On the Activity of the Corticostriatal Networks during Spike-and-Wave Discharges in a Genetic Model of Absence Epilepsy

SWD properties

The intra-SWD frequency, revealed by spectral analysis of the EEG signal, ranged from 7 to 9.7 Hz (7.9 ± 0.1 Hz; n = 517 SWDs from 13 rats) (Fig. 1C). SWDs had a duration of 7 ± 1.3 sec (from 0.6 to 23.1 sec) and recurred once every 9-260 sec (55.4 ± 26 sec). These temporal properties of SWDs, as well as the shape of individual spike-wave complexes, are similar to those described previously under analogous experimental conditions (Pinault et al., 1998; Charpier et al., 1999; Slaght et al., 2002a,b; Pinault, 2003) and in freely moving GAERS (Marescaux et al., 1992).

Intracellular activity of CS neurons

We recorded from CS neurons (n = 4) located in the orofacial motor cortex. The depth of intracellular recordings, between 1200 and 2000 μm from the cortical surface, suggested that CS neurons were located in the deep part of layer III and in layer V. This laminar localization is consistent with previous findings obtained from intracellular labeling of crossed CS neurons (Cowan and Wilson, 1994). CS neurons were electrophysiologically identified by antidromic activation after electrical stimulation of the contralateral striatum (Fig. 1A). Antidromic latencies ranged between 3.2 and 8.6 msec (6.4 ± 1.15 msec; n = 4), and the passive membrane properties of CS neurons, measured during interictal periods, included a membrane potential of -61.3 ± 3.1 mV (n = 4) and an apparent input resistance of 23 ± 3.4 MΩ (n = 4). Action potentials of CS neurons were overshooting, and the firing pattern evoked by intracellular injection of positive square current pulses (Fig. 1A, inset) was characteristic of “regular-spiking” neocortical neurons (Connors and Gutnick, 1990). These electrophysiological properties of CS neurons agree with those measured previously in crossed CS neurons from nonepileptic rats (Cowan and Wilson, 1994; Mahon et al., 2001).

During SWDs, CS neurons exhibited sudden rhythmic membrane depolarizations (Fig. 1B), which were temporally correlated with the spike-wave complexes (Fig. 1E, inset). These repetitive membrane depolarizations, which could trigger action potentials, were superimposed on a tonic membrane hyperpolarization (5-15 mV in amplitude) that lasted for the entire SWD (Fig. 1B). The mean spontaneous firing frequency of CS neurons (Fig. 1D) was increased during the SWDs compared with the interictal periods (interictal, 3.3 ± 1.3 Hz vs SWDs, 4.4 ± 1 Hz; n = 4 cells; p < 0.05). To assess the temporal relationship between the firing of CS neurons and the cortical paroxysms during absence seizures, we measured the timing of individual action potentials in CS neurons relative to the peak negativity of the corresponding spike component in the EEG (Fig. 1E, inset). Action potentials in CS neurons typically preceded the EEG spike component (Fig. 1E) with a mean latency of -19 ± 1.9 msec (n = 200 action potentials from four cells).

Extracellular recording of SONs

Changes in the electrical activity of SONs during SWDs were first examined by extracellular recordings (n = 10 cells). These cells, which form the majority of striatal neurons (Chang et al., 1982), were morphologically identified by juxtacellular injection of neurobiotin (see Materials and Methods). Labeled cells (Fig. 2A1) were located in the striatal projection field of the orofacial motor cortex and displayed the distinctive morphological features of SONs, i.e., those of the type I medium spiny cells (Chang et al., 1982; Kawaguchi, 1993). Briefly, they possessed somata of diameter between 10 and 20 μm, and their dendrites were densely covered with spines apart from their most proximal regions.

The spontaneous firing rate of SONs was dramatically reduced during SWDs. Between cortical paroxysms, SONs discharged action potentials intermittently at a mean frequency of 1.41 ± 1.97 Hz (from 0.022 to 4.9 Hz; n = 10 cells) (Fig. 2A2,A4,B). This firing rate was decreased by 79.4% during SWDs, reaching a mean value of 0.29 ± 0.22 Hz (from 0.02 to 0.68 Hz; n = 10 cells) (Fig. 2A2,A4,B), which corresponded to a probability of firing in association with individual spike-wave complexes of 0.04 ± 0.003 (from 0.004 to 0.1; n = 10 cells). In most neurons (n = 6 of 10), the strong reduction in firing during the ictal epochs was followed by a rebound of excitation manifested by either single or short bursts of two to five action potentials (Fig. 2A2, oblique arrow). As illustrated in Figure 2, A2 and A3, SWDs were accompanied by small-amplitude rhythmic striatal field potentials. This finding strongly suggested that the ictal cortical oscillations, which were associated with rhythmic firing in CS neurons, generated synchronized synaptic oscillations within the striatum. This hypothesis was also supported by the temporal relationships and the degree of association between SWDs and intracellular oscillations in SONs (see below).

Our extracellular recordings provide little information on the behavior of inactive neurons. However, we never recorded extracellular units that discharged action potentials exclusively during SWDs, providing evidence against the recruitment of a striatal neuron population during paroxysmal activity. This hypothesis was confirmed by the intracellular recordings of SONs (see below).

Intracellular activity of SONs

Because action potential firing in SONs is naturally and consistently initiated by excitatory synaptic potentials generated by synchronized corticostriatal inputs (Wilson, 1995; Wilson and Kawaguchi, 1996; Stern et al., 1997; Mahon et al., 2001), this absence of firing during SWDs was unexpected. Therefore, we performed intracellular records (n = 10) to explore cellular and synaptic mechanisms controlling SON activity during absence seizures. We first examined the membrane I-V relationship of SONs to test for changes in their intrinsic excitability, which could account for their apparent hypoexcitability during SWDs. The membrane I-V relationship was obtained, during interictal periods, by measuring membrane potential changes in response to a series of intracellular square current pulses (Fig. 3A). The apparent input resistance, measured from the linear portion of the I-V plot (Fig. 3B, dashed line) was 33 ± 2MΩ (from 25 to 40 MΩ; n = 10 cells), and a marked inward rectification was consistently observed in response to current pulses of increasingly negative intensity (Fig. 3B). Membrane time constants ranged between 3.1 and 7.7 msec (4.9 ± 0.5 msec; n = 10 cells). Suprathreshold positive current pulses evoked a slow ramp-like membrane depolarization (Fig. 3A, arrow) that led to a delay in the first action potential discharge. Action potentials of SONs had an amplitude and duration of 55.4 ± 1.9 mV and 1.6 ± 0.1 msec (n = 10 cells), respectively, and a voltage threshold of -50 ± 0.2 mV (n = 10 cells). Membrane potential of SONs, measured between SWDs, was -76.8 ± 1.4 mV (n = 10 cells). These membrane properties are similar to those described previously in vivo from nonepileptic rats (Wilson, 1995; Charpier et al., 1999; Mahon et al., 2001, 2003). This finding suggests that changed neuronal membrane properties did not underlie the inability of GAERS striatal output neurons to fire action potentials during SWDs. Furthermore, no striking differences were evident in synaptic events that impinged on SONs between paroxysms. Spontaneous events were composed of high-frequency, small-amplitude, depolarizing synaptic potentials (Figs. 4A, 5A, 6A) resembling those recorded in vivo in normal rats under fentanyl (Mahon et al., 2001). This intracellular activity of SONs noticeably differs from the two-state behavior observed under ketamine-xylazine anesthesia, i.e., characteristic shifts of membrane potential between a hyperpolarized downstate and depolarized upstate (Wilson, 1995; Wilson and Kawaguchi, 1996; Stern et al., 1997; Mahon et al., 2001), which are associated with rhythmic large-amplitude EEG waves (Mahon et al., 2001).

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Figure 4.

Intracellular activity of SONs during SWDs. A, The occurrence of an SWD in the EEG (top trace) was accompanied, in the recorded SON (bottom trace), by rhythmic membrane depolarizations superimposed on a sustained hyperpolarization (dashed line). A momentary interruption of the SWD was concomitant with an attenuation of the tonic hyperpolarization and a decrease in amplitude of the rhythmic depolarizations (vertical arrow). The end of ictal activity was succeeded by a rebound of membrane depolarization (oblique arrow). B, The rhythmic membrane depolarizations remained subthreshold for spike discharge. The rhythmic depolarizations (B1, bottom trace) associated with the SWD (B1, top trace) did not cause the cell to fire. The voltage firing threshold (-50 mV) was measured from the same cell during an interictal period (B2, middle trace), by intracellular injection of a positive square current pulse (1 nA) (B2, bottom trace) from a membrane potential corresponding to the preictal level of cell polarization (-79 mV). C, Properties of membrane depolarizations. C1, Expansion (asterisk in B1) of a spike-wave complex (top trace) and its cellular correlate in the simultaneously recorded SON (bottom trace). The large-amplitude depolarization was progressively sculpted by the summation of high-frequency depolarizing events (oblique lines). The full amplitude of the membrane depolarizations (ΔV) was measured (double arrow) from the baseline (dashed line) to the peak potential. C2, Pooled distribution of the amplitude of membrane depolarizations associated with spike-wave complexes (n = 888 from 80 SWDs; bin size, 1 mV; n = 10 cells). Amplitude of cellular depolarizations was measured as indicated in C1. The amplitude distribution was Gaussian (r² = 0.92) and distributed around a mean of 22.8 mV (± 0.18 mV). Records shown in A and B-C1 are from two different neurons that did not spontaneously discharge action potentials.

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Figure 5.

Temporal evolution of the relationships between EEG and intracellular activities of SONs. A, Typical EEG paroxysm (top trace) and its intracellular correlate in a simultaneously recorded SON (bottom trace). B, Time course of the strength of association between the cortical and striatal activities shown in A. The nonlinear correlation indices h² (EEG → SON, dashed line; SON → EEG, continuous line) were calculated for successive 2 sec windows with a temporal overlapping of 98%. C, Corresponding time delays. The time lag values fluctuated before and after the SWD but were relatively constant during the seizure. This result indicates a unidirectional coupling, from the cortex to the SONs, with a delay of ∼15 msec. The traces in A and the corresponding analysis in B and C are temporally aligned.

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Figure 6.

Decrease in SON excitability during SWD-associated rhythmic depolarizations. A, Voltage responses of an SON (middle trace) to repetitive (every 1.5 sec) injection of square current pulses (0.8 nA; bottom trace) applied during interictal (B, C) and ictal (D) periods in the EEG (top trace). B-D, expansion of the traces indicated by the corresponding letters in A. The injected current initiated action potential discharge during the interictal epoch (B, C) but was subthreshold when coincident with a large membrane depolarization (arrow) occurring during the SWD (D). The dashed lines indicate the membrane potential at the start of the pulse. E, The number of current-evoked action potentials during SWDs was significantly decreased compared with interictal periods (n = 10 pulses for each period; p < 0.001). Inset, Superimposition of the records depicted in C and D. Although the current pulse was injected, during the SWD, from a membrane potential slightly more depolarized than that of the interictal period (compare membrane potentials in C and D), it caused a smaller depolarization that remained subthreshold for action potential generation. The action potential evoked during the interictal activity is truncated. Data shown in A-E are from the same cell.

Analysis of the intracellular firing pattern of SONs during SWDs confirmed and extended the observations made during extracellular recordings. Of the 10 intracellularly recorded cells, seven were silent throughout the recording session, including the epochs of cortical paroxysms. In the three remaining neurons, which exhibited a low interictal mean firing rate (from 0.04 to 1 Hz), two were silenced by the cortical SWDs and one displayed a probability of firing, as low as 0.01, in association with the spike-wave complexes. Membrane potential and apparent input resistance of silent and weakly active SONs did not differ significantly (p > 0.1 for each parameter), indicating that the slight difference of firing among SONs was not attributable to distinct intrinsic membrane properties. In addition, the fact that extracellularly recorded SONs can be silenced by the SWDs (Fig. 2A2) demonstrates that cell impalement was not responsible for the inability of SONs to generate action potentials during SWDs.

In all intracellularly recorded SONs, the start of the SWD in the EEG was correlated with an abrupt membrane hyperpolarization of mean amplitude 8.2 ± 0.8 mV (range, 6-12 mV; n = 80 SWDs from 10 cells) (Figs. 4A,B1, 5A, 6A), which persisted throughout the cortical seizure. As illustrated in Figure 4A (vertical arrow), this negative shift in membrane potential could be reduced during a transient attenuation of the ictal EEG waves, demonstrating its functional link with the paroxysmal cortical oscillations. Rhythmic membrane depolarizations were superimposed on the tonic hyperpolarization (Figs. 4A,B1, 5A, 6A) and, despite their large amplitude (22.8 ± 0.2 mV; range, 9-39 mV; n = 888 from 80 SWDs in 10 cells) (Fig. 4A,B1,C1,C2), did not reach the voltage firing threshold, as measured by the intracellular injection of a threshold current pulse during interictal periods (Fig. 4B1,B2). Two major arguments suggest that these rhythmic membrane depolarizations originate synaptically (see also Discussion): (1) they were progressively sculpted by the temporal summation of high-frequency depolarizing potentials (Fig. 4C1), and (2) their frequency corresponded closely to that of spike-wave complexes and was unchanged when membrane potential was modified by maintained current injection (data not shown).

The end of the SWD in the cortical EEG coincided with the cessation of intracellular striatal oscillations and removal of the tonic hyperpolarization (Fig. 4A). In six SONs, this postictal membrane repolarization was immediately followed by an abrupt depolarization leading to a membrane potential more positive than that preceding the ictal activity (Fig. 4A, oblique arrow). This postictal depolarization, measured from the preictal membrane potential to its peak depolarization, had an amplitude of 13.8 ± 1.9 mV (range, 10-23 mV; n = 37 SWDs from six cells) and decayed to preictal potentials within 1 ± 0.2 sec. In two cells, one to three action potentials were generated corresponding to the postictal firing seen in extracellular recordings of SONs (Fig. 2A2).

Association strength and temporal relationship between striatal intracellular oscillations and SWDs

The strength of association (h²; see Material and Methods) between the EEG signal and intracellular activity in SONs increased as absence seizure developed. As illustrated in Figure 5B, the seizure onset was characterized by a gradual increase in the degree of association between the signals, and, after reaching a maximum that was maintained until the end of the crisis, h² progressively recovered its preictal value. In records from five SONs during 11 seizures, the average degree of association between intracellular and EEG signals during the first second of the SWD (h² = 0.47 ± 0.08; n = 11 SWDs) was significantly higher than before the ictal event (h² = 0.22 ± 0.07; p < 0.001). Furthermore, values for time delay extracted by nonlinear correlation of EEG waves, and the intracellular activity of SONs were profoundly modified by the absence seizure (Fig. 5C). Before SWDs, low values for h² indicated that the time delays were highly variable. In contrast, from the first second of the seizure, a clear time lag emerged between intracellular oscillations in SONs and cortical EEG waves and was maintained throughout the SWD (Fig. 5C). For 11 seizures analyzed, the mean delay was 11.4 ± 4.7 msec.

Chloride-dependent inhibition of SONs and activity of striatal GABAergic interneurons

Why did SWD-associated rhythmic membrane depolarizations in SONs not generate action potentials? This could result from an insufficient excitatory synaptic drive to bring the cell to its firing threshold and/or from a shunting inhibition concomitant with the rhythmic excitation. One way to test for the presence of shunting inhibition is to search for changes in membrane input resistance associated with the activation of shunting synapses (Douglas et al., 1988; Pei et al., 1991). Thus, we applied repetitive (every 1.5 sec) positive square current pulses (0.8-1 nA), and we examined, in two SONs, the voltage responses obtained from ictal and interictal periods (Fig. 6A). Because the I-V relationship of the SONs membrane is not linear (Nisenbaum and Wilson, 1995) (Fig. 3), we compared responses induced from similar membrane potentials (Fig. 6B-D). According to this criterion, and given the tonic hyperpolarization of SONs during SWDs, the responses measured during SWDs were superimposed on the rhythmic membrane depolarizations (Fig. 6D). In the experiment illustrated in Figure 6E, the mean number of current-evoked action potentials was significantly (p < 0.001) reduced during the SWD-associated rhythmic depolarizations (0.9 ± 0.1 spikes; n = 10 stimulations) compared with the interictal epochs (2.35 ± 0.9 action potentials; n = 10 stimulations). A similar result was obtained in another cell in which the number of current-induced action potentials was reduced by 76% during absence seizures (interictal, 1.78 ± 0.15 spikes vs ictal, 0.43 ± 0.2 spikes; p < 0.0001). This decrease in current-evoked firing and the associated reduction of the underlying membrane depolarization (Fig. 6E, inset) show that membrane input resistance was decreased during the SWD-associated membrane depolarizations.

To test for a possible role of a chloride-dependent conductance in the decrease of the membrane resistance of SONs during rhythmic depolarizations, intracellular recordings were performed with KCl-filled electrodes (n = 5 SONs). The interictal membrane potential of these neurons was more depolarized (-63 ± 0.12 mV; n = 5 cells; p < 0.001; one-way ANOVA) than that recorded with KAc-filled electrodes, although their apparent input resistance was lower (23.7 ± 0.7 MΩ; n = 3 cells; p < 0.001). Action potential amplitude (57.3 ± 3.8 mV; n = 5 cells) and duration (1.38 ± 0.1 msec; n = 5 cells) of Cl^--loaded SONs were similar to those recorded with KAc-filled electrodes (p > 0.2 for each parameter). As observed with KAc electrodes, Cl^--loaded SONs displayed during the cortical paroxysms a tonic membrane hyperpolarization (5-15 mV) that lasted throughout the SWD (Fig. 7A). The postictal rebound of membrane depolarization was still observed and could trigger one (Fig. 7A, oblique arrow) or multiple action potentials. A remarkable finding was the conversion of the subthreshold oscillations into large-amplitude rhythmic depolarizations (n = 3475 from 84 SWDs) that could generate up to five action potentials (1.6 ± 0.3 spikes per depolarization; n = 3475 depolarizations from five SONs) with a probability of firing of 0.6 ± 0.3 (Fig. 7A,C). The mean frequency and duration of the bursting activity, measured for at least two successive action potentials on a single depolarization, was 172 ± 15 Hz (range, 131-201 Hz; n = 1181 bursts from five SONs) and 9.8 ± 1.2 msec (range, 6.4-13.3 msec; n = 1181 bursts from five SONs), respectively. To assess the temporal relationship between the Cl^--dependent depolarization in SONs and the cortical paroxysms, we compared the timing of all action potentials, generated by the depolarizing Cl^--dependent potentials, with that of the associated EEG spike (Fig. 7B, inset). The corresponding distribution of action potential delays was normally distributed around a mean value of 7.8 ± 0.2 msec (n = 2357 spikes from five SONs) (Fig. 7B).

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Figure 7.

Intracellular activity of SONs recorded with KCl-filled microelectrodes. A, The start and the end of an SWD recorded from a Cl^--loaded SON. In contrast with the recordings obtained with KAc electrodes (see Figs. 4, 5, 6), SONs recorded with KCl electrodes (bottom trace) had an interictal membrane potential (dotted line) slightly more depolarized and, during SWDs (top trace), exhibited rhythmic depolarizing potentials that could generate bursts of action potentials. Two features were preserved: the tonic membrane hyperpolarization (dashed line) and the postictal rebound (oblique arrow). B, Distribution of the delays of the first action potential relative to the EEG spike component (inset). The pooled distribution (n = 2357 action potentials from 5 cells; bin size, 2 msec; mean = 7.8 ± 0.2 msec) was fitted bya Gaussian curve (r² = 0.98). C, Intracellular activity of a Cl^--loaded SON, at the start of an SWD, recorded from the resting potential (C1) and during maintained hyperpolarization to -84 mV by DC injection (C2). The frequency of the rhythmic depolarizations did not depend on the membrane polarization, and the cell still fired action potentials despite the substantial hyperpolarization. Records in A and C are from two different neurons.

When Cl^--loaded SONs were hyperpolarized by direct current (DC) injection (n = 2 SONs), at membrane potentials similar to or more negative than that of SONs recorded with KAc electrodes, the rhythmic depolarizations increased in amplitude and still induced action potential discharge (Fig. 7C). Thus, the rhythmic firing during SWDs did not result from the slight membrane depolarization observed in Cl^--loaded SONs (see above).

Our findings strongly suggest that the decrease in membrane resistance of SONs during SWDs resulted from an increase in Cl^- conductance. Because SONs were normally silenced by the SWDs, this increase could not result from GABAergic Cl^--dependent interactions between SONs. It might instead originate from striatal GABAergic interneurones, which are known to produce a robust Cl^--dependent inhibition able to block the generation of action potentials in SONs (Plenz and Kitai, 1998; Koós and Tepper, 1999). We tested this hypothesis by performing extracellular recordings of striatal GABAergic interneurons (n = 5) during cortical SWDs. These cells were identified by their distinctive short-duration action potentials (<1 msec) and their morphological characteristics, including a round soma (15-20 μm in diameter) and three to eight smooth primary dendrites (Kawaguchi, 1993) (Fig. 8A1). The irregular interictal firing of striatal GABAergic interneurons was converted during SWDs into high-frequency bursts (2.15 ± 0.3 action potentials per burst; n = 5 cells) in-phase with the spike-wave complexes (Fig. 8A2,A3). The duration of bursts was 9.8 ± 1.5 msec (range, 4.6-12.5 msec; n = 5 cells), with an internal frequency of 224.4 ± 23.1 Hz (range, 155-284 Hz; n = 5 cells). The latency of the first action potential measured from the EEG spike component of the corresponding spike-wave complex was 1 ± 0.3 msec (n = 1402 spikes from five cells) (Fig. 8B). A similar analysis using all action potentials in a burst indicated a mean latency to the EEG spike of 4.1 ± 0.2 msec (n = 3175 spikes from 1402 bursts; n = 5 cells) (Fig. 8C).

Sequential electrical events in the corticostriatal pathway during SWDs

The occurrence of SWDs in the EEG was concomitant in CS neurons with an abrupt modification of activity similar to that previously described from a population of GAERS layer V cortical neurons with unidentified subcortical targets (Charpier et al., 1999). This finding indicates that the GAERS CS neurons were not a functionally distinct subset of cells but rather suggests that cortical output neurons exhibit a homogenous electrical behavior during SWDs. The small-amplitude irregular synaptic activity observed in CS neurons during the interictal period was converted during the SWD into rhythmic suprathreshold depolarizations. This rhythmic activity had the same frequency as that of the spike-wave complexes and was concomitant with a tonic membrane hyperpolarization that lasted for the whole duration of the SWD. Because this hyperpolarizing envelope was associated with an increase in the membrane input resistance (Charpier et al., 1999; Slaght et al., 2002b), it was probably attributable to a synaptic disfacilitation, i.e., a reduction of the tonic excitatory synaptic drive, rather than an active synaptic inhibition that should instead increase the membrane conductance (Paré et al., 1998; Timofeev et al., 2001). The frequency of membrane oscillations in CS neurons during SWDs was not modified by DC-induced changes in membrane potential, and their amplitude increased with membrane hyperpolarization (Charpier et al., 1999; Slaght et al., 2002b), strongly suggesting that they are generated by rhythmic depolarizing synaptic inputs. Because large-amplitude EEG waves reflect a synchronization of synaptic potentials in many cortical neurons, and assuming a relatively homogeneous behavior of CS neurons during SWDs, our findings indicate that CS neurons are engaged, during absence seizures, in a tightly synchronized rhythmic excitation leading to coherent firing.

Both extracellular and intracellular recordings revealed that action potential discharge in SONs, which occurred irregularly and with a low frequency during interictal periods, was transiently interrupted during SWDs. This lack of firing was associated with subthreshold rhythmic depolarizations superimposed on a tonic hyperpolarization that lasted for the entirety of the cortical paroxysm. It is very unlikely that the sustained hyperpolarization resulted from a GABAergic synaptic inhibition because it was observed at membrane potentials more negative than the equilibrium potential of Cl^- as measured in SONs (∼-60 mV) (Plenz, 2003) and was not reversed in polarity in chloride-loaded neurons. Alternatively, it could originate from a cessation of excitatory synaptic activity in SONs resulting from a transition in firing of their cortical afferents. The tonic firing of CS neurons during interictal activity could produce a sustained depolarization in SONs attributable to the summation of spontaneous excitatory synaptic events. The phasic synchronized firing in CS neurons during an SWD might allow, between spike-and-wave complexes, a return to the SON resting potential, which depends strongly on inwardly rectifying potassium channels in the absence of strong depolarizing inputs (Nisenbaum and Wilson, 1995; Wilson and Kawaguchi, 1996).

Several lines of evidence strongly suggest that the synaptic oscillations in SONs during SWDs mainly result from rhythmic synchronized discharges in their cortical afferents. First, CS neurons provide powerful excitatory synaptic inputs to SONs (Kincaid et al., 1998; Zheng and Wilson, 2002) responsible for the large spontaneous synaptic depolarizations observed in these neurons in vivo (Wilson, 1995; Wilson and Kawaguchi, 1996; Stern et al., 1997; Charpier et al., 1999; Mahon et al., 2001). Second, the strength of association between cortical field potentials and synaptic depolarizations in SONs was dramatically increased during SWDs. Third, intracellular oscillations in SONs were delayed by ∼30 msec with respect to the discharge of CS neurons (Fig. 9), a temporal shift consistent with the time required for integration of cortical synaptic potentials in SONs (Mahon et al., 2001). Although the activity of the excitatory thalamostriatal neurons (Wilson et al., 1983) during SWDs remains unknown, we cannot exclude their participation in the generation of synaptic oscillations in SONs.

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Figure 9.

Temporal relationship between SWDs and associated cellular activities in the corticostriatal pathway. A, Examples of two successive EEG spike-wave complexes (top trace) and the corresponding intracellular activity in a CS neuron, an SON recorded with a KAc-filled electrode, an SON recorded with a KCl-filled electrode, and the extracellular activity of a striatal GABAergic interneuron (bottom traces). The interneuron and the SON recorded with the KAc-filled electrode were recorded in the same animal. B, Averaged latencies (mean ± SD) of all action potentials related to the EEG spike component used as the zero-time reference (A, dashed line). Because SONs recorded with KAc-filled electrodes were silent during SWDs, we used the time delays measured from the EEG and the intracellular waveforms (Fig. 5C). The number of computed neuron-EEG pairs is indicated in the figure.

An unexpected finding was that the rhythmic striatal depolarizations did not reach the firing threshold. These membrane oscillations were associated with an increase in membrane conductance and, in records with KCl-filled electrodes, were increased in amplitude and could evoke bursts of action potentials. This Cl^--dependent conductance in SONs, concomitant with the rhythmic excitatory inputs, may well result from the activity of striatal GABAergic interneurons that provide strong “feedforward” inhibition to SONs (Plenz and Kitai, 1998; Koós and Tepper, 1999). As illustrated in Figure 9, during SWDs, these interneurons discharged bursts of action potentials that followed the spiking of CS neurons at latencies of ∼20 msec and coincided with the firing of Cl^--loaded SONs. Thus, it is likely that the powerful excitatory synaptic drive provided by the synchronized cortical inputs during SWDs generated bursting patterns in striatal interneurons. However, because thalamic inputs to these cells have been described (Sidibé and Smith, 1999), excitatory thalamostriatal connections may also be involved in the rhythmic excitation of striatal GABAergic interneurons during ictal activity.

Because the membrane potential reached in SONs during SWDs is markedly more negative than the equilibrium potential of Cl^- (see above), GABA-mediated currents could, in principle, have a depolarizing action and therefore facilitate the rhythmic excitatory inputs originating from the cerebral cortex. However, it has been shown recently in pyramidal cortical neurons that such positive interactions between excitatory synaptic potentials and GABA-mediated depolarizing potentials is limited to time periods when the inhibitory conductance change has terminated but the associated membrane depolarization has not yet returned to rest (Gulledge and Stuart, 2003). In contrast, depolarizing inhibitory synaptic potentials have a powerful inhibitory action when excitatory inputs occur simultaneously with the GABA-mediated conductance change (Staley and Mody, 1992; Gulledge and Stuart, 2003). This was the case in the present study in which we found a close temporal overlap during SWDs, between firing in GABAergic interneurons and the rising phase of the rhythmic depolarizations in SONs (Fig. 9). Therefore, although GABA responses in SONs can be depolarizing during SWDs, the increase in membrane conductance, coincident with the cortical inputs, might produce a shunting inhibition that limits membrane depolarization positive to the equilibrium potential of Cl^- and, consequently, prevents SON firing.

The present experiments do not provide a straightforward explanation for the rebound of excitation observed in SONs at the end of the SWDs. However, it could result from a postictal increase in excitability in thalamic and/or cortical neurons (Grenier et al., 1998; Timofeev et al., 2002) afferent to the striatum.

Possible implications for the control of absence seizures

The main finding of the present study is the transient interruption of firing in SONs during absence seizures, which presumably decreases the synaptic inhibition of their targets in the SNr. This should modify the balance between synaptic inhibition and excitation of the SNr neurons, tending to reinforce the actions of excitatory synaptic inputs arising from the subthalamic nucleus (Kita, 1994). This hypothesis is supported by a recent study in freely moving GAERS showing a synchronized bursting firing of SNr neurons in-phase with the spike-wave complexes in the EEG (Deransart et al., 2003).

The possibility that absence seizures are modulated by the GABAergic striatonigral pathway has emerged from pharmacological studies in GAERS, suggesting that an increase or a decrease in the activity of the striatum could reduce or amplify, respectively, the occurrence of SWDs (Depaulis et al., 1988, 1989; Danober et al., 1998; Deransart et al., 1998, 2000, 2001; Deransart and Depaulis, 2002) (see Introduction). Changes in the output of SONs associated with the occurrence and the termination of the SWD could be part of endogenous mechanisms controlling the maintenance and the duration of the abnormal oscillations in the thalamocortical loops during absence seizures. First, the lack of spike discharge in SONs during SWDs should facilitate the bursting activity of SNr neurons and so tend to prolong the seizure. Second, the rebound of excitation that we observed in SONs at the end of SWDs, which is consistent with the decrease of SNr activity at the end of cortical paroxysms (Deransart et al., 2003), could contribute to seizure termination.