Effects of kainic and other amino acids on synaptic excitation in rat hippocampal slices: 1. Extracellular analysis

The actions of kainic acid on excitatory synaptic responses in rat hippocampal slices have been investigated and compared with the effects of other excitatory amino acids. Kainate administered iontophoretically or via the superfusate produced a large and long lasting potentiation of the population spike evoked in the CA1 region by Schaffer collateral-commissural stimulation. This potentiation was associated with a reduction in the field EPSP recorded in the dendritic region (stratum radiatum) but with no change in the presynaptic fibre volley or with any long lasting alteration in the antidromic population spike. The results suggest that one effect of kainate may be to produce dendritic depolarisation in CA1 pyramidal neurones. Kainate in equivalent amounts elicited similar potentiations of the population spike recorded in the dentate gyrus in response to either lateral or medial perforant path stimulation. Smaller amounts of kainate than those required to affect either CA1 or dentate pathways were able to potentiate the mossy fibre-evoked population spike in the CA3 region. Folic acid, which shares kainate's ability to produce seizures and distant brain damage when injected into the brain, elicited similar potentiations of synaptic excitation. Higher doses of folate than of kainate were required which is consistent with its weaker epileptogenic actions in vivo. In contrast, N-methyl-aspartate, ibotenate, L-glutamate and L-aspartate were unable to mimic kainate's potentiating action. In higher doses the substances depressed the population spike for long periods. These data suggest that potentiation of synaptic events may underlie the ability of kainate (and folate) to elicit seizures and distant brain damage.     

Contributions of mossy fiber and CA1 pyramidal cell sprouting to dentate granule cell hyperexcitability in kainic acid-treated hippocampal slice cultures

Axonal sprouting like that of the mossy fibers is commonly associated with temporal lobe epilepsy, but its significance remains uncertain. To investigate the functional consequences of sprouting of mossy fibers and alternative pathways, kainic acid (KA) was used to induce robust mossy fiber sprouting in hippocampal slice cultures. Physiological comparisons documented many similarities in granule cell responses between KA- and vehicle-treated cultures, including: seizures, epileptiform bursts, and spontaneous excitatory postsynaptic currents (sEPSCs) >600 pA. GABAergic control and contribution of glutamatergic synaptic transmission were similar. Analyses of neurobiotin-filled CA1 pyramidal cells revealed robust axonal sprouting in both vehicle- and KA-treated cultures, which was significantly greater in KA-treated cultures. Hilar stimulation evoked an antidromic population spike followed by variable numbers of postsynaptic potentials (PSPs) and population spikes in both vehicle- and KA-treated cultures. Despite robust mossy fiber sprouting, knife cuts separating CA1 from dentate gyrus virtually abolished EPSPs evoked by hilar stimulation in KA-treated but not vehicle-treated cultures, suggesting a pivotal role of functional afferents from CA1 to dentate gyrus in KA-treated cultures. Together, these findings demonstrate striking hyperexcitability of dentate granule cells in long-term hippocampal slice cultures after treatment with either vehicle or KA. The contribution to hilar-evoked hyperexcitability of granule cells by the unexpected axonal projection from CA1 to dentate in KA-treated cultures reinforces the idea that axonal sprouting may contribute to pathologic hyperexcitability of granule cells.     

Prolonged field bursts in the dentate gyrus: dependence on low calcium, high potassium, and nonsynaptic mechanisms.

1. The dentate gyrus has been proposed to be a gate for entry of neuronal activity into the hippocampus. This function would give it a critical role in the propagation of seizure activity in that region. The hallmark of epileptiform activity in the dentate itself, often referred to as "maximal dentate activation" (MDA), has not been reproduced previously in vitro. 2. With the use of rat hippocampal slices, bath [Ca2+] was decreased, and [K+] was increased concurrently to simulate conditions found during intense neuronal activity in vivo. Both evoked and spontaneous field bursts were observed in the dentate granule cell layer under these conditions. These bursts were similar to MDA, consisting of a prolonged negative shift in extracellular potential with large-amplitude population spikes. 3. In 0.5 mM bath [Ca2+], single stimuli applied to the perforant path could evoke prolonged field bursts in the dentate only when bath [K+] was > or = 9 mM. However, repetitive stimulation (10 Hz) of the perforant path could elicit similar dentate responses when bath [K+] was as low as 5 mM. 4. In 0.5 mM bath [Ca2+], interictal-type bursts appeared spontaneously in CA1 and CA3 when bath [K+] was > or = 5 mM but were lost when [K+] was > 9 mM. Spontaneous seizurelike activity in the dentate required a higher minimum bath [K+] (9 mM) and persisted at [K+] of 11 mM. 5. Stimulation-evoked field bursts in the dentate altered epileptiform activity in CA3. At bath [K+] insufficient to cause spontaneous CA3 bursts, CA3 was activated transiently when prolonged field bursts occurred in the dentate. At higher bath [K+] in which spontaneous CA3 bursts did occur, they were depressed during the dentate bursts. 6. Deletion of Ca2+ from the bath; the addition of 30 microM each of bicuculline methiodide, D,L-2-amino-5-phosphonopentanoate (AP-5), and 6,7-dinitroquinoxaline-2,3-dione (DNQX); or the combination of both manipulations did not block antidromically evoked or spontaneous prolonged field bursts in the dentate. Thus the mechanisms maintaining and propagating these events did not require fast amino acid-mediated synaptic transmission. 7. The concurrent alteration of [K+] and [Ca2+] required to produce prolonged field bursts in the dentate underscores the positive feedback relationship between neuronal excitation and extracellular ionic concentrations, whereas the ability of synaptic stimulation to trigger nonsynaptic seizurelike events such as these prolonged field bursts may be relevant to the transition from interictal to ictal activity in vivo.(ABSTRACT TRUNCATED AT 400 WORDS)     

Abnormal neuronal excitability in hippocampal slices from kindled rats

To determine if electrophysiological properties of hippocampal pathways are altered in kindled rats, extracellular recordings were made from hippocampal slices of rats kindled in the lateral entorhinal cortex and compared with those from implanted but unstimulated controls. Studies were made either 24 h or 28 days after the last kindled seizure and done in normal (3.5 mM) or elevated (7 mM) K+. The preparation of slices, data accumulation, and data analyses were done blind. One day or 28 days after the last kindled seizure, the proportion of slices with spontaneous epileptiform bursts recorded from the CA2/3 region in elevated K+ was significantly (P less than 0.001) increased in the kindled animals. The frequency of spontaneous burst firing was also increased and reached significance (P less than 0.02) at 28 days following the last kindling stimulus. One day after the last kindling stimulus, paired-pulse (GABAergic) inhibition in the CA1 region was decreased (P less than 0.001). Several measures suggested an increased synaptic inhibition in the dentate gyrus of slices from the kindled groups 1 day after kindling. Paired-pulse inhibition was increased (P less than 0.01), the current required to evoke a near-threshold population spike was increased (P less than 0.05), and the population spike amplitude was reduced for a given field excitatory postsynaptic potential (EPSP) (P less than 0.01). Twenty-eight days after the last kindling stimulus, however, paired-pulse inhibition in the dentate was slightly less in slices from kindled rats (P less than 0.005). In other respects the CA1 and dentate regions did not differ between kindled and control groups within 24 h of the last stage V seizure. Thus the maximum amplitudes of presynaptic fiber volley, population spike, and field-excitatory postsynaptic potential (EPSP) slope, and the number of population spikes evoked by a near-maximally effective afferent stimulus, were unchanged. In the CA1 region the input-output curve of field EPSP versus population spike, and the current intensity required to evoke a near-threshold population spike were also unchanged. In addition, no spontaneous bursts were recorded from CA1 in 3.5 mM K+. We conclude that either synapses or neurons intrinsic to the hippocampus are altered by kindling stimuli applied outside this brain area. The transient increase in inhibition in the dentate gyrus suggests that it may reflect a compensatory reaction to kindled seizures. In contrast, the long-lasting (at least 28 days) increase in burst firing in CA2/3 may represent a mechanism for the initiation or propagation of kindled seizures.(ABSTRACT TRUNCATED AT 400 WORDS)     

Induction of paroxysmal discharges in the dentate gyrus: frequency dependence and relationship to afterdischarge production

1. Electrical-stimulus trains activated hippocampal circuits in urethan-anesthetized rats. Responses were monitored with recordings of extracellular potentials and with measurements of the extracellular potassium ([K+]o). Stimulating electrodes were placed in the CA3 region contralateral to the recording electrode (cCA3) and in the ipsilateral angular bundle (AB) while recording in the CA1 pyramidal-cell layer or the granule-cell layer of the dentate gyrus. Patterns of maximal activation were identified. 2. In the CA1 region, maximal activation was indicated by the presence of 10- to 15-mV population spikes, a smooth rise in [K+]o of 5-6 mM above base-line levels of 3 mM, and a negative shift of the sustained DC potential of 2-4 mV. The dentate gyrus was considered to be maximally activated when bursts of large-amplitude (20-40 mV) population spikes were present. These population spikes were associated with a secondary rise in [K+]o to 6-8 mM above base line and an abrupt negative shift of the DC potential of 5-8 mV. 3. Maximal activation depended on the stimulus intensity, frequency, and duration. Trains of 10-Hz stimuli were used to determine the lowest stimulus intensity needed to elicit complete activation in CA1 and in the dentate gyrus. At this intensity, afterdischarges were not produced, and these responses could be elicited repeatedly. 4. By the use of this threshold stimulus intensity, cCA3 stimulation produced a rapid and progressive augmentation of CA1 responses until maximal activation was reached. In comparison, stimulation of the AB could produce the same end result, but the appearance of CA1 population spikes was delayed. Stimulation of the AB produced a steady increase in the evoked granule-cell population spike until the appearance of the paroxysmal large-amplitude population spikes. In contrast, cCA3 stimulation produced two positive evoked responses in the dentate gyrus before the paroxysms of population spikes began. 5. The frequency dependence of the responses was determined by giving stimulus trains ranging from 2 to 100 Hz. Stimulation of cCA3 produced population spikes in CA1 up to 100 Hz that were all associated with the same peak rise in [K+]o. AB stimulation only produced CA1 responses with stimulation frequencies between 15 and 25 Hz. The paroxysmal dentate population spikes were generated by stimulus frequencies between 15 and 30 Hz with both AB and cCA3 stimulation. 6. Unilateral colchicine injections caused a loss of dentate granule cells and a loss of maximal dentate activation on the side of the injection.(ABSTRACT TRUNCATED AT 400 WORDS)     

Does a unique type of CA3 pyramidal cell in primates bypass the dentate gate?

The predominant excitatory synaptic input to the hippocampus arises from entorhinal cortical axons that synapse with dentate granule cells, which in turn synapse with CA3 pyramidal cells.Thus two highly excitable brain areas--the entorhinal cortex and the CA3 field--are separated by dentate granule cells, which have been proposed to function as a gate or filter. However, unlike rats, primates have "dentate" CA3 pyramidal cells with an apical dendrite that extends into the molecular layer of the dentate gyrus, where they could receive strong, monosynaptic, excitatory synaptic input from the entorhinal cortex. To test this possibility, the dentate gyrus molecular layer was stimulated while intracellular recordings were obtained from CA3 pyramidal cells in hippocampal slices from neurologically normal macaque monkeys. Stimulus intensity of the outer molecular layer of the dentate gyrus was standardized by the threshold intensity for evoking a dentate gyrus field potential population spike. Recorded proximal CA3 pyramidal cells were labeled with biocytin, processed with diaminobenzidine for visualization, and classified according to their dendritic morphology. In response to stimulation of the dentate gyrus molecular layer, action potential thresholds were similar in proximal CA3 pyramidal cells with different dendritic morphologies. These findings do not support the hypothesis that dentate CA3 pyramidal cells receive stronger synaptic input from the entorhinal cortex than do other proximal CA3 pyramidal cells.     

Synaptic connections from multiple subfields contribute to granule cell hyperexcitability in hippocampal slice cultures

Limbic status epilepticus and preparation of hippocampal slice cultures both produce cell loss and denervation. This commonality led us to hypothesize that morphological and physiological alterations in hippocampal slice cultures may be similar to those observed in human limbic epilepsy and animal models. To test this hypothesis, we performed electrophysiological and morphological analyses in long-term (postnatal day 11; 40-60 days in vitro) organotypic hippocampal slice cultures. Electrophysiological analyses of dentate granule cell excitability revealed that granule cells in slice cultures were hyperexcitable compared with acute slices from normal rats. In physiological buffer, spontaneous electrographic granule cell seizures were seen in 22% of cultures; in the presence of a GABA(A) receptor antagonist, seizures were documented in 75% of cultures. Hilar stimulation evoked postsynaptic potentials (PSPs) and multiple population spikes in the granule cell layer, which were eliminated by glutamate receptor antagonists, demonstrating the requirement for excitatory synaptic transmission. By contrast, under identical recording conditions, acute hippocampal slices isolated from normal rats exhibited a lack of seizures, and hilar stimulation evoked an isolated population spike without PSPs. To examine the possibility that newly formed excitatory synaptic connections to the dentate gyrus contribute to granule cell hyperexcitability in slice cultures, anatomical labeling and electrophysiological recordings following knife cuts were performed. Anatomical labeling of individual dentate granule, CA3 and CA1 pyramidal cells with neurobiotin illustrated the presence of axonal projections that may provide reciprocal excitatory synaptic connections among these regions and contribute to granule cell hyperexcitability. Knife cuts severing connections between CA1 and the dentate gyrus/CA3c region reduced but did not abolish hilar-evoked excitatory PSPs, suggesting the presence of newly formed, functional synaptic connections to the granule cells from CA1 and CA3 as well as from neurons intrinsic to the dentate gyrus. Many of the electrophysiological and morphological abnormalities reported here for long-term hippocampal slice cultures bear striking similarities to both human and in vivo models, making this in vitro model a simple, powerful system to begin to elucidate the molecular and cellular mechanisms underlying synaptic rearrangements and epileptogenesis.     

Synchronized afterdischarges in the hippocampus: Contribution of local synaptic interactions

In the presence of picrotoxin, spontaneous synchronized bursts followed by afterdischarges were recorded from all pyramidal cell regions of the guinea pig hippocampal slice. Excitatory synaptic potentials, which reversed at approx ?5 mV, were found to be associated with both the initial burst and each afterdischarge. Afterdischarges were reversibly blocked, leaving the initial synchronized burst intact, by the application of several excitatory amino acid antagonists or by increasing Mg²⁺ so that the efficacy of synaptic transmission was reduced. All synchronized activity was suppressed by applying an increased concentration of antagonist or by raising Mg²⁺ and lowering Ca²⁺ so that synaptic transmission was completely blocked.This synchronized neuronal activity occurred spontaneously in the CA2–3 region when isolated from the CA1 pyramidal cell area and the dentate gyrus. When CA2 was separated from CA3 a synchronized rhythm of single bursts was observed in CA2, while a different, slower, synchronized population discharge consisting of initial bursts followed by afterdischarges occurred in CA3. The smallest completely isolated segments of the CA3 field which spontaneously generated synchronized afterdischarges, comparable to those observed in the intact slice, measured 500–700 μm along the stratum pyramidable.It is concluded that these afterdischarges depend on local neuronal interactions mediated by chemical synaptic mechanisms which may occur within a single population of as few as 1000 CA3 pyramidal cells. The results are consistent with a repeated activation of the same group of synapses, which may release an excitatory amino acid neurotransmitter, being responsible for the initiation of each afterdischarge.     

NMDA receptor-dependent plasticity of granule cell spiking in the dentate gyrus of normal and epileptic rats

Because granule cells in the dentate gyrus provide a major synaptic input to pyramidal neurons in the CA3 region of the hippocampus, spike generation by granule cells is likely to have a significant role in hippocampal information processing. Granule cells normally fire in a single-spike mode even when inhibition is blocked and provide single-spike output to CA3 when afferent activity converging into the entorhinal cortex from neocortex, brainstem, and other limbic regions increases. The effects of enhancement of N-methyl-D-aspartate (NMDA) receptor-dependent excitatory synaptic transmission and reduction in gamma-aminobutyric acid-A (GABA(A)) receptor-dependent inhibition on spike generation were examined in granule cells of the dentate gyrus. In contrast to the single-spike mode observed in normal bathing conditions, perforant path stimulation in Mg(2+)-free bathing conditions evoked graded burst discharges in granule cells which increased in duration, amplitude, and number of spikes as a function of stimulus intensity. After burst discharges were evoked during transient exposure to bathing conditions that relieve the Mg(2+) block of the NMDA receptor, there was a marked increase in the NMDA receptor-dependent component of the EPSP, but no significant increase in the non-NMDA receptor-dependent component of the EPSP in normal bathing medium. Supramaximal perforant path stimulation still evoked only a single spike, but granule cell spike generation was immediately converted from a single-spike firing mode to a graded burst discharge mode when inhibition was then reduced. The induction of graded burst discharges in Mg(2+)-free conditions and the expression of burst discharges evoked in normal bathing medium with subsequent disinhibition were both blocked by DL-2-amino-4-phosphonovaleric acid (APV) and were therefore NMDA receptor dependent, in contrast to long-term potentiation (LTP) in the perforant path, which is induced by NMDA receptors and is also expressed by alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionate (AMPA) receptors. The graded burst discharge mode was also observed in granule cells when inhibition was reduced after a single epileptic afterdischarge, which enhances the NMDA receptor-dependent component of evoked synaptic response, and in the dentate gyrus reorganized by mossy fiber sprouting in kindled and kainic acid-treated rats. NMDA receptor-dependent plasticity of granule cell spike generation, which can be distinguished from LTP and induces long-term susceptibility to epileptic burst discharge under conditions of reduced inhibition, could modify information processing in the hippocampus and promote epileptic synchronization by increasing excitatory input into CA3.     

The GABAergic projection of the dentate gyrus to hippocampal area CA3 of the rat: pre- and postsynaptic actions after seizures

The glutamatergic granule cells of the dentate gyrus transiently express GABAergic markers after seizures. Here we show that when this occurs, their activation produces (i) GABA_A receptor-mediated synaptic field responses in CA3, with the physiological and pharmacological characteristics of mossy fibre transmission, and (ii) GABA_A receptor-mediated collateral inhibition. Control hippocampal slices present, on stimulation of the dentate gyrus, population responses in stratum lucidum, which are blocked by ionotropic glutamate receptor antagonists. By contrast, in slices from rats subjected to seizures in vivo , dentate activation additionally produces GABA_A receptor-mediated field synaptic responses in the presence of glutamate receptor antagonists. One-dimensional current source density analysis confirmed the spatial coincidence of the glutamatergic and GABAergic dendritic currents. The GABA_A receptor-mediated field responses show frequency-dependent facilitation and strong inhibition during activation of metabotropic glutamate receptors. In the presence of glutamate receptor blockers, a conditioning pulse delivered to one site of the dentate gyrus inhibits the population synaptic response and the afferent volley provoked by the activation of a second site, in a bicuculline-sensitive manner. In accordance with this, antidromic responses evoked by mossy fibre activation were enhanced by perfusion of bicuculline. Our results suggest that, for GABA receptor-dependent field potentials to be detected, a considerable number of boutons of a well-defined GABAergic pathway should simultaneously release GABA to act on a large number of receptors. Therefore, putative GABA release from the mossy fibres acts on pre- and postsynaptic sites to affect hippocampal activity at the network level after seizures.     

Extracellular pH responses in CA1 and the dentate gyrus during electrical stimulation, seizure discharges, and spreading depression

Since neuronal excitability is sensitive to changes in extracellular pH and there is regional diversity in the changes in extracellular pH during neuronal activity, we examined the activity-dependent extracellular pH changes in the CA1 region and the dentate gyrus. In vivo, in the CA1 region, recurrent epileptiform activity induced by stimulus trains, bicuculline, and kainic acid resulted in biphasic pH shifts, consisting of an initial extracellular alkalinization followed by a slower acidification. In vitro, stimulus trains also evoked biphasic pH shifts in the CA1 region. However, in CA1, seizure activity in vitro induced in the absence of synaptic transmission, by perfusing with 0 Ca(2+)/5 mM K(+) medium, was only associated with extracellular acidification. In the dentate gyrus in vivo, seizure activity induced by stimulation to the angular bundle or by injection of either bicuculline or kainic acid was only associated with extracellular acidification. In vitro, stimulus trains evoked only acidification. In the dentate gyrus in vitro, recurrent epileptiform activity induced in the absence of synaptic transmission by perfusion with 0 Ca(2+)/8 mM K(+) medium was associated with extracellular acidification. To test whether glial cell depolarization plays a role in the regulation of the extracellular pH, slices were perfused with 1 mM barium. Barium increased the amplitude of the initial alkalinization in CA1 and caused the appearance of alkalinization in the dentate gyrus. In both CA1 and the dentate gyrus in vitro, spreading depression was associated with biphasic pH shifts. These results demonstrate that activity-dependent extracellular pH shifts differ between CA1 and dentate gyrus both in vivo and in vitro. The differences in pH fluctuations with neuronal activity might be a marker for the basis of the regional differences in seizure susceptibility between CA1 and the dentate gyrus.     

Excitatory synaptic input to granule cells increases with time after kainate treatment

Temporal lobe epilepsy is usually associated with a latent period and an increased seizure frequency following a precipitating insult. After kainate treatment, the mossy fibers of the dentate gyrus are hypothesized to form recurrent excitatory circuits between granule cells, thus leading to a progressive increase in the excitatory input to granule cells. Three groups of animals were studied as a function of time after kainate treatment: 1-2 wk, 2-4 wk, and 10-51 wk. All the animals studied 10-51 wk after kainate treatment were observed to have repetitive spontaneous seizures. Whole cell patch-clamp recordings in hippocampal slices showed that the amplitude and frequency of spontaneous excitatory postsynaptic currents (EPSCs) in granule cells increased with time after kainate treatment. This increased excitatory synaptic input was correlated with the intensity of the Timm stain in the inner molecular layer (IML). Flash photolysis of caged glutamate applied in the granule cell layer evoked repetitive EPSCs in 10, 32, and 66% of the granule cells at the different times after kainate treatment. When inhibition was reduced with bicuculline, photostimulation of the granule cell layer evoked epileptiform bursts of action potentials only in granule cells from rats 10-51 wk after kainate treatment. These data support the hypothesis that kainate-induced mossy fiber sprouting in the IML results in the progressive formation of aberrant excitatory connections between granule cells. They also suggest that the probability of occurrence of electrographic seizures in the dentate gyrus increases with time after kainate treatment.     

Tetanus-induced re-activation of evoked spiking in the post-ischemic dentate gyrus

This study aimed at investigating and influencing the basic electrophysiological functions and neuronal plasticity in the dentate gyrus in freely moving rats at several time-points after global ischemia. Although neuronal death was induced selectively in the cornu ammonis, subfield 1 (CA1)-region of the hippocampus, we found an additional loss of the population spike in the dentate gyrus after stimulation of the perforant path. Input/output-measurements revealed that as early as 1 day post-ischemia population spike generation in the granular cell layer is greatly decreased when compared with pre-ischemic values and to sham-operated animals, despite an apparently intact morphology of granular cells as evidenced by Nissl-staining. In contrast, the synaptic transmission (excitatory postsynaptic field potential) shows no significant difference when comparing values before and after ischemia and ischemic and sham-operated animals. Despite reduced output function, indicated by very small population spike amplitudes, long lasting potentiation can be induced 10 days after ischemia. Surprisingly, even "silent" populations of neurons, which appear selectively post-ischemia and do not show any evoked population spike, can be re-activated by tetanisation which is followed by a normal appearing long-term potentiation. However, this functional recovery seems to be partial and transient under current conditions: population spike-values do not reach pre-ischemic values and return to the low pre-tetanic baseline values the next day. Electrophysiological measurements ex vivo after ischemia indicate that the neuronal dysfunction in the dentate gyrus is not due to locally destroyed structures but that the activity of granular cells is merely suppressed only under in vivo conditions. In summary, global ischemia leaves a neighboring morphologically intact input area, functionally impaired. However, neuronal function can be partially regenerated by electrophysiological tetanic stimulation.     

Evoked responses of the dentate gyrus during seizures in developing gerbils with inherited epilepsy

When they are 1-2 mo old, domesticated Mongolian gerbils begin having initially mild seizures which become more severe with age. To evaluate the development of this increasing seizure severity, we obtained field potential responses of the dentate gyrus to paired-pulse stimulation of the perforant path during seizures. In 18 gerbils that were 1.5-8.0 mo old, 73 seizures were analyzed. We measured population spike amplitude, the slope of the field excitatory postsynaptic potential (fEPSP), and the population spike amplitude ratio (2nd/1st) to evaluate excitatory and inhibitory synaptic processes. In gerbils <2 mo old, exposure to a novel environment was followed by an increase in population spike amplitude and then by seizure onset, but population spike amplitude ratio and fEPSP slope remained at baseline levels, and multiple population spikes were never evoked. As previously reported for chronically epileptic gerbils, these findings provide little evidence of a disinhibitory seizure-initiating mechanism in the dentate gyrus when young gerbils begin having seizures. In young gerbils evoked responses changed little during the behaviorally mild seizures. In contrast, most seizures in older gerbils included generalized convulsions, postictal depression, and evoked responses that changed dramatically. In older gerbils, shortly after seizure onset the dentate gyrus became hyperexcitable. Population spike amplitude and fEPSP slope peaked, and multiple population spikes were evoked, suggesting that mechanisms for seizure amplification and spread are more developed in older gerbils. Next, dentate gyrus excitability decreased precipitously, and population spike amplitude and fEPSP slope diminished. This period of hypoexcitability began before the end of the seizure, suggesting it may contribute to seizure termination. After the convulsive phase of the seizure, older gerbils remained motionless during a period of postictal depression, and population spike amplitude remained suppressed until the abrupt switch to normal exploratory activity. These findings suggest that the mechanisms of postictal depression may suppress granule cell excitability. The population spike amplitude ratio peaked after the convulsive phase and then gradually returned to the baseline level an average of 12 min after seizure onset, suggesting that granule cell inhibition recovers within minutes after a spontaneous seizure. Although it is unclear whether the seizure-related changes in evoked responses are a cause or an effect of increased seizure severity in older gerbils, their analysis provides clues about developmental changes in the mechanisms of seizure spread and termination.     

Age-dependent reduction of gamma oscillations in the mouse hippocampus in vitro

Normal brain ageing is associated with a decline in hippocampal memory functions. Neuronal oscillations in the gamma frequency band have been implicated in various cognitive tasks. In this study we test the effect of normal brain ageing on gamma oscillations in the mouse hippocampus in vitro. gamma Oscillations were evoked by either 10 microM carbachol or 100 nM kainate in ventral hippocampus slices from young (>5 month) and aged (>22 month) C57Bl/J6 mice. In slices from young mice carbachol-induced gamma oscillations were more regular and more coherent than those induced by kainate. Compared with young, the power in the 20-80 Hz frequency range in area CA3 of slices from aged mice was reduced to 14% for kainate-induced oscillations and to 7% for carbachol-induced oscillations, whereas waveform, dominant frequency and coherence of the oscillation were unchanged. Local network properties were assessed by paired-pulse stimulation of Schaffer collateral/commissural fibers. The excitatory synaptic response in stratum radiatum of CA3 was reduced, in correlation with the antidromic population spike, but functional inhibition in CA3 and CA1 was unaffected. Changes in local network properties could not explain the reduced gamma oscillation strength. Since oscillations driven by two different pathways are similarly affected with age, an age-dependent effect on tonic depolarizing drive of principal cells is unlikely to explain the current results. Other mechanisms, including a change with age in the use-dependent modulation of synaptic strength, should account for the impaired gamma oscillations in the aged hippocampus that may contribute to age-dependent memory impairment.     

Apical and basal orthodromic population spikes in hippocampal CA1 in vivo show different origins and patterns of propagation.

There is controversy concerning whether orthodromic action potentials originate from the apical or basal dendrites of CA1 pyramidal cells in vivo. The participation of the dendrites in the initialization and propagation of population spikes in CA1 of urethan-anesthetized rats in vivo was studied using simultaneously recorded field potentials and current source density (CSD) analysis. CSD analysis revealed that the antidromic population spike, evoked by stimulation of the alveus, invaded in succession, the axon initial segment (stratum oriens), cell body and approximately 200 microm of the proximal apical dendrites. Excitation of the basal dendrites of CA1, following stimulation of CA3 stratum oriens, evoked an orthodromic spike that started near the cell body or initial segment and then propagated approximately 200 microm into the proximal apical dendrites. In contrast, the population spike that followed excitation of the apical dendrites of CA1 initiated at the proximal apical dendrites, 50-100 microm distal to the cell body layer, and then propagated centripetally to the cell body and the proximal basal dendrites. A late apical dendritic spike may arise in the mid-apical dendrites (250-300 microm from the cell layer) and propagated distally. The origin or the pattern of propagation of each population spike type was similar for near-threshold to supramaximal stimulus intensities. In summary, population spikes following apical dendritic and basal dendritic excitation in vivo appeared to originate from different locations. Apical dendritic excitation evoked a population spike that initiated in the proximal apical dendrites while basal dendritic excitation evoked a spike that started near the initial segment or cell body. An original finding of this study is the propagation of the population spike from basal to apical dendrites in vivo or vice versa. This backpropagation from one dendritic tree to the other may play an important role in the synaptic plasticity among a network of CA3 to CA1 neurons.     

Action-potential discharge in hippocampal CA1 pyramidal neurons: current source-density analysis

1. The site of origin of evoked action-potential discharge in hippocampal CA1 pyramidal neurons was investigated using the in vitro rat hippocampal slice preparation. 2. Action-potential discharge in pyramidal cells was evoked by stimulation of efferent pyramidal cell fibers in the alveus (antidromic) or afferent synaptic inputs in stratum oriens (SO) or stratum radiatum (SR). Laminar profiles of evoked extracellular field potentials were recorded at 25-micron intervals along the entire dendrosomatic axis of the pyramidal cell and a one-dimensional current source-density analysis was applied. 3. Suprathreshold stimulation of the alveus evoked an antidromic population spike response and current sink with the shortest peak latency in stratum pyramidale or proximal stratum oriens. A biphasic positive/negative potential associated with a current source/sink was recorded in dendritic regions, with both components increasing in peak latency with distance from the border of stratum pyramidale. 4. Suprathreshold stimulation of SO or SR evoked a population spike response superimposed upon the underlying synaptic depolarization at all levels of the dendrosomatic axis. The shortest latency population spike and current sink were recorded in stratum pyramidale or proximal stratum oriens. In dendritic regions, a biphasic positive/negative potential and current source/sink conducted with increasing latency from the border of stratum pyramidale. 5. A direct comparison of alvear- and SR-evoked responses revealed a basic similarity in population spike potentials and associated sink/source relationships at both the somatic and dendritic level and a similar shift in peak latency of spike components along the pyramidal cell axis. 6. It is concluded that the initial site for generation of a spike along the dendrosomatic axis of the pyramidal cell following antidromic or orthodromic stimulation is in the region of the cell body layer (soma or axon hillock). Action-potential discharge in dendritic regions then occurs as the result of a subsequent retrograde spike invasion of basal and apical dendritic arborizations.     

Chemical modulation of ephaptic activation of CA3 hippocampal pyramids

In rats under urethane anaesthesia, antidromic population spikes were evoked in CA3 pyramidal layer by fimbrial/commissural stimulation at a very low frequency (approximately 0.5 Hz). Submaximal population spikes--between 20 and 90% of maximum--were enhanced by 8-38% by applications of acetylcholine and bicuculline, or by medial septal stimulation. Noradrenaline had a less pronounced and regular facilitatory action, whereas gamma-aminobutyrate and glutamate only depressed population spikes. Maximal enhancement by acetylcholine or bicuculline was observed when the antidromic population spike was initially at 38-53% of maximum amplitude. A simple explanation of these results is that acetylcholine and bicuculline, by raising their excitability, facilitate the excitation of non-invaded pyramidal cells by antidromic field potentials. They are fully in keeping with previous intracellular observations on ephaptic interactions between CA3 neurons, and provide a further illustration, in situ, of the importance of increased excitability and disinhibition--whether caused by drugs or synaptic action--in promoting synchronized excitation by ephaptic currents.     

Differences of neuronal sensitivity to amino acids and related compounds in the rat hippocampal slice

The perfusion of slices of rat hippocampus with solutions containing N-methyl-DL-aspartic acid (NMA), kainic acid, ibotenic acid or quinolinic acid produced a reduction in the size of antidromically evoked population spikes in the CA1 pyramidal cells or dentate gyrus granule cells. The relative potencies of these compounds on CA1 cells compared with granule cells were kainate 3.65, quinolinate 3.46, NMA 2.19 and ibotenate 1.50. Since the former two compounds are known to show a degree of selective toxicity towards the CA1 cells, whereas NMA and ibotenate do not, these results are consistent with the excitotoxic hypothesis that excitation and neurotoxicity are related.     

Immunocytochemical localization of serotonin in intracellularly analyzed and dye-injected ganglion cells of the turtle retina

Epileptiform bursts of population spikes were evoked in the CA1 region of slices of the hippocampus in which the CA3 region had been previously lesioned with kainic acid. D-2-amino-5-phosphonovalerate (D-APV), a specific N-methyl-D-aspartate (NMDA) antagonist, would markedly reduce the number of spikes in the burst but had no effects on the primary population spike or the amplitude of the field excitatory postsynaptic potential (EPSP). In unlesioned control slices only a single population spike was evoked and D-APV had no effect on this response or the field EPSP. Multiple population spike bursts evoked following application of bicuculline to control slices were much less attenuated by D-APV. The results suggest that activation of NMDA receptors contributes to the production of epileptiform activity in the kainic acid-lesioned hippocampus.