The Amyloid Precursor Protein Regulates Synaptic Transmission at Medial Perforant Path Synapses.

The perforant path provides the primary cortical excitatory input to the hippocampus. Because of its important role in information processing and coding, entorhinal projections to the dentate gyrus have been studied in considerable detail. Nevertheless, synaptic transmission between individual connected pairs of entorhinal stellate cells and dentate granule cells remains to be characterized. Here, we have used mouse organotypic entorhino-hippocampal tissue cultures of either sex, in which the entorhinal cortex (EC) to dentate granule cell (GC; EC-GC) projection is present, and EC-GC pairs can be studied using whole-cell patch-clamp recordings. By using cultures of wild-type mice, the properties of EC-GC synapses formed by afferents from the lateral and medial entorhinal cortex were compared, and differences in short-term plasticity were identified. As the perforant path is severely affected in Alzheimer's disease, we used tissue cultures of amyloid precursor protein (APP)-deficient mice to examine the role of APP at this synapse. APP deficiency altered excitatory neurotransmission at medial perforant path synapses, which was accompanied by transcriptomic and ultrastructural changes. Moreover, presynaptic but not postsynaptic APP deletion through the local injection of Cre-expressing adeno-associated viruses in conditional APPflox/flox tissue cultures increased the neurotransmission efficacy at perforant path synapses. In summary, these data suggest a physiological role for presynaptic APP at medial perforant path synapses that may be adversely affected under altered APP processing conditions.SIGNIFICANCE STATEMENT The hippocampus receives input from the entorhinal cortex via the perforant path. These projections to hippocampal dentate granule cells are of utmost importance for learning and memory formation. Although there is detailed knowledge about perforant path projections, the functional synaptic properties at the level of individual connected pairs of neurons are not well understood. In this study, we investigated the role of APP in mediating functional properties and transmission rules in individually connected neurons using paired whole-cell patch-clamp recordings and genetic tools in organotypic tissue cultures. Our results show that presynaptic APP expression limits excitatory neurotransmission via the perforant path, which could be compromised in pathologic conditions such as Alzheimer's disease.

Latent toxoplasmosis impairs learning and memory yet strengthens short-term and long-term hippocampal synaptic plasticity at perforant pathway-dentate gyrus, and Schaffer collatterals-CA1 synapses.

Investigating long-term potentiation (LTP) in disease models provides essential mechanistic insight into synaptic dysfunction and relevant behavioral changes in many neuropsychiatric and neurological diseases. Toxoplasma (T) gondii is an intracellular parasite causing bizarre changes in host's mind including losing inherent fear of life-threatening situations. We examined hippocampal-dependent behavior as well as in vivo short- and long-term synaptic plasticity (STP and LTP) in rats with latent toxoplasmosis. Rats were infected by T. gondii cysts. Existence of REP-529 genomic sequence of the parasite in the brain was detected by RT-qPCR. Four and eight weeks after infection, spatial, and inhibitory memories of rats were assessed by Morris water maze and shuttle box tests, respectively. Eight weeks after infection, STP was assessed in dentate gyrus (DG) and CA1 by double pulse stimulation of perforant pathway and Shaffer collaterals, respectively. High frequency stimulation (HFS) was applied to induce LTP in entorhinal cortex-DG (400 Hz), and CA3-CA1 (200 Hz) synapses. T. gondii infection retarded spatial learning and memory performance at eight weeks post-infection period, whereas inhibitory memory was not changed. Unlike uninfected rats that normally showed paired-pulse depression, the infected rats developed paired-pulse facilitation, indicating an inhibitory synaptic network disruption. T. gondii-infected rats displayed strengthened LTP of both CA1-pyramidal and DG-granule cell population spikes. These data indicate that T. gondii disrupts inhibition/excitation balance and causes bizarre changes to the post-synaptic neuronal excitability, which may ultimately contribute to the abnormal behavior of the infected host.

Autophagy ameliorates cognitive impairment through activation of PVT1 and apoptosis in diabetes mice.

The underlying mechanisms of cognitive impairment in diabetes remain incompletely characterized. Here we show that the autophagic inhibition by 3-methyladenine (3-MA) aggravates cognitive impairment in streptozotocin-induced diabetic mice, including exacerbation of anxiety-like behaviors and aggravation in spatial learning and memory, especially the spatial reversal memory. Further neuronal function identification confirmed that both long term potentiation (LTP) and depotentiation (DPT) were exacerbated by autophagic inhibition in diabetic mice, which indicating impairment of synaptic plasticity. However, no significant change of pair-pulse facilitation (PPF) was recorded in diabetic mice with autophagic suppression compared with the diabetic mice, which indicated that presynaptic function was not affected by autophagic inhibition in diabetes. Subsequent hippocampal neuronal cell death analysis showed that the apoptotic cell death, but not the regulated necrosis, significantly increased in autophagic suppression of diabetic mice. Finally, molecular mechanism that may lead to cell death was identified. The long non-coding RNA PVT1 (plasmacytoma variant translocation 1) expression was analyzed, and data revealed that PVT1 was decreased significantly by 3-MA in diabetes. These findings show that PVT1-mediated autophagy may protect hippocampal neurons from impairment of synaptic plasticity and apoptosis, and then ameliorates cognitive impairment in diabetes. These intriguing findings will help pave the way for exciting functional studies of autophagy in cognitive impairment and diabetes that may alter the existing paradigms.

High-frequency stimulation induces gradual immediate early gene expression in maturing adult-generated hippocampal granule cells.

Increasing evidence shows that adult neurogenesis of hippocampal granule cells is advantageous for learning and memory. We examined at which stage of structural maturation and age new granule cells can be activated by strong synaptic stimulation. High-frequency stimulation of the perforant pathway in urethane-anesthetized rats elicited expression of the immediate early genes c-fos, Arc, zif268 and pCREB133 in almost 100% of mature, calbindin-positive granule cells. In contrast, it failed to induce immediate early gene expression in immature doublecortin-positive granule cells. Furthermore, doublecortin-positive neurons did not react with c-fos or Arc expression to mild theta-burst stimulation or novel environment exposure. Endogenous expression of pCREB133 was increasingly present in young cells with more elaborated dendrites, revealing a close correlation to structural maturation. Labeling with bromodeoxyuridine revealed cell age dependence of stimulation-induced c-fos, Arc and zif268 expression, with only a few cells reacting at 21 days, but with up to 75% of cells activated at 35-77 days of cell age. Our results indicate an increasing synaptic integration of maturing granule cells, starting at 21 days of cell age, but suggest a lack of ability to respond to activation with synaptic potentiation on the transcriptional level as long as immature cells express doublecortin.

Low-frequency stimulation induces long-term depression and slow onset long-term potentiation at perforant path-dentate gyrus synapses in vivo.

The expression of homosynaptic long-term depression (LTD) is thought to mediate a crucial role in sustaining memory function. Our in vivo investigations of LTD expression at lateral (LPP) and medial perforant path (MPP) synapses in the dentate gyrus (DG) corroborate prior demonstrations that PP-DG LTD is difficult to induce in intact animals. In freely moving animals, LTD expression occurred inconsistently among LPP-DG and MPP-DG responses. Interestingly, following acute electrode implantation in anesthetized rats, low-frequency stimulation (LFS; 900 pulses, 1 Hz) promotes slow-onset LTP at both MPP-DG and LPP-DG synapses that utilize distinct induction mechanisms. Systemic administration of the N-methyl-d-aspartate (NMDA) receptor antagonist (+/-)-cyclopiperidine-6-piperiperenzine (CPP; 10 mg/kg) 90 min before LFS selectively blocked MPP-DG but not LPP-DG slow onset LTP, suggesting MPP-DG synapses express a NMDA receptor-dependent slow onset LTP whereas LPP-DG slow onset LTP induction is NMDA receptor independent. In experiments where paired-pulse LFS (900 paired pulses, 200-ms paired-pulse interval) was used to induce LTD, paired-pulse LFS of the LPP resulted in rapid onset LTP of DG responses, whereas paired-pulse LFS of the MPP induced slow onset LTP of DG responses. Although LTD observations were very rare following acute electrode implantation in anesthetized rats, LPP-DG LTD was demonstrated in some anesthetized rats with previously implanted electrodes. Together, our data indicate in vivo PP-DG LTD expression is an inconsistent phenomenon that is primarily observed in recovered animals, suggesting perturbation of the dentate through surgery-related tissue trauma influences both LTD incidence and LTP induction at PP-DG synapses in vivo.

Endogenous opioid peptides contribute to associative LTP in the hippocampal CA3 region.

The medial and lateral perforant path projections to the hippocampal CA3 region display distinct mechanisms of long-term potentiation (LTP) induction, N-methyl-d-aspartate (NMDA) and opioid receptor dependent, respectively. However, medial and lateral perforant path projections to the CA3 region display associative LTP with coactivation, suggesting that while they differ in receptors involved in LTP induction they may share common downstream mechanisms of LTP induction. Here we address this interaction of LTP induction mechanisms by evaluating the contribution of opioid receptors to the induction of associative LTP among the medial and lateral perforant path projections to the CA3 region in vivo. Local application of the opioid receptor antagonists naloxone or Cys2-Tyr3-Orn5-Pen7-amide (CTOP) normally block induction of lateral perforant path-CA3 LTP. However, these opioid receptor antagonists failed to block associative LTP in lateral perforant path-CA3 synapses when it was induced by strong coactivation of the medial perforant pathway which displays NMDAR-dependent LTP. Thus strong activation of non-opioidergic afferents can substitute for the opioid receptor activation required for lateral perforant path LTP induction. Conversely, medial perforant path-CA3 associative LTP was blocked by opioid receptor antagonists when induced by strong coactivation of the opioidergic lateral perforant path. These data indicate endogenous opioid peptides contribute to associative LTP at coactive synapses when induced by strong coactivation of an opioidergic afferent system. These data further suggest that associative LTP induction is regulated by the receptor mechanisms of the strongly stimulated pathway. Thus, while medial and lateral perforant path synapses differ in their mechanisms of LTP induction, associative LTP at these synapses share common downstream mechanisms of induction.

Addictive nicotine alters local circuit inhibition during the induction of in vivo hippocampal synaptic potentiation.

The drug addiction process shares many commonalities with normal learning and memory. Addictive drugs subvert normal synaptic plasticity mechanisms, and the consequent synaptic changes underlie long-lasting modifications in behavior that accrue during the progression from drug use to addiction. Supporting this hypothesis, it was recently shown that nicotine administered to freely moving mice induces long-term synaptic potentiation of the perforant path connection to granule cells of the dentate gyrus. The perforant path carries place and spatial information that links the environment to drug taking. An example of that association is the nicotine-induced synaptic potentiation of the perforant path that was found to underlie nicotine-conditioned place preference. The present study examines the influence of nicotine over local GABAergic inhibition within the dentate gyrus during the drug-induced synaptic potentiation. In vivo recordings from freely moving mice suggested that both feedforward and feedback inhibition onto granules cells were diminished by nicotine during the induction of synaptic potentiation. In vitro brain slice studies indicated that nicotine altered local circuit inhibition within the dentate gyrus leading to disinhibition of granule cells. These changes in local inhibition contributed to nicotine-induced in vivo synaptic potentiation, thus, likely contributed to drug-associated memories. Through this learning process, environmental features become cues that motivate conditioned drug-seeking and drug-taking behaviors.

The antiepileptogenic effect of electrical stimulation at different low frequencies is accompanied with change in adenosine receptors gene expression in rats.

PURPOSE: Previous studies have shown that the anticonvulsant effects of low-frequency stimulation (LFS) can be affected by activation of adenosine receptors. In the present study, the effect of LFS at different frequencies on kindling rate and adenosine receptors gene expression was investigated. METHODS: Animals were kindled by perforant path stimulation in a rapid kindling manner. LFS (0.5, 1, and 5 Hz) was applied after termination of each kindling stimulation. Seizure severity was measured according to behavioral and electrophysiologic parameters. At the end of the experiments, adenosine A(1) and A(2A) receptor gene expression were measured. RESULTS: The inhibitory effect of LFS on kindling acquisition was observed at all frequencies. In addition, the inhibitory action of LFS on enhancement of field excitatory postsynaptic potential slope and population spike amplitude during kindling acquisition was not affected by the LFS frequency. However, the effects of LFS on paired-pulse recordings were greater at frequency of 5 Hz. Application of LFS during kindling acquisition also prevented the kindling induced decrease in the A(1) receptor gene expression and attenuated the level of A(2A) receptor gene expression in the dentate gyrus. These effects were also greater at the frequency of 5 Hz. DISCUSSION: According to these data, it may be suggested that the antiepileptogenic effects of LFS, developed through inhibition of synaptic transmission in the dentate gyrus, is mediated somehow through preventing the decrease of A(1) receptor and through attenuating the A(2A) receptor gene expression. These effects might be dependent on the frequency of LFS.

Intracerebroventricular injection of oxidant and antioxidant molecules affects long-term potentiation in urethane anaesthetized rats.

Production of superoxide anions in the incubation medium of hippocampal slices can induce long-term potentiation (LTP). Other reactive oxygen species (ROS) such as hydrogen peroxide are able to modulate LTP and are likely to be involved in aging mechanisms. The present study explored whether intracerebro-ventricular (ICV) injection of oxidant or antioxidant molecules could affect LTP in vivo. With this aim in mind, field excitatory post-synaptic potentials (fEPSPs) elicited by stimulation of the perforant pathway were recorded in the dentate gyrus of the hippocampal formation in urethane-anesthetized rats. N-acetyl-L-cysteine, hydrogen peroxide (H2O2) or hypoxanthine/xanthine-oxidase solution (a superoxide producing system) were administrated by ICV injection. The control was represented by a group injected with saline ICV. Ten minutes after the injection, LTP was induced in the granule cells of the dentate gyrus by high frequency stimulation of the perforant pathway. Neither the H(2)O(2) injection or the N-acetyl-L-cysteine injection caused any variation in the fEPSP at the 10-min post-injection time point, whereas the superoxide generating system caused a significant increase in the fEPSP. Moreover, at 60 min after tetanic stimulation, all treatments attenuated LTP compared with the control group. These results show that ICV administration of oxidant or antioxidant molecules can modulate LTP in vivo in the dentate gyrus. Particularly, a superoxide producing system can induce potentiation of the synaptic response. Interestingly, ICV injection of oxidants or antioxidants prevented a full expression of LTP compared to the saline injection.

Cytoarchitecture of fibroblast growth factor receptor 2 (FGFR-2) immunoreactivity in astrocytes of neurogenic and non-neurogenic regions of the young adult and aged rat brain.

Fibroblast growth factors (FGFs) are polypeptides that exert diverse biological effects on many cell types and tissues during embryogenesis and adulthood. In the adult brain, FGF-2 is primarily expressed by astrocytes and select groups of neurons. It has been shown that FGF-2 is neuroprotective and can stimulate proliferation of NSCs in neurogenic regions of the adult mammalian brain. Cellular responses to FGFs are mediated through membrane-spanning tyrosine kinase receptors in conjunction with low affinity binding to heparin sulfate proteoglycans. Four FGF receptors (FGFR1-4) have been cloned and characterized to date. In this study, we describe the anatomical distribution of FGFR-2 in young and aged rat brains. We demonstrate that the olfactory bulb, hippocampus, and cerebellum display the most robust FGFR-2 expression and observed age-related decrease in FGFR-2 levels in some but not all brain regions. In addition, we identified astrocytes as the primary source of FGFR-2 expression using immunofluorescence confocal microscopy. The astrocyte populations in the neurogenic areas, the subventricular zone (SVZ) and the subgranular zone (SGZ) of the dentate gyrus, express high levels of FGFR-2 protein, which points to its possible involvement in neurogenesis. We also explored the role of FGFR-2 in response to perforant pathway lesion and observed enhanced FGFR-2 expression by astrocytes surrounding the lesion. Thus, FGF-2 biological effects on astrocytes appear to be mediated through FGFR-2-dependent mechanisms, and this may provide an indirect route by which FGF-2 acts on neuronal populations.

Levetiracetam: antiepileptic properties and protective effects on mitochondrial dysfunction in experimental status epilepticus.

PURPOSE: To assess the anticonvulsant activity of the novel antiepileptic drug, levetiracetam (LEV) in a model of self-sustaining limbic status epilepticus, and to measure the consequence of LEV treatment on the pattern of mitochondrial dysfunction known to occur after status epilepticus (SE). METHODS: The rat perforant pathway was stimulated for 2 h to induce self-sustaining status epilepticus (SSSE). Stimulated rats were assigned to one of three treatment groups, receiving intraperitoneal injections of saline, 200 mg/kg LEV, or 1,000 mg/kg LEV, 15 min into SSSE and at 3 times over the next 44-h period. All animals received diazepam after 3-h SSSE to terminate seizures. Forty-four hours later, the hippocampi were extracted and prepared for electrochemical high-performance liquid chromatography (HPLC), to measure reduced glutathione levels, and for spectrophotometric assays to measure activities of mitochondrial enzymes (aconitase, alpha-ketoglutarate dehydrogenase, citrate synthase, complex I, and complex II/III). These parameters were compared between treatment groups and with sham-operated rats. RESULTS: LEV administration did not terminate seizures or have any significant effect on spike frequency, although rats that received 1,000 mg/kg LEV did exhibit improved behavioral seizure parameters. Significant biochemical changes occurred in saline-treated stimulated rats compared with shams: with reductions in glutathione, alpha-ketoglutarate dehydrogenase, aconitase, citrate synthase, and complex I activities. Complex II/III activities were unchanged throughout. Rats that received 1,000 mg/kg LEV had significantly improved biochemical parameters, in many instances, comparable to sham control levels. CONCLUSIONS: Despite continuing seizures, administration of LEV (1,000 mg/kg) protects against mitochondrial dysfunction, indicating that in addition to its antiepileptic actions, LEV may have neuroprotective effects.

Subcortical deafferentation impairs behavioral reinforcement of long-term potentiation in the dentate gyrus of freely moving rats.

Long-term potentiation is a form of neural functional plasticity which has been related with memory formation and recovery of function after brain injury. Previous studies have shown that a transient early-long-term potentiation can be prolonged by direct stimulation of distinct brain areas, or behavioral stimuli with a high motivational content. The basolateral amygdala and other subcortical structures, like the medial septum and the locus coeruleus, are involved in mediating the reinforcing effect. We have previously shown that the lesion of the fimbria-fornix--the main entrance of subcortical afferents to the hippocampus--abolishes the reinforcing basolateral amygdala-effects on long-term potentiation in the dentate gyrus in vivo. It remains to be investigated, however, if such subcortical afferents may also be important for behavioral reinforcement of long-term potentiation. Young-adult (8 weeks) Sprague-Dawley male rats were fimbria-fornix-transected under anesthesia, and electrodes were implanted at the dentate gyrus and the perforant path. One week after surgery the freely moving animals were studied. Fimbria-fornix-lesion reduced the ability of the animals to develop long-term potentiation when a short pulse duration was used for tetanization (0.1 ms per half-wave of a biphasic stimulus), whereas increasing the pulse duration to 0.2 ms per half-wave during tetanization resulted in a transient early-long-term potentiation lasting about 4 h in the lesioned animals, comparable to that obtained in non-lesioned or sham-operated control rats. In water-deprived (24 h) control animals, i.e. in non-lesioned and sham-operated rats, early-long-term potentiation could be behaviorally reinforced by drinking 15 min after tetanization. However, in fimbria-fornix-lesioned animals long-term potentiation-reinforcement by drinking was not detected. This result indicates that the effect of behavioral-motivational stimuli to reinforce long-term potentiation is mediated by subcortical, heterosynaptic afferents.

Functional properties of ES cell-derived neurons engrafted into the hippocampus of adult normal and chronically epileptic rats.

PURPOSE: Embryonic stem (ES) cell-based therapy strategies are thought to bear considerable promise in chronic neurologic disorders. Nonetheless, studies addressing the functional properties of ES cell-derived progeny after transplantation into the adult, pathologically modified CNS are scarce. METHODS: We therefore transplanted ES cell-derived neural precursors expressing enhanced green fluorescent protein only in neuronal progeny bilaterally into the hippocampi of pilocarpine-treated chronically epileptic and sham-control rats. Whole-cell patch-clamp recordings of identified ES cell-derived neurons (ESNs) in hippocampal slices were performed 13 to 34 days after transplantation. RESULTS: Most ESNs were found in clusters at the transplant site and did not migrate into host tissue. However, they gave rise to a dense network of processes extending over large distances into the host tissue. All ESNs possessed the ability to generate action potentials and expressed voltage-gated Na+ and K+ currents, as well as hyperpolarization-activated currents. Likewise, most ESNs received non-N-methyl-D-aspartate (NMDA) and gamma-aminobutyric acid (GABA)A receptor-mediated synaptic input. Both types of synapses displayed intact short-term plasticity. An unusual feature of the majority of ESNs was the occurrence of spontaneous pacemaking activity at frequencies approximately 3 Hertz. No obvious differences were found between the functional properties of ESNs in sham-control and in pilocarpine-treated rats. CONCLUSIONS: After transplantation into adult control and epileptic rats, ESNs displayed intrinsic and synaptic properties characteristic of neurons. Even though ESNs remained close to the transplant site, the formation of extensive networks of graft-derived processes may be useful for ES cell-based substance delivery.

Age-related impairments of synaptic plasticity in the lateral perforant path input to the dentate gyrus of galanin overexpressing mice.

In the present study, electrophysiological recordings were made from hippocampal slices obtained from mice overexpressing galanin under the promoter for the platelet-derived growth factor-B (GalOE mice). In these mice, a particularly strong galanin expression is seen in the granule cell layer/mossy fibers. Paired-pulse facilitation (PPF) of excitatory postsynaptic field potentials (fEPSPs) at the lateral perforant path (LPP)-dentate gyrus synapses was elicited in the dentate gyrus after stimulation with different interpulse intervals. Slices from young adult wild-type (WT) animals showed significant PPF of the 2nd EPSP evoked with paired-pulse stimuli, while PPF was reduced in slices from young adult GalOE mice, as well as aged WT mice, but were not observed at all in slices from aged GalOE animals. Application of the putative galanin antagonist M35 increased PPF in slices from aged WT mice as well as from adult and aged GalOE mice, but had no effect in slices taken from young adult WT mice. These data indicate that galanin is involved in hippocampal synaptic plasticity, in particular in age-related reduction of synaptic plasticity in the LPP input to the dentate gyrus. Galaninergic mechanisms may therefore represent therapeutic targets for treatment of age-related memory deficits and Alzheimer's disease.

Synergism between topiramate and budipine in refractory status epilepticus in the rat.

PURPOSE: To evaluate the antiepileptic and neuroprotective properties of topiramate (TPM) alone and with coadministration of the N-methyl-D-aspartate (NMDA)-receptor antagonist budipine in a rat model of refractory status epilepticus. METHODS: Male Sprague-Dawley rats had electrodes implanted into the perforant path and dentate granule cell layer of the hippocampus under halothane anesthesia. Approximately 1 week after surgery, the perforant path of each animal was electrically stimulated for 2 h to induce self-sustaining status epilepticus. Successfully stimulated rats were given intraperitoneally vehicle (n = 6), TPM (20-320 mg/kg; n = 28), budipine (10 mg/kg; n = 5), or budipine (10 mg/kg) and TPM (80 mg/kg; n = 6) 10 min after the end of the stimulation and monitored behaviorally and electroencephalographically for a further 3 h. The animals were killed 14 days later, and histopathology was assessed. RESULTS: Neither budipine alone nor TPM at any dose terminated status epilepticus. Despite this, TPM resulted in various degrees of neuroprotection at doses between 40 and 320 mg/kg. Coadministration of budipine with TPM terminated the status epilepticus in all rats. This combination also significantly improved the behavioral profile and prevented status-induced cell death compared with control. CONCLUSIONS: Budipine and TPM are an effective drug combination in stopping self-sustained status epilepticus, and TPM alone was neuroprotective, despite the continuation of seizure activity.

Distinct properties of presynaptic group II and III metabotropic glutamate receptor-mediated inhibition of perforant pathway-CA1 EPSCs.

I have compared the effects of group II or III metabotropic glutamate receptor (mGluR) activation on monosynaptic excitatory responses recorded intracellularly from CA1 pyramidal neurons of rat hippocampus and evoked by perforant pathway stimulation in vitro. The excitatory postsynaptic currents (EPSCs) were reduced either by the group II mGluR agonist LY354740 (500 nM, 31 +/- 6% of control) or by the group III agonist L-AP4 (400 microM, 53 +/- 5% of control). Both drugs enhanced EPSC paired-pulse facilitation (range 125-189% of control). These effects were blocked by the broad-spectrum mGluR antagonist LY341495 (1 or 20 microM) which when applied alone did not significantly change the EPSCs elicited at low (0.1-0.2 Hz) or higher (1-100 Hz) frequency of stimulation. Prior reduction of the EPSCs induced by L-AP4 did not occlude the subsequent inhibition elicited by LY354740. The effect of LY354740, but not that of L-AP4, was blocked in the presence of the cAMP analogue Sp-cAMPS (20 microM) and with the K(+) channel antagonist alpha-dendrotoxin (125 nM). In contrast, the effect of L-AP4, but not that of LY354740, was prevented by the calmodulin inhibitor ophiobolin A (25 microM) and with the N-type Ca(2+) channel antagonist omega-conotoxin-GVIA (1 microM). In the presence of the P/Q type Ca(2+) channel antagonist omega-agatoxin-IVA (400 nM), the EPSCs were depressed either by LY354740 or by L-AP4. Groups II and III mGluRs are segregated at the presynaptic terminal, and there are distinct differences between the properties of the presynaptic inhibition mediated by these two groups of receptors.

Perforant path activation of ectopic granule cells that are born after pilocarpine-induced seizures.

Granule cells in the dentate gyrus are born throughout life, and various stimuli can affect their development in the adult brain. Following seizures, for instance, neurogenesis increases greatly, and some new cells migrate to abnormal (ectopic) locations, such as the hilus. Previous electrophysiological studies of this population have shown that they have intrinsic properties that are similar to normal granule cells, but differ in other characteristics, consistent with abnormal integration into host circuitry. To characterize the response of ectopic hilar granule cells to perforant path stimulation, intracellular recordings were made in hippocampal slices from rats that had pilocarpine-induced status epilepticus and subsequent spontaneous recurrent seizures. Comparisons were made with granule cells located in the granule cell layer of both pilocarpine- and saline-treated animals. In addition, a few ectopic hilar granule cells were sampled from saline-treated rats. Remarkably, hilar granule cells displayed robust responses, even when their dendrites were not present within the molecular layer, where perforant path axons normally terminate. The evoked responses of hilar granule cells were similar in several ways to those of normally positioned granule cells, but there were some differences. For example, there was an unusually long latency to onset of responses evoked in many hilar granule cells, especially those without molecular layer dendrites. Presumably this is due to polysynaptic activation by the perforant path. These results indicate that synaptic reorganization after seizures can lead to robust activation of newly born hilar granule cells by the perforant path, even when their dendrites are not in the terminal field of the perforant path. Additionally, the fact that these cells can be found in normal tissue and develop similar synaptic responses, suggests that seizures, while not necessary for their formation, strongly promote their generation and the development of associated circuits, potentially contributing to a lowered seizure threshold.

Functional integration of embryonic stem cell-derived neurons in hippocampal slice cultures.

The generation of neurons and glia from pluripotent embryonic stem (ES) cells represents a promising strategy for the study of CNS development and repair. ES cell-derived neural precursors have been shown to develop into morphologically mature neurons and glia when grafted into brain and spinal cord. However, there is a surprising shortage of data concerning the functional integration of ES cell-derived neurons (ESNs) into the host CNS tissue. Here, we use ES cells engineered to express enhanced green fluorescent protein (EGFP) only in neuronal progeny to study the functional properties of ESNs during integration into long-term hippocampal slice cultures. After incorporation into the dentate gyrus, EGFP+ donor neurons display a gradual maturation of their intrinsic discharge behavior and a concomitant increase in the density of voltage-gated Na+ and K+ channels. Integrated ESNs express AMPA and GABA(A) receptor subunits. Most importantly, neurons derived from ES cells receive functional glutamatergic and GABAergic synapses from host neurons. Specifically, we demonstrate that host perforant path axons form synapses onto integrated ESNs. These synapses between host and ES cell-derived neurons display pronounced paired-pulse facilitation indicative of intact presynaptic short-term plasticity. Thus, ES cell-derived neural precursors generate functionally active neurons capable of integrating into the brain circuitry.

Stimulation-induced reset of hippocampal theta in the freely performing rat.

Previous research has suggested that visual and auditory stimuli in a working memory task have the ability to reset hippocampal theta, perhaps allowing an organism to encode the incoming information optimally. The present study examined two possible neural pathways involved in theta resetting. Rats were trained on a visual discrimination task in an operant chamber. At the beginning of a trial, a light appeared over a centrally located lever that the rat was required to press to receive a water reward. There was a 30-s intertrial interval before the next light stimulus appeared. After learning the task, all rats received surgical implantation of stimulating electrodes in both the fornix and the perforant path and recording electrodes, bilaterally in the hippocampus. After surgery, theta was recorded before and after the light stimulus to determine whether resetting to the visual stimulus occurred. During the intertrial interval, rats received single-pulse electrical stimulation of either the fornix or perforant path. Theta was recorded both before and after the electrical stimulation to determine whether resetting occurred. In this experiment, hippocampal theta was reset after all three stimulus conditions (light, perforant path, and fornix stimulation), with the greatest degree of reset occurring after the fornix stimulation. The results suggest that activation of the perforant path and fornix may underlie theta reset and provide a mechanism by which the hippocampus may enhance cognitive processing.

Oxytocin induces long-term depression on the rat dentate gyrus: possible ATPase and ectoprotein kinase mediation.

We studied the effects of the neuropeptide oxytocin (OT) on the long-term potentiation (LTP) paradigm in the dentate gyrus (DG) of urethane anesthetized rats. Intracerebroventricular injection of 1 microg of the hormone in 1 microl of physiological solution 2min before tetanization produced a significant decrease in both components of the perforant path evoked potentials (EP) in the DG. The effects appeared right after the tetanization stimuli and were more pronounced in the excitatory postsynaptic components of the EPs. The decrements lasted for the 2h of recording time. We concluded that OT induced and maintained long-term depression on the DG. In contrast, injection of OT in the absence of tetanic stimulation did not significantly affect perforant path EP in the DG. The results are discussed taking particular consideration of the inhibitory effects the OT has on (Ca(2+)+Mg(2+)) ATPase at membrane levels and the potential interference that this action may have with phosphorylation processes via an ectoprotein kinase isolated from membranes of hippocampal pyramidal neurons. Blocking of this ectoprotein kinase in vitro significantly impairs establishment and maintenance of LTP.