Na(+) current contribution to the diastolic depolarization in newborn rabbit SA node cells

Isolated newborn, but not adult, rabbit sinoatrial node (SAN) cells exhibit spontaneous activity that (unlike adult) are highly sensitive to the Na(+) current (I(Na)) blocker TTX. To investigate this TTX action on automaticity, cells were voltage clamped with ramp depolarizations mimicking the pacemaker phase of spontaneous cells (-60 to -20 mV, 35 mV/s). Ramps elicited a TTX-sensitive current in newborn (peak density 0.89 +/- 0.14 pA/pF, n = 24) but not adult (n = 5) cells. When depolarizing ramps were preceded by steplike depolarizations to mimic action potentials, ramp current decreased 54.6 +/- 8.0% (n = 3) but was not abolished. Additional experiments demonstrated that ramp current amplitude depended on the slope of the ramp and that TTX did not alter steady-state holding current at pacemaker potentials. This excluded a steady-state Na(+) window component and suggested a kinetic basis, which was investigated by measuring TTX-sensitive I(Na) during long step depolarizations. I(Na) exhibited a slow but complete inactivation time course at pacemaker voltages (tau = 33.9 +/- 3.9 ms at -50 mV), consistent with the rate-dependent ramp data. The data indicate that owing to slow inactivation of I(Na) at diastolic potentials, a small TTX-sensitive current flows during the diastolic depolarization in neonatal pacemaker myocytes.     

依托咪酯对成年大鼠脊髓胶状质局部突触传递的作用

应用盲插全细胞膜片钳技术,在成年大鼠脊髓薄片上观察依托咪酯(etomidate,ET)对脊髓胶状质局部突触传递的影响。实验结果显示,在钳制电压为-70mV时,500μmol/L的ET对微小兴奋性突触后电流(mEPSC)的持续时间、频率和幅度都无明显的作用。在钳制电压为0mV时,50μmol/L的ET使GABA能微小抑制性突触后电流(mIPSC)的持续时间延长45.57±12.46%(P<0.05),但对其频率和幅度无影响。同样在钳制电压为0mV的情况下,50μmol/L的ET对甘氨酸能mIPSC的持续时间、频率及幅度均无作用。以上结果表明,在成年大鼠的脊髓胶状质,ET主要通过延长GABA能mIPSC的持续时间,即延长受体通道的开放时间发挥作用,ET对于兴奋性的突触传递没有直接的作用。     

A voltage-dependent persistent sodium current in mammalian hippocampal neurons

Currents generated by depolarizing voltage pulses were recorded in neurons from the pyramidal cell layer of the CA1 region of rat or guinea pig hippocampus with single electrode voltage-clamp or tight-seal whole-cell voltage-clamp techniques. In neurons in situ in slices, and in dissociated neurons, subtraction of currents generated by identical depolarizing voltage pulses before and after exposure to tetrodotoxin revealed a small, persistent current after the transient current. These currents could also be recorded directly in dissociated neurons in which other ionic currents were effectively suppressed. It was concluded that the persistent current was carried by sodium ions because it was blocked by TTX, decreased in amplitude when extracellular sodium concentration was reduced, and was not blocked by cadmium. The amplitude of the persistent sodium current varied with clamp potential, being detectable at potentials as negative as -70 mV and reaching a maximum at approximately -40 mV. The maximum amplitude at -40 mV in 21 cells in slices was -0.34 +/- 0.05 nA (mean +/- 1 SEM) and -0.21 +/- 0.05 nA in 10 dissociated neurons. Persistent sodium conductance increased sigmoidally with a potential between -70 and -30 mV and could be fitted with the Boltzmann equation, g = gmax/(1 + exp[(V' - V)/k)]). The average gmax was 7.8 +/- 1.1 nS in the 21 neurons in slices and 4.4 +/- 1.6 nS in the 10 dissociated cells that had lost their processes indicating that the channels responsible are probably most densely aggregated on or close to the soma. The half-maximum conductance occurred close to -50 mV, both in neurons in slices and in dissociated neurons, and the slope factor (k) was 5-9 mV. The persistent sodium current was much more resistant to inactivation by depolarization than the transient current and could be recorded at greater than 50% of its normal amplitude when the transient current was completely inactivated. Because the persistent sodium current activates at potentials close to the resting membrane potential and is very resistant to inactivation, it probably plays an important role in the repetitive firing of action potentials caused by prolonged depolarizations such as those that occur during barrages of synaptic inputs into these cells.     

Properties of action potentials carried by divalent cations in identified leech neurons

Properties of divalent cation potentials carried by either Sr2+ or Ca2+ ions in Na+-free, TEA-Ringer solution were characterized in identified neurons of two species of leeches (Macrobdella and Haementeria). In Macrobdella, the overshoot of the potentials varied logarithmically with [Sr2+]0 (28.5 mV per 10-fold change). The overshoot, Vmax, and duration of the potentials increased with increasing divalent cation concentration and saturated at about 20 to 30 mM [Sr2+]0. The Vmax, amplitude, and duration of the potentials were reversibly blocked by Co2+ and Mn2+. The block by Mn2+ could be well-fitted by a reverse Langmuir-curve with an apparent KI of 100 micromolar. The local anesthetic procaine also reversibly inhibited the Vmax and duration of the potentials. The inhibition was greater at alkaline pH suggesting that procaine blocks the calcium channel from inside the membrane. The identified leech neurons examined in Macrobdella varied considerably in their ability to sustain somatic divalent cation potentials. Stimulation of T cells and most motoneurons produced no or only weak potentials, whereas stimulation of Retzius, N, Nut, and AP cells evoked overshooting potentials of several seconds' duration. Stimulation of the ALG cell of Haementeria in normal Ringer solution evoked a slowly-rising, purely Ca2+-dependent potential of approximately 100 ms duration. This response was TTX-resistant, unaffected by complete removal of Na+ from the Ringer solution, and abolished by 1 mM Mn2+. The overshoot varied logarithmically with a slope of 28 mV/decade change in [Ca2+]0.     

三叉神经中脑核神经元膜的电学特性

用大鼠脑干脑片,给三叉神经中脑核79个神经元作了细胞内记录,测算了20个神经元膜的电学特性:静息电位-60.3±5.6mV;输入阻抗为10.5±5.4MΩ;时间常数1.3±0.5ms。电刺激可诱发动作电位,测算32个神经元的有关参数:阈电位-50—-55mV;波幅69.5±6.1mV;超射11.9±3.6mV;波宽0.8±0.2ms。TTX(0.3μmol/L)或无钠使之消失。通以长时程矩形波电流可引起200—250Hz的2—15个重复放电,但在通电停止前终止,TEA或4-AP可延长放电。膜电位-60—-55mV时在动作电位之后可看到阈下电位波动,它不受TTX的影响,无钙时消失,TEA或4-AP使波幅增大。静息电位去极化可使45个神经元中的40个发生外向整流作用,并被TEA,4-AP或无钙抑制,超极化则发生内向整流作用,Cs或无钠抑制之。灌流液中加入各种钾通道阻断药时神经元的稳态I-V曲线发生相应变化,提示I_(DR),l_A,I_(K(Ca))及I_Q可能都与静息时的膜电导有关。     

The saxitoxin/tetrodotoxin binding site on cloned rat brain IIa Na channels is in the transmembrane electric field.

The rat brain IIa (BrIIa) Na channel alpha-subunit and the brain beta 1 subunit were coexpressed in Xenopus oocytes, and peak whole-oocyte Na current (INa) was measured at a test potential of -10 mV. Hyperpolarization of the holding potential resulted in an increased affinity of STX and TTX rested-state block of BrIIa Na channels. The apparent half-block concentration (ED50) for STX of BrIIa current decreased with hyperpolarizing holding potentials (Vhold). At Vhold of -100 mV, the ED50 was 2.1 +/- 0.4 nM, and the affinity increased to a ED50 of 1.2 +/- 0.2 nM with Vhold of -140 mV. In the absence of toxin, the peak current amplitude was the same for all potentials negative to -90 mV, demonstrating that all of the channels were in a closed conformation and maximally available to open in this range of holding potentials. The Woodhull model (1973) was used to describe the increase of the STX ED50 as a function of holding potential. The equivalent electrical distance of block (delta) by STX was 0.18 from the extracellular milieu when the valence of STX was fixed to +2. Analysis of the holding potential dependence of TTX block yielded a similar delta when the valence of TTX was fixed to +1. We conclude that the guanidinium toxin site is located partially within the transmembrane electric field. Previous site-directed mutagenesis studies demonstrated that an isoform-specific phenylalanine in the BrIIa channel is critical for high affinity toxin block.(ABSTRACT TRUNCATED AT 250 WORDS)     

Voltage-dependent and odorant-regulated currents in isolated olfactory receptor neurons of the channel catfish.

The electrical properties of olfactory receptor neurons, enzymatically dissociated from the channel catfish (Ictalurus punctatus), were studied using the whole-cell patch-clamp technique. Six voltage-dependent ionic currents were isolated. Transient inward currents (0.1-1.7 nA) were observed in response to depolarizing voltage steps from a holding potential of -80 mV in all neurons examined. They activated between -70 and -50 mV and were blocked by addition of 1 microM tetrodotoxin (TTX) to the bath or by replacing Na+ in the bath with N-methyl-D-glucamine and were classified as Na+ currents. Sustained inward currents, observed in most neurons examined when Na+ inward currents were blocked with TTX and outward currents were blocked by replacing K+ in the pipette solution with Cs+ and by addition of 10 mM Ba2+ to the bath, activated between -40 and -30 mV, reached a peak at 0 mV, and were blocked by 5 microM nimodipine. These currents were classified as L-type Ca2+ currents. Large, slowly activating outward currents that were blocked by simultaneous replacement of K+ in the pipette with Cs+ and addition of Ba2+ to the bath were observed in all olfactory neurons examined. The outward K+ currents activated over approximately the same range as the Na+ currents (-60 to -50 mV), but the Na+ currents were larger at the normal resting potential of the neurons (-45 +/- 11 mV, mean +/- SD, n = 52). Four different types of K+ currents could be differentiated: a Ca(2+)-activated K+ current, a transient K+ current, a delayed rectifier K+ current, and an inward rectifier K+ current. Spontaneous action potentials of varying amplitude were sometimes observed in the cell-attached recording configuration. Action potentials were not observed in whole-cell recordings with normal internal solution (K+ = 100 mM) in the pipette, but frequently appeared when K+ was reduced to 85 mM. These observations suggest that the membrane potential and action potential amplitude of catfish olfactory neurons are significantly affected by the activity of single channels due to the high input resistance (6.6 +/- 5.2 G omega, n = 20) and low membrane capacitance (2.1 +/- 1.1 pF, n = 46) of the cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

Inhibitory miniature synaptic potentials in rat motoneurons

In the newborn rat spinal cord, spontaneous potentials were recorded, with KCl electrodes, from motoneurons in the presence of tetrodotoxin (10(-6) g ml-1) to abolish nerve impulses. These potentials occurred at low frequencies (less than 2 Hz), and their mean amplitude was a fraction of 1 mV. An increase of osmolarity with sucrose or an increase of extracellular K+, increased the frequency of miniature synaptic potentials. The amplitude of the spontaneous potentials was increased by intracellular injection of Cl-. Strychnine (2-25 microM) completely abolished the spontaneous potentials. It is suggested that these potentials are produced by the spontaneous release of packages of inhibitory transmitter at synapses on motoneurons.     

The auditory evoked potentials of the brain stem in the guinea pig

The examination of the standard waves' amplitude and latency of the brain stem auditory evoked response (BAEP) was performed in 20 guinea pigs (males and females, weighing 250 to 300 g). According with the relative loudness of stimuli (90, 70, 50, 30, 10 dB SPL), the latency of BAEP waves was larger (t1 = 0.2 msec), but the conductance time between P1 to P5 was constant (3.1 to 3.6 msec). The highest wave of BAEP was P2 with an amplitude: 90 dB SPL, U = 6.5 +/- 1.2 microV; 70 dB SPL, U = 4.3 +/- 1.0 microV; 50 dB SPL, U = 3.5 +/- 0.6 microV; 30 dB SPL, U = 2.0 +/- 0.4 microV.     

Sodium and calcium currents in action potentials of rat somatotrophs: their possible functions in growth hormone secretion

We report that both Na+ and Ca2+ currents are involved in the action potentials and in the hormone release from rat somatotrophs in primary culture. Single somatotrophs were identified by reverse hemolytic plaque assay (RHPA) and transmembrane voltage and currents were recorded using the whole-cell mode of the patch-clamp technique. Somatotrophs displayed a mean resting potential of -80mV and an average input resistance of 5.7G omega. Most of the cells showed spontaneous or evoked action potentials. Single action potentials or the initial spike in a burst were characterized by their high amplitude and short duration. Tetrodotoxin (TTX, 1 microM) blocked single action potentials and the initial spikes in a burst, whereas action potentials of long duration and low amplitude persisted. Cobalt (2 mM) plus TTX (1 microM) blocked all the action potentials. Voltage-clamp experiments confirmed the presence of both a TTX-sensitive Na+ current and Co2(+)-sensitive Ca2+ currents. TTX or Na(+)-free medium slightly decreased the basal release of GH but did not markedly modify hGRF-stimulated GH release. However, Co2+ (2 mM), which partially decreased the basal release, totally blocked hGRF-stimulated release. We conclude that (1) Na+ currents which initiate rapid action potentials may participate in spontaneous GH release; (2) Ca2+ currents, which give rise to long duration action potentials and membrane voltage fluctuation, are probably involved in both basal and hGRF-stimulated GH releases.     

Saxitoxin and tetrodotoxin. Electrostatic effects on sodium channel gating current in crayfish axons.

The effects of extracellular saxitoxin (STX) and tetrodotoxin (TTX) on gating current (IgON) were studied in voltage clamped crayfish giant axons. At a holding potential (VH) of -90 mV, integrated gating charge (QON) was found to be 56% suppressed when 200 nM STX was added to the external solution, and 75% suppressed following the addition of 200 nM TTX. These concentrations of toxin are sufficiently high to block greater than 99% of sodium channels. A smaller suppression of IgON was observed when 1 nM STX was used (KD = 1-2 nM STX). The suppression of IgON by external toxin was found to be hold potential dependent, with only minimal suppression observed at the most hyperpolarized hold potentials, -140 to -120 mV. The maximal effect of these toxins on IgON was observed at hold potentials where the QON vs. VH plot was found to be steepest, -100 to -80 mV. The suppression of IgON induced by TTX is partially relieved following the removal of fast inactivation by intracellular treatment with N-bromoacetamide (NBA). The effect of STX and TTX on IgON is equivalent to a hyperpolarizing shift in the steady state inactivation curve, with 200 nM STX and 200 nM TTX inducing shifts of 4.9 +/- 1.7 mV and 10.0 +/- 2.1 mV, respectively. Our results are consistent with a model where the binding of toxin displaces a divalent cation from a negatively charged site near the external opening of the sodium channel, thereby producing a voltage offset sensed by the channel gating apparatus.     

Ca2+-dependent depolarization and burst firing of rat CA1 pyramidal neurones induced by N-methyl-D-aspartic acid and quinolinic acid: antagonism by 2-amino-5-phosphonovaleric and kynurenic acids

The excitatory effects of microiontophoretically applied quisqualic (QUIS), N-methyl-D-aspartic (NMDA), and quinolinic (QUIN) acids were investigated using intracellular recording from CAl pyramidal neurones in slices of rat hippocampus. QUIS evoked only simple action potentials superimposed upon a depolarization which attained a clear plateau. When this level had been reached, increased ejecting currents did not produce further depolarization. By contrast, with low currents NMDA and QUIN elicited small membrane depolarizations which triggered bursts of action potentials superimposed upon rhythmically occurring depolarizing shifts. Larger currents caused depolarization which if sufficiently large completely blocked spike activity. Tetrodotoxin (TTX) prevented the spikes evoked by QUIS and the bursts of action potentials seen with NMDA and QUIN, and the rhythmic depolarizing shifts then appeared as broad spikes of up to 50 mV in amplitude. These and the underlying membrane depolarization were blocked by Co2+, by the NMDA antagonist D(-)-2-amino-5-phosphonovaleric acid (DAPV), and by kynurenic acid (KYNU). It thus appears that the depolarization and burst firing of rat CAl pyramidal neurones elicited by NMDA and QUIN are Ca2+ dependent while the actions of QUIS are not.     

Inter-operator agreement in decomposition of motor unit firings from high-density surface EMG.

High-density surface EMG can be used to obtain a spatially selective representation of several motor unit action potentials. Recently, a decomposition of the signal into the underlying motor neuron firing patterns has been described. The reliability of the algorithm has not yet been tested. Eleven healthy subjects participated. High-density surface EMG was recorded from the vastus lateralis muscle during an isometric knee extension. Two independent operators analyzed the signals. After operator-supervised cluster analysis of spikes, motor unit action potential templates were constructed and an automatic template matching was performed. The decomposition was adjusted by hand. Agreement between operators was calculated for the number of coincident firings. Bland-Altman plots of peak-to-peak amplitude were constructed and limits of agreement were calculated. For completely decomposed motor unit action potential trains the between-operator agreement of firing events was very high. The peak-to-peak amplitude of monopolar motor unit action potentials was 115microV (SD 74microV). The agreement was within 3microV and independent of amplitude. With partial decomposition agreement within 26microV was achieved. For bipolarly derived motor unit action potentials the peak-to-peak amplitude was 54microV (SD 49microV), the agreement was within 3microV. Only for recordings obtained from a force level below 5% of the maximum voluntary contraction full decomposition was possible. It was concluded that when full decomposition is achieved, two independent operators are likely to arrive at nearly identical firing patterns.     

The sodium current underlying action potentials in guinea pig hippocampal CA1 neurons

Neurons were acutely dissociated from the CA1 region of hippocampal slices from guinea pigs. Whole-cell recording techniques were used to record and control membrane potential. When the electrode contained KF, the average resting potential was about -40 mV and action potentials in cells at -80 mV (current-clamped) had an amplitude greater than 100 mV. Cells were voltage-clamped at 22-24 degrees C with electrodes containing CsF. Inward currents generated with depolarizing voltage pulses reversed close to the sodium equilibrium potential and could be completely blocked with tetrodotoxin (1 microM). The amplitude of these sodium currents was maximal at about -20 mV and the amplitude of the tail currents was linear with potential, which indicates that the channels were ohmic. The sodium conductance increased with depolarization in a range from -60 to 0 mV with an average half-maximum at about -40 mV. The decay of the currents was not exponential at potentials more positive than -20 mV. The time to peak and half-decay time of the currents varied with potential and temperature. Half of the channels were inactivated at a potential of -75 mV and inactivation was essentially complete at -40 to -30 mV. Recovery from inactivation was not exponential and the rate varied with potential. At lower temperatures, the amplitude of sodium currents decreased, their time course became longer, and half-maximal inactivation shifted to more negative potentials. In a small fraction of cells studied, sodium currents were much more rapid but the voltage dependence of activation and inactivation was very similar.     

Sodium channels in cultured chick dorsal root ganglion neurons

Isolated Na currents were studied in cultured chick sensory neurons using the patch clamp technique. On membrane depolarization, whole cell currents showed the typical transient and voltage-dependent time course as in nerve fibres. Na currents appeared at about-40 mV and reached maximum amplitude at around-10 mV. At low voltages (-30 to 0 mV), their turning-on was sigmoidal and inactivation developed exponentially. The ratio of inactivation time constants was found to be smaller than in squid axons and comparable to that of mammalian nodes of Ranvier. Peak conductance and steady-state inactivation were strongly voltage-dependent, with maximum slopes at-17 and-40 mV, respectively. The reversal potential was close to the Nernst equilibrium potential, indicating a high degree of ion-selectivity for the channel. Addition of 3M TTX, or replacement of Na by Choline in the external bath, abolished these currents. Internal pronase (1 mg/ml) and N-bromoacetamide (0.4 mM) made inactivation incomplete, with little effect on its rate of decay.Single Na channel currents were studied in outside-out membrane patches, at potentials between-50 and-20 mV. Their activation required large negative holding potentials (-90 mV). They were fully blocked by addition of TTX (3 M) to the external bath. At-40 mV their mean open time was about 2ms and the amplitude distribution could be fitted by a single Gaussian curve, indicating the presence of a homogeneous population of channels with a conductance of 11±2 pS. Probability of opening increased and latency to first opening decreased with increasing depolarization. Inactivation of the channel became faster with stronger depolarizations, as measured from the inactivation time course of sample averages. Internal pronase (0.1 mg/ml) produced effects on inactivation comparable to those on whole cell currents. Openings of the channel had a tendency to occur in bursts and showed little inactivation during pulses of 250 ms duration. The open lifetime of the channel at low potentials (-50,-40 mV) was only three times larger than in control patches, suggesting that Na channels in chick sensory neurons can close several times before entering an inactivating absorbing state.     

Electrophysiological aspects of synaptic transmission at the electromotor junction of Torpedo marmorata

Miniature and stimulus-evoked electroplaque potentials were recorded in Torpedo electrocytes intracellularly and extracellularly and analysed quantitatively. Tetrodotoxin reversibly blocked stimulus evoked potentials but hardly affected spontaneous miniature potentials in amplitude and frequency. The quantum content of stimulus-evoked potentials varied between 150 and 400 in normal saline and decreased in low Ca2+ high Mg2+ solution. Quantal release conformed to binomial statistics and allowed determination of the release parameters p and n. Analysis of the time constant of decay of spontaneous miniature electroplaque currents showed variation around a mean of 0.75 +/- 0.16 msec (SD) which was greatly prolonged by application of neostigmine.     

Voltage- and calcium-activated currents in cultured olfactory receptor neurons of male Mamestra brassicae (Lepidoptera)

Insect olfactory receptor neurons (ORNs) grown in primary cultures were studied using the patch-clamp technique in both conventional and amphotericin B perforated whole-cell configurations under voltage-clamp conditions. After 10-24 days in vitro, ORNs had a mean resting potential of -62 mV and an average input resistance of 3.2 GOmega. Five different voltage-dependent ionic currents were isolated: one Na(+), one Ca(2+) and three K(+) currents. The Na(+) current (35-300 pA) activated between -50 and -30 mV and was sensitive to 1 microM tetrodotoxin (TTX). The sustained Ca(2+) current activated between -30 and -20 mV, reached a maximum amplitude at 0 mV (-4.5 +/- 6.0 pA) that increased when Ba(2+) was added to the bath and was blocked by 1 mM Co(2+). Total outward currents were composed of three K(+) currents: a Ca(2+)-activated K(+) current activated between -40 and -30 mV and reached a maximum amplitude at +40 mV (605 +/- 351 pA); a delayed-rectifier K(+) current activated between -30 and -10 mV, had a mean amplitude of 111 +/- 67 pA at +60 mV and was inhibited by 20 mM tetraethylammonium (TEA); and, finally, more than half of ORNs exhibited an A-like current strongly dependent on the holding potential and inhibited by 5 mM 4-aminopyridine (4-AP). Pheromone stimulation evoked inward current as measured by single channel recordings.     

Hyperpolarizing potentials induced by Ca-mediated K-conductance increase in hamster submandibular ganglion cells.

The mechanisms of three types of hyperpolarizing electrogenesis in hamster submandibular ganglion cells were analyzed with intracellular microelectrodes. These included (1) spike-induced hyperpolarizing afterpotential (S-HAP), (2) spontaneous transient hyperpolarizing potential (HP), and (3) the hyperpolarizing (H) phase of postsynaptic potential (PSP). Most of these hyperpolarizing potentials were due to conductance increases and reversed polarity at membrane potential (Em) between -70 and -85 mV, which was close to the K-equilibrium potential. The average resting potential of ganglion cells was -53 mV. Action potential overshoot increased slightly in high [Ca2+]0 and decreased in low [Ca2+]0. In most neurons action potentials were completely suppressed by 10(-7)-M tetrodotoxin (TTX). The S-HAP has an initial component due to delayed rectification and a late component. The late component is enhanced by increasing [Ca2+]0, or by applying Ca-ionophore (A23187), TEA, caffeine, or dibutyryl cyclic (DBc-) AMP; it is suppressed by decreasing [Ca2+]0, or by applying Mn2+. Perfusion with Cl--free saline reduced membrane potential slightly but did not modify the S-HAP. Depolarizing pulses also induced hyperpolarizing afterpotential (D-HAP), similar to the S-HAP. Spontaneous transient HPs occurred in some neurons at irregular intervals. HPs were insensitive to TTX but were suppressed by Mn2+. Caffeine induced low frequency rhythmic HPs in many neurons, often alternating with periods of repetitive spiking. The PSP was a monophasic depolarizing (D-) potential in some neurons, but in others the D-phase was followed by a small H-phase. Perfusion with A23187, caffeine or DBc-AMP increased the H-phase of the PSP. Perfusion with K+-free saline or treatment with 10(-5)M ouabain did not abolish the H-phase of PSPs. These membrane potential-dependent phenomena appear to be induced mainly by Ca-mediated K-conductance increases. This mechanism contributes to the regulation of low-frequency repetitive firing in submandibular ganglion cells.     

Sodium currents during differentiation in a human neuroblastoma cell line

The electrophysiological properties of a human neuroblastoma cell line, LA-N-5, were studied with the whole-cell configuration of the patch clamp technique before and after the induction of differentiation by retinoic acid, a vitamin A metabolite. Action potentials could be elicited from current clamped cells before the induction of differentiation, suggesting that some neuroblasts of the developing sympathetic nervous system are excitable. The action potential upstroke was carried by a sodium conductance, which was composed of two types of sodium currents, described by their sensitivity to tetrodotoxin (TTX) as TTX sensitive and TTX resistant. TTX-sensitive and TTX-resistant sodium currents were blocked by nanomolar and micromolar concentrations of TTX, respectively. The voltage sensitivity of activation and inactivation of TTX-resistant sodium current is shifted -10 to -30 mV relative to TTX-sensitive sodium current, suggesting that TTX-resistant sodium current could play a role in the initiation of action potentials. TTX-sensitive current comprised greater than 80% of the total sodium current in undifferentiated LA-N-5 cells. The surface density of total sodium current increased from 24.9 to 57.8 microA/microF after cells were induced to differentiate. The increase in total sodium current density was significant (P less than 0.05). The surface density of TTX-resistant sodium current did not change significantly during differentiation, from which we conclude that an increase in TTX-sensitive sodium current underlies the increase in total current.     

Characteristics of sodium tail currents in Myxicola axons. Comparison with membrane asymmetry currents.

Sodium currents after repolarization to more negative potentials after initial activation were digitally recorded in voltage-clamped Myxicola axons compensated for series resistance. The results are inconsistent with a Hodgkin-Huxley-type kinetic scheme. At potentials more negative than -50 mV, the Na+ tails show two distinct time constants, while at more positive potentials only a single exponential process can be resolved. The time-course of the tail currents was totally unaffected when tetrodotoxin (TTX) was added to reduce gNa to low values, demonstrating the absence of any artifact dependent on membrane current. Tail currents were altered by [Ca++] in a manner consistent with a simple alteration in surface potential. Asymmetry current "off" responses are well described by a single exponential. The time constant for this response averaged 2.3 times larger than that for the rapid component of the Na+ repolarization current and was not sensitive to pulse amplitude or duration, although it did vary with holding potential. Other asymmetry current observations confirm previous reports on Myxicola.