Ultrastructural Correlates of Presynaptic Functional Heterogeneity in Hippocampal Synapses.

Although similar in molecular composition, synapses can exhibit strikingly distinct functional transmitter release and plasticity characteristics. To determine whether ultrastructural differences co-define this functional heterogeneity, we combine hippocampal organotypic slice cultures, high-pressure freezing, freeze substitution, and 3D-electron tomography to compare two functionally distinct synapses: hippocampal Schaffer collateral and mossy fiber synapses. We find that mossy fiber synapses, which exhibit a lower release probability and stronger short-term facilitation than Schaffer collateral synapses, harbor lower numbers of docked synaptic vesicles at active zones and a second pool of possibly tethered vesicles in their vicinity. Our data indicate that differences in the ratio of docked versus tethered vesicles at active zones contribute to distinct functional characteristics of synapses.

Temporal dynamics of hippocampal neurogenesis in chronic neurodegeneration.

The study of neurogenesis during chronic neurodegeneration is crucial in order to understand the intrinsic repair mechanisms of the brain, and key to designing therapeutic strategies. In this study, using an experimental model of progressive chronic neurodegeneration, murine prion disease, we define the temporal dynamics of the generation, maturation and integration of new neurons in the hippocampal dentate gyrus, using dual pulse-chase, multicolour γ-retroviral tracing, transmission electron microscopy and patch-clamp. We found increased neurogenesis during the progression of prion disease, which partially counteracts the effects of chronic neurodegeneration, as evidenced by blocking neurogenesis with cytosine arabinoside, and helps to preserve the hippocampal function. Evidence obtained from human post-mortem samples, of both variant Creutzfeldt-Jakob disease and Alzheimer's disease patients, also suggests increased neurogenic activity. These results open a new avenue into the exploration of the effects and regulation of neurogenesis during chronic neurodegeneration, and offer a new model to reproduce the changes observed in human neurodegenerative diseases.

Neuroanatomical clues to altered neuronal activity in epilepsy: from ultrastructure to signaling pathways of dentate granule cells.

The dynamic aspects of epilepsy, in which seizures occur sporadically and are interspersed with periods of relatively normal brain function, present special challenges for neuroanatomical studies. Although numerous morphologic changes can be identified during the chronic period, the relationship of many of these changes to seizure generation and propagation remains unclear. Mossy fiber sprouting is an example of a frequently observed morphologic change for which a functional role in epilepsy continues to be debated. This review focuses on neuroanatomically identified changes that would support high levels of activity in reorganized mossy fibers and potentially associated granule cell activation. Early ultrastructural studies of reorganized mossy fiber terminals in human temporal lobe epilepsy tissue have identified morphologic substrates for highly efficacious excitatory connections among granule cells. If similar connections in animal models contribute to seizure activity, activation of granule cells would be expected. Increased labeling with two activity-related markers, Fos and phosphorylated extracellular signal-regulated kinase, has suggested increased activity of dentate granule cells at the time of spontaneous seizures in a mouse model of epilepsy. However, neuroanatomical support for a direct link between activation of reorganized mossy fiber terminals and increased granule cell activity remains elusive. As novel activity-related markers are developed, it may yet be possible to demonstrate such functional links and allow mapping of seizure activity throughout the brain. Relating patterns of neuronal activity during seizures to the underlying morphologic changes could provide important new insights into the basic mechanisms of epilepsy and seizure generation.

Critical involvement of postsynaptic protein kinase activation in long-term potentiation at hippocampal mossy fiber synapses on CA3 interneurons.

Hippocampal mossy fiber (MF) synapses on area CA3 lacunosum-moleculare (L-M) interneurons are capable of undergoing a Hebbian form of NMDA receptor (NMDAR)-independent long-term potentiation (LTP) induced by the same type of high-frequency stimulation (HFS) that induces LTP at MF synapses on pyramidal cells. LTP of MF input to L-M interneurons occurs only at synapses containing mostly calcium-impermeable (CI)-AMPA receptors (AMPARs). Here, we demonstrate that HFS-induced LTP at these MF-interneuron synapses requires postsynaptic activation of protein kinase A (PKA) and protein kinase C (PKC). Brief extracellular stimulation of PKA with forskolin (FSK) alone or in combination with 1-Methyl-3-isobutylxanthine (IBMX) induced a long-lasting synaptic enhancement at MF synapses predominantly containing CI-AMPARs. However, the FSK/IBMX-induced potentiation in cells loaded with the specific PKA inhibitor peptide PKI(6-22) failed to be maintained. Consistent with these data, delivery of HFS to MFs synapsing onto L-M interneurons loaded with PKI(6-22) induced posttetanic potentiation (PTP) but not LTP. Hippocampal sections stained for the catalytic subunit of PKA revealed abundant immunoreactivity in interneurons located in strata radiatum and L-M of area CA3. We also found that extracellular activation of PKC with phorbol 12,13-diacetate induced a pharmacological potentiation of the isolated CI-AMPAR component of the MF EPSP. However, HFS delivered to MF synapses on cells loaded with the PKC inhibitor chelerythrine exhibited PTP followed by a significant depression. Together, our data indicate that MF LTP in L-M interneurons at synapses containing primarily CI-AMPARs requires some of the same signaling cascades as does LTP of glutamatergic input to CA3 or CA1 pyramidal cells.

Identification and characterization of nCLP2, a novel C1q family protein expressed in the central nervous system.

The C1q family is characterized by the C-terminally conserved globular C1q (gC1q) domain. Although more than 30 C1q family proteins have been identified in mammals, many of them remain ill-defined with respect to their molecular and biological properties. Here, we report on a novel C1q family protein specifically expressed in the central nervous system (CNS), which we designated neural C1q-like protein (nCLP) 2. nCLP2 was secreted as disulphide-bonded multimers comprising trimeric units. The multimers were stabilized by interchain disulphide bonds involving the cysteine residues in the N-terminal variable region and the C-terminal gC1q domain. The expression of nCLP2 was restricted to several brain regions and retina, including regions associated with memory formation (i.e. hippocampus, entorhinal cortex, anterodorsal thalamic nucleus). Immunoelectron microscopy revealed that nCLP2 was localized in the mossy fibre axons of hippocampal granule cells and their synaptic boutons and clefts, implying that nCLP2 was anterogradely transported in mossy fibres and secreted from the presynaptic termini. These results suggest that nCLP2 plays roles in synaptic function and maintenance in the CNS.

Nicotine-induced and depolarisation-induced glutamate release from hippocampus mossy fibre synaptosomes: two distinct mechanisms.

Hippocampus mossy fibre terminals activate CA3 pyramidal neurons via two distinct mechanisms, both quantal and glutamatergic: (i) rapid excitatory transmission in response to afferent action potentials and (ii) delayed and prolonged release following nicotinic receptor activation. These processes were analysed here using rat hippocampus mossy fibres synaptosomes. The relationships between synaptosome depolarisation and glutamate release were established in response to high-KCl and gramicidin challenges. Half-maximal release corresponded to a 52 mV depolarisation step. KCl-induced release was accompanied by transient dissipation of the proton gradient across synaptic vesicle membrane. Nicotine elicited a substantial glutamate release from mossy fibre synaptosomes (EC(50) 3.14 microM; V(max) 12.01 +/- 2.1 nmol glutamate/mg protein; Hill's coefficient 0.99). However, nicotine-induced glutamate release was not accompanied by any change in the membrane potential or in the vesicular proton gradient. The effects of acetylcholine (200 microM) were similar to those of nicotine (25 microM). Nicotinic alpha7 receptors were evidenced by immuno-cytochemistry on the mossy fibre synaptosome plasma membrane. Therefore, the same terminals can release glutamate in response to two distinct stimuli: (i) rapid neurotransmission involving depolarisation-induced activation of voltage-gated Ca(2+) channels and (ii) a slower nicotinic activation which does not involve depolarisation or dissipation of the vesicular proton gradient.

Effect of hormonal replacement therapy in the hippocampus of ovariectomized epileptic female rats using the pilocarpine experimental model.

Amado and Cavalheiro [Amado, D., Cavalheiro, E.A., 1998. Hormonal and gestational parameters in female rats submitted to the pilocarpine model of epilepsy. Epilepsy Res. 32, 266-274], studying the establishment of the pilocarpine epilepsy model in female rats observed that the estrous cycle was dramatically altered during the three periods of this experimental model. This work was delineated to study the function of sexual hormones in the development of the epilepsy model induced by pilocarpine in ovariectomized rats. Experimental groups were: (a) control animals during estrus phase of the estrous cycle (E) and ovariectomized female rats (OVX) treated with saline instead of pilocarpine in the same volume, (b) experimental animals, that developed status epilepticus (SE) and were studied during the chronic phase of this model: intact chronic rats (CHRON) and ovariectomized chronic rats (OVX+CHRON) and (c) ovariectomized chronic rats, that were submitted to hormonal replacement therapy treated with: medroxyprogesterone (OVX+CHRON+MPA); 17beta-estradiol (OVX+CHRON+E2), or both (OVX+CHRON+E2+MPA). All ovariectomized animals showed genital atrophy 4 days after the surgical procedure. Moreover, all animals that developed SE and survived showed spontaneous recurrent seizures during the chronic phase. Concerning to seizure frequency, animals receiving medroxyprogesterone associated with 17beta-estradiol showed decreased seizures' number. However, animals that received only medroxyprogesterone therapy also showed reduction in the number of seizures. In addition, hormonal treatment was also able to stabilize the mossy fibers sprouting process, showing the importance of these hormones in the development of the epilepsy in female rats.

Neurons born in the adult dentate gyrus form functional synapses with target cells.

Adult neurogenesis occurs in the hippocampus and the olfactory bulb of the mammalian CNS. Recent studies have demonstrated that newborn granule cells of the adult hippocampus are postsynaptic targets of excitatory and inhibitory neurons, but evidence of synapse formation by the axons of these cells is still lacking. By combining retroviral expression of green fluorescent protein in adult-born neurons of the mouse dentate gyrus with immuno-electron microscopy, we found output synapses that were formed by labeled terminals on appropriate target cells in the CA3 area and the hilus. Furthermore, retroviral expression of channelrhodopsin-2 allowed us to light-stimulate newborn granule cells and identify postsynaptic target neurons by whole-cell recordings in acute slices. Our structural and functional evidence indicates that axons of adult-born granule cells establish synapses with hilar interneurons, mossy cells and CA3 pyramidal cells and release glutamate as their main neurotransmitter.

Adenosine A2A receptors are essential for long-term potentiation of NMDA-EPSCs at hippocampal mossy fiber synapses.

The physiological conditions under which adenosine A2A receptors modulate synaptic transmission are presently unclear. We show that A2A receptors are localized postsynaptically at synapses between mossy fibers and CA3 pyramidal cells and are essential for a form of long-term potentiation (LTP) of NMDA-EPSCs induced by short bursts of mossy fiber stimulation. This LTP spares AMPA-EPSCs and is likely induced and expressed postsynaptically. It depends on a postsynaptic Ca2+ rise, on G protein activation, and on Src kinase. In addition to A2A receptors, LTP of NMDA-EPSCs requires the activation of NMDA and mGluR5 receptors as potential sources of Ca2+ increase. LTP of NMDA-EPSCs displays a lower threshold for induction as compared with the conventional presynaptic mossy fiber LTP; however, the two forms of LTP can combine with stronger induction protocols. Thus, postsynaptic A2A receptors may potentially affect information processing in CA3 neuronal networks and memory performance.

[Correlation between hippocampal mossy fiber sprouting and synaptic reorganization and mechanisms of temporal lobe epilepsy].

OBJECTIVE: To explore the effects of the ultrastructural features of sprouted mossy fiber synapses in the mechanism of temporal lobe epilepsy. To explore the correlation between axon guidance molecule-netrin-1 gene expression and mossy fiber synaptic reorganization. METHODS: Sixty-one SD rats underwent intraperitoneal injection of lithium chloride and pilocarpine to establish models of status epilepticus characterized with temporal lobe epilepsy. Nineteen rats were used as controls. One, 2, and 4 weeks after the injection, a certain numbers of rat were killed with their brains taken out. The sprouted mossy fiber synaptic terminals were labeled by Timm histochemistry and the ultrastructure of new synapses were observed by electron microscopy. By in situ hybridization, the mRNA expression of netrin-1 gene was observed. RESULTS: The sprouted mossy fiber synapses in epileptic rats most commonly formed asymmetric synapses with dendritic spines and occasionally with granule cell somata. Seven days after the injection, up-regulation of netrin-1 mRNA expression was seen in the dentate granule cell layers of hippocampus and continued to 4 weeks after the injection. The time course of the increase of netrin-1 mRNA in the dentate granule cell layers was correlated with the time course of mossy fiber sprouting and synaptic reorganization in hippocampus. CONCLUSION: The ultrastructural features of sprouted mossy fiber synapses support the viewpoint that the reorganization of synapses prominently involves the formation of recurrent excitatory circuits. The axon guidance molecule- netrin-1 plays an important role in the process of mossy fiber axonal outgrowth and synaptogenesis in the hippocampal dentate gyrus.

Physiological and morphological characterization of dentate granule cells in the p35 knock-out mouse hippocampus: evidence for an epileptic circuit.

There is a high correlation between pediatric epilepsies and neuronal migration disorders. What remains unclear is whether there are intrinsic features of the individual dysplastic cells that give rise to heightened seizure susceptibility, or whether these dysplastic cells contribute to seizure activity by establishing abnormal circuits that alter the balance of inhibition and excitation. Mice lacking a functional p35 gene provide an ideal model in which to address these questions, because these knock-out animals not only exhibit aberrant neuronal migration but also demonstrate spontaneous seizures. Extracellular field recordings from hippocampal slices, characterizing the input-output relationship in the dentate, revealed little difference between wild-type and knock-out mice under both normal and elevated extracellular potassium conditions. However, in the presence of the GABA(A) antagonist bicuculline, p35 knock-out slices, but not wild-type slices, exhibited prolonged depolarizations in response to stimulation of the perforant path. There were no significant differences in the intrinsic properties of dentate granule cells (i.e., input resistance, time constant, action potential generation) from wild-type versus knock-out mice. However, antidromic activation (mossy fiber stimulation) evoked an excitatory synaptic response in over 65% of granule cells from p35 knock-out slices that was never observed in wild-type slices. Ultrastructural analyses identified morphological substrates for this aberrant excitation: recurrent axon collaterals, abnormal basal dendrites, and mossy fiber terminals forming synapses onto the spines of neighboring granule cells. These studies suggest that granule cells in p35 knock-out mice contribute to seizure activity by forming an abnormal excitatory feedback circuit.

Ecto-nucleotidases and nucleoside transporters mediate activation of adenosine receptors on hippocampal mossy fibers by P2X7 receptor agonist 2'-3'-O-(4-benzoylbenzoyl)-ATP.

The ionotropic and cytolytic P2X7 receptor is typically found on immune cells, where it is involved in the release of cytokines. Recently, P2X7 receptors were reported to be localized to presynaptic nerve terminals and to modulate transmitter release. In the present study, we reassessed this unexpected role of P2X7 receptors at hippocampal mossy fiber-CA3 synapses. In agreement with previous findings, the widely used P2X7 agonist 2'-3'-O-(4-benzoylbenzoyl)-adenosine-5'-triphosphate (BzATP) clearly depressed field potentials (fEPSPs); however, no evidence for an involvement of P2X7 receptors could be obtained. First, depression of fEPSPs by BzATP was unchanged in P2X7-/- mice. Second, experiments using P2X7-/- mice, immunohistochemistry, and electron microscopy showed that the antigen detected by frequently used P2X7 antibodies is not compatible with a plasmalemmal P2X7 receptor. Third, BzATP did not alter Ca2+ levels in synaptic terminals. In contrast, the depression of fEPSPs by BzATP was fully blocked by adenosine (A1) receptor antagonists. Furthermore, the application of BzATP also activated postsynaptic A1 receptor-coupled K+ channels. This effect of BzATP was mimicked by ATP and adenosine and was completely prevented by enzymes specifically degrading adenosine. Activation of A1-coupled K+ channels by BzATP was dependent on ecto-nucleotidases, extracellular enzymes that convert ATP to adenosine. Moreover, the opening of A1-coupled K+ channels by BzATP was dependent on nucleoside transporters. Taken together, our results indicate that BzATP is extracellularly catabolized to Bz-adenosine and subsequently hetero-exchanged for intracellular adenosine and then depresses mossy fiber fEPSPs through presynaptic A1 receptors rather than through P2X7 receptors. Thus, the present study casts doubts on the neuronal localization of P2X7 receptors in rodent hippocampus.

Phosphorylation of hippocampal Erk-1/2, Elk-1, and p90-Rsk-1 during contextual fear conditioning: interactions between Erk-1/2 and Elk-1.

The phosphorylation of proteins involved in the MAP kinase signal transduction pathway was investigated during associative learning of C57BL/6J mice. Context-dependent fear conditioning, consisting of a single exposure of mice to a context followed by foot shock, was employed as a learning paradigm. Control groups consisted of mice exposed to context only or an immediate shock in the context. Coincident up-regulation of phosphorylated Erk-1/2 and Elk-1 was observed in the CA3 hippocampal subfield and dentate gyrus 30 min after fear conditioning but not after the control paradigms. Phosphorylated Erk-1/2 and Elk-1 were associated and predominantly colocalized in the mossy fibers. In vitro kinase assays showed that hippocampal Erk-1/2 phosphorylates Elk-1. Notably, Elk-1 in turn enhances the phosphorylation of Erk-1/2 and its downstream target p90Rsk-1. Increased phosphorylation and nuclear translocation of p90Rsk-1 was also demonstrated in the CA3 hippocampal area in vivo during contextual fear conditioning. The observed interactions between hippocampal Elk-1 and Erk-1/2 proteins may affect the consolidation of contextual memories through activation of the downstream nuclear targets of Erk-1/2, such as p90Rsk-1, without requiring nuclear translocation of Elk-1 and Erk-1/2.

Nectin: an adhesion molecule involved in formation of synapses.

The nectin-afadin system is a novel cell-cell adhesion system that organizes adherens junctions cooperatively with the cadherin-catenin system in epithelial cells. Nectin is an immunoglobulin-like adhesion molecule, and afadin is an actin filament-binding protein that connects nectin to the actin cytoskeleton. Nectin has four isoforms (-1, -2, -3, and -4). Each nectin forms a homo-cis-dimer followed by formation of a homo-trans-dimer, but nectin-3 furthermore forms a hetero-trans-dimer with nectin-1 or -2, and the formation of each hetero-trans-dimer is stronger than that of each homo-trans-dimer. We show here that at the synapses between the mossy fiber terminals and dendrites of pyramidal cells in the CA3 area of adult mouse hippocampus, the nectin-afadin system colocalizes with the cadherin-catenin system, and nectin-1 and -3 asymmetrically localize at the pre- and postsynaptic sides of puncta adherentia junctions, respectively. During development, nectin-1 and -3 asymmetrically localize not only at puncta adherentia junctions but also at synaptic junctions. Inhibition of the nectin-based adhesion by an inhibitor of nectin-1 in cultured rat hippocampal neurons results in a decrease in synapse size and a concomitant increase in synapse number. These results indicate an important role of the nectin-afadin system in the formation of synapses.

Vesicular GABA transporter mRNA expression in the dentate gyrus and in mossy fiber synaptosomes.

In the normal granule cells of the dentate gyrus, glutamate and both gamma-aminobutyric acid (GABA) and glutamic acid decarboxylase (GAD) coexist. GAD expression is increased after seizures, and simultaneous glutamatergic and GABAergic neurotransmission from the mossy fibers to CA3 appears, supporting the hypothesis that GABA can be released from the mossy fibers. To sustain GABAergic neurotransmission, the amino acid must be transported into synaptic vesicles. To address this, using RT-PCR we looked for the presence and regulation of expression of the vesicular GABA transporter (VGAT) mRNA in the dentate gyrus and in mossy fiber synaptosomes of control and kindled rats. We found trace amounts of VGAT mRNA in the dentate gyrus and mossy fiber synaptosomes of control rats. In the dentate gyrus of kindled rats with several seizures and of control rats subject to one acute seizure, no changes were apparent either 1 or 24 h after the seizures. However, repetitive synaptic or antidromic activation of the granule cells in slices of control rats in vitro induces an activity-dependent enhancement of VGAT mRNA expression in the dentate. Surprisingly, in the mossy fiber synaptosomes of seizing rats, the levels of VGAT mRNA were significantly higher than in controls. These data show that the granule cells and their mossy fibers, besides containing machinery for the synthesis of GABA, also contain the elements that support its vesiculation. This further supports the notion that local synaptic molecular changes enable mossy fibers to release GABA in response to enhanced excitability.

Immunoelectron microscopic localization of the neural recognition molecules L1, NCAM, and its isoform NCAM180, the NCAM-associated polysialic acid, beta1 integrin and the extracellular matrix molecule tenascin-R in synapses of the adult rat hippocampus.

We have investigated the possibility that morphologically different excitatory glutamatergic synapses of the "trisynaptic circuit" in the adult rodent hippocampus, which display different types of long-term potentiation (LTP), may express the immunoglobulin superfamily recognition molecules L1 and NCAM, the extracellular matrix molecule tenascin-R, and the extracellular matrix receptor constituent beta1 integrin in a differential manner. The neural cell adhesion molecules L1, NCAM (all three major isoforms), NCAM180 (the largest major isoform with the longest cytoplasmic domain), beta1 integrin, polysialic acid (PSA) associated with NCAM, and tenascin-R were localized by pre-embedding immunostaining procedures in the CA3/CA4 region (mossy fiber synapses) and in the dentate gyrus (spine synapses) of the adult rat hippocampus. Synaptic membranes of mossy fiber synapses where LTP is expressed presynaptically did not show detectable levels of immunoreactivity for any of the molecules/epitopes studied. L1, NCAM, and PSA, but not NCAM180 or beta1 integrin, were detectable on axonal membranes of fasciculating mossy fibers. In contrast to mossy fiber synapses, spine synapses in the outer third of the molecular layer of the dentate gyrus, which display postsynaptic expression mechanisms of LTP, were both immunopositive and immunonegative for NCAM, NCAM180, beta1 integrin, and PSA. Those spine synapses postsynaptically immunoreactive for NCAM or PSA also showed immunoreactivity on their presynaptic membranes. NCAM180 was not detectable presynaptically in spine synapses. L1 could not be found in spine synapses either pre- or postsynaptically. Also, the extracellular matrix molecule tenascin-R was not detectable in synaptic clefts of all synapses tested, but was amply present between fasciculating axons, axon-astrocyte contact areas, and astrocytic gap junctions. Differences in expression of the membrane-bound adhesion molecules at both types of synapses may reflect the different mechanisms for induction and/or maintenance of synaptic plasticity.

A grading system for hippocampal sclerosis based on the degree of hippocampal mossy fiber sprouting.

In patients suffering from temporal lobe epilepsy (TLE) a highly variable degree of hippocampal sclerosis (HS) can be observed. For standard neuropathological evaluation after hippocampal resection, neuronal cell loss in the hippocampal subareas is assessed (Wyler score 0-4) [Wyler et al. (1992) J Epilepsy 5: 220-225]. Other marked morphological changes in the sclerotic hippocampus are gliosis and loss of mossy fibers in the hilus and mossy fiber sprouting in the supragranular layer. In this study we quantified changes in mossy fiber density using Timm's stain in resected hippocampal tissue from patients with various degrees of sclerosis. We found that tissue specimens from patients without sclerosis (W0) show almost no mossy fiber sprouting. Patients with moderate sclerosis show sprouting without fiber loss in the hilus, whereas specimens from patients with severe sclerosis show sprouting as well as fiber loss in the hilus. Thus, analysis of mossy fiber abundance in hilus and supragranular layer by the rapid and simple Timm's stain is a sensitive measure for hippocampal sclerosis. It provides a reliable rapid tool for neuropathological evaluation, even if the tissue only contains dentate gyrus due to the sectioning procedure.

Abnormal morphological and functional organization of the hippocampus in a p35 mutant model of cortical dysplasia associated with spontaneous seizures.

Cortical dysplasia is a major cause of intractable epilepsy in children. However, the precise mechanisms linking cortical malformations to epileptogenesis remain elusive. The neuronal-specific activator of cyclin-dependent kinase 5, p35, has been recognized as a key factor in proper neuronal migration in the neocortex. Deletion of p35 leads to severe neocortical lamination defects associated with sporadic lethality and seizures. Here we demonstrate that p35-deficient mice also exhibit dysplasia/ heterotopia of principal neurons in the hippocampal formation, as well as spontaneous behavioral and electrographic seizures. Morphological analyses using immunocytochemistry, electron microscopy, and intracellular labeling reveal a high degree of abnormality in dentate granule cells, including heterotopic localization of granule cells in the molecular layer and hilus, aberrant dendritic orientation, occurrence of basal dendrites, and abnormal axon origination sites. Dentate granule cells of p35-deficient mice also demonstrate aberrant mossy fiber sprouting. Field potential laminar analysis through the dentate molecular layer reflects the dispersion of granule cells and the structural reorganization of this region. Similar patterns of cortical disorganization have been linked to epileptogenesis in animal models of chronic seizures and in human temporal lobe epilepsy. The p35-deficient mouse may therefore offer an experimental system in which we can dissect out the key morphological features that are causally related to epileptogenesis.

Expression of type I adenylyl cyclase in intrinsic pathways of the hippocampal formation of the macaque (Macaca nemestrina).

The mossy fiber pathway of the hippocampal formation and type 1 adenylyl cyclase (AC1) have been implicated in long-term potentiation and memory function. Using immunohistochemical labeling and light microscopy we demonstrated intense labeling of AC1 in the mossy fibers and less intense labeling in the molecular layers of both the dentate gyrus and fields CA1, CA2 and CA3 of the hippocampus, i.e. in terminal fields of the perforant pathway. These findings indicate that, in the non-human primate, AC1 is found in the mossy fibers and in terminal fields of the perforant pathway where it may play a role in long term potentiation similar to that demonstrated in the rodent.

Regulation of GFRalpha-1 and GFRalpha-2 mRNAs in rat brain by electroconvulsive seizure.

The influence of both acute and chronic electroconvulsive seizure (ECS) or antidepressant drug treatments on expression of mRNAs encoding glial cell line-derived neurotrophic factor (GDNF) and its receptors, GFRalpha-1, GFRalpha-2, and c-Ret proto-oncogene (RET) in the rat hippocampus was examined by in situ hybridization. Two hours after acute ECS, levels of GFRalpha-1 mRNA in the dentate gyrus were significantly increased. This increase peaked to nearly 3-fold at 6 h after acute ECS and returned to basal levels 24 h after treatment. Chronic (once daily for 10 days) ECS significantly increased the expression of GFRalpha-1 mRNA nearly 5-fold after the last treatment. Levels of GFRalpha-2 mRNA in the dentate gyrus were also significantly increased by acute and chronic ECS, although this effect was less than that observed for GFRalpha-1. Maximum induction of GFRalpha-2 was 30% and 70% compared to sham in response to acute or chronic ECS, respectively. Levels of GDNF and RET mRNAs were not significantly changed following either acute or chronic ECS treatment at the time points examined. Chronic (14 days) administration of different classes of antidepressant drugs, including tranylcypromine, desipramine, or fluoxetine, did not significantly affect the GDNF, GFRalpha-1, GFRalpha-2, or RET mRNA levels in CA1, CA3, and dentate gyrus areas of hippocampus. The results demonstrate that acute ECS increases the expression of GFRalpha-1 and GFRalpha-2 and that these effects are enhanced by chronic ECS. The results also imply that regulation of the binding components of GDNF receptor complex may mediate the adaptive responses of the GDNF system to acute and chronic stimulation.