microelectrodes. The increase in intracellular sodium activity caused by luminal hyperosmolar exposure was mainly due to cell shrinkage. Inhibition of Na+/K+ ATPase or Na+-K+-2Cl- cotransporter increased hyperosmotic cell shrinkage (-52 +/- 5%, -85 +/- 19%, and -77 +/- 9% for control, ouabain, and bumetanide, respectively). Inhibition of Na+/K+ ATPase increased intracellular sodium activity (from 18 +/- 4 to 52 +/- 12 mM). Cell volume regulation in gastric epithelial surface cells during mucosal hyperosmolar exposure is maintained by the basolateral Na+-K+-2Cl- cotransporter, while Na+/K+ ATPase maintains sodium balance, but Na+/H+ antiport seems to have a less important role.

Suppressive effect of volatile anesthetics on U wave in anesthetized patients

Although several hypotheses have been proposed to explain the late deflection immediately after T wave in the ECG [1], the cellular mechanism of the genesis of U wave remains uncertain [2]. Early and delayed afterdepolarizations, due to alteration in transmembrane ionic currents and intracellular calcium, might also play a role in the formation of U wave [3,4]. Volatile anesthetics exert antiarrhythmic actions via inhibitory effects on various transmembrane ionic currents and intracellular sodium and calcium activities in cardiomyocytes [5]. It has been demonstrated that isoflurane and halothane could suppress digitalis or catecholamine-mediated triggered activity in canine Purkinje fibers and ventricular muscles [6]. It is interesting, therefore, to see whether volatile anesthetics affect the amplitude of U wave. The dynamic changes in T–U wave using routine perioperative ECG recording (lead II) were analyzed in the 186 anesthetized gynecologic patients (ASA class I, 20–70 years old). This prospective, observational, non-invasive clinical study was approved by local IRB and informed consent was obtained from the patients. All ECG recordings were amplified (5 mm/0.1 mV) and all selected ECG strips were printed out at a paper speed of 25 mm/s. The U wave was defined as a positive or negative deflection immediately following the T wave. The amplitudes and the duration of intervals of ECG waveforms were measured manually by

Age-related effects of acidosis in isolated cardiac muscle.

Acidosis is associated with myocardial ischemia and several reports indicate the greater vulnerability of the aged heart to ischemic dysfunction. We investigated the effects of hypercapnic acidosis on isolated heart (n = 14) and papillary muscle (n = 10) from adult and senescent rats. Acidosis (pH from 7.36 to 6.91) induced a decrease in left ventricular developed pressure together with an increase in left ventricular end-diastolic pressure, but was significantly more evident in senescent than in adult hearts (p < .01). The return to normal pH induced a further increase in the end-diastolic pressure parallel to the development of arrhythmias that were greater in senescent than in adult hearts. In isolated papillary muscle, acidosis confirmed its greater negative inotropic effect on senescent than adult muscles (p < .01), while intracellular sodium activity (aNai) increased to a similar extent in both adult and senescent papillary muscles (p = NS). 5-(N,N-dimethyl)-amiloride hydrochloride (DMA), a specific inhibitor of Na+/H+ exchanger, produced similar modification of tension and aNai in both adult and senescent muscles. When DMA was superfused in acidotic solution, the contractility was markedly compromised in senescent than in adult muscles (p < .01), but the aNai modifications were similar in adult and senescent muscles (p = NS). Our results show that acidosis induced a greater reduction of contractility in senescent than in adult hearts. The similarity of contractility during DMA administration between adult and senescent muscle and of modifications of aNai suggests that depression of contractility with acidosis may be related to pathophysiologic mechanisms other than the Na+/H+ exchanger.

Interaction between Na+ and H+ ions on Na-H exchange in sheep cardiac Purkinje fibers.

The interaction between Na+ and H+ ions upon Na-H exchange (NHE) was examined in sheep cardiac Purkinje fibers. Acid equivalent fluxes through NHE were examined using recordings of intracellular pH and Na+ in isolated preparations measured with ion selective microelectrodes. The extent of acid-extrusion by NHE was estimated from pH(i) recovery-rate, multiplied by beta(i) (intracellular buffering power) in response to an internal acid load induced by 20 mm NH4Cl removal (nominally HCO3- free media). A mixed inhibitory effect was found of extracellular H+ on external Na+-activation of NHE (i.e. an increase, at low pH(o), in the apparent Michaelis constant for external Na+ ions [K(Nao)(0.5)] and a decrease in the maximum transport rate [V(Nao)(max)]). In addition, we confirmed that the stoichiometry of Na(o) binding is unaffected by the pH(o) (between 7.5 and 6.5), showing a Hill coefficient close to one. The interaction between Na+ and H+ ions at the internal face of the cardiac NHE was also studied. Our evidence suggests that an increase in the intracellular Na+ ion concentration ([Na+]i) inhibits acid efflux and that this inhibition can be approximated by the decrease in thermodynamic driving force caused by reducing the transmembrane Na+ gradient. It appears, however, that small variations in [Na+]i from the normal resting level (intracellular sodium activity, a(i)Na = 7 to 13 mm) have little or no effect on acid efflux, suggesting that variation of a(i)Na is not a physiologically important controller of NHE activity in heart.

The effect of the sodium channel blocker QX-314 on recovery after acute spinal cord injury.

There is evidence that elevated intracellular sodium ([Na+]i) activity potentiates spinal cord injury (SCI) and the hypoxic/ischemic cell death. In this study, we examined the effect of QX-314, a potent Na+ channel blocker, on recovery after SCI in vivo. QX-314 (2.0 and 10 nmol) or vehicle was microinjected (2 microL) into the injury site 15 min after SCI. Injury was performed by compression of the spinal cord at C7-T1 for 1 min with a modified aneurysm clip exerting a closing force of 35 g. Neurological function was assessed 1 day after injury and weekly thereafter until 6 weeks by the inclined plane method and by the modified Tarlov technique. After 6 weeks of injury, the origin of descending axons at the injury site was determined by retrograde labeling with fluorogold (FG), and a computer-assisted morphometric assessment of the injury site was performed. There was a significant improvement in counts of retrogradely labeled neurons in the red nucleus and rostral ventrolateral medulla (RVLM) in rats treated with either 2 nM (1338 +/- 366 and 28.8 +/- 16) or 10 nM (1390 +/- 511 and 46.3 +/- 31) QX-314 as compared to vehicle (902 +/- 403 and 13.8 +/- 8). There was a trend to increased neuronal counts in the sensorimotor cortex (170.8 +/- 226.8) and vestibular nuclei (1096.2 +/- 970.2) with QX-314 (10 nM) as compared to the vehicle-treated group. There was no significant difference in the extent of neurological recovery between the control and treated groups. Our results suggest that the Na+ channel blocker QX-314 partially preserves the integrity of descending motor axons after SCI. However, in this study, the effects were insufficient to result in sustained improvements in behavioral neurological function.

Lack of inhibition by inhalational anesthetics of myocardial contraction dependent on intracellular sodium activity.

When cardiac muscle is stimulated at a high frequency after rest, the peak force increases rapidly at first and then slowly. The slow phase reflects Ca2+ accumulation dependent on an increase in intracellular Na+ activity. The positive inotropic effect of high concentrations of cardiac glycosides also involves Ca2+ accumulation dependent on an increase in intracellular Na+ activity. The effect of isoflurane on the component of myocardial contraction dependent on an increase in intracellular Na+ activity is not well understood. The isometric force of rabbit papillary muscle contractions was measured. The authors studied the effects of isoflurane on the force increase that occurs when the muscles are stimulated after a rest and on the force increase induced by ouabain. Isoflurane (1.5%, 2.4%) had no statistically significant effect on the magnitude of the slow phase of force increase when the muscles were stimulated at 2 Hz after a rest. Isoflurane (1.5%) did not decrease the magnitude of force increase induced by ouabain (1 microM). Isoflurane has little effect on myocardial contractions that depend on Ca2+ accumulation after an increase in intracellular Na+ activity. This may partly account for the smaller cardiac depressant effect of isoflurane observed at high frequencies rather than at low frequencies. The results of the present study and an earlier study with halothane suggest that the lack of inhibition of contractions dependent on Ca2+ accumulation after an increase in intracellular Na+ activity may be a common property of inhalational anesthetics.

The role of aging on the control of contractile force by Na(+)-Ca2+ exchange in rat papillary muscle.

Sarcolemmal Na(+)-Ca2+ exchange system is believed to be fundamental to the control of cardiac contractility. However, the relation between Na(+)-Ca2+ exchange and the control of contractile force has not been studied in senescent myocardium. The role of Na(+)-Ca2+ exchange in the regulation of the cardiac muscle's contractile force was studied in adult and senescent papillary muscles by simultaneously measuring intracellular sodium activity (aNai), action potential, and contractile force while varying extracellular concentration of sodium and/or calcium. Reduction of [Na+]o decreased aNai from 8.0 +/- 1.8 to 4.1 +/- 0.8 in adult (-3.9 mM) and from 8.7 +/- 1.9 to 4.7 +/- 0.9 in senescent (-4.0 mM) papillary muscles, while developed tension (DT) increased by 80.2% in adult and by 135.6% in senescent papillary muscles (p < .01 vs adult). During low [Ca2+]o and high [Na+]o, aNai and DT modifications were similar both in adult and senescent papillary muscles. During high [Ca2+]o, aNai decreased to a similar extent in both adult and senescent papillary muscles, while DT increased by 37.8% in adult and by 67.8% in senescent (p < .05 vs adult). Simultaneous reduction of [Na+]o and [Ca2+]o decreased aNai from 8.1 +/- 1.2 to 6.8 +/- 1.1 mM in adult (-1.3 mM), and from 8.4 +/- 1.0 to 7.2 +/- 1.0 mM in senescent (-1.2 mM) papillary muscles while DT decreased by 22.1% in adult and by only 12.0% in senescent (p < .01 vs adult) papillary muscles. Simultaneous increase of [Na+]o and [Ca2+]o similarly increased aNai in both adult senescent papillary muscles, but decreased DT by 28.5% in adult and by 11.7% in senescent (p < .01 vs adult). After [Na+]o modifications, the equilibration time for the ratio of external and internal sodium ion activities was slowed in senescent papillary muscles (i.e., in low [Na+]o solution the equilibration time was 4.6 +/- 0.9 min in adult and 6.3 +/- 1.2 min in senescent papillary muscles, p < .001). Similar changes of aNai during transmembrane Na+ and Ca2+ gradients modifications associated to changes in contractile force seem to demonstrate that Na(+)-Ca2+ exchange is probably not modified by the aging process. However, the slow equilibration time for the ratio of Na+ activities might reflect an age-related reduction of the Na(+)-K+ pump activity.

The voltage dependence of contraction at different stimulation rates in guinea-pig ventricular myocytes.

Ventricular myocytes, isolated from the guinea-pig, were stimulated to contract by 100 ms long voltage clamp pulses from -80 to 0 mV at 0.5 and 3 Hz. An increase in frequency from 0.5 to 3 Hz led to a positive inotropic effect. Contraction-voltage relationships (CVR) were determined at each frequency. The CVR at 0.5 Hz was bell shaped and peaked between 0 and +20 mV, displaying a voltage dependence similar to the L-type Ca2+ current (ICa). At 3 Hz, contractions continued to increase at positive voltages, giving a more sigmoidal CVR. At 0.5 Hz, TTX reduced the size of steady-state contractions to 91 +/- 2% of control values, but had no effect on the shape of the CVR. At 3 Hz, TTX significantly reduced (P < 0.05) the magnitude of contractions at positive voltages (> or = +20 mV) but had no significant effect on contractions at voltages negative to 0 mV. These data illustrate that intracellular sodium activity (aNa(i)) and, in particular, Na+ entry due to the sodium current (INa) are important in determining the voltage dependence of contraction at positive voltages. Thapsigargin (2.5 microM), a blocker of the sarcoplasmic reticulum (SR) Ca(2+)-ATPase, reduced the size of steady-state contractions at 0 mV to 65 +/- 7% at 0.5 Hz. Increasing frequency to 3 Hz abolished the positive inotropy seen under control conditions. With thapsigargin present, contractions at 0.5 Hz were reduced at all potentials and the CVR was bell shaped. At 3 Hz the CVR was sigmoidal in shape. Contractions were significantly inhibited by thapsigargin at all potentials, but most significantly at more positive potentials (> or = +20 mV). These data show that, at normal body temperature, the shape of the CVR of guinea-pig ventricular myocytes changes with stimulation rate. Due to the voltage dependence of ICa, contractions evoked at positive voltages at 3 Hz must be supported by other mechanisms. The sensitivity of such contractions to TTX and thapsigargin suggests the involvement of both a Na(+)-dependent process and the SR. One possibility is that when aiNa and the Ca2+ content of the SR are raised at higher stimulation rates, enhanced Ca2+ entry via reverse Na(+)-Ca2+ exchange leads to a direct activation of the myofilaments and, to a lesser extent, the release of Ca2+ from the SR.

The role of the Na(+)‐Ca2+ exchanger in the rate‐dependent increase in contraction in guinea‐pig ventricular myocytes.

1. The intracellular sodium activity (alpha Na1), contraction and membrane current were recorded simultaneously in voltage-clamped guinea-pig ventricular myocytes. 2. Increasing the frequency (from 0.5 to 3 Hz) of voltage clamp pulses to 0 mV from a holding potential of -80 mV led to an increase in both alpha Na1 and contraction. The rate-dependent increase in contraction was reduced by 25 microM tetrodotoxin (TTX) and abolished with a holding potential of -40 mV. There was no rate-dependent rise in alpha Na1 with a holding potential of -40 mV. These results suggest an important role for alpha Na1 and in particular Na+ influx via Na+ channels during rate-dependent changes in contraction. 3. After an increase in frequency from 0.5 to 3 Hz, membrane current at the end of voltage clamp pulses became progressively more outward and the tail current upon at repolarization became progressively more inward compared with those recorded at 0.5 Hz. TTX reduced the magnitude of both the outward and inward rate-dependent shifts of current. 4. The addition of extracellular CsCl blocked the inward rectifier potassium current (IK.1) and the delayed rectifier (IK), but did not change the rate-dependent shift in current. 5. The difference between current-voltage relationships at 0.5 and 3 Hz showed that the rate-dependent outward shift of current at the end of voltage clamp pulses was small at potentials negative to -20 mV, was larger at more positive potentials and was reduced by TTX at most potentials. The TTX-sensitive component reversed at -47 mV. 6. These results are consistent with a net increase in outward Na(+)-Ca2+ exchange current during a voltage clamp pulse in response to the rise of alpha Na1. The increase in outward current (resulting from either enhanced Ca2+ influx or reduced Ca2+ efflux) will augment the Ca2+ load of the cell and contribute to the rate-dependent increase in contraction.

Effects of Hyperosmolar Solution on the Twitch Force, Membrane Potential, and Intracellular Sodium Activity in Purkinje Fibers and Ventricular Muscles

Effects of Hyperosmolar Solution on the Twitch Force, Membrane Potential, and Intracellular Sodium Activity in Purkinje Fibers and Ventricular Muscles

Classification of Positive Inotropic Actions Based on Electrophysiologic Characteristics

A number of new positive inotropic agents with diverse mechanisms of action have been discovered over the past 20 years. Most of these cardiotonic drugs exhibit characteristic electrophysiologic profiles. This prompted us to propose a classification scheme based on electrophysiologic principles, modifying the categories recently suggested by another author. Class I actions designate positive inotropic mechanisms that enhance the transmembrane calcium current by various means, such as beta-receptor stimulation (dobutamine, class I/A), phosphodiesterase inhibition (milrinone, class I/B), direct stimulation of adenylate cyclase (forskolin, class I/C), or direct modulation of calcium channel gating (BAY K 8644, class I/D). Class II action includes mechanisms that lead to elevation of intracellular sodium activity either by inhibiting the Na,K pump (digitalis, class II/A) or by increasing transmembrane sodium influx (DPI 201-106, class II/B). Class III action involves a mechanism by which sensitivity of the myofilaments to calcium increases (EMD 53998, levosimendan). This mechanism is not associated with apparent electrophysiologic manifestations. Positive inotropism due to lengthening of the cardiac repolarization (almokalant) is considered as class IV action. The possible clinical implications of the various positive inotropic mechanisms are also discussed.

Effects of caffeine on intracellular sodium activity in cardiac Purkinje fibres: relation to force.

1. An increase in cytoplasmic calcium by caffeine would lead to Ca extrusion via the Na/Ca exchange. The hypotheses were investigated that, as a consequence, caffeine might increase intracellular sodium activity (aiNa) and that the relation between aiNa and force might be conditioned by the Ca load. 2. Action potential, aiNa and contractile force were recorded in sheep Purkinje fibres during exposure to caffeine under conditions that decrease or increase the Ca load by different mechanisms. 3. In Tyrode solution, caffeine (8 mM) increased aiNa from 8.05 +/- 0.20 to 10.52 +/- 0.40 mM (+30.5%) and had a triphasic effect on force: an initial transient increase (+93.6%), a subsequent decrease (-37.1%) (negative inotropy) and slow partial recovery (+8.9%). 4. Decreasing the Ca load by means of manganese (1 mM) decreased aiNa and force. Adding caffeine re-increased aiNa and no longer caused a negative inotropic action. Cadmium (0.2 mM) also decreased aiNa, and caffeine reincreased it although far less than in Tyrode solution. 5. High [K]o (10 mM) and tetrodotoxin (5 microM) decreased aiNa as well as force. In their presence, caffeine re-increased aiNa and no longer had a negative inotropic action. 6. Increasing the Ca load by means of high [Ca]o (8.1 mM) increased force (+195%) and decreased aiNa, (-20.3%). Adding caffeine re-increased aiNa (+28.1%), but immediately decreased force (-32.3%). 7. Addition of pyruvate (10 mM) to caffeine increased force, as it does in the presence of Ca overload. 8. Noradrenaline (0.1-1 microM) decreased aiNa and increased contractile force. In its presence, caffeine decreased aiNa further and increased force. 9. It is concluded that caffeine increases aiNa, even during the negative inotropic effect. The decrease in force appears to depend on Ca load. Thus, caffeine no longer decreases force under conditions that decrease Ca load (Mn, high [K]0, TTX) and immediately decreases force when the Ca load is increased(high [Ca]0). However, in the presence of noradrenaline, caffeine decreases aiNa and markedly increases force, as the Ca load is increased, but Ca can be removed from the cytoplasm into the SR.

Changes of contractile responses due to simulated weightlessness in rat soleus muscle

Some contractile and electrophysiological properties of muscle fibers isolated from the slow-twitch soleus (SOL) and fast-twitch extensor digitorum longus (EDL) muscles of rats were compared with those measured in SOL muscles from suspended rats. In suspendede SOL (21 days of tail-suspension) membrane potential (Em), intracellular sodium activity (aiNa) and the slope of the relationship between Em and log [K] o were typical of fast-twitch muscles. The relation between the maximal amplitude of K-contractures vs Em was steeper for control SOL than for EDL and suspended SOL muscles. After suspension, in SOL muscles the contractile threshold and the inactivation curves for K-contractures were shifted to more positive Em. Repriming of K-contractures was unaffected by suspencion. The exposure of isolated fibers to perchlorate (ClO 4 −)-containing (6–40 mM) solutions resulted ina similar concentration-dependent shift to more negative Em of activation curves for EDL and suspended SOL muscles. On exposure to a Na-free TEA solution, SOL from control and suspended rats, in contrast to EDL muscles, generated slow contractile responses. Suspended SOL showed a reduced sensitivity to the contracture-producing effect of caffeine compared to control muscles. These results suggested that the modification observed due to suspension could be encounted by changes in the characteristics of muscle fibers from slow to fast-twitch type.

Role of Intracellular Sodium Activity in the Control of Contraction in Cardiac Purkinje Fibers

Role of Intracellular Sodium Activity in the Control of Contraction in Cardiac Purkinje Fibers

Role of Intracellular Sodium Activity in the Control of Contraction in Cardiac Purkinje Fibers.

The role of intracellular sodium activity (a(i)(Na)) in the control of force was studied in sheep cardiac Purkinje fibers exposed to norepinephrine (NE) and high [Ca](o) in the absence and presence of overdrive or of a low concentration of strophanthidin. Both NE and high [Ca](o) decrease a(i)(Na) and increae force, while overdrive increases and low strophanthidin decreases both parameters. In the presence of NE, overdrive increases a(i)(Na) less than force and is followed by a more pronounced undershoot in a(i)(Na) and force. In contrast, in high [Ca](o) overdrive increases a(i)(Na) more than force and is followed by a less pronounced undershoot in a(i)(Na) and force than in NE. High [Ca](o) increases force to a peak, but then the decreasing a(i)(Na) reduces force. In all these conditions, a(i)(Na) determines force changes during recovery from overdrive. NE and high [Ca](o) decrease a(i)(Na) less and increase force more in low strophanthidin. Thus, changes in a(i)(Na) modulate the increase in force due to increased Ca influx and control force development when Ca influx is either unchanged (low strophanthidin) or has reached a steady state (high [Ca](o), recovery from overdrive). Copyright 1994 S. Karger AG, Basel

The modulatory effects of endothelin-1, carbachol and isoprenaline upon Na(+)-H+ exchange in dog cardiac Purkinje fibres.

1. The modulatory effects of carbachol, endothelin-1 and isoprenaline upon Na(+)-H+ exchange were examined in dog cardiac Purkinje fibres. Intracellular pH (pHi) and intracellular sodium activity (aiNa) were recorded using pH and Na(+)-selective microelectrodes. Acid extrusion via Na(+)-H+ exchange was estimated from the pHi recovery rate (multiplied by intrinsic buffering power (beta i) and adding mean background acid load) in response to an internal acid load induced by the removal of 20 mM NH4Cl. All experiments in this work were performed in Hepes-buffered solutions at 37 degrees C. 2. beta i was estimated at various values of pHi in the range of 7.4-6.4 and was calculated from the fall of pHi induced by the addition and removal of NH4Cl. Experiments were performed when Na(+)-H+ exchange was blocked. The values of beta i in this tissue were only slightly dependent on pHi in the range of 7.4-6.4 with an empirical relationship: beta i = -4.69 pHi + 64.59. 3. Endothelin-1 (10(-8) M) alkalinized the resting pHi by approximately 0.1 pH unit and accelerated acid extrusion, by approximately 96%, at pHi approximately 6.9. A reduction of background acid loading within the cell cannot account for the augmentation of pHi recovery, since the rate of acid extrusion was not changed either at resting pHi or at internal acidification in Na(+)-free solution (a measure of background loading) by the addition of endothelin-1. The protein kinase C inhibitors staurosporin (10(-6) M) and 1-(5-isoquinolinylsulphonyl)-2-methyl-piperazine (H-7, 50 microM) could not block the effect of endothelin-1 on the antiporter. 4. At pHi approximately 6.8, carbachol (7.5 x 10(-4) M) accelerated pHi recovery by approximately 68% and alkalinized the resting pHi by approximately 0.1 pH unit. This stimulatory effect of carbachol was completely blocked by pretreatment with atropine (10(-4) M) and staurosporine 10(-6) M. The background acid load was not reduced by adding carbachol, since the acid extrusion during pHi recovery or at the resting state was not affected by the addition of carbachol to a sodium-free solution. 5. Isoprenaline (10(-6) M) slowed pHi recovery by approximately 45% measured at pHi 6.9 with no change in resting pHi. A rise in background acid loading could not account for the reduction of acid extrusion. Pretreated with atenolol (10(-6) M), a beta 1-selective antagonist, completely blocked the effect of isoprenaline.(ABSTRACT TRUNCATED AT 400 WORDS)

The effect of tri-iodothyronine on the membrane potential and intracellular sodium and potassium activities of rat soleus and extensor digitorum longus muscles.

Tri-iodothyronine (T3) induces a considerable increase in the number of Na/K pump units in rat skeletal muscles, but this effect is more pronounced in red muscles, such as soleus, than in white muscles such as extensor digitorum longus (EDL). In the present study, the effect of T3 on the membrane potential (Em) and the intracellular activities of K+ and Na+ (aKi and aNai) in the two muscle types has been studied using microelectrodes. Hyperthyroidism was induced by seven subcutaneous injections of 50 micrograms T3 per 100 g body weight on alternate days. Following treatment of rats with T3, the mean T3 concentration in the serum was 8.4 +/- 0.46 nmol/l (S.D.) compared with 1.1 +/- 0.16 nmol/l in the serum of untreated control rats, and the heart weight/body weight ratio of the treated rats had increased from 0.29 +/- 0.01 in control rats to 0.39 +/- 0.02. In soleus muscle fibres from the T3-treated rats, the Em, aKi and aNai were unchanged, whereas, in EDL muscle fibres from these rats, the Em had depolarized by 2 mV (P < 0.01) and aNai had increased by 2 mmol/l (P < 0.05). Since T3 induces not only an increase in the number of pump units and active movement of Na+ and K+ but also an increase in the passive fluxes of these ions, the results are discussed in terms of these two opposing ionic movements.

Intracellular sodium activity and its regulation in guinea-pig atrial myocardium.

1. Intracellular Na+ activity (aNai) and membrane resting potential were studied in quiescent guinea-pig atrial and papillary muscles by means of Na(+)-sensitive and conventional microelectrodes. The effects of the cardioactive steroid dihydroouabain (DHO) on aiNa, force of contraction and sarcolemmal Na+, K(+)-ATPase activity were also investigated. 2. In thirty atria and twenty-two papillary muscles, aNai amounted to 8.0 +/- 0.2 and 4.7 +/- 0.3 mM, respectively (mean +/- S.E.M.). When both tissues were from the same animal, with the same ion-sensitive microelectrode mean aNai values of 7.9 +/- 0.2 and 5.1 +/- 0.5 mM (P < 0.01) were obtained from eight atrial and eight papillary muscles, respectively. 3. Membrane resting potentials (Em) were significantly (P < 0.001) more negative in the papillary muscles (-83.5 +/- 0.7 mV; n = 8) than in the atrium (-78.1 +/- 0.5 mV; n = 8). Deviation of Em from EK (determined by K(+)-sensitive microelectrodes) was 3.0 +/- 0.2 mV in ventricular (P < 0.05) and 6.1 +/- 0.3 mV in atrial preparations (P < 0.05). 4. Inhibition of the Na+ pump by DHO increased aNai of the atrium within 10 min by 0.6 +/- 0.1 (n = 7), 1.3 +/- 0.1 (n = 5) and 3.2 +/- 0.2 mM (n = 5) at 5, 10 and 30 microM, respectively. In the papillary muscle, 10 microM DHO was without effect while aNai rose by 1.0 +/- 0.1 (n = 5) and 2.9 +/- 0.2 mM (n = 6) at 30 and 120 microM DHO. 5. Consistent with the aNai measurements, the potency of DHO to increase force of the isometric contraction was three times higher in atrium than in papillary muscle (stimulation frequency 0.2 Hz). 6. Hydrolytic activity of sarcolemmal Na+,K(+)-ATPase isolated from atria amounted to only one third of that detected in ventricles (0.07 +/- 0.01, n = 6, versus 0.2 +/- 0.01 mumol phosphate released min-1 (g tissue)-1, n = 5). The inhibitory potencies of DHO on sarcolemmal Na+,K(+)-ATPase preparations were found to be identical in the enzymes from either tissue. 7. It is concluded that a lower Na+ pump density is responsible for the higher aNai and for the lower resting membrane potential in atrial as compared to ventricular cells. The regulation of cellular Na+ homeostasis in atrial muscle appears to be closer to the limits of its capacity than in ventricle, explaining the higher sensitivity of the atrium to interventions which impede Na+ pump activity.