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)     

Repetitive perforant-path stimulation induces epileptiform bursts in minislices of dentate gyrus from rats with kainate-induced epilepsy

The epileptic hippocampus has an enhanced propensity for seizure generation, but how spontaneous seizures start is poorly understood. Using whole cell and field-potential recordings, this study explored whether repetitive perforant-path stimulation at physiological frequencies could induce epileptiform bursts in dentate gyrus minislices from rats with kainate-induced epilepsy. Control slices from saline-treated rats responded to single perforant-path stimulation with an excitatory postsynaptic potential (EPSP) and a single population spike in normal medium, and repetitive stimulation at different frequencies (0.1, 1, 2, 5, 10 Hz) did not cause significant increases in the responses. Most minislices (82%) from rats with kainate-induced epilepsy also responded to single perforant-path stimulation with an EPSP and a single population spike/action potential, but some slices (18%) had a more robust response with a prolonged duration and negative DC shift or responses with two to three population spikes. Repetitive perforant-path stimulation at 5-10 Hz, however, transformed the single-spike responses into epileptiform bursts with multiple spikes in half (52%) of the slices, while lower frequency (e.g., ≤ 1 Hz) stimulation failed to produce these changes. The emergence of epileptiform bursts was consistently associated with a negative field-potential DC shift and membrane depolarization. The results suggest that compared with the controls, the "gate" function of the dentate gyrus is compromised in rats with kainate-induced epilepsy, and epileptiform bursts (but not full-length seizure events) can be induced in minislices by repetitive synaptic stimulation at physiological frequencies in the range of hippocampal theta rhythm (i.e., 5-10 Hz).     

Locus coeruleus bursts induced by glutamate trigger delayed perforant path spike amplitude potentiation in the dentate gyrus

Summary Glutamate pressure ejections in the vicinity of locus coeruleus (LC) neurons have been shown to produce both short and long-lasting potentiation of perforant path (PP) evoked population spike amplitude in the dentate gyrus (DG). These effects of LC-glutamate activation resemble those produced by direct application of NE in vitro or in vivo. The present study monitored the cellular response of LC neurons to local glutamate ejections concomitant with stimulation of the PP evoked potential. Double barrel micropipettes or 33 ga cannulaelectrode assemblies permitted LC unit recording and glutamate ejection at or near the same site in urethane anesthetized rats. Glutamate ejections produced a burst of LC activity lasting 250–400 ms and followed by a depression of unit activity lasting 4.6 min. The maximal spike potentiation produced by LC activation was 158%. The first spike to exceed the control range occurred 34 s after the LC burst. Comparable silent intervals in LC unit activity induced by systemic clonidine were not accompanied by population spike amplitude potentiation. The mean duration of potentiation was 4.4 min except in four cases where responses remained potentiated for the duration of the experiment. The duration of potentiation was not correlated with the termination of LC depression. LC units recovered to baseline rates following glutamate induced depression of activity. The occurrence of potentiation appeared to require that glutamate activation reach a critical number of LC neurons since small glutamate ejections could produce a local burst without eliciting potentiation. Long-lasting changes were also related to larger glutamate volumes (100 nl). EPSP slope increases were briefer and occurred less frequently than spike amplitude changes suggesting EPSP-spike dissociation. The delay between the burst of LC activity and amplitude increases in the DG supports a model of NE action in which there are both rapid and slowly developing effects of NE release in the DG. In summary, brief, but intense, activation of LC neurons produces a delayed potentiation of DG responses lasting minutes to hours following the LC burst.     

齿状回层的形成

中枢神经系统大部分结构内的神经元及其纤维成层排列,尤以大脑皮质、小脑皮质和海马(hippocampus)最为明显.这种层状结构对中枢神经系统完成其功能非常重要,皮质神经元排列紊乱的突变小鼠表现出功能上的障碍.我们曾以联合器官型脑片培养技术(organotypic slice co-culture)为主要手段对海马结构内齿状回(dentate gyrus)层的形成进行了深入细致的研究,结果表明,齿状回内细胞和纤维层的形成受不同因子的控制和调节.     

Newborn granule cells in the ageing dentate gyrus

The dentate gyrus of the hippocampus generates neurons throughout life, but adult neurogenesis exhibits a marked age-dependent decline. Although the decrease in the rate of neurogenesis has been extensively documented in the ageing hippocampus, the specific characteristics of dentate granule cells born in such a continuously changing environment have received little attention. We have used retroviral labelling of neural progenitor cells of the adult mouse dentate gyrus to study morphological properties of neurons born at different ages. Dendritic spine density was measured to estimate glutamatergic afferent connectivity. Fully mature neurons born at the age of 2 months display ∼2.3 spines μm⁻¹ and maintain their overall morphology and spine density in 1-year-old mice. Surprisingly, granule cells born in 10-month-old mice, at which time the rate of neurogenesis has decreased by ∼40-fold, reach a density of dendritic spines similar to that of neurons born in young adulthood. Therefore, in spite of the sharp decline in cell proliferation, differentiation and overall neuronal number, the ageing hippocampus presents a suitable environment for new surviving neurons to reach a high level of complexity, comparable to that of all other dentate granule cells.     

Cholecystokinin modulates neurotransmission through the dentate gyrus

The sulfated and unsulfated cholecystokinin (CCK) octapeptide sequences and the pancreatic CCK antagonists, CR 1409 and benzotript, were administered iontophoretically while dentate gyrus granule cell activity was recorded in the anesthetized rat. During application of the compounds, the peri-stimulus time histogram (PSTH) was constructed of granule cell activity coupled to stimulation of the sciatic nerve. CCK and CR 1409, but not benzotript, were found to change significantly the PSTH by enhancing and prolonging the response to sensory stimulation. These results are interpreted as indicating that CCK can modulate impulse flow through the dentate gyrus.     

Electrophysiology of dentate gyrus granule cells

The orthodromic synaptic responses, membrane properties, and responses of dentate gyrus granule cells (DGCs) to several convulsant agents were studied in the in vitro hippocampal slice preparation. Orthodromic stimulation via the perforant pathway (PP) evoked excitatory-inhibitory postsynaptic potentials (EPSP-IPSP) sequences in 27 of 34 DGCs studied. In the majority, only one action potential could be evoked by supramaximal orthodromic stimulation. In recordings from DGC somata, overshooting spikes could be evoked either orthodromically or by current injections. Small-amplitude, fast transients were seen in 5 of 34 DGCs. The current/voltage (I-V) characteristic of most DGCs was linear throughout a range of membrane potentials between 15 and 20 mV negative and 5 and 15 mV positive to the resting potential. At the extremes of this range nonohmic behavior was noted. Exposure of slices to agents that block IPSPs, such as penicillin, bicuculline, picrotoxin, and media containing low Cl- concentrations, eliminated PP-evoked hyperpolarizations in DGCs and prolonged the repolarizing phase of the PP EPSP. In contrast to findings in hippocampal pyramidal cells and neocortical neurons, blockade of IPSPs did not lead to the development of orthodromically evoked slow depolarizations and burst discharges. After slices were exposed to 5 mM tetraethylammonium, current pulses evoked slow spikes, which were resistant to tetrodotoxin and presumably mediated by Ca2+. Spontaneous burst discharges or bursts evoked by brief depolarizing pulses did not occur under these conditions. Substitution of Ba2+ for Ca2+ in the perfusion solution resulted in development of spontaneous slow membrane depolarizations and burst discharges in DGCs. Burst discharges could be directly evoked and spikes were prolonged and resistant to tetrodotoxin (TTX). After hyperpolarizations lasting 200-1,000 ms, associated with a conductance increase and presumably due to a Ca2+-activated K+ conductance, followed directly evoked spike trains in 5 of 20 DGCs. These data suggest that Ca2+ conductances may be evoked in DGCs under certain circumstances but are not prominent during activation of DGCs under standard in vitro recording conditions.(ABSTRACT TRUNCATED AT 400 WORDS)     

Distribution of calretinin immunoreactivity in the mouse dentate gyrus

Calretinin containing elements were visualized with immunocytochemistry in the adult mouse dentate gyrus (DG). In the ventral DG calretinin immunoreactive (CR-IR) large multipolar cells were clustered; they extended between two and four thick cylindrical dendrites which further branched into several thinner processes. Characteristic grape like spiny appendages were occasionally observed on these thick and thinner dendritic processes. On the basis of these structural features these large CR-IR cells were identified as hilar mossy cells. At the supragranular zone a dense CR-IR band was seen, where numerous CR-IR punctae and fibers were packed tightly among putative granule cell dendrites. In the granule cell layer, especially at the dorsal DG, numerous faintly CR-IR cells were located at the interface with the hilus. They were triangular in shape and neither calbindin D28k nor GABA positive, but were immunoreactive for highly polysialylated neural cell adhesion molecule (NCAM-H) and thus considered as newly generated neurons. In the molecular layer CR-IR cells were also scattered; they were mainly located near the pial surface and the hippocampal fissure, small in size, ovoid in shape and usually gave rise to one very thin axon like and one thin cylindrical dendritic process. These cells were assumed to be Cajal-Retzius cells. Throughout the layers, that is, the molecular layer, the granule cell layer and the hilus, CR-IR multipolar and/or fusiform cells were encountered. They resembled those reported in the rat DG in their structural features and usually extended smooth or varicose or sparsely spiny dendritic processes; some of them were confirmed to be GABA-like immunoreactive and/or glutamic acid decarboxylase immunoreactive. The present study showed that CR immunoreactivity in the mouse DG differed significantly from that in the rat and monkey dentate gyri reported previously.     

Synaptic and non-synaptic mechanisms underlying low calcium bursts in the in vitro hippocampal slice

Summary 1. The epileptiform activity generated by lowering extracellular [Ca⁺⁺] was studied in the CA1 subfield of rat hippocampal slices maintained in vitro at 32° C. Extracellular and intracellular recordings were performed with NaCl and KCl filled microelectrodes. 2. Synaptic potentials evoked by stimulation of the stratum radiatum and alveus were blocked upon perfusion with artificial cerebrospinal fluid (ACSF) containing 0.2 mM Ca⁺⁺, 4 mM Mg⁺⁺. Blockade of synaptic potentials was accompanied by the appearance of synchronous field bursts which either occurred spontaneously or could be induced by stimulation of the alveus. 3. Both spontaneous and stimulus-induced low Ca⁺⁺ bursts recorded extracellularly in stratum pyramidale consisted of a negative potential shift with superimposed population spikes. This extracellular event was closely associated with intracellularly recorded action potentials rising from a prolonged depolarization shift. Steady hyperpolarization of the cell membrane potential decreased the amplitude of the depolarizing shift suggesting that synaptic conductance were not involved in the genesis of the low Ca⁺⁺ burst. 4. Spontaneous depolarizing inhibitory potentials recorded in normal ACSF with KCl filled microelectrodes were reduced in size in low Ca⁺⁺ ACSF. However, small amplitude potentials could still be observed at a time when low Ca⁺⁺ bursts were generated by hippocampal CA1 pyramidal neurons. 5. Bicuculline methiodide, an antagonist of -aminobutyric acid (GABA), was capable of modifying the frequency of occurrence and the shape of synchronous field bursts. The effects evoked by bicuculline methiodide were, however, not observed when 81–100% of NaCl was replaced with Na-Methylsulphate. Hence, it was concluded that in low Ca⁺⁺ ACSF even though large release of transmitter such as those following electrical activation of stratum radiatum or alveus cannot be observed, small spontaneous release of the inhibitory transmitter GABA seems to persist. 6. Substitution of NaCl with Na-Methylsulphate also caused changes in the synchronous field bursts which were different from those observed following application of bicuculline methiodide. These findings suggest that in low Ca⁺⁺ ACSF, in addition to residual GABAergic Cl^- mechanisms, non-synaptic Cl^- conductances might play a role in controlling the excitability of hippocampal neurons.Supported by grants from the MRC of Canada (MA-8109) and Sick Children Foundation to MA     

Dendritic atrophy in the dentate gyrus of the senescent rat

Quantitative electron microscopic analysis of the supragranular zone of the dentate gyrus molecular layer has shown that the number, volume fraction and surface area of dendritic shaft profiles are significantly decreased in senescent rats, relative to young adults. These modifications of dendritic morphology, which are not associated with age-related changes in dimensions of the molecular layer or in numbers of granule cells, may result from a decrease in the number and/or length of dendrites. In either case, the decreases in the number, volume fraction and surface area of dendritic shaft profiles found in the dentate gyrus of senescent rats signify an age-related atrophy of dendrites. Comparison of changes in the number and volume fraction of dendritic shaft profiles has demonstrated that age-related dendritic atrophy involves predominantly smaller dendritic branches.     

NMDA receptor dependent paroxysmal discharges in the dentate gyrus

The N-methyl-D-aspartic acid subclass of glutamate receptors is thought to be involved in events that depend on repetitive activation of neurons. The present study shows that repetitive stimulation of CA3 can produce epileptiform discharges in the contralateral dentate gyrus. This distinctive paroxysmal response was blocked by the NMDA antagonists, ketamine and MK-801. The MK-801 blockade required previous activation of the dentate gyrus in the presence of the drug, thus demonstrating, in vivo, a use-dependent effect. These studies show that repetitive, low intensity stimulation of hippocampal circuits can produce NMDA-mediated epileptiform discharges in the dentate gyrus.     

Phaclofen antagonizes post-tetanic disinhibition in the rat dentate gyrus

Tetanic mossy fiber stimulation transiently reduced recurrent inhibition in the rat dentate gyrus. The post-tetanic depression of inhibition was maximal 200 ms after the tetanus and typically lasted for about 2 s. Phaclofen, a selective gamma-aminobutyric acid-B (GABAB) receptor antagonist, significantly increased the post-tetanic level of inhibition. These results suggest that GABAB receptor activation is important for the development of post tetanic disinhibition. We suggest that GABA released during repetitive firing acts on GABAB receptors on inhibitory interneurons to suppress recurrent inhibition.     

Dynamics of neurogenesis in the dentate gyrus of adult rats

Hippocampal neurogenesis declines steadily over the first year of life in the rodent, but the process persists into senescence despite a dramatic drop in the number of neurons it produces. At this point though, the survival and development patterns exhibited by new granule cells in the aging brain remain unclear in relation to patterns observed in the younger brain. The present study was carried out in order to obtain a direct quantitative comparison of hippocampal neurogenesis in juvenile and middle-aged rats with a high degree of temporal resolution, and to compare the survival and differentiation of the new cells over time. Thirty-eight-day-old and 12-month-old, male Sprague--Dawley rats were injected with 5-bromo-2'-deoxyuridine (BrdU) in order to label cells dividing in the dentate gyrus over a 24-h period, and immunohistochemical labeling was performed in order to record cell production and survival at eight different time points over the following two-month period. Using a marker of neuronally committed precursors and immature neurons (doublecortin; DCX), as well as a marker of mature neurons (calbindin d-28K; CaBP), the extent and timeline of neuronal differentiation, maturation, and migration of the new cells were also characterized. Results indicated that 12-month-old rats experienced a nearly 94% reduction in neurogenesis relative to juveniles, due almost entirely to a 92% drop in cell production. A largely preserved course of development and migration in the remaining newborn cells suggests treatments that enhance cell proliferation could be crucial in reversing the age-related decline in neurogenesis.     

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.     

Fast network oscillations in the rat dentate gyrus in vitro

The dentate gyrus is a prominent source of gamma frequency activity in the hippocampal formation in vivo. Here we show that transient epochs of gamma frequency network activity (67 +/- 12 Hz) can be generated in the dentate gyrus of rat hippocampal slices, following brief pressure ejections of a high-molarity potassium solution onto the molecular layer. Oscillatory activity remains synchronized over distances >300 microm and is accompanied by a modest rise in [K(+)](o). Gamma frequency oscillations were abolished by a GABA(A) receptor antagonist demonstrating their dependence on rhythmic inhibition. However, in many cases, higher frequency oscillations (>80 Hz) remained in the absence of synaptic transmission, thus demonstrating that nonsynaptic factors may underlie fast oscillatory activity.     

Loss of synapses in the dentate gyrus of the senescent rat

Synapses were counted in electron micrographs of the middle third of the molecular layer of the dentate gyrus of Fischer 344 rats, 3 months and 25 months of age. A 27% decrease in the number of synapses was found in senescent animals compared with young adults. This loss of synapses could not be correlated with changes in synaptic size, tissue volume or number of postsynaptic granule cells.     

不同发育阶段大鼠齿状回nNOS的表达

目的 :研究nNOS在生后不同发育阶段大鼠齿状回的表达。方法 :免疫组化ABC法及图象分析。结果 :nNOS免疫阳性细胞面数密度 :2周龄组最高 ,其次为 1周龄组 ,2周以后随着年龄的增加逐渐下降 ,3周龄组和 4周龄组之间差异不显著 ;其余各组间差异均显著。nNOS免疫阳性细胞IOD值 :2周龄组最高 ,2周以后随着年龄的增加逐渐下降 ,1周组IOD与 3月组相近。结论 :不同发育阶段大鼠齿状回的nNOS表达的表现出明显的动态变化特征 ,提示NO与齿状回的发育及功能具有密切关系。     

GABAergic signaling to newborn neurons in dentate gyrus

Neurogenesis in the dentate gyrus begins before birth but then continues into adulthood. Consequently, many newborn granule cells must integrate into a preexisting hippocampal network. Little is known about the timing of this process or the characteristics of the first established synapses. We used mice that transiently express enhanced green fluorescent protein in newborn granule cells to examine their synaptic input. Although newborn granule cells had functional glutamate receptors, evoked and spontaneous synaptic currents were exclusively GABAergic with immature characteristics including slow rise and decay phases and depolarized reversal potentials. Synaptic currents in newborn granule cells were relatively insensitive to the GABA(A) receptor modulator zolpidem compared with neighboring mature granule cells. Consistent with the kinetics and pharmacology, newborn granule cells isolated by fluorescent cell sorting lacked the alpha1 GABA(A) receptor subunit. Our results indicate that newborn granule cells initially receive only GABAergic synapses even in the adult.     

Ethanol enhances recurrent inhibition in the dentate gyrus of the hippocampus

Intraperitoneal injection of ethanol (2 g/kg) substantially augmented recurrent inhibition in the dentate gyrus, as measured by population responses to paired-pulse stimulation of the perforant path. In contrast, this dose of ethanol had no significant effect on singly evoked (or conditioning) population spikes. These data indicate that the increased recurrent inhibition by ethanol was not due to a generalized depressant effect, and suggests that at these doses and time points ethanol can selectively alter synaptic transmission in the hippocampus.