Differing actions of baclofen and 3-amino-propylphosphinic acid in rat neocortical slices

Rat neocortical slices maintained in Mg2(+)-free Krebs medium developed spontaneous paroxysmal discharges which were attenuated or suppressed by the gamma-aminobutyric acid-B (GABAB) receptor agonist baclofen, occasionally accompanied by a slight hyperpolarisation, and antagonised by the specific GABAB-receptor antagonist, 2-OH-saclofen. Over the same dose range, the GABA-analogue 3-amino-propylphosphinic acid (3-APA) caused a marked, prompt hyperpolarisation with little or no effect on the frequency of the discharges, although their amplitude was attenuated. In the presence of 2-OH-saclofen, 3-APA still induced a hyperpolarisation but the amplitude of the discharges was no longer affected. This marked difference in action between baclofen and 3-APA in the rat neocortical slices suggests there may be a heterogeneity of GABAB-receptors.     

Suppression of epileptiform burst discharges in CA3 neurons of rat hippocampal slices by the organic calcium channel blocker,verapamil

Summary We studied the effects of the organic calcium channel blocker, verapamil, on spontaneous and bicuculline-induced epileptiform burst discharges in CA3 pyramidal cells of hippocampal slices. A transient increase of burst discharge rate was observed in most cells within 30 min after the addition of verapamil (100 M) to the perfusing medium. Prolonged verapamil perfusions gradually reduced the rate and duration of burst discharges, then abolished them in all tested slices (over periods of 50–150 min) without blocking synaptic transmission. Responses to intracellular injections of current pulses were also gradually affected by verapamil: Action potential amplitude was decreased, action potential duration increased, frequency adaptation increased, amplitude of the fast hyperpolarization following a single action potential decreased, and amplitude and duration of the slow afterhyperpolarization markedly reduced. The amplitude of calcium spikes elicited in slices perfused with tetrodotoxin-containing medium was not affected by verapamil, but the mean velocity of depolarization near the peak of the calcium spike was decreased. Membrane resting potential and input resistance were not affected by verapamil. These results confirm that verapamil is able to suppress epileptiform activity, but suggest that this effect is rather non-specific, due to inhibition of both postsynaptic sodium and calcium conductances.     

Tissue resistance changes and the profile of synchronized neuronal activity during ictal events in the low-calcium model of epilepsy

Population spikes vary in size during prolonged epileptic ("ictal") discharges, indicating variations in neuronal synchronization. Here we investigate the role of changes in tissue electrical resistivity in this process. We used the rat hippocampal slice, low-Ca(2+) model of epilepsy and measured changes in pyramidal layer extracellular resistance during the course of electrographic seizures. During each burst, population spike frequency decreased, whereas amplitude and spatial synchronization increased; after the main discharge, there could be brief secondary discharges that, in contrast with those in the primary discharge, started with high-amplitude population spikes. Mean resistivity increased from 1,231 Omega.cm immediately before the burst to a maximum of 1,507 Omega.cm during the burst. There was no significant increase during the first 0.5-1 s of the field burst, but resistance then increased (tau approximately 5 s), reaching its peak at the end of the burst, and then decayed slowly (tau approximately 10 s). In further experiments, we modulated the efficacy of electrical field effects by changing perfusate osmolarity. Reducing osmolarity by 40-70 mOsm increased preburst resistivity by 19%; it reduced minimum population spike frequency (x0.6-0.7) and increased both maximum population spike amplitude (x1.5-2.3) and spatial synchronization (x1.4-2.5, cross-correlation over 0.5 mm) during bursts. Increasing osmolarity by 20-40 mOsm had the opposite effects. These results suggest that, during each field burst, field effects between neurons gradually become more effective as cells swell, thereby modulating burst dynamics and facilitating the rapid synchronization of secondary discharges.     

Role of EPSPs in initiation of spontaneous synchronized burst firing in rat hippocampal neurons bathed in high potassium

1. Spontaneous discharges that resemble interictal spikes arise in area CA3 b/c of rat hippocampal slices bathed in 8.5 mM [K+]o. Excitatory postsynaptic potentials (EPSPs) also appear at irregular intervals in these cells. The role of local synaptic excitation in burst initiation was examined with intracellular and extracellular recordings from CA3 pyramidal neurons. 2. Most (70%) EPSPs were small (less than 2 mV in amplitude), suggesting that they were the product of quantal release or were evoked by a single presynaptic action potential in another cell. It is unlikely that most EPSPs were evoked by a presynaptic burst of action potentials. Indeed, intrinsic burst firing was not prominent in CA3 b/c pyramidal cells perfused in 8.5 mM [K+]o. 3. The likelihood of occurrence and the amplitude of EPSPs were higher in the 50-ms interval just before the onset of each burst than during a similar interval 250 ms before the burst. This likely reflects increased firing probability of CA3 neurons as they emerge from the afterhyperpolarization (AHP) and conductance shunt associated with the previous burst. 4. Perfusion with 2 microM 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), a potent quisqualate receptor antagonist, decreased the frequency of EPSPs in CA3 b/c neurons from 3.6 +/- 0.9 to 0.9 +/- 0.3 (SE) Hz. Likewise, CNQX reversibly reduced the amplitude of evoked EPSPs in CA3 b/c cells. 5. Spontaneous burst firing in 8.5 mM [K+]o was abolished in 11 of 31 slices perfused with 2 microM CNQX.(ABSTRACT TRUNCATED AT 250 WORDS)     

Activation of postsynaptic GABAB receptors modulates the bursting pattern and synaptic activity of olfactory bulb juxtaglomerular neurons

Olfactory bulb glomeruli are formed by a network of three major types of neurons collectively called juxtaglomerular (JG) cells, which include external tufted (ET), periglomerular (PG), and short axon (SA) cells. There is solid evidence that gamma-aminobutyric acid (GABA) released from PG neurons presynaptically inhibits glutamate release from olfactory nerve terminals via activation of GABA(B) receptors (GABA(B)-Rs). However, it is still unclear whether ET cells have GABA(B)-Rs. We have investigated whether ET cells have functional postsynaptic GABA(B)-Rs using extracellular and whole cell recordings in olfactory bulb slices. In the presence of fast synaptic blockers (CNQX, APV, and gabazine), the GABA(B)-R agonist baclofen either completely inhibited the bursting or reduced the bursting frequency and increased the burst duration and the number of spikes/burst in ET cells. In the presence of fast synaptic blockers and tetrodotoxin, baclofen induced an outward current in ET cells, suggesting a direct postsynaptic effect. Baclofen reduced the frequency and amplitude of spontaneous EPSCs in PG and SA cells. In the presence of sodium and potassium channel blockers, baclofen reduced the frequency of miniature EPSCs, which were inhibited by the calcium channel blocker cadmium. All baclofen effects were reversed by application of the GABA(B)-R antagonist CGP55845. We suggest that activation of GABA(B)-Rs directly inhibits ET cell bursting and decreases excitatory dendrodendritic transmission from ET to PG and SA cells. Thus the postsynaptic GABA(B)-Rs on ET cells may play an important role in shaping the activation pattern of the glomeruli during olfactory coding.     

钙、锶、铜和铁离子对蜗牛食道下神经节放电细胞电生理特性的影响

细胞内微电极电位记录实验法表明:钙离子和锶离子能抑制蜗牛神经细胞的自发放电,延长放电的间隔时间,增加动作电位后超极化电位,但对动作电位的振幅和持续时间无明显影响。二价铜离子抑制动作电位的振幅,延长放电的间隔时间,增加动作电位的持续时间。二价铁离子能使自发放电的神经细胞产生爆发性放电(burst discbarges)。爆发性放电与正常放电相比,在振幅、持续时间和间隔时间等方面都有着明显的差别。这些二价阳离子对神经细胞的静息电位无明显影响。结果表明亚铁离子诱发爆发波可作为在细胞水平研究癫痫发生机制的一个模型。     

GABA(B) modulation of GABA(A) and glycine receptor-mediated synaptic currents in hypoglossal motoneurons

We studied the effects of GABA(B) receptor activation on either glycine or GABA(A) receptor-mediated synaptic transmission to hypoglossal motoneurons (HMs, P8-13) using a rat brainstem slice preparation. Activation of GABA(B) receptors with baclofen, a GABA(B) receptor agonist, inhibited the amplitude of evoked glycine and GABA(A) receptor-mediated inhibitory postsynaptic currents. Additionally, with blockade of postsynaptic GABA(B) receptors baclofen decreased the frequency of both glycine and GABA(A) receptor-mediated spontaneous miniature inhibitory postsynaptic currents (mIPSCs), indicating a presynaptic site of action. Conversely, the GABA(B) receptor antagonist CGP 35348 increased the frequency of glycine receptor-mediated mIPSCs. Application of the GABA transport blocker SKF 89976A decreased the frequency of glycinergic mIPSCs. Lastly, we compared the effects of baclofen on the frequency of glycine and GABA(A) receptor-mediated mIPSC during HM development. At increased postnatal ages (P8-13 versus P1-3) mIPSC frequency was more strongly reduced by baclofen. These results show that presynaptic GABA(B) receptors inhibits glycinergic and GABAergic synaptic transmission to HMs, and the presynaptic sensitivity to baclofen is increased in P8-13 versus P1-3 HMs. Further, endogenous GABA is capable of modulating inhibitory synaptic transmission to HMs.     

Epileptiform burst activity induced by potassium in the hippocampus and its regulation by GABA-mediated inhibition

Intracellular and extracellular recordings were made from pyramidal neurons in hippocampal slices in order to study spontaneous paroxysmal bursting induced by raising the extracellular potassium concentration from 3.5 to 8.5 mM. Extracellular recordings from all hippocampal subfields indicated that spontaneous bursts appeared to originate in region CA3c or CA3b as judged by burst onset. Burst intensity was also greatest in regions CA3b and CA3c and became progressively less toward region CA2. Intracellular recordings indicated that in 8.5 mM potassium, large spontaneous excitatory postsynaptic potentials (EPSPs), large burst afterhyperpolarizations, and rhythmic hyperpolarizing-depolarizing waves of membrane potential were invariably present in CA3c neurons. High potassium (8.5 mM) induced a positive shift (+9 mV) in the reversal potential of GABAergic inhibitory postsynaptic potentials (IPSPs) in CA3c neurons without changing input resistance or resting potential. This resulted in a drastic reduction in amplitude of the IPSP. Reduction of IPSP amplitude occurred before the onset of spontaneous bursting and was reversible upon return to normal potassium. A new technique to quantify the relative intensity of interictal-like burst discharges is described. Pentobarbital, diazepam, and GABA uptake inhibitors, which enhance GABA-mediated synaptic inhibition, reduced the intensity of potassium-induced bursts, whereas the GABA antagonist bicuculline increased burst intensity. Diphenylhydantoin and phenobarbital, anticonvulsants that have little effect on GABAergic inhibition, were without effect on spontaneous bursts. Burst frequency was reduced by bicuculline and 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol but was unaffected by other drugs. Reduction of slice temperature from 35 to 19 degrees C dramatically reduced burst intensity but did not markedly affect burst frequency. We hypothesize that high potassium induces a rise in intracellular chloride concentration, possibly by activating an inward KCl pump or by a passive Donnan effect, which results in a decreased IPSP amplitude. With inhibition suppressed, the large spontaneous EPSPs that appear in high potassium cause individual CA3c neurons to fire. A combination of synaptic and electrical interactions among CA3c cells then synchronizes discharges into interictal spike bursts.     

Presynaptic GABA(B) receptors inhibit synaptic inputs to rat subthalamic neurons.

Effects of baclofen on synaptic transmission were studied in rat subthalamic neurons using whole-cell patch clamp recording from brain slices. Focal electrical stimulation of the brain slice evoked GABAergic inhibitory postsynaptic currents and glutamatergic excitatory postsynaptic currents. Baclofen reduced the amplitude of evoked inhibitory postsynaptic currents in a concentration-dependent manner with an IC(50) of 0.6+/-0.2 microM. Evoked excitatory postsynaptic currents were also reduced by baclofen concentration-dependently (IC(50) of 1.6+/-0.2 microM), but baclofen was more potent at reducing the GABA(A) receptor inhibitory postsynaptic currents. The GABA(B) receptor antagonist CGP 35348 blocked these inhibitory effects of baclofen on evoked inhibitory and excitatory postsynaptic currents. Baclofen increased the paired-pulse ratios of evoked inhibitory and excitatory postsynaptic currents. Furthermore, baclofen reduced the frequency of spontaneous miniature excitatory postsynaptic currents, but had no effect on their amplitude.These results provide evidence for presence of presynaptic GABA(B) receptors that modulate both GABA and glutamate release from afferent terminals in the subthalamus.     

Initial burst of primary endings of isolated mammalian muscle spindles.

The initial burst has been studied in primary endings of isolated mammalian muscle spindles subject to controlled ramp-and-hold stretch. Near the onset of ramp stretch the primary ending discharges at a frequency dependent on stretch velocity. The initial burst is reduced or abolished by repetitive stretch. After block of impulse activity by tetrodotoxin, the receptor potential of primary endings shows an initial component, a rapid depolarization which occurs near the onset of ramp stretch at the same time as the initial burst. This initial component depends, in rate of rise and amplitude, on stretch velocity. It is also reduced or abolished by repetitive stretch. Recording of tension development by the isolated spindle in response to ramp-and-hold stretch shows an early rise in tension associated with the initial burst and the initial component of the receptor potential. This tension rise is also dependent on stretch velocity and is reduced or abolished by repetitive stretch. The results provide direct evidence that the initial burst results from mechanical factors, probably from cross bridge formation between thick and thin filaments as has been suggested (3).     

Differential behavioral state-dependence in the burst properties of CA3 and CA1 neurons

Brief bursts of fast high-frequency action potentials are a signature characteristic of CA3 and CA1 pyramidal neurons. Understanding the factors determining burst and single spiking is potentially significant for sensory representation, synaptic plasticity and epileptogenesis. A variety of models suggest distinct functional roles for burst discharge, and for specific characteristics of the burst in neural coding. However, little in vivo data demonstrate how often and under what conditions CA3 and CA1 actually exhibit burst and single spike discharges. The present study examined burst discharge and single spiking of CA3 and CA1 neurons across distinct behavioral states (awake-immobility and maze-running) in rats. In both CA3 and CA1 spike bursts accounted for less than 20% of all spike events. CA3 neurons exhibited more spikes per burst, greater spike frequency, larger amplitude spikes and more spike amplitude attenuation than CA1 neurons. A major finding of the present study is that the propensity of CA1 neurons to burst was affected by behavioral state, while the propensity of CA3 to burst was not. CA1 neurons exhibited fewer bursts during maze running compared with awake-immobility. In contrast, there were no differences in burst discharge of CA3 neurons. Neurons in both subregions exhibited smaller spike amplitude, fewer spikes per burst, longer inter-spike intervals and greater spike amplitude attenuation within a burst during awake-immobility compared with maze running. These findings demonstrate that the CA1 network is under greater behavioral state-dependent regulation than CA3. The present findings should inform both theoretic and computational models of CA3 and CA1 function.     

Changes in electrical properties of rat myometrium during gestation and following hormonal treatments.

1. The membrane properties of the rat myometrium, during gestation and following ovarian hormone treatment, have been investigated with the micro-electrode technique. 2. Spontaneously generated bursts of electrical activity alternating with silent periods were recorded from non-pregnant, pregnant and post-partum myometria. The membrane potential was highest during the middle stage of gestation, but the spike amplitude within a burst was not uniform. In the final stage of gestation and during parturition, the membrane potential was low and the spikes within a burst were of low frequency and uniform amplitude. 3. During parturition and post-partum, a gradual depolarization of the membrane, accompanied by an increase in membrane resistance, occurred before the generation of a burst. 4. Excitability of the membrane fluctuated from a peak just before the generation of a burst to a low after the cessation of a burst. 5. Displacement of the membrane potential by electrical current or by lowering the temperature modified the slope spontaneous depolarization, but the fluctuations of excitability persisted. The Q10 value for the frequency of spontaneous bursts, measured between 36 and 30 degrees C, was 3-8. 6. Hyperpolarization of the membrane increased the maximum rate of rise of the spike, but beyond -70 mV, the rate of rise was reduced. Half-inactivation of spike generation of spike generation occurred at a membrane potential less negative than the interburst potential, indicating that the current carrying system was not fully activated during parturition. 7. In both normal and spayed rats, oestradiol hyperpolarized the membrane and the burst of spikes was generated hyperpolarized the membrane and the burst of spikes was generated on a sustained depolarization. Progesterone slightly hyperpolarized the membrane and burst discharges occurred without a sustained depolarization. Simultaneous treatment with progesterone and oestradiol produced a plateau potential of long duration during burst discharges. 8. The thickness of the muscle layer, length constant of the tissue and time constant of the membrane were measured during gestation and from spayed rats under various hormonal conditions. The length constant of the tissue was increased by oestradiol and was further increased by simultaneous treatment withoestradiol and progesterone. The increase in tissue thickness appeared to have the most marked influence on the length constant. 9. The resting and active membrane properties of the progresterone treated myometrium were similar to those observed during the middle stages of gestation. The oestradiol-treated myometrium did not resemble that during the last stages of gestation and parturition, which was simulated by combination of the two hormones, oestradiol preceding progesterone.     

Synaptically activated calcium responses in dendrites of hippocampal oriens-alveus interneurons

Activation of metabotropic glutamate receptors (mGluRs) by agonists increases intracellular calcium levels ([Ca(2+)](i)) in interneurons of stratum oriens/alveus (OA) of the hippocampus. We examined the mechanisms that contribute to dendritic Ca(2+) increases in these interneurons during agonist activation of mGluRs and during synaptically evoked burst discharges, using simultaneous whole cell recordings and confocal Ca(2+) imaging in rat hippocampal slices. First, we found that the group I/II mGluR agonist 1S,3R-1-aminocyclopentane-1,3-dicarboxylic acid (ACPD; 100 microM) increased dendritic [Ca(2+)](i) and depolarized OA interneurons. Dendritic Ca(2+) responses were correlated with membrane depolarizations, but Ca(2+) responses induced by ACPD were larger in amplitude than those elicited by equivalent somatic depolarization. Next, we used linescans to measure changes in dendritic [Ca(2+)](i) during synaptically evoked burst discharges and somatically elicited repetitive firing in disinhibited slices. Dendritic Ca(2+) signals and electrophysiological responses were stable over repeated trials. Peak Ca(2+) responses were linearly related to number and frequency of action potentials in burst discharges for both synaptic and somatic stimulation, but the slope of the relationship was steeper for responses evoked somatically. Synaptically evoked [Ca(2+)](i) rises and excitatory postsynaptic potentials were abolished by antagonists of ionotropic glutamate receptors. The group I/II mGluR antagonist S-alpha-methyl-4-carboxyphenylglycine (500 microM) produced a significant partial reduction of synaptically evoked dendritic Ca(2+) responses. The mGluR antagonist did not affect synaptically evoked burst discharges and did not reduce either Ca(2+) responses or burst discharges evoked somatically. Therefore ionotropic glutamate receptors appear necessary for synaptically evoked dendritic Ca(2+) responses, and group I/II mGluRs may contribute partially to these responses. Dendritic [Ca(2+)](i) rises mediated by both ionotropic and metabotropic glutamate receptors may be important for synaptic plasticity and the selective vulnerability to excitotoxicity of OA interneurons.     

Chloride-cotransport blockade desynchronizes neuronal discharge in the "epileptic" hippocampal slice

Antagonism of the chloride-cotransport system in hippocampal slices has been shown to block spontaneous epileptiform (i.e., hypersynchronized) discharges without diminishing excitatory synaptic transmission. Here we test the hypotheses that chloride-cotransport blockade, with furosemide or low-chloride (low-[Cl(-)](o)) medium, desynchronizes the firing activity of neuronal populations and that this desynchronization is mediated through nonsynaptic mechanisms. Spontaneous epileptiform discharges were recorded from the CA1 and CA3 cell body layers of hippocampal slices. Treatment with low-[Cl(-)](o) medium led to cessation of spontaneous synchronized bursting in CA1 >/=5-10 min before its disappearance from CA3. During the time that CA3 continued to burst spontaneously but CA1 was silent, electrical stimulation of the Schaffer collaterals showed that hyperexcited CA1 synaptic responses were maintained. Paired intracellular recordings from CA1 pyramidal cells showed that during low-[Cl(-)](o) treatment, the timing of action potential discharges became desynchronized; desynchronization was identified with phase lags in firing times of action potentials between pairs of neurons as well as a with a broadening and diminution of the CA1 field amplitude. Continued exposure to low-[Cl(-)](o) medium increased the degree of the firing-time phase shifts between pairs of CA1 pyramidal cells until the epileptiform CA1 field potential was abolished completely. Intracellular recordings during 4-aminopyridine (4-AP) treatment showed that prolonged low-[Cl(-)](o) exposure did not diminish the frequency or amplitude of spontaneous postsynaptic potentials. CA3 antidromic responses to Schaffer collateral stimulation were not significantly affected by prolonged low-[Cl(-)](o) exposure. In contrast to CA1, paired intracellular recordings from CA3 pyramidal cells showed that chloride-cotransport blockade did not cause a significant desynchronization of action potential firing times in the CA3 subregion at the time that CA1 synchronous discharge was blocked but did reduce the number of action potentials associated with CA3 burst discharges. These data support our hypothesis that the anti-epileptic effects of chloride-cotransport antagonism in CA1 are mediated through the desynchronization of population activity. We hypothesize that interference with Na(+),K(+),2Cl(-) cotransport results in an increase in extracellular potassium ([K(+)](o)) that reduces the number of action potentials that are able to invade axonal arborizations and varicosities in all hippocampal subregions. This reduced efficacy of presynaptic action potential propagation ultimately leads to a reduction of synaptic drive and a desynchronization of the firing of CA1 pyramidal cells.     

Modulation by GABAB receptors of spontaneous synchronous activities induced by 4-aminopyridine in the rat hippocampus.

Field potential recordings were made in the CA3 and/or CA1 subfields of rat hippocampal slices maintained in vitro during perfusion with medium containing the convulsant drug 4-aminopyridine (4AP, 50 microM). In this experimental condition, spontaneous interictal epileptiform discharges caused by the activation of excitatory amino acid receptors and synchronous, GABA-mediated potentials occurred regularly in both areas. Interictal discharges and GABA-mediated potentials were reduced and eventually abolished in both subfields by bath application of baclofen (0.5-100 microM), which is a selective agonist for the GABAB receptor. However, interictal epileptiform events disappeared in the presence of baclofen concentrations (i.e., 4.75 +/- 0.7 microM; IC50 = 3.4 microM; n = 9) that were lower than those required for abolishing the GABA-mediated potential (i.e., 96.1 +/- 19.4 microM; IC50 = 9.8 microM; n = 12). The effects of baclofen were antagonized by the GABAB receptor antagonists saclofen (1 mM; n = 3 slices) or CGP 35348 (0.2-1 mM; IC50 = 240 microM; n = 12 slices). Our findings indicate that activation of GABAB receptors by baclofen inhibits both types of 4AP-induced activities in the rat hippocampus although epileptiform discharges appear to be more sensitive than GABA-mediated potentials to this pharmacological procedure. Although our data do not indicate whether the action of baclofen is pre- or postsynaptic, they demonstrate that this mechanism is sensitive to available GABAB receptor antagonists.     

Medium-voltage 5-9-Hz oscillations give rise to spike-and-wave discharges in a genetic model of absence epilepsy: in vivo dual extracellular recording of thalamic relay and reticular neurons

In humans with absence epilepsy, spike-and-wave discharges develop in the thalamocortical system during quiet immobile wakefulness or drowsiness. The present study examined the initial stage of the spontaneous development of spike-and-wave discharges in Genetic Absence Epilepsy Rats from Strasbourg. Bilateral electrocorticograms were recorded in epileptic and non-epileptic rats under freely moving and undrugged conditions and under neuroleptanalgesia. Short-lasting episodes of medium-voltage 5-9-Hz (mean=6-Hz) oscillations usually emerged spontaneously from a desynchronized electrocorticogram and in bilateral synchrony in both rat strains. These oscillations were distinguishable from sleep spindles regarding their internal frequency, duration, morphology, and moment of occurrence. Spontaneous spike-and-wave discharges developed from such synchronized medium-voltage oscillations, the spike-and-wave complex occurring at the same frequency as the 5-9-Hz wave. Because the thalamus is thought to play a significant role in the generation of spike-and-wave discharges, dual extracellular recording and juxtacellular labelling of relay and reticular neurons were conducted to study the thalamic cellular mechanisms associated with the generation of spike-and-wave discharges. During medium-voltage 5-9-Hz oscillations, discharges of relay and reticular cells had identical patterns in epileptic and non-epileptic rats, consisting of occasional single action potentials and/or bursts (interburst frequency of up to 6-8 Hz) in relay cells, and of rhythmic bursts (up to 12-15 Hz) in reticular neurons, these discharging in the burst mode almost always before relay neurons. The discharge frequency of reticular bursts decelerated to 6 Hz by the beginning of the spike-and-wave discharges. During these, relay and reticular neurons usually fired in synchrony a single action potential or a high-frequency burst of two or three action potentials and a high-frequency burst, respectively, about 12 ms before the spike component of the spike-and-wave complexes. The frequency of these corresponded to the maximal frequency of the thalamocortical burst discharges associated with 5-9-Hz oscillations. The patterns of relay and reticular phasic cellular firings associated with spike-and-wave discharges had temporal characteristics similar to those associated with medium-voltage 5-9-Hz oscillations, suggesting that these normal and epileptic oscillations are underlain by similar thalamic cellular mechanisms. In conclusion, medium-voltage 5-9-Hz oscillations in the thalamocortical loop give rise to spike-and-wave discharges. Such oscillations are not themselves sufficient to initiate spike-and-wave discharges, meaning that genetic factors render thalamocortical networks prone to generate epileptic electrical activity, possibly by decreasing the excitability threshold in reticular cells. While these GABAergic neurons play a key role in the synchronization of glutamatergic relay neurons during seizures, relay cells may participate significantly in the regulation of the recurrence of the spike-and-wave complex. Furthermore, it is very likely that synchronization of relay and reticular cellular discharges during absence seizures is generated in part by corticothalamic inputs.     

Pertussis toxin pretreatment discriminates between pre- and postsynaptic actions of baclofen in rat dorsal raphe nucleus in vitro

Intracellular recordings were made from rat dorsal raphe neurons in vitro. Baclofen (30 microM) and 5-carboxamidotryptamine (5-CT, 300 nM to 1 microM) hyperpolarized these neurons by 10 and 13 mV, respectively. Depolarizing synaptic potentials (DSPs) were evoked by single shocks: baclofen reduced the amplitude of the DSP by 81%, but 5-CT reduced it by only 23%. The somatic response to iontophoretically applied glutamate pulses was reduced by 12% by baclofen, and 23% by 5-CT. In slices from rats pretreated with intracerebroventricular pertussis toxin (PTX), the ability of baclofen to reduce the DSP was almost unchanged, although the hyperpolarizing action of baclofen, and both actions of 5-CT were virtually eliminated. We conclude that it is possible to distinguish the pre- and postsynaptic actions of baclofen with PTX, and that the actions of 5-CT are both blocked.     

Modulation of burst frequency, duration, and amplitude in the zero-Ca(2+) model of epileptiform activity

Incubation of hippocampal slices in zero-Ca(2+) medium blocks synaptic transmission and results in spontaneous burst discharges. This seizure-like activity is characterized by negative shifts (bursts) in the extracellular field potential and a K(+) wave that propagates across the hippocampus. To isolate factors related to seizure initiation, propagation, and termination, a number of pharmacological agents were tested. K(+) influx and efflux mechanisms where blocked with cesium, barium, tetraethylammonium (TEA), and 4-aminopyridine (4-AP). The effect of the gap junction blockers, heptanol and octanol, on zero-Ca(2+) bursting was evaluated. Neuronal excitability was modulated with tetrodotoxin (TTX), charge screening, and applied electric fields. Glial cell function was examined with a metabolism antagonist (fluroacetate). Neuronal hyperpolarization by cation screening or applied fields decreased burst frequency but did not affect burst amplitude or duration. Heptanol attenuated burst amplitude and duration at low concentration (0.2 mM), and blocked bursting at higher concentration (0.5 mM). CsCl(2) (1 mM) had no effect, whereas high concentrations (1 mM) of BaCl(2) blocked bursting. TEA (25 mM) and low concentration of BaCl(2) (300 microM) resulted in a two- to sixfold increase in burst duration. Fluroacetate also blocked burst activity but only during prolonged application (>3 h). Our results demonstrate that burst frequency, amplitude, and duration can be independently modulated and suggest that neuronal excitability plays a central role in burst initiation, whereas potassium dynamics establish burst amplitude and duration.     

GABA(B) receptor activation promotes seizure activity in the juvenile rat hippocampus.

We analyzed how the GABA(B) receptor agonist baclofen (10-50 microM) influences the activity induced by 4-aminopyridine (4-AP, 50 microM) in the CA3 area of hippocampal slices obtained from 12- to 25-day-old rats. Interictal and ictal discharges along with synchronous GABA-mediated potentials occurred spontaneously in the presence of 4-AP. Baclofen abolished interictal activity (n = 29 slices) and either disclosed (n = 21/29) or prolonged ictal discharges (n = 8/29), whereas GABA-mediated potentials occurred at a decreased rate. The N-methyl-D-aspartate (NMDA) receptor antagonist 3,3-(2-carboxypiperazine-4-yl)-propyl-1-phosphate (CPP, 10 microM, n = 8) did not modify the GABA-mediated potentials or the ictal events recorded in 4-AP + baclofen. In contrast ictal, activity, but not GABA-mediated potentials, was blocked by the non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 microM, n = 5). Most baclofen effects were reversed by the GABA(B) receptor antagonist CGP 35348 (1 mM; n = 4). Baseline and transient increases in [K(+)](o) associated with the 4-AP-induced synchronous activity were unaffected by baclofen. Baclofen hyperpolarized CA3 pyramids (n = 8) recorded with K-acetate-filled electrodes by 4.8 +/- 1.3 mV and made spontaneous, asynchronous hyperpolarizing and depolarizing potentials disappear along with interictal depolarizations. GABA-mediated synchronous long-lasting depolarizations (LLDs) and asynchronous depolarizations were also studied with KCl-filled electrodes in 4-AP + CPP + CNQX (n = 6); under these conditions baclofen did not reduce LLD amplitude but abolished the asynchronous events. Dentate hilus stimulation at 0. 2-0.8 Hz suppressed the ictal activity recorded in 4-AP + baclofen (n = 8). Our data indicate that GABA(B) receptor activation by baclofen decreases transmitter release leading to disappearance of interictal activity along with asynchronous excitatory and inhibitory potentials. By contrast, GABA-mediated LLDs and ictal events, which reflect intense action potential firing invading presynaptic inhibitory and excitatory terminals respectively, are not abolished. We propose that the proconvulsant action of baclofen results from 1) block of asynchronous GABA-mediated potentials causing disinhibition and 2) activity-dependent changes in hippocampal network excitability.     

Low-magnesium epilepsy in rat hippocampal slices: inhibitory postsynaptic potentials in the CA1 subfield

Spontaneous, synchronous epileptiform discharges were recorded in both CA3 and CA1 subfields of rat hippocampal slices perfused with Mg2+-free medium. Surgical separation of the two areas abolished the spontaneous discharges only in the CA1 subfield. However, epileptiform responses in the isolated CA1 subfield could still be evoked by orthodromic stimulation. Intracellularly these stimulus-induced responses were characterized by a depolarization associated with a burst of action potentials. Stimulation of the alveus still evoked a hyperpolarizing potential, presumably a recurrent inhibitory postsynaptic potential (IPSP) in CA1 pyramidal cells. Both spontaneous and stimulus-induced epileptiform discharges were blocked by the selective antagonist of N-methyl-D-aspartate (NMDA) receptors DL-2-amino-phosphonovalerate (APV). APV also reduced the amplitude and duration of the IPSP induced by alveus stimulation. Thus, epileptiform discharges evoked by lowering Mg2+ in the CA1 subfield are associated with a preservation of inhibitory mechanisms. Furthermore the effects exerted by APV upon the IPSP implicate that NMDA receptors might be involved in the neuronal circuit responsible for the hyperpolarizing IPSP generated by CA1 pyramidal neurons.