Activity-dependent release of adenosine contributes to short-term depression at CA3-CA1 synapses in rat hippocampus

High-frequency stimulation results in a transient, presynaptically mediated decrease in synaptic efficacy called short-term depression (STD). Stimulation of Schaffer-collateral axons at 10 Hz for 5 s resulted in approximately 75% depression of excitatory postsynaptic current (EPSC) slope recorded from CA1 cells in rat organotypic slice cultures. An adenosine A(1) receptor antagonist decreased the magnitude of STD elicited with 10-Hz stimulation by approximately 30%. The A(1) receptor antagonist had no effect on STD elicited with 3-Hz stimulation. The activation of A(1) receptors during 10-Hz stimulation was not due to the extracellular conversion of released ATP to adenosine, because block of 5'-ectonucleotidases did not significantly affect STD. The adenosine transport inhibitor dipyridamole did not reduce STD, indicating that adenosine was not released by facilitated transport. We conclude that 10-Hz, but not 3-Hz, stimulation causes the vesicular release of adenosine and the rapid (<3 s) activation of presynaptic inhibitory A(1) receptors, which account for approximately 40% of homosynaptic EPSC depression.     

Age-related changes in glutamate release in the CA3 and dentate gyrus of the rat hippocampus

The present studies employed a novel microelectrode array recording technology to study glutamate release and uptake in the dentate gyrus, CA3 and CA1 hippocampal subregions in anesthetized young, late-middle aged and aged male Fischer 344 rats. The mossy fiber terminals in CA3 showed a significantly decreased amount of KCl-evoked glutamate release in aged rats compared to both young and late-middle-aged rats. Significantly more KCl-evoked glutamate release was seen from perforant path terminals in the DG of late-middle-aged rats compared young and aged rats. The DG of aged rats developed an increased glutamate uptake rate compared to the DG of young animals, indicating a possible age-related change in glutamate regulation to deal with increased glutamate release that occurred in late-middle age. No age-related changes in resting levels of glutamate were observed in the DG, CA3 and CA1. Taken together, these data support dynamic changes to glutamate regulation during aging in subregions of the mammalian hippocampus that are critical for learning and memory.     

Developmental changes of glutamate receptors in the rat cerebral cortex and hippocampus

We studied the immunohistochemial localization of the glutamate receptors (GluR-1, -2, and -3,) in the developing rat cerebral cortex and hippocampus using antibodies to GluR1 and to an epitope common to GluR2 and GluR3 (GluR2/3) subunits. In the cerebral cortex, GluR1 immunoreactivity appeared in the neurons from postnatal day (PND) 0, increased with maturation, was highest at PND?10, decreased until PND 30, and thereafter remained at the same level as on PND?0. GluR2/3 immunoreactivity appeared earlier in scattered neurons on embryonal day (ED) 18, increased with maturation and reached a peak between PND?10 and PND?15, after which the immunoreactivity gradually decreased and reached a plateau at PND?30. For both GluR1 and GluR2/3, some of the pyramidal neurons showed intense staining. In the pyramidal layers of the hippocampus, GluR1 and GluR2/3 immunoreactivity was found in all the pyramidal neurons of the CA1–4 area from ED?20. In the dentate gyrus of the hippocampus, GluR1 and GluR2/3 immunoreactivity was found in the neurons of the granule cells after PND?0. Immunoreactivity in the neurons of the subiculum was found after PND?5 and that of the polymorphic cell layers was found after PND?15–20. Our results indicate that the development of glutamate receptor subunits in the rat cerebral cortex and hippocampus is expressed in different spatial patterns and distinct temporal patterns throughout development and is scheduled during the early postnatal period, when synaptic plasticity or synaptic connection occurs in these regions.     

Developmental changes of glutamate receptors in the rat cerebral cortex and hippocampus

 We studied the immunohistochemial localization of the glutamate receptors (GluR-1, -2, and -3,) in the developing rat cerebral cortex and hippocampus using antibodies to GluR1 and to an epitope common to GluR2 and GluR3 (GluR2/3) subunits. In the cerebral cortex, GluR1 immunoreactivity appeared in the neurons from postnatal day (PND) 0, increased with maturation, was highest at PND 10, decreased until PND 30, and thereafter remained at the same level as on PND 0. GluR2/3 immunoreactivity appeared earlier in scattered neurons on embryonal day (ED) 18, increased with maturation and reached a peak between PND 10 and PND 15, after which the immunoreactivity gradually decreased and reached a plateau at PND 30. For both GluR1 and GluR2/3, some of the pyramidal neurons showed intense staining. In the pyramidal layers of the hippocampus, GluR1 and GluR2/3 immunoreactivity was found in all the pyramidal neurons of the CA1–4 area from ED 20. In the dentate gyrus of the hippocampus, GluR1 and GluR2/3 immunoreactivity was found in the neurons of the granule cells after PND 0. Immunoreactivity in the neurons of the subiculum was found after PND 5 and that of the polymorphic cell layers was found after PND 15–20. Our results indicate that the development of glutamate receptor subunits in the rat cerebral cortex and hippocampus is expressed in different spatial patterns and distinct temporal patterns throughout development and is scheduled during the early postnatal period, when synaptic plasticity or synaptic connection occurs in these regions. Accepted: 13 June 1996     

Frequency-dependent shift from paired-pulse facilitation to paired-pulse depression at unitary CA3-CA3 synapses in the rat hippocampus

Paired recordings between CA3 interconnected pyramidal neurons were used to study the properties of short-term depression occurring in these synapses under different frequencies of presynaptic firing (   n = 22  ). In stationary conditions (0.05-0.067 Hz) pairs of presynaptic action potentials (50 ms apart) evoked EPSCs whose amplitude fluctuated from trial to trial with occasional response failures. In 15/20 cells, paired-pulse ratio (PPR) was characterized by facilitation (PPF) while in the remaining five by depression (PPD). Increasing stimulation frequency from 0.05-0.067 Hz to 0.1-1 Hz induced low frequency depression (LFD) of EPSC amplitude with a gradual increase in the failure rate. Overall, 9/12 cells at 1 Hz became almost 'silent'. In six cells in which the firing rate was sequentially shifted from 0.05 to 0.1 and 1 Hz, changes in synaptic efficacy were so strong that PPR shifted from PPF to PPD. The time course of depression of EPSC1 could be fitted with single exponentials with time constants of 98 and 36 s at 0.1 and 1 Hz, respectively. In line with the inversion of PPR at 1 Hz, the time course of depression of EPSC2 was faster than EPSC1 (7 s). Recovery from depression could be obtained by lowering the frequency of stimulation to 0.025 Hz. These results could be explained by a model that takes into account two distinct release processes, one dependent on the residual calcium and the other on the size of the readily releasable pool of vesicles.     

BK potassium channels control transmitter release at CA3–CA3 synapses in the rat hippocampus

Augmentation is a component of short-term synaptic plasticity with a gradual onset and duration in seconds. To investigate this component at the corticogeniculate synapse, whole cell patch-clamp recordings were obtained from principal cells in a slice preparation of the rat dorsal lateral geniculate nucleus. Trains with 10 stimuli at 25 Hz evoked excitatory postsynaptic currents (EPSCs) that grew in amplitude, primarily from facilitation. Such trains also induced augmentation that decayed exponentially with a time constant τ= 4.6 ± 2.6 s (mean ± standard deviation). When the trains were repeated at 1–10 s intervals, augmentation markedly increased the size of the first EPSCs, leaving late EPSCs unaffected. The magnitude of augmentation was dependent on the number of pulses, pulse rate and intervals between trains. Augmented EPSCs changed proportionally to basal EPSC amplitudes following alterations in extracellular calcium ion concentration. The results indicate that augmentation is determined by residual calcium remaining in the presynaptic terminal after repetitive spikes, competing with fast facilitation. We propose that augmentation serves to maintain a high synaptic strength in the corticogeniculate positive feedback system during attentive visual exploration.     

Modulation of LTP at rat hippocampal CA3-CA1 synapses by direct current stimulation

Transcranial direct current stimulation (tDCS) can produce a lasting polarity-specific modulation of cortical excitability in the brain, and it is increasingly used in experimental and clinical settings. Recent studies suggest that the after-effects of tDCS are related to molecular mechanisms of activity-dependent synaptic plasticity. Here we investigated the effect of DCS on the induction of one of the most studied N-methyl-d-aspartate receptor-dependent forms of long-term potentiation (LTP) of synaptic activity at CA3-CA1 synapses in the hippocampus. We show that DCS applied to rat brain slices determines a modulation of LTP that is increased by anodal and reduced by cathodal DCS. Immediate early genes, such as c-fos and zif268 (egr1/NGFI-A/krox24), are rapidly induced following neuronal activation, and a specific role of zif268 in the induction and maintenance of LTP has been demonstrated. We found that both anodal and cathodal DCS produce a marked subregion-specific increase in the expression of zif268 protein in the cornus ammonis (CA) region, whereas the same protocols of stimulation produce a less pronounced increase in c-fos protein expression in the CA and in dentate gyrus regions of the hippocampus. Brain-derived neurotrophic factor expression was also investigated, and it was found to be reduced in cathodal-stimulated slices. The present data demonstrate that it is possible to modulate LTP by using DCS and provide the rationale for the use of DCS in neurological diseases to promote the adaptive and suppress the maladaptive forms of brain plasticity.     

Chronic restraint stress induces changes in synapse morphology in stratum lacunosum-moleculare CA1 rat hippocampus: a stereological and three-dimensional ultrastructural study

Chronic restraint stress is known to affect the morphology and synaptic organization of the hippocampus, predominantly within CA3 but also in CA1 and dentate gyrus. In this study, we provide the first evidence for specific ultrastructural alterations affecting asymmetric axo-spinous synapses in CA1 stratum lacunosum-moleculare following chronic restraint stress (6 h/day, 21 days) in the rat. The structure of asymmetric axo-spinous post-synaptic densities was investigated using serial section three-dimensional reconstruction procedures in control (n=4) and chronic restraint stress (n=3) animals. Dendritic spine profiles (spine head+neck) associated with the sampled synaptic contacts (30 per animal) were also reconstructed in three-dimensions. Morphometric analyses revealed a significant increase in post-synaptic density surface area (+36%; P=0.03) and a highly significant increase in post-synaptic density volume (+79%; P=0.003) in the chronic restraint stress group. These changes were directly associated with 'non-macular' (perforated, complex and segmented) post-synaptic densities. A highly significant overall increase in the 'post-synaptic density surface area/spine surface area' ratio was also detected in the chronic restraint stress group (+27%; P=0.002). In contrast, no quantitative changes in spine parameters were found between groups. The Cavalieri method was used to assess the effects of chronic restraint stress exposure upon CA1 hippocampal volume. The mean volume of total dorsal anterior CA1 hippocampus was significantly lower in the chronic restraint stress group (-16%; P=0.036). However, when corrected for volume changes, no significant alteration in a relative estimate of the mean number of asymmetric axo-spinous synapses was detected in CA1 stratum lacunosum-moleculare between control and chronic restraint stress groups. The data indicate a structural remodeling of excitatory axo-spinous synaptic connectivity in rat CA1 stratum lacunosum-moleculare as a result of chronic restraint stress.     

Subicular lesions cause dendritic atrophy in CA1 and CA3 pyramidal neurons of the rat hippocampus

The subiculum is a major source of output projections from hippocampus to cortical and subcortical regions. Our previous studies have demonstrated the selective loss of CA1 pyramidal neurons of the hippocampus, and operant and spatial learning impairment in subicular lesioned rats [Govindaiah et al. (1997) Brain Res. 745, 121-126; Laxmi et al. (1999) Brain Res. 816, 245-148]. In the present study, the effect of ibotenate lesions of the subiculum on the dendritic morphology of CA1 and CA3 pyramidal neurons of the hippocampus was investigated in 30-day-old male Wistar rats. The ventral subiculum was lesioned bilaterally with multiple injections of ibotenic acid, stereotaxically. The dendritic branching points and intersections were studied in apical and basal dendrites up to 320 and 160 microm, respectively, in Golgi-impregnated CA1 and CA3 pyramidal neurons of the hippocampus. The results revealed a significant (P<0.001) decrease in the number of dendritic branching points, intersections and total number of dendrites in both apical and basal dendrites of CA1, as well as CA3 pyramidal neurons of the hippocampus. It is surprising that the subicular lesions caused dendritic atrophy of CA3 neurons without affecting the cell density.The results of the present study demonstrate the dendritic atrophy of hippocampal neurons following selective subicular lesions. This might be responsible for the impairments in operant and spatial learning tasks in these rats as observed in our earlier studies. In addition, hippocampal damage is also associated with an impairment in the process of the active monitoring of movements in space, rather than place learning per se [Whishaw (1998) Neurosci. biobeh. Rev. 22, 209-220]. Accordingly, further studies are required to correlate the differential effect of subicular lesions on impairments in learning and movement in space in rats.     

Muscarinic inhibition of endogenous glutamate release from rat hippocampus synaptosomes

The effects of acetylcholine (ACh) on the depolarization-evoked release of endogenous glutamic acid (Glu) have been studied using synaptosomes prepared from rat hippocampus and depolarized in superfusion with 15 mM KCl. Acetylcholine inhibited Glu release in a concentration-dependent way. The natural agonist was particularly effective causing 50% inhibition of Glu release at 10 microM in the absence of acetylcholinesterase (AChE) inhibitors. The inhibitory effect of ACh on the K+-evoked release of Glu was antagonized by the selective muscarinic receptor antagonist atropine but not by the nicotinic receptor antagonist mecamylamine. The data represent the first demonstration that muscarinic receptors located on Glu axon terminals in rat hippocampus may modulate the release of Glu.     

Voltage-clamp analysis of posttetanic potentiation of the mossy fiber to CA3 synapse in hippocampus

1. Short-term changes in synaptic efficacy were studied at the mossy fiber (MF) to CA3 (MF-CA3) synapse in the in vitro hippocampus. Monosynaptic excitatory postsynaptic currents (EPSCs) were recorded before and during posttetanic potentiation (PTP) with the use of intracellular recording and single-electrode voltage-clamp (SEVC) techniques. 2. Repetitive stimulation (100 Hz for 1 s) of the MF synaptic inputs to CA3 pyramidal cells resulted in PTP averaging 170 +/- 19% (SE, n = 42) over control and decaying with a time constant (tau p) of 59.7 +/- 5 s(n = 23). Reproducible episodes of PTP could be recorded if low stimulus intensities were used. Also, after MF tetanization, a faster component, termed augmentation, preceded PTP but could not be accurately resolved within the experimental protocol; only estimates of this component are included. 3. Biophysical parameters of the EPSC that were monitored before and during PTP included synaptic conductance (G), synaptic reversal potential (Erev), decay time constant (tau EPSC), and input resistance of the postsynaptic cell. During PTP the EPSC synaptic conductance increased from 9.8 to 32.7 nS (P less than 0.02, n = 6), whereas there was no statistical change in Erev (-6.0 compared with -6.7 mV, n = 6), tau EPSC (4.3 compared with 4.5 ms, n = 9), or postsynaptic input resistance (59 compared with 63 M omega, n = 12). 4. A presynaptic contribution to PTP was studied directly by observing changes in transmitter release during PTP. Presynaptic mechanisms were assessed by determining the ratio of evoked synaptic excitatory postsynaptic potentials (EPSPs) over the total number of stimuli (EPSP-to-stimuli ratio). The ratio of EPSP to stimuli changed from 0.64 to 0.90 (P less than 0.01, n = 7) during PTP. A reduction in the number of synaptic failures can only be explained by a presynaptic mechanism. No assumptions concerning the statistical distribution of transmitter release were necessary because no statistical parameters were determined. 5. Changes in postsynaptic cell properties do not appear to contribute to PTP studied under the present experimental conditions. Direct stimulation of the postsynaptic neuron via the intracellular recording electrode (20-100 Hz/1 s) failed to produce potentiation of the EPSC; in fact, a slight depression was observed at 50 and 100 Hz direct stimulation. Likewise, the postsynaptic input resistance and synaptic Erev did not change during PTP. 6. The specific N-methyl-D-aspartate (NMDA) receptor antagonist D-2-amino-5-phosphonovaleric acid (APV, 20 microM) had no effect on either the magnitude or duration of PTP.(ABSTRACT TRUNCATED AT 400 WORDS)     

大鼠海马CA3区胆碱能神经元的衰老变化

目的:探讨大鼠海马CA3区胆碱能神经元增龄性变化的规律.方法:SD大鼠随机分为1～2月龄、4～5月龄,11～12月龄、≥24月龄4组,常规石蜡包埋海马连续冠状切片,尼氏染色、乙酰胆碱转移酶(ChAT)免疫组织化学显色,图像分析仪测量ChAT免疫反应阳性产物的光密度及其阳性神经元的多种形态学参数.结果:随着年龄的增长,大...     

Morphine modulates glutamate release in the hippocampal CA1 area in mice

Opiate abuse is associated with long-lasting neural adaptative changes in the brain. Increasing evidence demonstrates that opiates significantly alter the function of the glutamatergic system, while how the system is regulated still remains elusive. In the present study, we studied the effect of morphine on extracellular glutamate concentration in the hippocampus, a nucleus rich of the glutamatergic neurons. The results showed that glutamate concentration in the extracellular fluid of the hippocampus was decreased following either acute or chronic treatment of morphine. However, naloxone-induced withdrawal increased glutamate concentration significantly. These results suggest an adaptation of the glutamatergic neuronal transmission in the hippocampus after morphine treatment.     

Synaptic modifications at the CA3–CA1 synapse after chronic AMPA receptor blockade in rat hippocampal slices

Maintenance of dendritic spines, the postsynaptic elements of most glutamatergic synapses in the central nervous system, requires continued activation of AMPA receptors. In organotypic hippocampal slice cultures, chronic blockade of AMPA receptors for 14 days induces a substantial loss of dendritic spines on CA1 pyramidal neurons. Here, using serial section electron microscopy, we show that loss of dendritic spines is paralleled by a significant reduction in synapse density. In contrast, we observed an increased number of asymmetric synapses onto the dendritic shaft, suggesting that spine retraction does not inevitably lead to synapse elimination. Functional analysis of the remaining synapses revealed that hippocampal circuitry compensates for the anatomical loss of synapses by increasing synaptic efficacy. Moreover, we found that the observed morphological and functional changes were associated with altered bidirectional synaptic plasticity. We conclude that continued activation of AMPA receptors is necessary for maintaining structure and function of central glutamatergic synapses.     

Desynchronization of glutamate release prolongs synchronous CA3 network activity

Periodic bursts of activity in the disinhibited in vitro hippocampal CA3 network spread through the neural population by the glutamatergic recurrent collateral axons that link CA3 pyramidal cells. It was previously proposed that these bursts of activity are terminated by exhaustion of releasable glutamate at the recurrent collateral synapses so that the next periodic burst of network activity cannot occur until the supply of glutamate has been replenished. As a test of this hypothesis, the rate of glutamate release at CA3 axon terminals was reduced by substitution of extracellular Ca(2+) with Sr(2+). Reduction of the rate of glutamate release reduces the rate of depletion and should thereby prolong bursts. Here we demonstrate that Sr(2+) substitution prolongs spontaneous bursts in the disinhibited adult CA3 hippocampal slices to 37.2 +/- 7.6 (SE) times the duration in control conditions. Sr(2+) also decreased the probability of burst initiation and the rate of burst onset, consistent with reduced synchrony of glutamate release and a consequent reduced rate of spread of excitation through the slice. These findings support the supply of releasable glutamate as an important determinant of the probability and duration of synchronous CA3 network activity.     

The effect of simulated ischaemia on spontaneous GABA release in area CA1 of the juvenile rat hippocampus

An early consequence of brain energy deprivation is an increase in the frequency of spontaneous inhibitory and excitatory postsynaptic currents (sIPSCs and sEPSCs), which may disrupt neural information processing. This increase in spontaneous transmitter release has been reported to occur in calcium-free solution and has been attributed either to calcium release from internal stores or to a direct effect of hypoxia on the transmitter release mechanism. Here we investigate the mechanism of the increase in sIPSC frequency that occurs in area CA1 of rat hippocampus during simulated ischaemia, by making patch-clamp recordings from CA1 pyramidal neurones. When recording in whole-cell mode, exposure to ischaemic solution increased the sIPSC frequency 30-fold (to 49 Hz) after 5 min, and doubled the sIPSC amplitude. Ischaemic sIPSCs were action potential independent, vesicular in origin and, contrary to the results of earlier studies which did not buffer extracellular calcium to a low level, dependent on extracellular calcium. The properties of the ischaemic sIPSCs were not affected by depleting intracellular stores of calcium or by blocking the neuronal GABA transporter GAT-1. Recording from neurones using gramicidin-perforated patch-clamping showed a 10-fold smaller, more transient increase in sIPSC frequency during ischaemia, with no change of sIPSC amplitude, suggesting that whole-cell clamp recording increases the ischaemia-induced sIPSC rate and amplitude by controlling the intracellular milieu.     

Membrane properties of interneurons in stratum oriens-alveus of the CA1 region of rat hippocampus in vitro

The membrane properties of interneurons situated near the border of stratum oriens and the alveus of the CA1 region were examined with intracellular recording and staining in rat hippocampal slices in vitro. Cellular staining with Lucifer Yellow indicated that the somata of these interneurons were multipolar and their dendrites projected horizontally along the alveus and vertically toward stratum lacunosum-moleculare. Intrinsic properties (input resistance, action potential amplitude, time constant) and spike after-potentials were typical of non-pyramidal cells. Action potential duration, however, was of relatively medium duration (1.15 ms) and slow afterhyperpolarizations followed depolarization-induced trains of action potentials. Spontaneous activity of interneurons was prominent and of either of two types: single action potentials or high frequency bursts of action potentials. Interneurons displayed marked, voltage- and time-dependent inward rectification and anodal break excitation. Analysis of the slope of the charging function of hyperpolarizing transients, suggested that these interneurons were electrically compact (dendrite to soma conductance ratio, p approximately 2.7; and electrotonic length constant, L approximately 1.1). Characteristically, interneurons sustained high frequency repetitive firing during long depolarizing pulses. The slope of the frequency-current relation was 442 Hz/nA for the first interspike interval and 117 Hz/nA for later intervals (no. 60), suggesting the presence of spike frequency adaptation. Physiologically, these interneurons resembled more closely basket cells of stratum pyramidale than stellate cells of stratum lacunosum-moleculare.