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C. Besta National Neurological Institute, Milan, Italy
Submitted 7 June 2005; accepted in final form 21 January 2006
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ABSTRACT |
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INTRODUCTION |
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; Stafstrom et al. 1985), it has been discovered that it is involved in modulating the functions of cortical neurons. Early voltage-clamp studies (Stafstrom et al. 1985) demonstrated that it contributes toward sustaining recurrent firing in response to membrane depolarization in layer V neurons, and subsequent experiments showed that it takes part in generating depolarizing afterpotentials (Azouz et al. 1996) and action potential (AP) bursts in cortical structures (Franceschetti et al. 1995; Kim and McCormick 1998; Mantegazza et al. 1998; Mattia et al. 1997; Menendez de la Prida et al. 2003). It also significantly modulates postsynaptic inputs to dendrite arborization (Mittmann et al. 1997; Poznanski and Bell 2000; Schwindt and Crill 1995) and contributes in generating subthreshold membrane oscillations in neuronal subsets of the hippocampus, subiculum, and both entorhinal and sensorimotor cortices (Agrawal et al. 2001; Amitai 1994; Hu et al. 2002). Finally, the peacemaking effect played by INaP in subthalamic neurons (Do and Bean 2003) and its contribution to setting the AP threshold for spontaneous discharges in tuberomammillary neurons (Taddese and Bean 2002) are further recent examples of its primary role in specific physiological functions.
A pathological enhancement of INaP may occur in case of dysfunctional events involving cortical neurons, such as epileptic discharges in acquired (Agrawal et al. 2003; Vreugdenhil et al. 2004) or genetically determined Na+ channelopathies (Lossin et al. 2002). The enhancement of INaP resulting from ischemia (Ju et al. 1996) may lead to neuronal damage and, more generally, increased Na+ flux throughout long depolarizations may play a role in neurodegenerative processes (Vajda 2002). Because INaP plays a role in sustaining epileptic discharges and long membrane depolarizations, it is a recognized target of antiepileptic and neuroprotective drugs. A number of traditional and new anticonvulsants or anesthetics (Chao and Alzheimer 1995; Gebhardt et al. 2001; Spadoni et al. 2002; Taverna et al. 1999) can reduce the persistent fraction of the Na+ current often at concentrations that do not inhibit the transient fraction of the current INaT (Lampl et al. 1998; Segal and Douglas 1997).
These and other findings seem to have sufficiently established the role of INaP in triggering and sustaining physiological and pathological depolarizing events. Its involvement in shaping and sustaining such events certainly arises from its particular activation characteristics. In fact, INaP begins to activate at rather negative membrane potentials that are close to the resting potential and below the firing threshold (French et al. 1990; Stafstrom et al. 1985). However, its inactivation properties can also play a crucial role in determining its functional effect on membrane excitability. The results of previous experiments using mouse layer V neurons indicate that INaP actually inactivates with a time constant of about 2 s (Fleidervish and Gutnick 1996). In the entorhinal cortex, an even longer time constant characterizes its kinetics of inactivation, assessed by single-channel recordings (Magistretti and Alonso 1999), thus suggesting complex inactivation kinetics (Magistretti and Alonso 2002).
The aim of this study was to characterize the voltage dependency of activation and inactivation and the kinetics of inactivation of INaP in sensorimotor neocortical slices and to assess whether different properties distinguish INaP in layer V and layer II/III pyramidal neurons, which are endowed with different firing properties and different functional implications.
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METHODS |
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Sprague–Dawley rats (Charles River, Florence, Italy) aged 13–18 days were anesthetized with ether and decapitated. Their brains were removed and placed in ice-cold artificial cerebrospinal fluid (standard ACSF) with (in mM): 124 NaCl, 26.5 NaHCO3, 2 CaCl2, 1.25 NaH2PO4, 2 MgSO4, 3.5 KCl, 10 glucose, and bubbled with 95% O2-5% CO2.
Coronal slices of 300 µm were prepared from the sensorimotor cortex, transferred to a submersion chamber kept at 35°C, and perfused with ACSF (see following text). All of the experimental procedures were carried out in compliance with the 86/609/UE law on animal research and the guidelines for animal care and management of the Ethics Committee of C. Besta Institute.
Electrophysiological recordings
The whole cell patch-clamp recordings were made at 35°C using an Axopatch 200B amplifier (Axon Instruments, Union City, CA) in layers II/III and V. Pyramidal neurons were directly visualized in brain slices with infrared differential-interference contrast microscopy using an upright microscope (Zeiss Axioscope) and a CCD camera (Hamamatsu).
The neurons were bath perfused with an external solution containing (in mM): 3 KCl, 102 NaCl, 5 MgCl2, 15 NaHCO3, 10 HEPES-NaOH, 10 glucose, 0.2 CaCl2, 0.3 NiCl2, 0.4 CdCl2, 30 tetraethylammonium-Cl (TEA-Cl), and 2 kynurenic acid, and bubbled with 95% O2-5% CO2; the electrodes were filled with a solution containing (in mM) 132 CsCl, 2 MgCl2, 1 CaCl2, 10 HEPES, 10 EGTA-CsOH, 2 Na2ATP, 10 phosphocreatine-diTris, 0.3 Na-GTP, and 20 U/ml creatine phosphokinase, pH 7.2.
In some experiments, CsCl in the internal solution was substituted with KGluconate and slices were perfused with standard ACSF to evaluate physiological firing characteristics with current-clamp recordings before Ca2+ and K+ currents blockade and voltage-clamp recordings of INaP. In these experiments, we began the recording of INaP after a long perfusion with extracellular blockers, when a satisfactory blockade of Ca2+ and K+ currents was achieved.
In control experiments, 1 µM tetrodotoxin (TTX) was added to the perfusing medium to rule out the contribution of TTX-resistant currents that could activate in the voltage range used to study INaP kinetics.
The data were digitized using a Digidata 1320 interface (Axon Instruments); pClamp 8.0 software (Axon Instruments) was used to generate stimulus protocols and acquire the signals. After seal formation and cell membrane rupturing, capacitance currents were minimized using the amplifier circuitry and 70–80% series resistance compensation was routinely applied.
INaP was evoked using slow (50 mV s–1) voltage ramps or, in control experiments, by means of depolarizing steps from a holding potential of –80 mV. Junction potential errors were not corrected. The sampling frequency was 5 kHz for the ramp protocols and current-clamp recordings and 10 kHz for the step protocol. The membrane currents were filtered at 1 kHz (voltage ramps) or 3 kHz (step protocol). The recordings with voltage-clamp errors (i.e., presence of unclamped action currents) were excluded from the analysis.
Conductance–voltage (g–V) relationships (activation curves) were calculated from the currents recorded using voltage ramps as g = INa/(V – ENa), where INa is the recorded Na+ current measured at potential V and ENa is the calculated equilibrium potential. Normalized activation curves were fitted to Boltzmann relationships in the form: G/Gmax = 1/{1 + exp[(V1/2 – V)/k]}, where Gmax is the maximal peak conductance, G is the peak conductance at each test voltage, V1/2 is the voltage at which half-maximal activation is reached, and k isthe slope factor. The patch clamp data were analyzed using pClamp8 (Axon Instruments) and Origin 7.5 software (MicroCal, Northampton, MA).
The data, presented as means ± SE, were statistically analyzed using nonparametric (Wilcoxon or Mann–Whitney) or ANOVA tests. Fits were compared using the F test to evaluate statistically the number of exponentials needed for the best fit.
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RESULTS |
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INaP in both layer V and layer II/III pyramidal neurons was elicited using a ramp stimulus protocol (50 mV s–1), whereas step pulses were applied in control experiments aimed at evaluating the correspondence between the amplitude and activation properties of INaP evoked by means of different stimulation protocols (Fig. 1, A and B). In both layers, INaP started to activate at potentials slightly negative of –60 mV, and reached a broad peak at membrane potentials ranging from –33 and –30 mV. In the presence of 1 µM TTX, the ramp protocols evoked only a small outward current, which activated at more positive membrane potentials than INaP (see Fig. 1A). Boltzmann fitting of the activation curves showed similar properties in the two layers (Fig. 1, C and D, and Table 1), and the comparison of INaP activation assessed on TTX-subtracted traces using slow voltage ramps and step protocols showed similar activation properties (V1/2: –43.5 ± 0.4 mV and –43.2 ± 0.6; k: 4.1 ± 0.2 and 4.2 ± 0.1; n = 6).
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) because the ages of rats from which we recorded layer V (15.6 ± 0.2 days) and II/III (15.8 ± 0.3 days) neurons were similar. Voltage dependency of steady-state INaP inactivation
We estimated the voltage dependency of INaP inactivation by delivering voltage ramp commands at the end of a 10-s depolarizing pulse at voltages ranging from –90 to –10 mV (see Fig. 2C, inset). To avoid the accumulation of slow inactivation, inactivating prepulses were delivered every 45 s.
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Boltzmann fitting of the mean normalized current amplitudes confirmed a significant difference in voltage dependencies of inactivation between the neurons of the two layers. In fact, the voltage dependency of inactivation was significantly shifted to the right (about 4.5 mV) in layer V (Fig. 2C; Table 1), and the slope factor reflected the significantly steeper inactivation curve (Table 1). On average, after a 10-s prepulse to –60 mV the inactivation was negligible or absent in layer V (4.9 ± 2.0%), but already substantial in layer II/III pyramidal neurons (15.6 ± 2.0% of the maximal current peak; P < 0.002).
In all of the pyramidal neurons of both layers V and II/III, a fraction of INaP remained after prepulses to –10 mV membrane potential, which was on average slightly larger in layer V (33.4 ± 2.3% of the maximum unconditioned current peak amplitude) than in layer II/III (27.9 ± 1.1%), but the difference was not statistically significant. The INaP amplitude also did not further decrease in the case of conditioning potentials to 0 mV (10 neurons, data not shown).
Kinetics of INaP development of inactivation and recovery from inactivation
The time dependency of inactivation was evaluated by applying depolarizing prepulses to –10 mV of increasing duration (from 0 to 40 s) and eliciting INaP by means of a standard voltage ramp (see stimulus protocol in Fig. 3A). In both layer II/III and layer V, the time dependencies of slow INaP inactivation were fitted by biexponential functions in the form y = y0 + A1 exp[–(x – x0)t1] + A2 exp[–(x – x0)t2]. The faster time constant measured in layer V varied widely from 158.0 to 1133.8 ms. On average (425.9 ± 80.5 ms) it was significantly longer than in layer II/III (145.8 ± 18.2 ms; range 63–157; P < 0.003). The second time constant was variable but on average of about 5,000 ms, without any significant differences between the two layers. The curve of development of INaP inactivation reached a plateau with prepulses lasting >25 s. In fact, a small current remained in all of the neurons even after the longest inactivation prepulses (corresponding to 13–26% of the unconditioned current peak), without any difference between the two layers.
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To verify the absence of unblocked currents possibly contaminating the traces, we evaluated the time-dependent inactivation properties in three layer V neurons in the presence of TTX. As shown in Fig. 3D, the time course of INaP inactivation measured on the original and TTX-subtracted traces completely overlapped, thus ruling out any effect of contaminating currents.
To evaluate the contribution of INaP to the depolarized plateau after APs observed when Ca2+ and K+ currents are blocked (Franceschetti et al. 1995; Stafstrom et al. 1985), we did current-clamp experiments with the solutions used for the voltage-clamp recordings of INaP in nine neurons: three layer II/III and six layer V pyramidal neurons. In all layer II/III neurons, the depolarizing plateau did not exceed 100 ms, whereas extremely long depolarizations lasting >1 s consistently followed the AP evoked in three of six layer V neurons. The remaining three layer V pyramidal neurons showed depolarized plateaus lasting from 22 to 220 ms. We analyzed the decay of these long depolarizing plateaus, as shown in Fig. 4A for a representative layer V pyramidal neuron, and compared it with the time course of development of inactivation of the INaP recorded in the same neuron (Fig. 4B). Both decays could be fitted using a biexponential function with a similar fast time constant (
1 values of 552.4 and 490.4 ms, respectively). The slow time constant of the voltage plateau decay was shorter (2 = 4,252.8 ms) than that of development of INaP inactivation (9,155.6 ms), probably because of the abrupt end (about 7 s from its onset) of the AP shoulder. We did not investigate the mechanism of the abrupt end of the depolarizing plateau, which is frequently observed in cells recorded under similar experimental conditions and may have arisen from residual unblocked voltage-dependent hyperpolarizing currents or K+ currents activated by persistent Na+ entry into the cell.
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With the aim of directly assessing the relationship between INaP properties and different physiological firing characteristics, in some pyramidal neurons we recorded in sequence with a KGluconate-based solution, the physiological firing (in standard ACSF), the firing with perfusion of Ca2+ and K+ channel blockers and, switching to voltage clamp, the properties of INaP.
In response to the injection of depolarizing pulses, all neurons recorded in layer II/III (n = 5) discharged with individual APs, each followed by postexcitatory hyperpolarization and showing a more or less robust firing frequency adaptation (Fig. 6A1). Three of the seven neurons recorded in layer V showed a firing behavior similar to that of layer II/III neurons (Fig. 6B1). Four pyramidal layer V neurons discharged with nonadapting individual APs, each followed by a prominent depolarized potential (Fig. 6C1, arrow), that were preceded by an early burst in two neurons. In these neurons, the AP bursts also appeared as an all-or-none response to short depolarizing stimuli threshold. During the perfusion of Ca2+ and K+ channel blockers recurrent firing disappeared and a depolarized plateau after an early AP was evident. The depolarized plateau had a shorter duration in adapting regular-spiking pyramidal neurons of both layer II/III and layer V (Fig. 6, A3 and B3), but exceeded 700 ms in both intrinsically bursting and nonadapting layer V pyramidal neurons (Fig. 6C3). In all of the neurons, the curve of the development of inactivation of INaP followed a biexponential decay. The values of the early time constant measured in layer II/III pyramidal neurons was similar to that assessed in neurons recorded directly in voltage-clamp configuration (between 115.0 and 268.3 ms, n = 3). In layer V neurons, the early time constant was substantially slower in intrinsically bursting and nonadapting neurons (between 662.6 and 1,256.3 ms; n = 4), whereas in adapting regular-spiking neurons it showed intermediate values (328.3–332.8 ms; n = 2).
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DISCUSSION |
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In the neocortex, as in other structures, the basic characteristics of INaP are its low threshold of activation and its persistence. The low threshold of activation allows INaP to exert its depolarizing effect at potentials that are more negative than firing levels, thus amplifying low depolarizing inputs such as small excitatory postsynaptic potentials (Schwindt and Crill 1995). The persistence of the current over time allows INaP to maintain its contribution during long depolarizations, sustaining recurrent neuronal firing (Crill 1996; Do and Bean 2003; Stafstrom et al. 1985) or long plateau potentials in hyperexcitable neurons (Bikson et al. 2003; Mantegazza et al. 1998; Segal and Douglas 1997). However, INaP undergoes a slow inactivation process that may critically define its ultimate effect on membrane excitability (Fleidervish and Gutnick 1996; French et al. 1990; Magistretti and Alonso 1999; Vreugdenhil et al. 2004).
Differential INaP properties in pyramidal neurons of layer II/III and V of the sensorimotor cortex
On the basis of findings obtained in cortical neurons (Alzheimer et al. 1993b; Brown et al. 1994), most studies suggest that INaP is generated by a fraction of Na+ channels that escape the fast transition to a nonconductive state affecting most activated Na+ channels. In agreement with this assumption, a small channel fraction remains capable of late openings in the "persistent" mode despite sustained membrane depolarizations. Our data indicate that the characteristics of INaP activation in both layers II/III and V of rat sensorimotor cortex are similar to those previously described in dissociated cortical neurons (Alzheimer et al. 1993b; Brown et al. 1994; Franceschetti et al. 2000; Taverna et al. 1999), neocortical slices (Fleidervish and Gutnick 1996), hippocampus (French et al. 1990), and entorhinal cortex (Magistretti and Alonso 1999), by activating at potentials that are a few millivolts more positive than the spontaneous resting potential. This finding suggests that in all these brain structures INaP plays a similar role in increasing subthreshold membrane excitability, thus facilitating neuronal AP generation. In our experiments, one significant difference between the neocortical layers was the amplitude of the current, which was larger in layer V than that in layer II/III. This difference could be attributed to a different cell size, although the current density calculated on the basis of the cell capacitance as an approximate measure of the membrane surface was also greater in layer V than that in layer II/III. Because of the different voltage dependencies of INaP inactivation in the two layers, at the holding potential of –70 mV used to study the activation properties, INaP is partially inactivated in layer II/III neurons (by 8.8 ± 1.6%; Fig. 2C). Thus we also compared the data applying a correcting factor for the recordings made in layer II/III, but the difference maintained the statistic significance. The larger current amplitude in layer V may well account for the particular firing properties and threshold of excitability of the pyramidal neurons in this layer. In fact, in rat somatosensory cortex, a considerable percentage of layer V neurons typically generate high-frequency bursts of APs that arise from a prominent subthreshold depolarization and that are produced in an all-or-none fashion in response to very small membrane depolarization (Connors and Gutnick 1990).
Unlike activation, INaP inactivation properties were significantly different in the pyramidal neurons of the two neocortical layers. Moreover, the parameters describing both the voltage and the time dependencies of inactivation were different from those previously reported in other neuronal populations. This undoubtedly partially depends on the applied stimulus protocols (i.e., the length of the conditioning pulses and imposed membrane depolarization), although some evidences suggest that different inactivation kinetics may differentiate the INaP recorded in different structures.
In the entorhinal cortex, INaP completely inactivates after a 15-s conditioning depolarization at a membrane potential of –10 mV (Magistretti and Alonso 1999) and is half-inactivated at about –49 mV. Our findings in neocortical neurons showed that INaP reached its inactivation midpoint at a more depolarized potential in layer V (about –42 mV), thus suggesting that a significant current fraction remains available for sustaining neuronal excitability at rather depolarized membrane potentials. Moreover, a current fraction corresponding to about 25% of maximum INaP amplitude did not inactivate despite long conditioning prepulses to 0 mV. A similar fraction also remained at the end of the longest conditioning pulses we used when evaluating the development of slow inactivation (40 s at –10 mV), which suggests that a fraction of INaP in neocortical neurons is a real "persistent current" and does not obviously inactivate regardless of the extent and duration of the conditioning depolarization.
Courses of time-dependent slow INaP inactivation and recovery from inactivation were fitted by biexponential functions in all of the neurons, thus further indicating that INaP inactivation occurs as complex transitions in sensorimotor cortex. Our inactivating protocol included more data points and inactivating prepulses with very long durations; this probably accounts for the difference between our results and those previously obtained in sensorimotor cortex (Fleidervish and Gutnick 1996) that showed a time-dependent decay well fitted by a simple exponential function. However, development of INaP slow inactivation is also well fitted by a single exponential function in entorhinal cortex (Magistretti and Alonso 1999), despite the use of a stimulus protocol that was quite similar to that used in our experiments. The presence in entorhinal cortex of a peculiar Na+ channel subtype that opens in the persistent mode only (Magistretti and Alonso 2002) may explain the different inactivating behavior of entorhinal neurons with respect to the sensorimotor ones. In neocortical layers, the expression of channel subunit isoforms is uneven (Gong et al. 1999; Whitaker et al. 2000) and the level of persistent current that they generate may be modulated (Mantegazza et al. 2005); this may account for the differences that we observed in slow inactivation in layers II/III and V, without the need to hypothesize the presence of novel and specific channel subtypes opening in persistent mode only.
Our experiments were designed to characterize INaP inactivation and compare its time course in different neocortical layers, but we did not investigate the similar process affecting INaT. However, it is well known that INaT also undergoes slow inactivation processes similar to those we studied for INaP (see Goldin 2003 for a review). Slow inactivation probably arises from conformational channel changes (mainly involving the channel pore), which substantially differ from those of fast inactivation and are likely to involve Na+ channels regardless of their "transient" or "persistent" opening. Slow Na+ channel inactivation modulates firing properties and oscillatory activities and seems to occur with different kinetics depending on channel location (i.e., cell soma versus dendrites) or molecular characteristics of channel subunits. Interestingly, abnormalities in slow Na+ channel inactivation in skeletal muscle and cardiac Na+ channels account for genetically determined human diseases, including hyperkalemic periodic paralysis (Bendahhou et al. 2002; Hayward et al. 1999) and Brugada syndrome (Veldkamp et al. 2000). Moreover, defective sodium channel slow inactivation may contribute to the inherited neuronal dysfunctions leading to epileptic disorders (Lossin et al. 2002; Spampanato et al. 2001). These observations underline the importance of slow inactivation processes and their specific kinetic properties in the neocortex, which may contribute to the specific ability of neocortical subpopulations to sustain neuronal synchronization and epileptogenesis (Chagnac-Amitai and Connors 1989).
In our experiments, we made whole cell recordings from the cell soma and we cannot exclude the possibility of a special contribution of dendritic Na+ channels to the INaP recorded in layer V. Moreover, neurons in this layer are morphologically and electrotonically more complex than those in layer II/III, and this in theory may bias the measurements. However, the properties of the activation and of the recovery from inactivation of INaP in the two layers are almost identical. Thus the differences observed in the other characteristics can be considered as specific features of INaP in different layers that can have considerable effects on the excitable properties of the superficial versus deep neocortical layers.
Physiological significance of the different properties of INaP in layers II/III and V
Pyramidal neurons are the principal elements of neocortical circuitry because they are recipients of the input system, the source of local excitatory circuits, and the sole output of the neocortex. The fine modulation of their firing properties is therefore fundamental to elaborating inputs and generating appropriate outputs.
Pyramidal cells can generate a series of individual APs in all neocortical layers of mammals, whereas a large subset of layer V pyramidal cells discharge with a burst of APs, which can be followed by recurrent bursts or by individual APs with a prominent depolarized afterpotential. Moreover, an intermediate class of layer V regular-spiking neurons discharge in response to both threshold and suprathreshold depolarizations with individual APs each followed by a depolarized after potential (RSDAP, according to the definition of Tseng and Prince 1993). These two neuronal phenotypes share similar morphological characteristics that are already visible in young rats (Franceschetti et al. 1998; Kasper et al. 1994) and contribute with their rhythmic firing behavior to intracortical synchronization (Silva et al. 1991). In both these neurons, the extremely long (TTX-sensitive) plateau potential observed in the presence of K+ and Ca2+ channel blockers (Franceschetti et al. 1995; Stafstrom et al. 1985) is consistent with the especially high contribution of INaP in sustaining their firing behavior. Most layer II/III pyramidal neurons and a subclass of layer V pyramidal neurons conversely discharge with individual APs not followed by a depolarizing afterpotential, and show fire frequency adaptation.
Differences in INaP amplitude and kinetics may dramatically influence the firing properties of these different neuronal subtypes, and the more heterogeneous neuronal population in layer V with respect to layer II/III may explain the more heterogeneous values of INaP amplitude and inactivation kinetics that we measured in neurons of this layer. Accordingly, control experiments that we performed to evaluate the firing properties, before evaluating the time course of INaP inactivation, confirmed the presence of a slower early inactivation time constant in layer V intrinsically bursting and nonadapting RS neurons, with respect to adapting RS neurons of layer V and layer II/III pyramidal neurons.
The APs forming a burst arise from depolarizing envelopes that reach a plateau at a membrane potential of about –50 mV, roughly corresponding to the firing level (Franceschetti et al. 1998). On the basis of our evaluation of the voltage dependency of inactivation in layer V, INaP amplitude is just slightly affected by membrane depolarizations at –50 mV, and this may suitably maintain the plateau potential from which the AP bursts arise. The voltage- and time-dependent properties of INaP inactivation may also contribute to maintaining the burst recurrence or the rhythmic (nonadapting) recurrence of AP characterizing both intrinsically bursting and nonadapting regular-spiking neurons. Conversely, the same mechanism may play a subsidiary role in the adaptation of regular-spiking neurons, which have a quite high firing frequency at the onset of membrane depolarization that quickly declines because of the strong adapting effect of various voltage-dependent and Ca2+-activated K+ currents (Sah and Davies 2000).
The level and duration of membrane depolarization that we imposed during conditioning pulses (many seconds) used to evaluate the kinetics of slow inactivation of INaP largely exceed the range of the real depolarizations expected to occur physiologically in neocortical neurons. However, large and long-lasting depolarizations do occur in the case of pathological events such as epileptic discharges that lead to extreme firing frequencies or long depolarizing plateaus. In this case, the whole range of INaP inactivation kinetics becomes important in determining the characteristics and duration of such "paroxysmal" events. INaP plays an important role in sustaining interictal and ictal epileptic events in various experimental preparations (Bikson et al. 2003; Segal and Douglas 1997; Timofeev et al. 2002, 2004), and similar mechanisms may occur in genetically determined (Lossin et al. 2002) or acquired epilepsies (Vreugdenhil et al. 2004). The extremely long Na+-dependent depolarizing plateau generated by layer V neurons after Ca2+ and K+ current blockade, which we found in the present and previous experiments (Franceschetti et al. 1995, 2000; Taverna et al. 1999), may be an example of a high degree of Na+-dependent hyperexcitability. The decay of these plateau potentials was best-fitted by a biexponential function, and exactly followed the kinetic properties of INaP slow inactivation assessed in the same neurons. Therefore an especially Na+ dependent excitability may account for the greater likelihood of epileptic discharges to originate from layer V neurons (Chagnac-Amitai and Connors 1989; Connors 1984; Hoffman et al. 1994).
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FOOTNOTES |
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Address for reprint requests and other correspondence: S. Franceschetti, C. Besta National Neurological Institute, Via Celoria 11, 20133 Milan, Italy (E-mail: franceschetti{at}istituto-besta.it)
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REFERENCES |
|---|
|
Agrawal N, Hamam BN, Magistretti J, Alonso A, and Ragsdale DS. Persistent sodium channel activity mediates subthreshold membrane potential oscillations and low-threshold spikes in rat entorhinal cortex layer V neurons. Neuroscience 102: 53–64, 2001.[CrossRef][Web of Science][Medline]
Alzheimer C, Schwindt PC, and Crill WE. Postnatal development of a persistent sodium current in pyramidal neurons from rat sensorimotor cortex. J Neurophysiol 69: 290–292, 1993a.
Alzheimer C, Schwindt PC, and Crill WE. Modal gating of Na+ channels as a mechanism of persistent Na+ current in pyramidal neurons from rat and cat sensorimotor cortex. Neuroscience 13: 660–673, 1993b.[Abstract]
Amitai Y. Membrane potential oscillations underlying firing patterns in neocortical neurons. J Neurosci Methods 63: 151–161, 1994.[Medline]
Azouz R, Jensen MS, and Yaary Y. Ionic basis of spike after-depolarization and burst generation in adult rat hippocampal CA1 pyramidal cells. J Physiol 492.3: 211–223, 1996.
Bendahhou S, Cummins TR, Kula RW, Fu YH, and Ptacek LJ. Impairment of slow inactivation as a common mechanism for periodic paralysis in DIIS4-S5. Neurology 58: 1266–1272, 2002.
Bikson M, Hahn PJ, Fox JE, and Jefferys JG. Depolarization block of neurons during maintenance of electrographic seizures. J Neurophysiol 90: 2402–2408, 2003.
Brown AM, Schwindt PC, and Crill WE. Different voltage dependence of transient and persistent Na+ currents is compatible with modal-gating hypothesis for sodium channels. J Neurophysiol 7: 2562–2565, 1994.
Chagnac-Amitai Y and Connors BW. Synchronized excitation and inhibition driven by intrinsically bursting neurones in neocortex. J Neurophysiol 62: 1149–1162, 1989.
Chao TI and Alzheimer C. Effects of phenytoin on the persistent Na+ current of mammalian CNS neurones. Neuroreport 6: 1778–1780, 1995.[Web of Science][Medline]
Connors BW. Initiation of synchronized neuronal bursting in neocortex. Nature 310: 685–687, 1984.[CrossRef][Medline]
Connors BW and Gutnick MJ. Intrinsic firing patterns of diverse neocortical neurons. Trends Neurosci 13: 99–104, 1990.[CrossRef][Web of Science][Medline]
Crill WE. Persistent sodium current in mammalian central neurons. Annu Rev Physiol 58: 349–362, 1996.[CrossRef][Web of Science][Medline]
Do MT and Bean BP. Subthreshold sodium currents and peacemaking of subthalamic neurons, modulation by slow inactivation. Neuron 39: 109–120, 2003.[CrossRef][Web of Science][Medline]
Fleidervish IA and Gutnick MJ. Kinetics of slow inactivation of persistent sodium current in layer V neurons of mouse neocortical slices. J Neurophysiol 76: 2125–2130,1996.
Franceschetti S, Guatteo E, Panzica F, Sancini G, Wanke E, and Avanzini G. Ionic mechanisms underlying burst firing in pyramidal neurons, intracellular study in rat sensorimotor cortex. Brain Res 696: 127–139, 1995.[CrossRef][Web of Science][Medline]
Franceschetti S, Sancini G, Panzica F, Radici C, and Avanzini G. Postnatal differentiation of firing properties and morphological characteristics in layer V pyramidal neurons of the sensorimotor cortex. Neuroscience 83: 1013–1024, 1998.[CrossRef][Web of Science][Medline]
Franceschetti S, Taverna S, Sancini G, Panzica F, Lombardi R, and Avanzini G. Protein kinase C-dependent modulation of Na+ currents increases the excitability of rat neocortical pyramidal neurones. J Physiol 528: 291–304, 2000.
French CR, Sah P, Buckett KJ, and Gage PW. A voltage-dependent persistent sodium current in mammalian hippocampal neurons. J Gen Physiol 95: 1139–1157, 1990.
Gebhardt C, Breustedt JM, Noldner M, Chatterjee SS, and Heinemann U. The antiepileptic drug losigamone decreases the persistent Na+ current in rat hippocampal neurons. Brain Res 920: 27–31, 2001.[CrossRef][Web of Science][Medline]
Goldin AL. Mechanisms of sodium channel inactivation. Curr Opin Neurobiol 13: 284–290, 2003.[CrossRef][Web of Science][Medline]
Gong B, Rhodes KJ, Bekele-Arcuri Z, and Trimmer JS. Type I and type II Na(+) channel alpha-subunit polypeptides exhibit distinct spatial and temporal patterning, and association with auxiliary subunits in rat brain. J Comp Neurol 412: 342–352, 1999.[CrossRef][Web of Science][Medline]
Hayward LJ, Sandoval GM, and Cannon SC. Defective slow inactivation of sodium channels contributes to familial periodic paralysis. Neurology 52: 1447–1453, 1999.
Hoffman SN, Salin PA, and Prince DA. Chronic neocortical epileptogenesis in vitro. J Neurophysiol 71: 1762–1773, 1994.
Hu H, Vervaeke K, and Storm JF. Two forms of electrical resonance at theta frequencies, generated by M-current, h-current and persistent Na+ current in rat hippocampal pyramidal cells. J Physiol 545: 783–805, 2002.
Ju YK, Saint DA, and Gage PW. Hypoxia increases persistent sodium current in rat ventricular myocytes. J Physiol 497: 337–347, 1996.
Kasper EM, Larkman AU, Lubke J, and Blakemore C. Pyramidal neurons in layer 5 of the rat visual cortex. I. Correlation among cell morphology, intrinsic electrophysiological properties, and axon targets. J Comp Neurol 339: 459–474, 1994.[CrossRef][Web of Science][Medline]
Kim U and McCormick DA. Functional and ionic properties of a slow afterhyperpolarization in ferret perigeniculate neurons in vitro. J Neurophysiol 80: 1222–1235, 1998.
Lampl I, Schwindt P, and Crill W. Reduction of cortical pyramidal neuron excitability by the action of phenytoin on persistent Na+ current. J Pharmacol Exp Ther 284: 228–237, 1998.
Lossin C, Wang DW, Rhodes TH, Vanoye CG, and George AL Jr. Molecular basis of an inherited epilepsy. Neuron 34: 877–884, 2002.[CrossRef][Web of Science][Medline]
Magistretti J and Alonso A. Biophysical properties and slow voltage-dependent inactivation of a sustained sodium current in entorhinal cortex layer-II principal neurons, a whole-cell and single-channel study. J Gen Physiol 114: 491–509, 1999.
Magistretti J and Alonso A. Fine gating properties of channels responsible for persistent sodium current generation in entorhinal cortex neurons. J Gen Physiol 120: 855–873, 2002.
Mantegazza M, Franceschetti S, and Avanzini G. Anemone toxin (ATX II)-induced increase in persistent sodium current, effects on the firing properties of rat neocortical pyramidal neurones. J Physiol 507: 105–116, 1998.
Mantegazza M, Yu FH, Powell AJ, Clare JJ, Catterall WA, and Scheuer T. Molecular determinants for modulation of persistent sodium current by G-protein betagamma subunits. J Neurosci 25: 3341–3349, 2005.
Mattia D, Kawasaki H, and Avoli M. Repetitive firing and oscillatory activity of pyramidal-like bursting neurons in the rat subiculum. Exp Brain Res 114: 507–517, 1997.[CrossRef][Web of Science][Medline]
Menendez de la Prida L, Suarez F, and Pozo MA. Electrophysiological and morphological diversity of neurons from the rat subicular complex in vitro. Hippocampus 13: 728–744, 2003.[CrossRef][Web of Science][Medline]
Mittmann T, Linton SM, Schwindt P, and Crill W. Evidence for persistent Na+ current in apical dendrites of rat neocortical neurons from imaging of Na+-sensitive dye. J Neurophysiol 78: 1188–1192, 1997.
Poznanski RR and Bell J. Theoretical analysis of the amplification of synaptic potentials by small clusters of persistent sodium channels in dendrites. Math Biosci 166: 123–147, 2000.[CrossRef][Web of Science][Medline]
Sah P and Davies P. Calcium-activated potassium currents in mammalian neurons. Clin Exp Pharmacol Physiol 27: 657–663, 2000.[CrossRef][Web of Science][Medline]
Schwindt PC and Crill WE. Amplification of synaptic current by persistent sodium conductance in apical dendrite of neocortical neurons. J Neurophysiol 74: 2220–2224, 1995.
Segal MM and Douglas AF. Late sodium channel openings underlying epileptiform activity are preferentially diminished by the anticonvulsant phenytoin. J Neurophysiol 77: 3021–3034, 1997.
Silva LR, Amitai Y, and Connors BW. Intrinsic oscillations of neocortex generated by layer 5 pyramidal neurons. Science 251: 432–435, 1991.
Spadoni F, Hainsworth AH, Mercuri NB, Caputi L, Martella G, Lavaroni F, Bernardi G, and Stefani A. Lamotrigine derivatives and riluzole inhibit INa, P in cortical neurons. Neuroreport 13: 1167–1170, 2002.[CrossRef][Web of Science][Medline]
Spampanato J, Escayg A, Meisler MH, and Goldin AL. Functional effects of two voltage-gated sodium channel mutations that cause generalized epilepsy with febrile seizures plus type 2. Neuroscience 21: 7481–7490, 2001.
Stafstrom CE, Schwindt PC, Chubb MC, and Crill WE. Properties of persistent sodium conductance and calcium conductance of layer V neurons from cat sensorimotor cortex in vitro. J Neurophysiol 53: 153–170, 1985.
Taddese A and Bean BP. Subthreshold sodium current from rapidly inactivating sodium channels drives spontaneous firing of tuberomammillary neurons. Neuron 33: 587–600, 2002.[CrossRef][Web of Science][Medline]
Taverna S, Sancini G, Mantegazza M, Franceschetti S, and Avanzini G. Inhibition of transient and persistent Na+ current fractions by the new anticonvulsant topiramate. J Pharmacol Exp Ther 288: 960–968, 1999.
Timofeev I, Bazhenov M, Sejnowski T, and Steriade M. Cortical hyperpolarization-activated depolarizing current takes part in the generation of focal paroxysmal activities. Proc Natl Acad Sci USA 99: 9533–9537, 2002.
Timofeev I, Grenier F, and Steriade M. Contribution of intrinsic neuronal factors in the generation of cortically driven electrographic seizures. J Neurophysiol 92: 1133–1143, 2004.
Tseng GF and Prince DA. Heterogeneity of rat corticospinal neurons. J Comp Neurol 335: 92–108, 1993.[CrossRef][Web of Science][Medline]
Vajda FJ. Neuroprotection and neurodegenerative disease. J Clin Neurosci 9: 4–8, 2002.[CrossRef][Web of Science][Medline]
Veldkamp MW, Viswanathan PC, Bezzina C, Baartscheer A, Wilde AA, and Balser JR. Two distinct congenital arrhythmias evoked by a multidysfunctional Na+ channel. Circ Res 86: E91–E97, 2000.
Vreugdenhil M, Hoogland G, van Veelen CW, and Wadman WJ. Persistent sodium current in subicular neurons isolated from patients with temporal lobe epilepsy. Eur J Neurosci 19: 2769–2778, 2004.[CrossRef][Web of Science][Medline]
Whitaker WR, Clare JJ, Powell AJ, Chen YH, Faull RL, and Emson PC. Distribution of voltage-gated sodium channel alpha-subunit and beta-subunit mRNAs in human hippocampal formation, cortex, and cerebellum. J Comp Neurol 422: 123–139, 2000.[CrossRef][Web of Science][Medline]
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