Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
Comparative Study
. 2005 Sep 21;25(38):8755-65.
doi: 10.1523/JNEUROSCI.1165-05.2005.

Presubiculum Stimulation in Vivo Evokes Distinct Oscillations in Superficial and Deep Entorhinal Cortex Layers in Chronic Epileptic Rats

Affiliations
Free PMC article
Comparative Study

Presubiculum Stimulation in Vivo Evokes Distinct Oscillations in Superficial and Deep Entorhinal Cortex Layers in Chronic Epileptic Rats

Else A Tolner et al. J Neurosci. .
Free PMC article

Abstract

The characteristic cell loss in layer III of the medial entorhinal area (MEA-III) in human mesial temporal lobe epilepsy is reproduced in the rat kainate model of the disease. To understand how this cell loss affects the functional properties of the MEA, we investigated whether projections from the presubiculum (prS), providing a main input to the MEA-III, are altered in this epileptic rat model. Injections of an anterograde tracer in the prS revealed bilateral projection fibers mainly to the MEA-III in both control and chronic epileptic rats. We further examined the prS-MEA circuitry using a 16-channel electrode probe covering the MEA in anesthetized control and chronic epileptic rats. With a second 16-channel probe, we recorded signals in the hippocampus. Current source density analysis indicated that, after prS double-pulse stimulation, afterdischarges in the form of oscillations (20-45 Hz) occurred that were confined to the superficial layers of the MEA in all epileptic rats displaying MEA-III neuronal loss. Slower oscillations (theta range) were occasionally observed in the deep MEA layers and the dentate gyrus. This kind of oscillation was never observed in control rats. We conclude that dynamical changes occur in an extensive network within the temporal lobe in epileptic rats, manifested as different kinds of oscillations, the characteristics of which depend on local properties of particular subareas. These findings emphasize the significance of the entorhinal cortex in temporal lobe epilepsy and suggest that the superficial cell layers could play an important role in distributing oscillatory activity.

Figures

Figure 1.
Figure 1.
Projection from the prS to the degenerated MEA in chronic epileptic rats. The inset on the top left shows the injection spot of the anterograde tracer BDA in the superficial layers of the prS (horizontal section counterstained by Nissl staining). A, B, Confocal images of horizontal brain sections showing projecting fibers from the prS (red) in the MEA of a control rat (A) and a chronic epileptic rat (B) (6 months after KA-induced SE). Counterstaining with the neuronal marker NeuN (green) reveals the massive neuronal loss (and tissue shrinkage) in the MEA-III that is observed in most chronic epileptic rats. Presubicular fibers predominantly target the MEA-III in both the control and epileptic rats, despite the neurodegeneration in layer III in the epileptic rats. C, Detail of the target area in the MEA-III from the chronic epileptic rat in B. Counterstaining with DAPI (blue) reveals the presence of non-neuronal cells (possibly glia) in the neurodegenerated MEA-III that may be targeted by fibers from the prS. D–F, Same projections as shown in A–C, counterstained for the presence of PV (green) instead of NeuN. No difference was found in the amount of PV-positive neurons in the superficial layers of the MEA in chronic epileptic rats in comparison with controls (for details, see Results). PV-positive neurons are present in the target area from the prS in the MEA-III and -II in both control rats (D) and chronic epileptic rats (E, F) (F, counterstaining with DAPI in blue). G–I, Neuronal loss is extensive in layer III of the MEA in chronic KA rats also in the dorsal part of the MEA. G, NeuN-stained sagittal brain section showing the MEA around the level of recording in the dorsal part of the MEA from a chronic KA rat (Fig. 2 B). H, I, Details of the MEA layers (rectangle in G) of a control rat (H) and a KA rat (I), revealing extensive loss of neurons in layer III for the chronic KA rat. Scale bars: A, B, D, E, 100 μm; C, F, 40 μm; (in I) H, I, 150 μm.
Figure 2.
Figure 2.
prS-evoked responses in the MEA of control rats. A, Nissl-stained sagittal section of the parahippocampal formation illustrating the stimulation site in the prS. Dotted lines indicate the borders of the prS. B, Nissl-stained sagittal section illustrating the recording track of a 16-channel silicon probe in the dorsal part of the MEA. Lesions were made at the 1st and 16th recording sites (arrowheads). Entorhinal layers are indicated with Roman numerals, with a separation between the deep layers (V and VI) and the superficial layers (I–III). The extent of the white matter and the borders of layer II are drawn as lines, and the lamina dissecans is drawn as a dashed line. C, Typical example of laminar profiles of field potential responses (average of 4 sweeps) in the MEA evoked by prS stimulation (indicated by an asterisk) in control rats (n = 8). The response to the test stimulus in the double-pulse protocol is shown. D, CSD laminar profile of the field responses shown in A. Relevant sinks are shaded or striped, and the early negative components n2 and n3 in the superficial layers are indicated. The main effect of prS stimulation is seen in the occurrence of a sharp current sink (n2) that is of small amplitude in the MEA-III (indicated by an asterisk) and is most prominent in the MEA-II. Then 2 sink is followed by a broad current sink (n3 wave; striped) at the border ofMEA-I and -II. See Results for details. The scheme of the layers of the entorhinal cortex on the left indicates the location of the recording sites, as reconstructed from histology. CA, Cornu ammonis; Ctrl, control; l.d., lamina dissecans; Sub, subiculum; sup, superficial; wm or w.m., white matter; 3pCSD, three-point CSD.
Figure 3.
Figure 3.
prS-evoked responses in the MEA of chronic epileptic rats. A, Typical example of the laminar profiles of field potential responses (average of 4 sweeps) in the MEA evoked by PrS stimulation (indicated by an asterisk) in a chronic epileptic rat (3 months after SE induction with KA, 30% remaining neurons in the MEA-III). B, CSD laminar profile of the field responses shown in A. C, Example of CSD laminar profile in a chronic epileptic rat (3 months after SE induction, 40% remaining neurons in the MEA-III) displaying an n2 component of small amplitude. Relevant components are depicted as in Figure 2. See Results for details. D, Comparison between the n3 current sink (wave) at the edge of the MEA-I and -II in controls and chronic epileptic rats. The ratio between the n3 and n2 component was significantly larger for chronic epileptic rats (n = 6) in comparison with controls (n = 5; p = 0.026; Mann–Whitney U test). The asterisk indicates early sink that is evoked in MEA-III after prS stimulation. Error bars indicate SE. ctrl, Control; l.d., lamina dissecans; w.m., white matter.
Figure 4.
Figure 4.
prS stimulation evokes oscillations in the MEA of chronic KA rats. Typical examples of laminar profiles of field potential responses (nonaveraged) in the MEA evoked by prS stimulation (indicated by an asterisk) in a control rat (A) and a chronic KA rat (B) (4 months after KA-induced SE, 40% remaining neurons in the MEA-III). The indication of the MEA layers is as in Figures 2 and 3. Ctrl, Control; l.d., lamina dissecans; w.m., white matter.
Figure 5.
Figure 5.
CSD laminar profiles of the field responses shown in Figure 4. Both in control rats (A) and in KA rats (B), prS stimulation evoked short-latency responses in layers I–III of the MEA. C1 and C2 showde tails (smaller rectangles in A and B) of this short-latency response after the test stimulus in the double-pulse protocol. After double-pulse stimulation of the prS (indicated by asterisks) in all KA rats that displayed significant neuronal loss in the MEA-III (n = 5 of 6), oscillations in the beta/gamma frequency range were evident in the superficial layers of the MEA. Such oscillations were never observed in controls or in the KA rat that did not show neuronal loss in the MEA-III. C3 displays a detail of the superficial oscillations shown in B, of which the sink-source configuration resembles that of the direct response to prS stimulation (compare with C1 and C2). D1 and D2 show details of the direct response to prS stimulation (dashed rectangle in A and B) with the presence of a long-latency response. The long-latency response consisted of a sharp current sink in the deep MEA followed by a broad wave in the more superficial layers. The relevant current sinks are shaded gray. Ctrl, Control; l.d., lamina dissecans; w.m., white matter. Asterisks in A, B, C1, C2, D1, and D2 represent time of prS stimulation.
Figure 6.
Figure 6.
Wavelet analysis of the oscillations in channel 3 in the CSD laminar profile of the KA rat from Figures 4 B and 5B, revealing the time–frequency characteristics of the superficial oscillations. Shown is the time–frequency z-score Gabor amplitude of the single-channel CSD during paired-pulse stimulation in the prS. The pulses were delivered at 100 and 200 ms relative to the beginning of the plot (horizontal axis) in one epileptic and five control animals. The current source densities of the second “superficial” channel were convoluted with a set of Gabor filters of logarithmically increasing frequencies (vertical axis) ranging from 4 to 150 Hz. The z-score, represented in pseudo-color coding, was computed as described in Materials and Methods using five control traces. The averaging, the SD, and the subtraction were performed independently for each time–frequency point. In this example, after prS double-pulse stimulation (indicated by the 2 asterisks at a 100 ms interval) in the KA rat, superficial-layer oscillations are evoked that are more or less continuous for ∼300 ms in the range of 20–50 Hz. freq, Frequency; sup, superficial.
Figure 7.
Figure 7.
Oscillations occur occasionally in deep layers of the MEA after double-pulse stimulation of the prS in chronic epileptic rats. A, A typical example of CSD laminar profiles of field potential responses (nonaveraged) in the MEA evoked by prS stimulation (indicated by an asterisk) in a chronic KA rat (3.5 months after SE induction with KA, 50% remaining neurons in the MEA-III). The indication of entorhinal cortex layers is as in previous figures. In addition to a pattern of beta/gamma oscillations in the superficial MEA layers, in the deep MEA some events were observed that were in the theta range (∼8 Hz). These deep-layer events were synchronous with events recorded in the DG and subiculum. B, C, Overlays of the events triggered at B, the start of the current sink observed in the MEA-V (arrow in channel 8), and at C, the peak of the wave in the MEA-II (arrow in the superficial MEA trace), did not reveal a relationship between the superficial oscillations and the slower events in the deep MEA/DG. The events observed in the deep MEA, however, are in phase with events in the DG and subiculum. The short time delay between the occurrence of these slower events is compatible with a monosynaptic relay between DG→subiculum→ MEA-V. For B, an overlay was made of the four events that were observed in the deep MEA/DG (see bottom boxed area in A). For C, an overlay was made of 10 superficial events observed in channel 3 (see top boxed area in A); the deep MEA taken at channel 8. D, Histology of the recording locations in the hippocampus. The Nissl-stained sagittal section illustrates the recording track of a 16-channel silicon probe in the dorsal part of the hippocampus. Lesions were made at the 1st (arrowhead) and 16th recording sites. CA, Cornu ammonis; gc, granule cell layer; ml, molecular layer; fis, fissure; l.d., lamina dissecans; Sub, subiculum; Sup, superficial; w.m., white matter.
Figure 8.
Figure 8.
Scheme that summarizes the possible connections with and within the superficial layers of the MEA of chronic epileptic rats with regard to input from the prS. Cells are depicted as principal neuron (PRIN) or interneuron (INTER) (PV, parvalbumin-positive). ▸, Excitatory synaptic connections; ▹, inhibitory synaptic connections. After chronic epilepsy in the KA model, neuronal loss affects mainly principal neurons in the MEA-III (gray shading indicating loss). Targets of the prS in the MEA in KA rats are presumed to be as follows. 1, Interneurons in the superficial MEA [targeted by excitatory and to a lesser extent by inhibitory presubicular fibers (van Haeften et al., 1997)]. 2, Dendrites of MEA-II neurons in the MEA-III and to a lesser extent in the MEA-I (Caballero-Bleda and Witter, 1994). 3, Dendrites of MEA-V neurons in the MEA-III (Wouterlood et al., 2004). 4, Remaining principal neurons in the MEA-III (Caballero-Bleda and Witter, 1994). 5, Connections between MEA-III and MEA-II neurons may be involved in the direct response to prS stimulation in the superficial MEA. 6, With loss of mainly principal neurons, PV-positive interneurons that control inhibition in layers II and III receive less activation after prS stimulation and may even be inhibited. 7, 8, This would result in decreased inhibition (7) and increased excitability among remaining principal neurons (8) (Eid et al., 1999; van Vliet et al., 2004; the present study). 9, Connections between deep and superficial neurons (Kloosterman et al., 2003b; van Haeften et al., 2003) might add to the hyperexcitability of the superficial layers. This basic scheme is adapted from Witter and Amaral (2004).

Similar articles

See all similar articles

Cited by 11 articles

See all "Cited by" articles

Publication types

Feedback