Development of potentiation in the dentate gyrus of rat: Physiology and anatomy

DUFFY, C. J. AND T. J. TEYLER. Development of potentiation in the dentate gyrus of rat: physiology and anatomy. BRAIN RES. BULL. 3(5) 425–430, 1978.—The physiological development of potentiating processes in the rat dentate gyrus were compared to morphological development. Rapid Golgi techniques were coupled with in vitro studies of dentate granule cell frequency potentiation, post-tetanic potentiation and long-term potentiation. Frequency potentiation and long-term potentiation exhibited a developmental progression between 7 and 210 days postnatal. Posttetanic potentiation remained constant across this period. The relation of these findings to synaptogenesis and dendritic spine formation are discussed.     

Hippocampal kindling induced paired-pulse depression in the dentate gyrus and paired-pulse facilitation in CA3

Rats received high frequency (100 Hz, 1 s) hippocampal stimulations that evoked afterdischarges (ADs) in a partial kindling model of epilepsy. Kindled rats were given 15 ADs over 3 days. Control rats received the same stimulation pulses at 0.17 Hz (low frequency stimulation, LFSs). Subsequent to in vivo stimulations, hippocampal slices were obtained from the rats and extracellular field responses were recorded in the CA3 cell layer following CA1 stratum radiatum stimulation, and in the granule cell layer of the dentate gyrus (DG) following perforant path stimulation. At the DG, the paired-pulse facilitation of the population spike was depressed in the kindled than the control group, on 1 day or 23 days post ADs/LFSs. The excitatory postsynaptic potential (EPSP) responses in DG following single or paired pulses were not different between kindled and control slices. At CA3, paired-pulse facilitation of both the population spike and the population EPSP was increased in the kindled compared to the control group.     

The development and decay of kindling-induced increases in paired-pulse depression in the dentate gyrus

Epileptogenic stimulation (kindling) leads to an increase in recurrent inhibition in the dentate gyrus, possibly due to an increase in benzodiazepine receptors. A second, late inhibitory component also potentiates as a result of kindling. In the present experiments, the time course of the development and decay of this kindling-induced increase in inhibition was studied. Rats were kindled by stimulation of the perforant path or by direct stimulation of the dentate gyrus. Paired-pulse stimulation was applied to the perforant path and field potential measures were taken within the dentate gyrus. These procedures allowed the monitoring of inhibitory events in chronic preparations over prolonged periods. Input/output measures of the baseline responses were also monitored during the course of, and after the completion of kindling. The increase in the early component (about 20-50+ ms) of paired-pulse depression was seen after the first kindling stimulation. The increase in the late component of depression (about 100-500+ ms) did not develop until after about 10 stimulations had been delivered. The late component then decayed more rapidly than the early component after the completion of kindling. The baseline response also showed some indication of depression, particularly in the dentate gyrus kindled group, raising the possibility that feedforward inhibition had also been potentiated.     

Field potential evidence for long-term potentiation of feed-forward inhibition in the rat dentate gyrus

Trains of high-frequency stimulation to the perforant path cause (i) long-term potentiation (LTP) of the population excitatory post-synaptic potential (EPSP), (ii) a lasting increase in the population spike, and (iii) a lasting alteration of the relationship between the EPSP and population spike (E-S relationship), consisting of a decreased x-intercept and decreased slope of the linear regression. To compare the thresholds of these changes, we applied a series of trains, increasing in duration from below LTP threshold. The EPSP potentiated with about the same low threshold as the reduction in E-S slope, whereas the reduction in E-S x-intercept required longer trains. In the second experiment, LTP of the EPSP was reduced by concurrent high-frequency stimulation of the commissural input and a lasting reduction of the population spike height was observed. In a third experiment, picrotoxin, an antagonist of gamma-aminobutyric acid (GABA)-mediated inhibition, blocked the decrease in slope of the E-S relationship which normally accompanies LTP. These results imply that perforant path/granule cell LTP is normally accompanied by long-term potentiation of a feed-forward inhibitory pathway which may involve interneurones.     

Effects of sub-chronic lithium treatment on synaptic plasticity in the dentate gyrus of rat hippocampal slices

Although it has long been suggested that lithium has robust neuroplastic actions, and these actions lead to an enhancement on synaptic plasticity, the effects of lithium treatment on synaptic plasticity have been rarely studied. This study examined the effects of sub-chronic lithium treatment on synaptic plasticity in the dentate gyrus (DG) of hippocampal slices in the rats. Young adult rats were intraperitoneally administered a daily dose of 1 mgEq LiCl or saline-vehicle for 14 days. Twelve hours after the last injections, the input/output (I/0) responses of field excitatory postsynaptic potentials (fEPSP) and the long-term potentiation (LTP) of fEPSP and population spikes (PS) were determined in the DG of hippocampal slices prepared from the animals treated with lithium or vehicle. Treatment of lithium for 14 days significantly increased the I/O responses of fEPSP and the LTP of fEPSP and PS. These results indicate that sub-chronic treatment of lithium increases the excitatory postsynaptic responses, synaptic strength and the cell firing of the granule cells in the DG of the hippocampus.     

A role for monomeric G-proteins in synaptic plasticity in the rat dentate gyrus in vitro

Recent studies have implicated Ras signalling in synaptic plasticity. In this study we have investigated a role for the low molecular weight G proteins Ras, Rap, Ra1 and Rac in long-term potentiation and depression using Clostridium Sordelli Lethal Toxin-82 (LT-82), which inactivates Ras, Rap, Ra1 and Rac, and manumycin A, a Ras inhibitor. Perfusion of hippocampal slices with LT-82 (200 ng/ml) attenuated LTP (83±10%, n=5, P<0.01, compared with controls of 160±11% at 60 min post HFS, n=5). LT-82 had no effect on LTD (63±1% at 100 ng/ml, n=5 and 66±1% at 200 ng/ml, n=4, compared to controls of 56±6%, n=6). Manumycin A (2μM) had no effect on LTP (162±2%, n=5, compared to controls of 167±13%, n=5), but significantly attenuated LTD (88±6%, n=5, P<0.01, compared to controls of 63±9%, n=7). LT-82 (200 ng/ml) significantly increased the amplitude of the isolated NMDA-EPSP at 60 min post-drug application (240±40%, n=5, P<0.01, compared with controls of 100±4%, n=5). However, manumycin A, had no significant effect on NMDAR-EPSP amplitude (92±2%, n=5, compared with controls). These results demonstrate an important role for Ras in LTD and a role for Rap, Ra1 and Rac in LTP.     

Neuropeptide Y inhibits glutamate release and long-term potentiation in rat dentate gyrus

The effect of intracerebroventricular injection of neuropeptide Y (NPY) was assessed on LTP in dentate gyrus. We report that NPY attenuated LTP and inhibited KCl-induced glutamate release in synaptosomes prepared from dentate gyrus. Activity of the stress-activated kinase, c-Jun NH2-terminal kinase (JNK) in synaptosomes was increased by incubation with NPY or following intracerebroventricular injection. Activation of JNK might underlie the inhibitory effect of NPY on LTP.     

Intravenously administered cell-permeant calcium buffer decreases evoked synaptic potentials in rat dentate gyrus in vivo

We examined the effects of the neuroprotective cell-permeant Ca²⁺ buffer, 2-aminophenol-N,N,O-triacetic acid acetoxymethyl ester (APTRA-AM, 20–40 mg/kg), on synaptically evoked potentials in the dentate gyrus of awake rats. Intravenous APTRA-AM (20 mg/kg) decreased the evoked potentials with peak effects ≈6 h after infusion, and recovery to control levels by 24 h. Peak decrease in the population spike (PS) amplitude was by 72±17% of control, and the excitatory postsynaptic potential (EPSP) slope was decreased by 31±12%. APTRA-AM (40 mg/kg), decreased the PS amplitude and EPSP slope by 58±7% and 31±6% of pre-drug levels, respectively. These effects were qualitatively similar to the presynaptically mediated decreases in synaptic potentials previously demonstrated in vitro with APTRA-AM. These results indicate that the cell-permeant Ca²⁺ buffer, APTRA-AM, attenuates hippocampal excitability in vivo, most likely by decreasing synaptic neurotransmission.     

Modulation of paired-pulse responses in the dentate gyrus: effects of prenatal protein malnutrition

Since our major hypothesis is that prenatal protein malnutrition significantly affects hippocampal neuroplasticity, this study examined the effects of prenatal protein malnutrition on the modulation of dentate granule cell excitability in freely moving rats at 15, 30 and 90 days of age across the vigilance states of quiet waking (QW), slow-wave sleep (SWS) and rapid eye movement (REM) sleep. Using paired-pulse stimulation, the paired-pulse index (PPI), a measure of the type and degree of modulation of dentate granule cell excitability elicited by stimulation of the medial perforant path, was obtained for each vigilance state at each stage of development. Four specific measures of granule cell excitability were computed, namely, PPI using both population spike amplitude (PSA) and EPSP slope measures, absolute values of PSA(1) and EPSP(1) slope. PPI values obtained at 15, 30 and 90 days of age, however, were altered during normal ontogenetic development, but not by vigilance state. At 15 days of age, the malnourished group exhibits greater early inhibition of the PPI using the PSA measure at IPIs between 20 and 30 ms regardless of vigilance state, while at 30 days of age, the malnourished group exhibits greater facilitation at IPIs between 50 and 70 ms during QW and SWS, but not during REM sleep. In the control adult (PND90) and juvenile (PND30) animal, PSA(1) values are significantly higher during SWS than in QW or REM sleep. However, for the younger malnourished animals (PND15 and PND30), PSA(1) values were found to be significantly greater during REM sleep rather than SWS. Therefore, as the animal matures, there appears to be a shift in vigilance state dependent synaptic transmission through the hippocampal trisynaptic circuit from REM sleep to SWS in both control and malnourished animals, with the change occurring later in malnourished animals when compared to control ones. Furthermore, our findings suggests that prenatal protein malnutrition significantly alters modulation of dentate granule cell excitability (i.e., PPI values using the PSA measure) during the earlier stages of development but not in adulthood.     

Lonf-term potentiation in the dentate gyrus is preferentially induced at theta rhythm periodicity

In urethane-anesthetized rats, high frequency stimulation was applied to the medial perforant pathway at various time intervals (50, 100, 200, 350 and 500 ms) following stimulation of the same pathway by a single pulase of equal intensity. Recordings of dentate gyrus granule cell evoked responses were made to investigate the range of stimuli that are effective in inducing long-term potentiation (LTP). LTP was induced almost exclusively at the 200 ms interval, corresponding to the periodicity if the theta rhythm. Taken in conjunction with similar findings reported in the CA1 field of the hippocampal slice, these results suggest that the correlation between theta rhythm periodicity and LTP is a general phenomenon within the hippocampal formation and lends further support to the hypothesis that the naturally occuring theta rhythm may play a modulatory role in the induction of LTP.     

Development of habituation in the dentate gyrus of rat: Physiology and anatomy

The physiological development of monosynaptic response habituation in the rat denate gyrus was compared to morphological development. Rapid Golgi techniques were coupled with in vitro studies of dentate granule cell habituation to several frequencies and intensities of monosynaptic excitation. Except for the youngest group, the degree of habituation increased as a function of age, paralleling the morphological development.     

Impairment of long-term potentiation and paired-pulse facilitation in rat hippocampal dentate gyrus following developmental lead exposure in vivo

Neonatal rats were exposed to lead from parturition to weaning via the milk of dams drinking 0.2% lead acetate solution. The alterations of long-term potentiation (LTP) and paired-pulse facilitation (PPF) of hippocampal dentate gyrus in adult rats (90–115 days) following developmental lead exposure were studied in vivo. Input/output (I/O) function, paired-pulse facilitation (PPF), excitatory postsynaptic potential (EPSP) and population spike (PS) amplitude were measured in the dentate gyrus (DG) in response to stimulation applied to the lateral perforant path. The results showed that LTP was induced in control rats with an average PS potentiation of 321.1±50.0% (n=18), which was significantly greater than the increase in PS potentiation (173.5±30.0%, n=17, p<0.001) in lead-exposed rats after tetanizing stimulation. The mean EPSP potentiation increased to 172.4±27.0% (n=18) in control and 138.8±21.4% (n=17) in lead-exposed rats after tetanizing stimulation. The lead-induced impairment of LTP of PS potentiation was more serious than that of EPSP potentiation. Following pairs stimulation of perforant fiber at 250 μA and an interpulse interval (IPI) of 10–1000 ms, the average peak facilitation of PS was 211.3±25.0% (n=13) in control and 187.7±23.0% (n=11) in lead-exposed rats. The average facilitation period duration of PS was 243.0±35.8 ms (n=13) in control and 138.0±24.4 ms (n=11) in lead-exposed rats. These results suggested that developmental lead exposure in neonatal rats caused impairments in LTP and PPF of hippocampal dentate gyrus.     

NDRG2通过Hes1参与癫痫发作后海马齿状回的神经发生

目的 探讨N-Myc下游调节基因2(N-Myc downstream regulated gene 2,NDRG2)与癫痫发作后海马齿状回神经发生的关系。方法 C57BL/6小鼠20只,随机分为癫痫组和对照组,每组又分为癫痫造模后1和7 d两个时间点,每个时间点5只,通过蛋白免疫印迹检测癫痫后海马齿状回NDRG2蛋白相对表达水平和mRNA相对表达水平变化; 使用双皮质素(DCX)染色标记未成熟神经元,神经巢蛋白(Nestin)标记神经干细胞,神经核蛋白(NeuN)标记成熟神经元,观察NDRG2对海马齿状回神经干细胞增殖影响; 采用RT-PCR检测发状分裂相关增强子1(hairy and enhancer of split 1,Hes 1)、NDRG2 mRNA相对表达表达水平,并分析两者之间的相关性; 观察NDRG2参与癫痫发作后神经发生的可能机制。结果 癫痫组与对照组比较,DCX、Nestin、NeuN、Hes1、NDRG2蛋白相对表达水平在1和7 d这2个时间点有显著性增高,并随时间逐渐递增。结论 癫痫发作后海马NDRG2蛋白相对表达水平增高,与癫痫发作后海马齿状回的神经细胞增值时间具有一致性和相关性,NDRG2可能参与癫痫发作后海马齿状回的神经发生过程; 同时发现海马NDRG2表达增加和Hes1分子表达增加具有相关性,故推测NDRG2可能通过Hes1参与癫痫发作后海马齿状回的神经发生。     

Anisomycin blocks the late phase of long-term potentiation in the dentate gyrus of freely moving rats

Freely moving rats, chronically implanted with stimulation electrodes in the medial entorhinal cortex and recording electrodes in the dentate gyrus, received two 400 micrograms intraventricular injections of anisomycin during a tetanization procedure that induced a long-lasting potentiation (72 hours) of the monosynaptic field potential. Inhibition of protein synthesis during the tetanization procedure did not immediately influence the induction of long-term potentiation (LTP). However, 3-4 hours after the beginning of tetanization the potentiation effect decayed progressively and was abolished totally during the remaining 7 day observation period. In control experiments anisomycin did not affect the slope of field EPSP's and produced a reversible depression of the population spike amplitude. These data indicate a relatively specific effect of the protein synthesis inhibitor on mechanisms involved in a late phase of LTP stabilization.     

The influence of long-term potentiation on the spatial relationship between astrocyte processes and potentiated synapses in the dentate gyrus neuropil of rat brain

The influence of long-term potentiation (induced by repeated high-frequency stimulation of the perforant pathway) on the distribution pattern of astrocyte processes in the neuropil of the hippocampal dentate area containing the potentiated synapses was investigated by quantitative electronmicroscopy. It has been found that significant changes occurred in the ramification of astrocyte processes as well as in their topographic relation to synaptic complexes. When comparing the results obtained in LTP animals with active control or sham-operated animals, we found significant higher numerical density, but smaller volume, higher surface density and closer apposition of astrocyte processes to the synaptic clefts, boutons terminaux or spines in the potentiated synapses containing neuropil. The glial reaction to synaptic activation has been seen most pronounced 8 h after the LTP induction. The results are pointing to a participation of the glia cells in the maintenance of the LTP effect as well as to a metabolic coupling between synaptic transmission and glia function for equilibrating the homeostasis by clearing the extracellular space next to the transmission zones.     

Single and repetitive paired-pulse suppression: A parametric analysis and assessment of usefulness in epilepsy research

Purpose :  The paired-pulse technique has been widely used as a convenient but indirect measure of "inhibition" in hippocampal circuits of normal and epileptic animals. Most investigators have used a single paired-pulse protocol, whereas others have utilized repetitive paired pulses. This study investigated which parameters influence results from paired-pulse tests, focusing on the repetitive paired-pulse technique; it aims to assess how this technique may be used in an unbiased and quantitative manner across animal preparations for comparisons of control and experimental epileptic animals.
Methods :  The perforant path was stimulated while field potentials were recorded from the granule cell layer under isoflurane anesthesia. Paired-pulse suppression was analyzed as a function of stimulation intensity and interpulse interval and frequency.
Results :  Paired-pulse suppression was greater with increased stimulus intensity and decreased interpulse interval (20–100 ms). During repetitive protocols, stimulation frequencies ≤1.0 Hz produced paired-pulse suppression similar to single paired-pulse responses, but caused more paired-pulse suppression between 1.0 and 4.0 Hz at all but the lowest intensities. The amplitude of the population spike produced by the conditioning pulse increased progressively during stimulation at higher frequencies (1.0–4.0 Hz).
Discussion :  The single paired-pulse technique is highly dependent on stimulation parameters, as is the repetitive paired-pulse protocol, which is more variable. To generate reliable, consistent, and unbiased data in comparisons of control and experimental epileptic groups, all parameters should be specified and controlled across experiments. Paired-pulse suppression is susceptible to alterations in many mechanisms, and, therefore, represents a circuit response rather than an assay of γ-aminobutyric acid (GABA)ergic inhibition in epilepsy research.     

Differential paired-pulse responses between the CA1 region and the dentate gyrus are related to altered CLC-2 immunoreactivity in the pilocarpine-induced rat epilepsy model

The epileptic hippocampus shows differential paired-pulse responses between the dentate gyrus and the CA1 region. However, little data are available to explain this phenomenon. In the present study, we identified the relationship between regional differences of paired-pulse response and voltage gated Cl(-) channel 2 (CLC-2)/vesicular GABA transport (VGAT) expression in a pilocarpine-induced rat model. During epileptogenic periods, paired-pulse inhibitions in the dentate gyrus and the CA1 region were markedly reduced. After recurrent seizure onset, paired-pulse inhibition in the dentate gyrus was markedly enhanced, while that in the CA1 region more reduced. Unlike VGAT, CLC-2 immunoreactivity was markedly reduced in the hippocampus during epileptogenic periods and was re-enhanced only in the dentate gyrus after recurrent seizure onset. Linear regression analysis showed an inverse proportional relationship between alterations in CLC-2 immunoreactivity and changes in normalized population spike amplitude ratio within the CA1 region and the dentate gyrus. Therefore, our findings suggest that the regionally specific alterations in CLC-2 immunoreactivity after SE may determine the properties of paired-pulse responses in the hippocampus of the pilocarpine-induced rat epilepsy model.     

Differential alteration of NMDA receptor subunits in the gerbil dentate gyrus and subiculum following seizure

In the present study, a chronological and comparative analysis of the immunoreactivities of N-methyl-D-aspartate (NMDA) receptor subunits in hippocampus of both seizure resistant (SR) and seizure sensitive (SS) gerbils was made in order to clarify the temporal and spatial alterations of NMDA receptor subunit expressions in the hippocampus complex. The changes in NMDA receptor immunoreactivity in the hippocampi of SS gerbils were restricted to both the dentate gyrus and the subiculum. At 30 min postictal, a decline in NMDA receptor subunit 1 (NR1) immunoreactivity in the suprablade of dentate gyrus was observed. This is in contrast to the enhancement of its immunodensity in the infrablade. At 3 h postictal the NR1 immunoreactivity in the infrablade also declined significantly. At 12 h postictal, its immunoreactivity in the hilar neurons was reduced. The NMDA receptor subunit 2A/B (NR2A/B) immunoreactivity did not alter until 12 h following seizure-onset, when it was slightly decreased in the granule cells and hilar neurons. In the subiculum, NR1 immunoreactivity was significantly decreased, and was almost undetectable in this region until 12 h postictal; in contrast the NR2A/B immunoreactivity in this region increased significantly in this time point. These results suggest that the altering NMDA receptor expression in both the dentate gyrus and subiculum may affect tissue excitability and have an important role in regulating seizure activity in SS gerbils.     

Early impairment and late recovery of synaptic transmission in the rat dentate gyrus following transient forebrain ischemia in vivo

Ischemic stroke causes various functional deficits in the brain such as memory impairment, and clinical reports have shown that the impaired brain functions may partially recover. However, there has been no experimental model suitable for studying cellular mechanisms of functional recovery following brain ischemia. Therefore, we investigated the long-term influence of transient forebrain ischemia on excitatory synaptic transmission in the rat dentate gyrus, a brain region relatively resistant to ischemia. Fifteen minutes of transient forebrain ischemia produced no apparent histological damage in dentate granule cells, but caused a significant reduction of basal synaptic potentials evoked by perforant path stimulation. Field excitatory postsynaptic potential remained reduced for at least 1 month after ischemia, while population spike recovered to control level in 1 month. The induction of long-term potentiation was also impaired after ischemia, but it showed faster recovery than basal synaptic potentials. In conclusion, we found that synaptic transmission in the dentate gyrus of the rat is impaired following transient forebrain ischemia, but has a potential to recover. These results may provide a good model for studying the mechanisms of impairment and recovery of brain function after transient ischemia.     

Field responses to perforant path stimulation in the rat dentate gyrus: role of corticosterone and NMDA-receptor activation

Recent studies showed that corticosterone and NMDA receptor activation suppress cell turn-over in the dentate gyrus through a common pathway, the NMDA receptor acting downstream of the corticosteroids. The present data show that in the absence of corticosteroids but not of NMDA receptor activation synaptic responses of dentate cells are reduced. The reduced synaptic responsiveness in the absence of corticosterone is therefore probably not caused by changes in cell turn-over.