Joachim Behr

Adult-Generated Hippocampal Neurons Allow the Flexible Use of Spatially Precise Learning Strategies

Despite enormous progress in the past few years the specific contribution of newly born granule cells to the function of the adult hippocampus is still not clear. We hypothesized that in order to solve this question particular attention has to be paid to the specific design, the analysis, and the interpretation of the learning test to be used. We thus designed a behavioral experiment along hypotheses derived from a computational model predicting that new neurons might be particularly relevant for learning conditions, in which novel aspects arise in familiar situations, thus putting high demands on the qualitative aspects of (re-)learning. In the reference memory version of the water maze task suppression of adult neurogenesis with temozolomide (TMZ) caused a highly specific learning deficit. Mice were tested in the hidden platform version of the Morris water maze (6 trials per day for 5 days with a reversal of the platform location on day 4). Testing was done at 4 weeks after the end of four cycles of treatment to minimize the number of potentially recruitable new neurons at the time of testing. The reduction of neurogenesis did not alter longterm potentiation in CA3 and the dentate gyrus but abolished the part of dentate gyrus LTP that is attributed to the new neurons. TMZ did not have any overt side effects at the time of testing, and both treated mice and controls learned to find the hidden platform. Qualitative analysis of search strategies, however, revealed that treated mice did not advance to spatially precise search strategies, in particular when learning a changed goal position (reversal). New neurons in the dentate gyrus thus seem to be necessary for adding flexibility to some hippocampus-dependent qualitative parameters of learning. Our finding that a lack of adult-generated granule cells specifically results in the animals inability to precisely locate a hidden goal is also in accordance with a specialized role of the dentate gyrus in generating a metric rather than just a configurational map of the environment. The discovery of highly specific behavioral deficits as consequence of a suppression of adult hippocampal neurogenesis thus allows to link cellular hippocampal plasticity to well-defined hypotheses from theoretical models.

Cellular and network properties of the subiculum in the pilocarpine model of temporal lobe epilepsy

The subiculum was recently shown to be crucially involved in the generation of interictal activity in human temporal lobe epilepsy. Using the pilocarpine model of epilepsy, this study examines the anatomical substrates for network hyperexcitability recorded in the subiculum. Regular‐ and burst‐spiking subicular pyramidal cells were stained with fluorescence dyes and reconstructed to analyze seizure‐induced alterations of the dendritic and axonal system. In control animals burst‐spiking cells outnumbered regular‐spiking cells by about two to one. Regular‐ and burst‐spiking cells were characterized by extensive axonal branching and autapse‐like contacts, suggesting a high intrinsic connectivity. In addition, subicular axons projecting to CA1 indicate a CA1‐subiculum‐CA1 circuit. In the subiculum of pilocarpine‐treated rats we found an enhanced network excitability characterized by spontaneous rhythmic activity, polysynaptic responses, and all‐or‐none evoked bursts of action potentials. In pilocarpine‐treated rats the subiculum showed cell loss of about 30%. The ratio of regular‐ and burst‐spiking cells was practically inverse as compared to control preparations. A reduced arborization and spine density in the proximal part of the apical dendrites suggests a partial deafferentiation from CA1. In pilocarpine‐treated rats no increased axonal outgrowth of pyramidal cells was observed. Hence, axonal sprouting of subicular pyramidal cells is not mandatory for the development of the pathological events. We suggest that pilocarpine‐induced seizures cause an unmasking or strengthening of synaptic contacts within the recurrent subicular network. J. Comp. Neurol. 483:476–488, 2005.

Entorhinal cortex entrains epileptiform activity in CA1 in pilocarpine-treated rats

Layer III neurons of the medial entorhinal cortex (mEC) project to CA1 via the temporoammonic pathway and exert a powerful feed-forward inhibition of CA1 pyramidal neurons. The present study evaluates the hypothesis that disrupted inhibition of CA1 pyramidal neurons causes an eased propagation of entorhinal seizures to the hippocampus via the temporoammonic pathway. Using a method to induce a confined epileptic focus in brain slices, we investigated the spread of epileptiform activity from the disinhibited mEC to CA1 in control and pilocarpine-treated rats that had displayed status epilepticus and spontaneous recurrent seizures. In pilocarpine-treated rats, the mEC showed a moderate layer III cell loss and an enhanced susceptibility to epileptiform discharges compared to control animals. Entorhinal discharges propagated to CA1 in pilocarpine-treated rats but not in controls. Disconnecting CA3 from CA1 did not affect the spread of epileptiform activity to CA1 excluding its propagation via the trisynaptic hippocampal loop. Mimicking the invasion of epileptiform discharges by repetitive stimulation of the temporoammonic pathway caused a facilitation of field potentials in CA1 that were contaminated by population spikes and afterdischarges in pilocarpine-treated but not control rats. Single cell recordings of CA1 pyramidal neurons revealed a dramatic loss of feed-forward inhibition and the occurrence of strong postsynaptic excitatory potentials in pilocarpine-treated rats. Excitatory responses in CA1 were characterized by multiple NMDA receptor-mediated afterdischarges and a strong paired-pulse facilitation in response to activation of the temporoammonic pathway. Our results suggest that, irrespective of the enhanced seizure-susceptibility of the mEC in epileptic rats, the loss of feed-forward inhibition and the enhanced NMDA receptor-mediated excitability CA1 pyramidal cells ease the spread of epileptiform activity from the mEC to CA1 via the temporoammonic pathway bypassing the classical trisynaptic hippocampal loop.

Loss of GABAergic neurons in the subiculum and its functional implications in temporal lobe epilepsy

Clinical and experimental evidence suggest that the subiculum plays an important role in the maintenance of temporal lobe seizures. Using the pilocarpine-model of temporal lobe epilepsy (TLE), the present study examines the vulnerability of GABAergic subicular interneurons to recurrent seizures and determines its functional implications. In the subiculum of pilocarpine-treated animals, the density of glutamic acid decarboxylase (GAD) mRNA-positive cells was reduced in all layers. Our data indicate a substantial loss of parvalbumin-immunoreactive neurons in the pyramidal cell and molecular layer whereas calretinin-immunoreactive cells were predominantly reduced in the molecular layer. Though the subiculum of pilocarpine-treated rats showed an increased intensity of GAD65 immunoreactivity, the density of GAD65 containing synaptic terminals in the pyramidal cell layer was decreased indicating an increase in the GAD65 intensity of surviving synaptic terminals. We observed a decrease in evoked inhibitory post-synaptic currents that mediate dendritic inhibition as well as a decline in the frequency of miniature inhibitory post-synaptic currents (mIPSCs) that are restricted to the perisomatic region. The decrease in mIPSC frequency (-30%) matched with the reduced number of perisomatic GAD-positive terminals (-28%) suggesting a decrease of pre-synaptic GABAergic input onto pyramidal cells in epileptic animals. Though cell loss in the subiculum has not been considered as a pathogenic factor in human and experimental TLE, our data suggest that the vulnerability of subicular GABAergic interneurons causes an input-specific disturbance of the subicular inhibitory system.

Subthreshold membrane potential oscillations in neurons of deep layers of the entorhinal cortex

Neuronal oscillations are important for information processing. The entorhinal cortex is one of the structures which is involved in generation of theta rhythm. The major role of the entorhinal cortex is to feed diverse sources of information both to and from the hippocampus. Far from simply being a funnel for this information it becomes clear that the entorhinal cortex has its own active properties that contribute to signal processing. Interestingly, stellate cells in layer II of the entorhinal cortex can intrinsically generate subthreshold, Na+-dependent membrane potential oscillations. Here, using intracellular and patch-clamp recordings, we report a similar phenomenon from neurons of the deep layers of the entorhinal cortex. In our in vitro slice preparation about two-thirds of recorded neurons were able to generate voltage-sensitive subthreshold membrane potential oscillations. At a membrane potential of about 50 mV the mean frequency of the voltage-oscillations was 8.1 Hz, whereby at slightly more positive potentials (-44 mV) the frequency of the membrane potential oscillations was 20 Hz and the oscillations became interrupted by clusters of non-adapting trains of spikes. Pharmacological experiments revealed that the oscillations were not affected by Cs+, but could be blocked by the fast Na+-channel blocker tetrodotoxin. We therefore conclude that voltage- and Na+-dependent subthreshold membrane potential oscillations are not only present in stellate cells of entorhinal cortex-layer II, but are also typical for neurons of the deep layers of the entorhinal cortex.

Spread of low Mg2+ induced epileptiform activity from the rat entorhinal cortex to the hippocampus after kindling studied in vitro

Extracellular recordings were performed in in vitro combined hippocampal-entorhinal cortex (HC-EC) slices obtained from control and amygdala kindled rats to investigate the spread of epileptiform activity from the entorhinal cortex (EC) to the hippocampus (HC). Epileptiform activity was induced by lowering extracellular Mg2+ concentration. In control slices epileptiform activity was in most slices characterized by intericatal discharges and short recurrent discharges in areas CA1 and CA3 and by early seizure like events and late recurrent discharges in the EC and the subiculum. In spite of well preserved anatomical pathways in the combined HC-EC slice in which most of the fibre connectivity between the EC and the dentate gyrus (DG) is intact, seizure like events and late recurrent discharges generated in the EC had only moderate effects on the epileptiform activity in areas CA3 and CA1. In contrast in HC-EC slices obtained from kindled rats epileptiform activity generated in the EC spread to the DG and the areas CA3 and CA1. Kindling facilitates the propagation of seizure like events and late recurrent discharges through the HC-EC slice and appears to alter the filtering function of the DG.

The subiculum: a potential site of ictogenesis in human temporal lobe epilepsy.

Summary:  Purpose: This study determines synaptic and intrinsic alterations of subicular pyramidal cells that are associated with activity recorded in patients suffering from temporal lobe epilepsy.

Low Mg2+ induced epileptiform activity in the subiculum before and after disconnection from rat hippocampal and entorhinal cortex slices☆

The subiculum is an area within the hippocampal complex which participates strongly in ictaform activity generated in the entorhinal cortex (EC). To study the properties of epileptiform activity with intra- and extracellular recording techniques in the subiculum, combined slices containing the EC, subiculum and hippocampus were prepared with and without surgical disconnection of the subiculum from the EC and area CA1. For induction of epileptiform activity extracellular magnesium was lowered. After acute disconnection of the subiculum from the cornu ammonis and the EC, seizure like events similar to those in the more intact preparation did develop. These were characterized by slow negative field potential shifts and, in intracellular recordings by sustained depolarization shifts lasting for 10-43 s. This activity could develop into late recurrent discharges of 1-2 s. These data indicate that the subiculum may be an important zone for epileptogenesis in temporal lobe epilepsy.

Two different forms of long-term potentiation at CA1–subiculum synapses

Distinct functional roles in learning and memory are attributed to certain areas of the hippocampus and the parahippocampal region. The subiculum as a part of the hippocampal formation is the principal target of CA1 pyramidal cell axons and serves as an interface in the information processing between the hippocampus and the neocortex. Subicular pyramidal cells have been classified as bursting and regular firing cells. Here we report fundamental differences in long‐term potentiation (LTP) between both cell types. Prolonged high‐frequency stimulation induced NMDA receptor‐dependent LTP in both cell types. While LTP relied on postsynaptic calcium in regular firing neurons, no increase in postsynaptic calcium was required in bursting cells. Furthermore, paired‐pulse facilitation revealed that the site of LTP expression was postsynaptic in regular firing neurons, while presynaptic in burst firing neurons. Our findings on synaptic plasticity in the subiculum indicate that regular firing and bursting cells represent two functional units with distinct physiological roles in processing hippocampal output.

Synaptic plasticity in the subiculum.

The subiculum is the principal target of CA1 pyramidal cells. It functions as a mediator of hippocampal-cortical interaction and has been proposed to play an important role in the encoding and retrieval of long-term memory. The cellular mechanisms of memory formation are thought to include long-term potentiation (LTP) and depression (LTD) of synaptic strength. This review summarizes the contemporary knowledge of LTP and LTD at CA1-subiculum synapses. The observation that the underlying mechanisms of LTP and LTD at CA1-subiculum synapses correlate with the discharge properties of subicular pyramidal cell reveals a novel and intriguing mechanism of cell-specific consolidation of hippocampal output.