Natalie L. M. Cappaert

The anatomy of memory: an interactive overview of the parahippocampal-hippocampal network

Converging evidence suggests that each parahippocampal and hippocampal subregion contributes uniquely to the encoding, consolidation and retrieval of declarative memories, but their precise roles remain elusive. Current functional thinking does not fully incorporate the intricately connected networks that link these subregions, owing to their organizational complexity; however, such detailed anatomical knowledge is of pivotal importance for comprehending the unique functional contribution of each subregion. We have therefore developed an interactive diagram with the aim to display all of the currently known anatomical connections of the rat parahippocampal–hippocampal network. In this Review, we integrate the existing anatomical knowledge into a concise description of this network and discuss the functional implications of some relatively underexposed connections.

Ethyl benzene-induced ototoxicity in rats: a dose-dependent mid-frequency hearing loss.

Rats were exposed to ethyl benzene at 0, 300, 400 and 550 ppm for 8 hours/day for 5 consecutive days. Three to six weeks after the exposure, auditory function was tested by measuring compound action potentials (CAP) in the frequency range of 1-24 kHz and 2f1-f2 distortion product otoacoustic emissions (DPOAEs) in the frequency range of 4-22.6 kHz. In addition, outer hair cell (OHC) loss was quantified by histological examination. The lowest concentration ethyl benzene had no effect on any of the above measures. At 400 ppm, auditory thresholds were increased by 15 and 16 dB at 12 and 16 kHz, respectively, and at 550 ppm by 24, 31, and 22 dB at 8, 12, and 16 kHz, respectively. DPOAE amplitude growth with stimulus level was affected only after 550 ppm at 5.6, 8, and 11.3 kHz. OHC loss was found in two of the five examined locations in the cochlea. At 400 ppm, 25% OHC loss was found at the 11- and 21-kHz region. The highest concentration evoked 40% and 75% OHC loss at the 11- and 21-kHz location, respectively. Thus, the mid-frequency region of rats is affected after exposure to relatively low concentrations of ethyl benzene (400-550 ppm). These results indicate that ethyl benzene is one of the most potent ototoxic organic solvents known today.

The ototoxic effects of ethyl benzene in rats

Exposure to organic solvents has been shown to be ototoxic in animals and there is evidence that these solvents can induce hearing loss in humans. In this study, the effects of inhalation of the possibly ototoxic solvent ethyl benzene on the cochlear function and morphology were evaluated using three complementary techniques: (1) reflex modification audiometry (RMA), (2) electrocochleography and (3) histological examination of the cochleas. Rats were exposed to either ethyl benzene (800 ppm, 8 h/day for 5 days) or to control conditions. The RMA threshold increased significantly by about 25 dB, 1 and 4 weeks after the exposure, irrespective of the stimulus frequency tested (4-24 kHz). Electrocochleography was performed between 8 and 11 weeks after exposure to the organic solvent. The threshold for the compound action potential increased significantly by 10-30 dB at all frequencies tested (1-24 kHz). Histological examination of the cochlea showed outer hair cell (OHC) loss, especially in the upper basal and lower middle turns (corresponding to the mid-frequency region) to an extent of 65%. We conclude that exposure to 800 ppm ethyl benzene for 8 h/day during 5 days induces hearing loss in rats due to OHC loss.

Simultaneous exposure to ethyl benzene and noise : Synergistic effects on outer hair cells

The effects on hearing of simultaneous exposure to the ototoxic organic solvent ethyl benzene and broad-band noise were evaluated in rats. The effects of three ethyl benzene concentrations (0, 300 or 400 ppm) and three noise levels (95 or 105 dB(lin) SPL or background noise at 65 dB(lin) SPL) and all their combinations were investigated for a 5 day exposure at 8 h/day. Distortion product otoacoustic emissions and compound action potentials were affected after 105 dB noise alone, and after 105 dB noise in combination with ethyl benzene (300 and 400 ppm). However, the amount of loss for these combinations did not exceed the loss for 105 dB noise alone. Outer hair cell (OHC) loss after exposure to 300 ppm ethyl benzene was located in the third row of OHCs. At 400 ppm, the loss spread out to the second and first row of OHCs. Noise alone hardly affected the OHC counts except for a minor loss in the first row of OHCs after 105 dB SPL. Noise at 105 dB in combination with ethyl benzene at 300 and 400 ppm, however, showed OHC loss greater than the sum of the losses induced by noise and ethyl benzene alone.

Differential susceptibility of rats and guinea pigs to the ototoxic effects of ethyl benzene

The present study was designed to compare the ototoxic effects of volatile ethyl benzene in guinea pigs and rats. Rats showed deteriorated auditory thresholds in the mid-frequency range, based on electrocochleography, after 550-ppm ethyl benzene (8 h/day, 5 days). Outer hair cell (OHC) loss was found in the corresponding cochlear regions. In contrast, guinea pigs showed no threshold shifts and no OHC loss after exposure to much higher ethyl benzene levels (2500 ppm, 6 h/day, 5 days). Subsequently, a limited study (four rats and four guinea pigs) was performed in an attempt to understand these differences in susceptibility. Ethyl benzene concentration in blood was determined in both species after exposure to 500-ppm ethyl benzene (8 h/day, 3 days). At the end of the first day, blood of the rats contained 23.2+/-0.8-microg/ml ethyl benzene, whereas the concentration in guinea pig blood was 2.8+/-0.1 microg/ml. After 3 days, the concentration in both species decreased with respect to the first day, but the ethyl benzene concentration in rat blood was still 4.3 times higher than that in guinea pig blood. Thus, the difference in susceptibility between the species may be related to the ethyl benzene concentration in blood.

Spatio-temporal dynamics of theta oscillations in hippocampal-entorhinal slices

Theta oscillations (4–12 Hz) are associated with learning and memory and are found in the hippocampus and the entorhinal cortex (EC). The spatio‐temporal organization of rhythmic activity in the hippocampal–EC complex was investigated in vitro. The voltage sensitive absorption dye NK3630 was used to record the changes in aggregated membrane voltage simultaneously from the neuronal networks involved. Oscillatory activity at 7.0 Hz (range, 5.8–8.2) was induced in the slice with the muscarinic agonist carbachol (75–100 μM) in the presence of bicuculline (5 μM). Time relations between all recording sites were analyzed using cross‐correlation functions which revealed systematic phase shifts in the theta oscillation recorded from the different entorhinal and hippocampal subregions. These phase shifts could be interpreted as propagation delays. The oscillation propagates over the slice in a characteristic spatio‐temporal sequence, where the entorhinal cortex leads, followed by the subiculum and then the dentate gyrus (DG), to finally reach the CA3 and the CA1 area. The delay from dentate gyrus to the CA3 area was 12.4 ± 1.1 ms (mean ± s.e.m.) and from the CA3 to the CA1 region it was 10.9 ± 1.9 ms. The propagation delays between the hippocampal subregions resemble the latencies of electrically evoked responses in the same subregions. Removing the entorhinal cortex from the slice changed the spatiotemporal pattern into a more clustered pattern with higher local synchrony. We conclude that in the slice, carbachol‐induced theta oscillations are initiated in the entorhinal cortex. The EC could serve to control the information flow through the neuronal network in the subregions of the hippocampus by synchronizing and/or entraining their responses to external inputs.

The functional and morphological characteristics of sciatic nerve degeneration and regeneration after crush injury in rats.

BACKGROUND Peripheral nerve damage induces a sequence of degeneration and regeneration events with a specific time course that leads to (partial) functional recovery. Quantitative electrophysiological analysis of degeneration and recovery over time is essential to understand the process. NEW METHOD The presented ex vivo neurophysiological method evaluates functional recovery of the propagation of the compound action potential after crush injury of the rat sciatic nerve. A 32 channel electrode array was used to monitor compound action potential propagation at time points between 1h and 35 days after semi-quantitative crush injury of the rat sciatic nerve. RESULTS The compound action potential was characterized by four measures: the latency, the duration, the amplitude and a measure that combined time and location. These four parameters reflected the subsequent steps in early axonal degradation, the transition to rapid degeneration followed by sprouting and the long period of remyelination that accompanied regeneration. COMPARISON WITH EXISTING METHODS The neurophysiology measures of the compound action potential were compared with the morphology of the nerve at representative time points and analysis of functional recovery of action potential propagation was compared with a behavioral test: the foot flick test. CONCLUSIONS Our data suggests that the ex vivo electrophysiological method is complementary to the classical behavioral foot flick test in that it allows a detailed time analysis of the degeneration and early regeneration phases at a high spatial and temporal sensitivity. The results were well-matched with observations made with immunohistochemical and morphological methods.

Delay and Impairment in Brain Development and Function in Rat Offspring After Maternal Exposure to Methylmercury

Maternal exposure to the neurotoxin methylmercury (MeHg) has been shown to have adverse effects on neural development of the offspring in man. Little is known about the underlying mechanisms by which MeHg affects the developing brain. To explore the neurodevelopmental defects and the underlying mechanism associated with MeHg exposure, the cerebellum and cerebrum of Wistar rat pups were analyzed by [(18)F]FDG PET functional imaging, field potential analysis, and microarray gene expression profiling. Female rat pups were exposed to MeHg via maternal diet during intrauterinal and lactational period (from gestational day 6 to postnatal day (PND)10), and their brain tissues were sampled for the analysis at weaning (PND18-21) and adulthood (PND61-70). The [(18)F]FDG PET imaging and field potential analysis suggested a delay in brain activity and impaired neural function by MeHg. Genome-wide transcriptome analysis substantiated these findings by showing (1) a delay in the onset of gene expression related to neural development, and (2) alterations in pathways related to both structural and functional aspects of nervous system development. The latter included changes in gene expression of developmental regulators, developmental phase-associated genes, small GTPase signaling molecules, and representatives of all processes required for synaptic transmission. These findings were observed at dose levels at which only marginal changes in conventional developmental toxicity endpoints were detected. Therefore, the approaches applied in this study are promising in terms of yielding increased sensitivity compared with classical developmental toxicity tests.

EFFICACY OF A NEW CHARGE-BALANCED BIPHASIC ELECTRICAL STIMULUS IN THE ISOLATED SCIATIC NERVE AND THE HIPPOCAMPAL SLICE

Most deep brain stimulators apply rectangular monophasic voltage pulses. By modifying the stimulus shape, it is possible to optimize stimulus efficacy and find the best compromise between clinical effect, minimal side effects and power consumption of the stimulus generator. In this study, we compared the efficacy of three types of charge-balanced biphasic pulses (CBBPs, nominal duration 100 μs) in isolated sciatic nerves and in in vitro hippocampal brain slices of the rat. Using these two models, we tested the efficacy of several stimulus shapes exclusively on axons (in the sciatic nerve) and compared the effect with that of stimuli in the more complex neuronal network of the hippocampal slice by considering the stimulus-response relation. We showed that (i) adding an interphase gap (IPG, range 100-500 μs) to the CBBP enhances stimulus efficacy in the rat sciatic nerve and (ii) that this type of stimuli (CBBP with IPG) is also more effective in hippocampal slices. This benefit was similar for both models of voltage and current stimulation. In our two models, asymmetric CBBPs were less beneficial. Therefore, CBBPs with IPG appear to be well suited for application to DBS, since they enhance efficacy, extend battery life and potentially reduce harmful side effects.

Chapter 20 – Hippocampal Formation

The hippocampal formation and parahippocampal region are prominent components of the rat nervous system and play a crucial role in learning, memory, and spatial navigation. Many new details regarding the entorhinal cortex have been discovered since the previous edition, and the growing interest in the area of CA2 has been covered in this chapter. Emphasis is on a conceptual change: instead of perceiving the hippocampal circuit as the standard sequential processing network, current insights favor the concept that multiple parallel networks are present. Many new facts, combined with a thorough restructuring of information and inclusion of pointers to relevant (online) resources, make this chapter relevant to both the novice and senior readership.