Neuropeptide Y regulates recurrent mossy fiber synaptic transmission less effectively in mice than in rats: Correlation with Y2 receptor plasticity

A unique feature of temporal lobe epilepsy is the formation of recurrent excitatory connections among granule cells of the dentate gyrus as a result of mossy fiber sprouting. This novel circuit contributes to a reduced threshold for granule cell synchronization. In the rat, activity of the recurrent mossy fiber pathway is restrained by the neoexpression and spontaneous release of neuropeptide Y (NPY). NPY inhibits glutamate release tonically through activation of presynaptic Y2 receptors. In the present study, the effects of endogenous and applied NPY were investigated in C57Bl/6 mice that had experienced pilocarpine-induced status epilepticus and subsequently developed a robust recurrent mossy fiber pathway. Whole cell patch clamp recordings made from dentate granule cells in hippocampal slices demonstrated that, as in rats, applied NPY inhibits recurrent mossy fiber synaptic transmission, the Y2 receptor antagonist (S)-N2-[[1-[2-[4-[(R,S)-5,11-dihydro-6(6H)-oxodibenz[b,e]azepin-11-yl]-1-piperazinyl]-2-oxoethyl]cyclopentyl]acetyl]-N-[2-[1,2-dihydro-3,5(4H)-dioxo-1,2-diphenyl-3H-1,2,4-triazol-4-yl]ethyl]-argininamide (BIIE0246) blocks its action and BIIE0246 enhances synaptic transmission when applied by itself. Y5 receptor agonists had no significant effect. Thus spontaneous release of NPY tonically inhibits synaptic transmission in mice and its effects are mediated by Y2 receptor activation. However, both NPY and BIIE0246 were much less effective in mice than in rats, despite apparently equivalent expression of NPY in the recurrent mossy fibers. Immunohistochemistry indicated greater expression of Y2 receptors in the mossy fiber pathway of normal mice than of normal rats. Pilocarpine-induced status epilepticus markedly reduced the immunoreactivity of mouse mossy fibers, but increased the immunoreactivity of rat mossy fibers. Mossy fiber growth into the inner portion of the dentate molecular layer was associated with increased Y2 receptor immunoreactivity in rat, but not in mouse. These contrasting receptor changes can explain the quantitatively different effects of endogenously released and applied NPY on recurrent mossy fiber transmission in mice and rats.     

Neuropeptide Y biosynthesis is markedly induced in mossy fibers during temporal lobe epilepsy of the rat

Neuropeptide Y (NPY) immunoreactivity and gene expression was investigated in the hippocampus after kainic acid-induced seizures and pentylenetetrazol kindling in the rat. Pronounced increases of NPY immunoreactivity were found in the terminal field of mossy fibers in both animal models. In kainic acid-treated rats the peptide progressively accumulated in the hilus and the stratum lucidum of CA3, 5-60 days after injection of the toxin and, at the later intervals, extended to the supragranular molecular layer of the dentate gyrus indicating sprouting of these neurons. Unilateral injection of colchicine into the hilus abolished NPY staining of the mossy fibers. Using in situ hybridization, in both animal models markedly enhanced expression of prepro-NPY mRNA was observed in the granular layer, containing the perikarya of the mossy fibers. It is suggested that sustained expression of the neuromodulatory neuropeptide NPY, in addition to the observed plastic changes, may contribute to altered excitability of hippocampal mossy fibers in epilepsy. Neither somatostatin immunoreactivity nor gene expression were enhanced in granule cells/mossy fibers.     

Limbic seizures cause pronounced changes in the expression of neurokinin B in the hippocampus of the rat.

Immunohistological and in situ hybridization techniques were used to study the influence of kainic acid-induced seizures and of pentylenetetrazol kindling on neurokinin B immunoreactivity and neurokinin B mRNA in the rat hippocampus. Pronounced increases in neurokinin B immunoreactivity were observed in the terminal field of mossy fibres 10-60 days after intraperitoneal injection of kainic acid. These slow but persistent increases in immunoreactivity were accompanied by markedly enhanced expression of neurokinin B mRNA in the granule cells and in hilar interneurons adjacent to the granule cell layer. These changes were preceded by transient increases in neurokinin B mRNA and immunoreactivity in CA1 pyramidal cell layer two and 10 days after kainic acid, which, however, subsided later on. Pentylenetetrazol kindling caused similar increases in neurokinin B mRNA expression in granule cells and in CA1 pyramidal cells, but not in hilar interneurons. In CA1, increased neurokinin B message was present two days after termination of the kindling procedure but not after 10 days. Sixty days after kainic acid injection, neurokinin B immunoreactivity extended to the inner-third of the molecular layer of the dentate gyrus. After pentylenetetrazol kindling, a neurokinin B-immunoreactive band was observed in the infrapyramidal region of CA3. Lesions of the dentate granule cells by local injection of colchicine in kainic acid-treated rats abolished the supragranular neurokinin B-positive staining, whereas it was almost unchanged after transection of the ventral hippocampal commissure. These observations suggest that neurokinin B immunoreactivity may be located in ipsilateral mossy fibres undergoing collateral sprouting to the inner molecular layer or to the infrapyramidal region in CA3, respectively. Preprotachykinin A mRNA, which encodes for neurokinin A and substance P, and substance P immunoreactivity were not changed in the hippocampus of epileptic rats compared with untreated animals. The observed changes in neurokinin B immunoreactivity and mRNA indicate that specific functional and morphological changes may be induced in hippocampal neurons by recurrent limbic seizures.     

The C5a anaphylatoxin receptor CD88 is expressed in presynaptic terminals of hippocampal mossy fibres

Background  

In the periphery, C5a acts through the G-protein coupled receptor CD88 to enhance/maintain inflammatory responses. In the brain, CD88 can be expressed on astrocytes, microglia and neurons. Previous studies have shown that the hippocampal CA3 region displays CD88-immunolabelling, and CD88 mRNA is present within dentate gyrus granule cells. As granule cells send dense axonal projections (mossy fibres) to CA3 pyramidal neurons, CD88 expression could be expressed on mossy fibres. However, the cellular location of CD88 within the hippocampal CA3 region is unknown.     

The persistent expression of a highly polysialylated NCAM in the dentate gyrus of the adult rat.

The expression of a highly polysialylated form of the neural cell adhesion molecule (NCAM-H), often termed 'embryonic NCAM', has been investigated in the hippocampal formation of developing and adult rats. To determine the immunohistochemical localization of NCAM-H, a monoclonal antibody that reacts with the polysialic acid portion of NCAM-H was used. In the late embryonic and early postnatal periods, immunoreactivity for NCAM-H was found throughout the hippocampal formation, except for the ventricular layer. Thereafter, the immunoreactivity gradually decreased and almost vanished in most parts in the adult. However, a strong immunoreactivity remained on a number of cells in the dentate gyrus of adult rats, particularly in the deepest part of the granular layer. The immunoreactive arborized dendrites, mostly arising from the primary apical pole of the granule cells, were found to enter the molecular layer. The mossy fibers also were positive. Electron-microscopic examination of the hilus portion showed that the immunoreactivity was detected on the plasma membrane of some axons in the mossy fiber bundles. Since postnatal neurogenesis is known to continue into adulthood in the deepest part of the granule cell layer of the dentate gyrus, these results suggest that, in the adult dentate gyrus, NCAM-H is expressed by newly generated granule cells, and that the NCAM-H-expressing new cells may participate in the formation of new neural circuits.     

Oral choline alfoscerate counteracts age-dependent loss of mossy fibres in the rat hippocampus.

Mossy fibres represent a major intrahippocampal associative pathway. They consist of axons of granule cells of the dentate gyrus and show an age-dependent loss as do the granule cells of the dentate gyrus. The present study was designed to assess whether long-term treatment of rats with choline alfoscerate in their drinking water would be effective in countering the loss of mossy fibres and of granule cells occurring with aging. Choline alfoscerate is a precursor in the biosynthesis of brain phospholipids and increases the bioavailability of choline in nervous tissue. Male Sprague-Dawley rats of 18 months of age were divided into two groups. One group received a daily dose of 100 mg/kg choline alfoscerate for 6 months; the other group was used as an untreated control. Twelve-month-old untreated animals were used as a reference group. The area occupied by mossy fibres, as well as their density, was significantly reduced in 24-month-old control rats in comparison with 12-month-old rats. The same is true for the density granule cells of the dentate gyrus which was decreased by about 20% in the oldest animals. In choline alfoscerate-treated rats both the area occupied by mossy fibres and their density were significantly higher than in age-matched controls. Moreover, the number of granule neurons of the hippocampus was higher by about 7% in choline alfoscerate-treated than in control 24-month-old rats. The above data suggest that choline alfoscerate treatment counteracts some anatomical changes of the rat hippocampus occurring in old age.     

Reassessment of the effects of cycloheximide on mossy fiber sprouting and epileptogenesis in the pilocarpine model of temporal lobe epilepsy

A feature of animal models of temporal lobe epilepsy and the human disorder is hippocampal sclerosis and Timm stain in the inner molecular layer (IML) of the dentate gyrus, which represents synaptic reorganization and may be important in epileptogenesis. We reassessed the hypothesis that pre-treatment with cycloheximide (CHX) prevents Timm staining in the IML following pilocarpine (PILO)-induced status epilepticus (a multifocal model of temporal lobe epilepsy), but allows epileptogenesis (i.e., chronic spontaneous seizures) after a latent period. Hippocampal slices from PILO-treated rats without Timm stain in the IML after CHX treatment were hypothesized to lack the electrophysiological abnormalities suggestive of recurrent excitation. The primary experimental groups were as follows: 1) CHX (1 mg/kg) 30-45 min prior to administration of PILO (320 mg/kg ip, 2) only PILO, and 3) only saline (0.5 ml, IP). The CHX pre-treatment significantly decreased the number of rats that responded to PILO with status epilepticus compared to rats that received only PILO. Pre-treatment with CHX did not significantly alter the spontaneous motor seizure rate post-treatment compared to treatment with PILO alone in those animals from each group that developed status epilepticus during PILO treatment. Timm stain in the IML was not significantly different between the PILO- and PILO+CHX-treated rats. Using quantitative methods, CHX did not prevent hilar, CA1, or CA3 neuronal loss compared to the PILO-treated rats. Extracellular responses to hilar stimulation in 30 microM bicuculline and 6 mM [K(+)](o) demonstrated all-or-none bursting in both the CHX+PILO- and PILO-treated rats but not in control rats. Whole cell recordings from granule cells, using glutamate flash photolysis to activate other granule cells, showed that both the CHX+PILO- and PILO-treated rats had excitatory synaptic interactions in the granule cell layer, which were not found after saline treatment. Some rats responded to PILO (with or without CHX pre-treatment) with only one or a few seizures at treatment, and some of these animals (n = 4) demonstrated spontaneous motor seizures within 2 mo after treatment. Timm staining and neuron loss in this group were not clearly different from saline-treated rats. These results suggest that in the PILO model, pre-treatment with CHX does not affect mossy fiber sprouting in the IML of epileptic rats and does not prevent the formation of recurrent excitatory circuits. However, the develoment of spontaneous motor seizures, in a small number of rats, could occur without detectable hippocampal neuron loss or mossy fiber sprouting, as assessed by the Timm stain method.     

Loss of dynorphin-mediated inhibition of voltage-dependent Ca2+ currents in hippocampal granule cells isolated from epilepsy patients is associated with mossy fiber sprouting

The endogenous kappa receptor selective opioid peptide dynorphin has been shown to inhibit glutamate receptor-mediated neurotransmission and voltage-dependent Ca2+ channels. It is thought that dynorphin can be released from hippocampal dentate granule cells in an activity-dependent manner. Since actions of dynorphin may be important in limiting excitability in human epilepsy, we have investigated its effects on voltage-dependent Ca2+ channels in dentate granule cells isolated from hippocampi removed during epilepsy surgery. One group of patients showed classical Ammon's horn sclerosis characterized by segmental neuronal cell loss and astrogliosis. Prominent dynorphin-immunoreactive axon terminals were present in the inner molecular layer of the dentate gyrus, indicating pronounced recurrent mossy fiber sprouting. A second group displayed lesions in the temporal lobe that did not involve the hippocampus proper. All except one of these specimens showed a normal pattern of dynorphin immunoreactivity confined to dentate granule cell somata and their mossy fiber terminals in the hilus and CA3 region. In patients without mossy fiber sprouting the application of the kappa receptor selective opioid agonist dynorphin A ([D-Arg6]1-13, 1 microM) caused a reversible and dose-dependent depression of voltage-dependent Ca2+ channels in most granule cells. These effects could be antagonized by the non-selective opioid antagonist naloxone (1 microM). In contrast, significantly less dentate granule cells displayed inhibition of Ca2+ channels by dynorphin A in patients with mossy fiber sprouting (Chi-square test, P < 0.0005). The lack of dynorphin A effects in patients showing mossy fiber sprouting compares well to the loss of kappa receptors on granule cells in Ammon's horn sclerosis but not lesion-associated epilepsy. Our data suggest that a protective mechanism exerted by dynorphin release and activation of kappa receptors may be lost in hippocampi with recurrent mossy fiber sprouting.     

移植的齿状回颗粒细胞在培养的大鼠海马组织上移行特征

为探讨齿状回颗粒细胞在培养的海马组织表面上的移行过程以及苔状纤维的投射特征,本实验从生后3d的绿色荧光蛋白(GFP)基因导入的SD大鼠海马组织切片中分离出齿状回颗粒细胞层,移植到培养的SD大鼠海马组织切片表面上,共同培养3～4d后,激光共聚焦显微镜观察。结果显示,97.6%的细胞移行到齿状回和CA3区,仅有2.4%细胞移行到CA2和CA1区。移行到齿状回颗粒细胞层的颗粒细胞投射苔状纤维到齿状回门区和CA3区的透明层,却不到CA2和CA1区。上述结果提示,在海马组织培养中移植的齿状回颗粒细胞的移行以及苔状纤维的投射方向明显受宿主的影响。     

海人酸诱导癫痫大鼠齿状回神经肽Y能苔状纤维侧枝发芽

目的：探讨海马苔状纤维侧枝发芽与癫痫发作敏感性形成的关系。材料与方法：在海人酸诱发大鼠出现癫痫发作后,采用免疫组织化学染色法观察大鼠海马齿状回内神经肽Ｙ能纤维的异常发芽。结果：首次证实在癫痫发作后７天时,在海马齿状回内分子层就已出现神经肽Ｙ能纤维的异常增生。结论：这可能是对癫痫发作后齿状回门区神经肽Ｙ能神经元缺失的一种代偿性变化。     

Expression of synaptopodin, an actin-associated protein, in the rat hippocampus after limbic epilepsy

Synaptopodin, a 100 kD protein, associated with the actin cytoskeleton of the postsynaptic density and dendritic spines, is thought to play a role in modulating actin-based shape and motility of dendritic spines during formation or elimination of synaptic contacts. Temporal lobe epilepsy in humans and in rats shows neuronal damage, aberrant sprouting of hippocampal mossy fibers and subsequent synaptic remodeling processes. Using kainic acid (KA) induced epilepsy in rats, the postictal hippocampal expression of synaptopodin was analyzed by in situ hybridization (ISH) and immunohistochemistry. Sprouting of mossy fibers was visualized by a modified Timm's staining. ISH showed elevated levels of Synaptopodin mRNA in perikarya of CA3 principal neurons, dentate granule cells and in surviving hilar neurons these levels persisted up to 8 weeks after seizure induction. Synaptopodin immunoreactivity in the dendritic layers of CA3, in the hilus and in the inner molecular layer of the dentate gyrus (DG) was initially reduced. Eight weeks after KA treatment Synaptopodin protein expression returned to control levels in dendritic layers of CA3 and in the entire molecular layer of the DG. The recovery of protein expression was accompanied by simultaneous supra- and infragranular mossy fiber sprouting. Postictal upregulation of Synaptopodin mRNA levels in target cell populations of limbic epilepsy-elicited damage and subsequent Synaptopodin protein expression largely co-localized with remodeling processes as demonstrated by mossy fiber sprouting. It may thus represent a novel postsynaptic molecular correlate of hippocampal neuroplasticity.     

Elevated voltage gated Cl- channel expression enhances fast paired-pulse inhibition in the dentate gyrus of seizure sensitive gerbil

The present study was performed to determine whether the voltage gated Cl- channel (CLC) expression is altered in the hippocampus of seizure sensitive (SS) gerbils, and to identify the strong fast paired-pulse inhibition in the dentate gyrus of SS gerbils is associated with altered CLC expression. In the hippocampal proper and the granule cell layer of the dentate gyrus of the SS gerbils, strong CLC-2 immunoreactivity was detected, as compared with seizure resistant (SR) gerbils. In addition, CLC-3 immunoreactivity was observed in the CA1-3 pyramidal cells, and the granule cell and the molecular layer of the dentate gyrus in the SS gerbils, whereas its immunoreactivity was rarely detected in the SR gerbils. However, CLC-3 immunoreactivity in the mossy fiber was reduced, as compared with SR gerbils. Moreover, infusion of the potential CLC inhibitor (4,4'-diisothiocyanostibene-2,2'-disulfanic acid, DIDS) reduced fast paired-pulse inhibition in the dentate gyrus of SS gerbils, although evoked responses in the dentate gyrus between SR and SS gerbils were similarly detected. These findings suggest that enhancement of CLC expression in the dentate gyrus of SS gerbils may be one of the compensatory responses for reduced GABA(A) receptor-mediated fast postsynaptic inhibitory potentials.     

Recurrent mossy fiber pathway in rat dentate gyrus: synaptic currents evoked in presence and absence of seizure-induced growth

A common feature of temporal lobe epilepsy and of animal models of epilepsy is the growth of hippocampal mossy fibers into the dentate molecular layer, where at least some of them innervate granule cells. Because the mossy fibers are axons of granule cells, the recurrent mossy fiber pathway provides monosynaptic excitatory feedback to these neurons that could facilitate seizure discharge. We used the pilocarpine model of temporal lobe epilepsy to study the synaptic responses evoked by activating this pathway. Whole cell patch-clamp recording demonstrated that antidromic stimulation of the mossy fibers evoked an excitatory postsynaptic current (EPSC) in approximately 74% of granule cells from rats that had survived >10 wk after pilocarpine-induced status epilepticus. Recurrent mossy fiber growth was demonstrated with the Timm stain in all instances. In contrast, antidromic stimulation of the mossy fibers evoked an EPSC in only 5% of granule cells studied 4-6 days after status epilepticus, before recurrent mossy fiber growth became detectable. Notably, antidromic mossy fiber stimulation also evoked an EPSC in many granule cells from control rats. Clusters of mossy fiber-like Timm staining normally were present in the inner third of the dentate molecular layer at the level of the hippocampal formation from which slices were prepared, and several considerations suggested that the recorded EPSCs depended mainly on activation of recurrent mossy fibers rather than associational fibers. In both status epilepticus and control groups, the antidromically evoked EPSC was glutamatergic and involved the activation of both AMPA/kainate and N-methyl-D-aspartate (NMDA) receptors. EPSCs recorded in granule cells from rats with recurrent mossy fiber growth differed in three respects from those recorded in control granule cells: they were much more frequently evoked, a number of them were unusually large, and the NMDA component of the response was generally much more prominent. In contrast to the antidromically evoked EPSC, the EPSC evoked by stimulation of the perforant path appeared to be unaffected by a prior episode of status epilepticus. These results support the hypothesis that recurrent mossy fiber growth and synapse formation increases the excitatory drive to dentate granule cells and thus facilitates repetitive synchronous discharge. Activation of NMDA receptors in the recurrent pathway may contribute to seizure propagation under depolarizing conditions. Mossy fiber-granule cell synapses also are present in normal rats, where they may contribute to repetitive granule cell discharge in regions of the dentate gyrus where their numbers are significant.     

Contributions of mossy fiber and CA1 pyramidal cell sprouting to dentate granule cell hyperexcitability in kainic acid-treated hippocampal slice cultures

Axonal sprouting like that of the mossy fibers is commonly associated with temporal lobe epilepsy, but its significance remains uncertain. To investigate the functional consequences of sprouting of mossy fibers and alternative pathways, kainic acid (KA) was used to induce robust mossy fiber sprouting in hippocampal slice cultures. Physiological comparisons documented many similarities in granule cell responses between KA- and vehicle-treated cultures, including: seizures, epileptiform bursts, and spontaneous excitatory postsynaptic currents (sEPSCs) >600 pA. GABAergic control and contribution of glutamatergic synaptic transmission were similar. Analyses of neurobiotin-filled CA1 pyramidal cells revealed robust axonal sprouting in both vehicle- and KA-treated cultures, which was significantly greater in KA-treated cultures. Hilar stimulation evoked an antidromic population spike followed by variable numbers of postsynaptic potentials (PSPs) and population spikes in both vehicle- and KA-treated cultures. Despite robust mossy fiber sprouting, knife cuts separating CA1 from dentate gyrus virtually abolished EPSPs evoked by hilar stimulation in KA-treated but not vehicle-treated cultures, suggesting a pivotal role of functional afferents from CA1 to dentate gyrus in KA-treated cultures. Together, these findings demonstrate striking hyperexcitability of dentate granule cells in long-term hippocampal slice cultures after treatment with either vehicle or KA. The contribution to hilar-evoked hyperexcitability of granule cells by the unexpected axonal projection from CA1 to dentate in KA-treated cultures reinforces the idea that axonal sprouting may contribute to pathologic hyperexcitability of granule cells.     

Enhanced but fragile inhibition in the dentate gyrus in vivo in the kainic acid model of temporal lobe epilepsy: a study using current source density analysis

Temporal lobe epilepsy is related to many structural and physiological changes in the brain. We used kainic acid in rats as an animal model of temporal lobe epilepsy, and studied the neural interactions of the dentate gyrus in urethane-anesthetized rats in vivo. Our initial hypothesis was that sprouting of mossy fibers, the axons of the granule cells, increases proximal dendritic excitatory currents in the inner molecular layer of the dentate gyrus. Extracellular currents were detected in vivo using current source density analysis. Backfiring the mossy fibers in CA3 or orthodromic excitation of the granule cells through the medial perforant path induced a current sink at the inner molecular layer. However, the sink or inferred excitation at the inner molecular layer was not increased in kainic acid-treated rats and the sink actually correlated negatively with the degree of mossy fiber sprouting. It is inferred that the latter sink was mediated mainly by association fibers and not by recurrent mossy fibers. After kainic acid treatment, paired-pulse inhibition of the population spikes in the dentate gyrus was increased. In contrast, reverberant activity that involved looping around an entorhinal-hippocampal circuit was increased in kainic acid-treated rats, compared to control rats. The increase of inhibition in kainic acid-treated rats was readily blocked by a small dose of GABA(A) receptor antagonist bicuculline. The latter dose of bicuculline induced paroxsymal spike bursts in kainic acid-treated but not control rats, demonstrating that the increased inhibition in dentate gyrus was fragile.In conclusion, after kainic acid induced seizures, the dentate gyrus in vivo showed an increase in inhibition that appeared to be fragile. The hypothesized increase in proximal dendritic excitation due to mossy fiber sprouting was not detected. However, the fragile inhibition could explain the seizure susceptibility in patients with temporal lobe epilepsy.     

Enkephalin-like immunoreactivity in the mossy fiber pathway of the hippocampal formation of the tree shrew (Tupaia glis)

The distribution of enkephalin-like immunoreactivity within the hippocampal formation of the tree shrew (Tupaia glis) has been investigated. By far the most conspicuous immunoreactive structures within the hippocampal formation were components of the mossy fiber pathway. Immunoreactive granule cells within the dentate gyrus could often be seen sending dendrites into the molecular layer and axons into the hilus of the dentate gyrus. Within the regio inferior of Ammon's horn, massive immunoreactive terminal swellings associated with thin fibers were located in the zone just above the pyramidal cell layer. While the mossy fiber pathway was the most intensely labeled portion of the hippocampal formation, occasionally sparse terminals and lightly immunoreactive cell bodies were noted in regio superior.These results indicate that the mossy fiber system may be an important pathway for opiate modulation of hippocampal activity.     

Electrophysiological evidence of monosynaptic excitatory transmission between granule cells after seizure-induced mossy fiber sprouting

Mossy fiber sprouting is a form of synaptic reorganization in the dentate gyrus that occurs in human temporal lobe epilepsy and animal models of epilepsy. The axons of dentate gyrus granule cells, called mossy fibers, develop collaterals that grow into an abnormal location, the inner third of the dentate gyrus molecular layer. Electron microscopy has shown that sprouted fibers from synapses on both spines and dendritic shafts in the inner molecular layer, which are likely to represent the dendrites of granule cells and inhibitory neurons. One of the controversies about this phenomenon is whether mossy fiber sprouting contributes to seizures by forming novel recurrent excitatory circuits among granule cells. To date, there is a great deal of indirect evidence that suggests this is the case, but there are also counterarguments. The purpose of this study was to determine whether functional monosynaptic connections exist between granule cells after mossy fiber sprouting. Using simultaneous recordings from granule cells, we obtained direct evidence that granule cells in epileptic rats have monosynaptic excitatory connections with other granule cells. Such connections were not obtained when age-matched, saline control rats were examined. The results suggest that indeed mossy fiber sprouting provides a substrate for monosynaptic recurrent excitation among granule cells in the dentate gyrus. Interestingly, the characteristics of the excitatory connections that were found indicate that the pathway is only weakly excitatory. These characteristics may contribute to the empirical observation that the sprouted dentate gyrus does not normally generate epileptiform discharges.     

Cholecystokinin in the mouse hippocampus: localization in the mossy fiber and dentate commissural systems

Summary The distribution and source of cholecystokinin-like immunoreactivity (CCK-I) in the hippocampus of the Swiss Webster mouse was analyzed using light microscopic immunocytochemical techniques. In agreement with what has been observed in other animals, CCK-I was localized within sparsely scattered neurons throughout the hippocampus proper, in axons that arborize within and around stratum pyramidale, and in fine axons and puncta in stratum lacunosum-moleculare of region CA1. In contrast to other animals, CCK-I was also localized within the mossy fiber system (including dentate gyrus granule cells), within a dense band which occupied the full septo-temporal extent of the dentate gyrus inner molecular layer, and within polymorph neurons of the central hilus. The presence of CCK-I within the latter two areas suggested localization within the dentate gyrus commissural system. This was verified by the combined use of retrograde fluorescent dye transport and CCK immunocytochemistry. Virtually all of the dyelabeled dentate commissural neurons within the hilus were CCK-I. These data demonstrate that while there is little change in the distribution of CCK-I within hippocampal local circuit neurons across animals, there are substantial interspecies differences in the localization of CCK-I within major axonal projections in the hippocampal formation.