Increased cortical and thalamic excitability in freely moving APPswe/PS1dE9 mice modeling epileptic activity associated with Alzheimer's disease.

Amyloid precursor protein transgenic mice modeling Alzheimer's disease display frequent occurrence of seizures peaking at an age when amyloid plaques start to form in the cortex and hippocampus. We tested the hypothesis that numerous reported interactions of amyloid-ß with cell surface molecules result in altered excitation-inhibition balance in brain-wide neural networks, eventually leading to epileptogenesis. We examined electroencephalograms (EEGs) and auditory-evoked potentials (AEPs) in freely moving 4-month-old APPswe/PS1dE9 (APdE9) and wild-type (WT) control mice in the hippocampus, cerebral cortex, and thalamus during movement, quiet waking, non-rapid eye movement sleep, and rapid eye movement (REM) sleep. Cortical EEG power was higher in APdE9 mice than in WT mice over a broad frequency range (5-100 Hz) and during all 4 behavioral states. Thalamic EEG power was also increased but in a narrower range (10-80 Hz). Furthermore, APdE9 mice displayed augmented cortical and thalamic AEPs. While power and theta-gamma modulation were preserved in the APdE9 hippocampus, REM sleep-related phase shift of theta-gamma modulation was altered. Our data suggest that at the early stage of amyloid pathology, cortical principal cells become hyperexcitable and via extensive cortico-thalamic connection drive thalamic cells. Minor hippocampal changes are most likely secondary to abnormal entorhinal input.

Orientation selective stimulation with tetrahedral electrodes of the rat infralimbic cortex to indirectly target the amygdala.

Introduction: Deep brain stimulation (DBS) is a rapidly developing therapeutic intervention with constantly expanding neurological and psychiatric indications. A major challenge for the approach is the precise targeting and limitation of the effect on the desired neural pathways. We have introduced a new approach, orientation selective stimulation (OSS) that allows free rotation of the induced electric field on a plane when using a probe with three parallel electrodes forming an equilateral triangle at the tip. Here, we expand the technique by introducing a tetrahedral stimulation probe that enables adjustment of the primary electric field direction freely at any angle in a 3D space around the stimulating probe. OSS in 3D will enable better targeting of the electric field according to the local brain anatomy. We tested its utility in a rat model of DBS for treatment-resistant depression. The stimulation directed to the subgenual anterior cingulate cortex (sgACC) has yielded dramatic improvement in individual patients suffering from therapy resistant depression, but no consistent benefit in larger series. This failure has been ascribed to the challenging anatomy of sgACC with several crossing neural tracts and individual differences in the local anatomy. Methods: We stimulated infralimbic cortex (IL), the rat analog of sgACC, and recorded local electrical responses in amygdala (AMG) that is monosynaptically connected to IL and plays a central role in emotional states. We further traced AMG-IL connections using a viral vector and tractography using diffusion magnetic resonance imaging (MRI). Finally, we mimicked the clinical situation by delivering sustained 130 Hz stimulation at IL at the most effective field orientation and followed changes in resting-state functional connectivity with IL using functional MRI. To help interpretation of responses in functional connectivity, we stimulated only the left IL, which we did not expect to evoke measurable changes in the rat behavior. Results: The AMG evoked responses depended systematically on the IL stimulation field orientation and yielded the maximum response in near vertical field orientation in accordance with tractography. Sustained 130 Hz stimulation at a field orientation yielding the strongest AMG evoked responses increased functional connectivity between IL and AMG on the stimulation side. Conclusion: These findings suggest that OSS in 3D provides a new approach to optimize the DBS for every individual patient with a single stimulation probe implantation.

Genetic ablation of tenascin-C expression leads to abnormal hippocampal CA1 structure and electrical activity in vivo.

Despite evidence that the extracellular matrix glycoprotein tenascin-C (TNC) is implicated in brain development and plasticity, its roles in the intact adult brain are unknown. Here we report that spontaneous local field potential (LFP) activity in freely moving adult TNC-deficient mice is abnormal. The power of cortical and hippocampal theta and gamma oscillations was enhanced in comparison to wild-type mice. The alteration in hippocampal gamma rhythm was subfield specific, such that CA1 gamma was accentuated while dentate gyrus gamma was normal. Similar to LFP, synaptic transmission and plasticity at perforant path synapses in the dentate gyrus were unaffected by the mutation. Morphological analyses revealed a subfield-specific reduction in the CA1 volume and a reduction in the numbers of somatostatin-positive interneurons in the hippocampus as potential structural substrates of the observed functional aberrations. These findings indicate a role for tenascin-C in structural organization of the CA1 hippocampal subfield and in shaping neural activity.

Characterization of Epileptic Spiking Associated With Brain Amyloidosis in APP/PS1 Mice.

Epileptic activity without visible convulsions is common in Alzheimer's disease (AD) and may contribute adversely to the disease progress and symptoms. Transgenic mice with amyloid plaque pathology also display epileptic seizures, but those are too infrequent to assess the effect of anti-epileptic treatments. Besides spontaneous seizures, these mice also display frequent epileptic spiking in epidural EEG recordings, and these have provided a means to test potential drug treatment to AD-related epilepsy. However, the origin of EEG spikes in transgenic AD model mice has remained elusive, which makes it difficult to relate electrophysiology with underlying pathology at the cellular and molecular level. Using multiple cortical and subcortical electrodes in freely moving APP/PS1 transgenic mice and their wild-type littermates, we identified several types of epileptic spikes among over 15 800 spikes visible with cortical screw electrodes based on their source localization. Cortical spikes associated with muscle twitches, cortico-hippocampal spikes, and spindle and fast-spindle associated spikes were present equally often in both APP/PS1 and wild-type mice, whereas pure cortical spikes were slightly more common in APP/PS1 mice. In contrast, spike-wave discharges, cortico-hippocampal spikes with after hyperpolarization and giant spikes were seen almost exclusively in APP/PS1 mice but only in a subset of them. Interestingly, different subtypes of spikes responded differently to anti-epileptic drugs ethosuximide and levetiracetam. From the translational point most relevant may be the giant spikes generated in the hippocampus that reached an amplitude up to ± 5 mV in the hippocampal channel. As in AD patients, they occurred exclusively during sleep. Further, we could demonstrate that a high number of giant spikes in APP/PS1 mice predicts seizures. These data show that by only adding a pair of hippocampal deep electrodes and EMG to routine cortical epidural screw electrodes and by taking into account underlying cortical oscillations, one can drastically refine the analysis of cortical spike data. This new approach provides a powerful tool to preclinical testing of potential new treatment options for AD related epilepsy.

Increased cortical beta power and spike-wave discharges in middle-aged APP/PS1 mice.

Amyloid plaque-forming transgenic mice display neuronal hyperexcitability, epilepsy, and sudden deaths in early adulthood. However, it is unknown whether hyperexcitability persists until middle ages when memory impairment manifests. We recorded multichannel video electroencephalography (EEG), local field potentials, and auditory evoked potentials in transgenic mice carrying mutated human amyloid precursor protein (APP) and presenilin-1 (PS1) genes and wild-type littermates at 14-16 months and compared the results with data we have earlier collected from 4-month-old mice. Furthermore, we monitored acoustic startle responses in other APP/PS1 and wild-type mice from 3 to 11 months of age. Independent of the age APP/PS1 mice demonstrated increased cortical power at 8-60 Hz. They also displayed over 5-fold increase in the occurrence of spike-wave discharges and augmented auditory evoked potentials compared with nontransgenic littermates. In contrast to evoked potentials, APP/PS1 mice showed normalization of acoustic startle responses with aging. Increased cortical power and spike-wave discharges provide powerful new biomarkers to monitor progression of amyloid pathology in preclinical intervention studies.

Amyloid Plaques Show Binding Capacity of Exogenous Injected Amyloid-ß.

Amyloid plaques, although inducing damage to the immediately surrounding neuropil, have been proposed to provide a relatively innocuous way to deposit toxic soluble amyloid-ß (Aß) species. Here we address this hypothesis by exploring spread and absorption of fluorescent Aß to pre-existing amyloid plaques after local application in wild-type mice versus APP/PS1 transgenic mice with amyloid plaques. Local intracortical or intracerebroventricular injection of fluorescently-labeled Aß in APP/PS1 mice with a high plaque density resulted in preferential accumulation of the peptide in amyloid plaques in both conventional postmortem histology and in live imaging using two-photon microscopy. These findings support the contention that amyloid plaques may act as buffers to protect neurons from the toxic effects of momentary high concentrations of soluble Aß oligomers.

Normal Amplitude of Electroretinography and Visual Evoked Potential Responses in AßPP/PS1 Mice.

Alzheimer's disease has been shown to affect vision in human patients and animal models. This may pose the risk of bias in behavior studies and therefore requires comprehensive investigation. We recorded electroretinography (ERG) under isoflurane anesthesia and visual evoked potentials (VEP) in awake amyloid expressing AßPPswe/PS1dE9 (AßPP/PS1) and wild-type littermate mice at a symptomatic age. The VEPs in response to patterned stimuli were normal in AßPP/PS1 mice. They also showed normal ERG amplitude but slightly shortened ERG latency in dark-adapted conditions. Our results indicate subtle changes in visual processing in aged male AßPP/PS1 mice specifically at a retinal level.

Increased Epileptiform EEG Activity and Decreased Seizure Threshold in Arctic APP Transgenic Mouse Model of Alzheimer's Disease.

Several Alzheimer model mice carrying transgenic amyloid precursor protein (APP) with the Swedish mutation have been reported to exhibit spontaneous seizures and/or increased epileptiform EEG activity. The primary cause for the epilepsy phenotype is still under debate. In contrast to mice with APPswe mutation that develop extracellular amyloid plaques, mice with APP Arctic mutation (E693G) have no bias toward ß-secretase cleavage and display intracellular amyloid deposits but not plaques. We conducted a systematic long-term video-EEG recording in three two-week sessions on 21 APParc and 11 wild-type control mice between 3.5 and 8 months of age. Spontaneous seizures were not detected more often in APParc mice than in their wild-type control mice. Long (1 - 5 s) epileptiform discharges were occasionally detected in both APParc and wild-type mice, but short (0.5 - <1 s) epileptiform discharges were more common in APParc mice than in wild-types. However, they were far less frequent than in 6 APPswe/PS1dE9 mice recorded in parallel. In pentylenetetrazole test for seizure susceptibility, APParc mice displayed a shorter latency to sharp-wave discharges than wildtype mice but no increase in seizure duration. These data speak for a relatively mild epilepsy phenotype in APParc mice compared to APPswe mice despite even higher extent of APP overexpression. Thus extracellular amyloid plaques or increased ß-secretase cleavage products appear important for the epilepsy phenotype in APPswe mice.

Altered Electroencephalographic Activity Associated with Changes in the Sleep-Wakefulness Cycle of C57BL/6J Mice in Response to a Photoperiod Shortening.

AIM: Under natural conditions diurnal rhythms of biological processes of the organism are synchronized with each other and to the environmental changes by means of the circadian system. Disturbances of the latter affect hormonal levels, sleep-wakefulness cycle and cognitive performance. To study mechanisms of such perturbations animal models subjected to artificial photoperiods are often used. The goal of current study was to understand the effects of circadian rhythm disruption, caused by a short light-dark cycle regime, on activity of the cerebral cortex in rodents. METHODS: We used electroencephalogram to assess the distribution of vigilance states, perform spectral analysis, and estimate the homeostatic sleep drive. In addition, we analyzed spontaneous locomotion of C57BL/6J mice under symmetric, 22-, 21-, and 20-h-long light-dark cycles using video recording and tracking methods. RESULTS AND CONCLUSIONS: We found that shortening of photoperiod caused a significant increase of slow wave activity during non-rapid eye movement sleep suggesting an elevation of sleep pressure under such conditions. While the rhythm of spontaneous locomotion was completely entrained by all light-dark cycles tested, periodic changes in the power of the θ- and γ-frequency ranges during wakefulness gradually disappeared under 22- and 21-h-long light-dark cycles. This was associated with a significant increase in the θ-γ phase-amplitude coupling during wakefulness. Our results thus provide deeper understanding of the mechanisms underlying the impairment of learning and memory retention, which is associated with disturbed circadian regulation.

Place cells of aged rats in two visually identical compartments.

Aged rats perform poorly on spatial learning tasks, a cognitive impairment which has been linked to the failure of hippocampal networks to fully encode changes in the external environment [Barnes CA, Suster MS, Shen J, McNaughton BL. Multistability of cognitive maps in the hippocampus of old rats. Nature 1997;388(6639):272-5; Wilson IA, Ikonen S, Gureviciene I, McMahan RW, Gallagher M, Eichenbaum H, et al. Cognitive aging and the hippocampus: how old rats represent new environments. J Neurosci 2004;24(15):3870-8]. To examine whether the impairment in hippocampal processing extends to conditions in which self-motion provides the cues for environmental change, we have analyzed spatial firing patterns of hippocampal pyramidal neurons in young and aged rats, as well as in young rats with selective cholinergic lesions, another model of cognitive aging. The rats walked between two visually identical environments, pitting self-motion cues that indicated environmental change against visual inputs that indicated no differences between environments. Our results indicated that place cells in both aged and cholinergic-lesioned rats were equally likely as those of young rats to create new spatial representations in the second compartment. These findings suggest that the hippocampal network of aged rats is able to process changes in internally generated cues without rigidity, but that incomplete processing of external landmark cues may lead to impaired spatial learning.

Alteration of cortical EEG in mice carrying mutated human APP transgene.

Transgenic mice expressing human APPswe and PS1-A264E mutations mimic certain neuropathological features of Alzheimer's disease (AD). These mice have elevated levels of the highly fibrillogenic amyloid beta1-42 peptide (Abeta42) and develop amyloid plaques around the age of 9 months. Our aim was to find whether these transgenic mice differ electrophysiologically from non-transgenic mice and whether the alteration in EEG activity progresses with the accumulation of Abeta. The APP/PS1 mice had reduced cortical theta activity and enhanced beta and gamma activity, but these changes were not age-dependent. APP single mutant mice had similar EEG alterations in theta, beta and gamma bands as APP/PS1 double mutant mice while PS1 single mutant mice did not differ from non-transgenic controls. Insoluble Abeta40 and Abeta42 levels were robustly increased in APP/PS1 double mutant mice and insoluble Abeta40 moderately increased also in APP single mutant mice. Soluble Abeta42 was found in all APP mutant mice but also in lower concentrations in PS1 single mutant mice. Plaques were deposited in 13-month-old APP/PS1 double mutant mice but not in 8-month-old double mutant or 13-month-old single mutant mice. We conclude that the alteration of EEG activity in APP/PS1 double mutant and APP single mutant mice is related to their APP genotype rather than to deposition of beta-amyloid in the brain.

Early retinal function deficit without prominent morphological changes in the R6/2 mouse model of Huntington's disease.

Huntington's disease (HD) is an inherited neurodegenerative disorder that primarily affects the medium-size GABAergic neurons of striatum. The R6/2 mouse line is one of the most widely used animal models of HD. Previously the hallmarks of HD-related pathology have been detected in photoreceptors and interneurons of R6/2 mouse retina. Here we aimed to explore the survival of retinal ganglion cells (RGCs) and functional integrity of distinct retinal cell populations in R6/2 mice. The pattern electroretinography (PERG) signal was lost at the age of 8 weeks in R6/2 mice in contrast to the situation in wild-type (WT) littermates. This defect may be attributable to a major reduction in photopic ERG responses in R6/2 mice which was more evident in b- than a-wave amplitudes. At the age of 4 weeks R6/2 mice had predominantly the soluble form of mutant huntingtin protein (mHtt) in the RGC layer cells, whereas the aggregated form of mHtt was found in the majority of those cells from the 12-week-old R6/2 mice and onwards. Retinal astrocytes did not contain mHtt deposits. The total numbers of RGC layer cells, retinal astrocytes as well as optic nerve axons did not differ between 18-week-old R6/2 mice and their WT controls. Our data indicate that mHtt deposition does not cause RGC degeneration or retinal astrocyte loss in R6/2 mice even at a late stage of HD-related pathology. However, due to functional deficits in the rod- and cone-pathways, the R6/2 mice suffer progressive deficits in visual capabilities starting as early as 4 weeks; at 8 weeks there is severe impairment. This should be taken into account in any behavioral testing conducted in R6/2 mice.

Short- and long-term habituation of auditory event-related potentials in the rat.

An auditory oddball paradigm in humans generates a long-duration cortical negative potential, often referred to as mismatch negativity. Similar negativity has been documented in monkeys and cats, but it is controversial whether mismatch negativity also exists in awake rodents. To this end, we recorded cortical and hippocampal evoked responses in rats during alert immobility under a typical passive oddball paradigm that yields mismatch negativity in humans. The standard stimulus was a 9 kHz tone and the deviant either 7 or 11 kHz tone in the first condition. We found no evidence of a sustained potential shift when comparing evoked responses to standard and deviant stimuli. Instead, we found repetition-induced attenuation of the P60 component of the combined evoked response in the cortex, but not in the hippocampus. The attenuation extended over three days of recording and disappeared after 20 intervening days of rest. Reversal of the standard and deviant tones resulted is a robust enhancement of the N40 component not only in the cortex but also in the hippocampus. Responses to standard and deviant stimuli were affected similarly. Finally, we tested the effect of scopolamine in this paradigm. Scopolamine attenuated cortical N40 and P60 as well as hippocampal P60 components, but had no specific effect on the deviant response. We conclude that in an oddball paradigm the rat demonstrates repetition-induced attenuation of mid-latency responses, which resembles attenuation of the N1-component of human auditory evoked potential, but no mismatch negativity.

Artificial theta stimulation impairs encoding of contextual fear memory.

Several experiments have demonstrated an intimate relationship between hippocampal theta rhythm (4-12 Hz) and memory. Lesioning the medial septum or fimbria-fornix, a fiber track connecting the hippocampus and the medial septum, abolishes the theta rhythm and results in a severe impairment in declarative memory. To assess whether there is a causal relationship between hippocampal theta and memory formation we investigated whether restoration of hippocampal theta by electrical stimulation during the encoding phase also restores fimbria-fornix lesion induced memory deficit in rats in the fear conditioning paradigm. Male Wistar rats underwent sham or fimbria-fornix lesion operation. Stimulation electrodes were implanted in the ventral hippocampal commissure and recording electrodes in the septal hippocampus. Artificial theta stimulation of 8 Hz was delivered during 3-min free exploration of the test cage in half of the rats before aversive conditioning with three foot shocks during 2 min. Memory was assessed by total freezing time in the same environment 24 h and 28 h after fear conditioning, and in an intervening test session in a different context. As expected, fimbria-fornix lesion impaired fear memory and dramatically attenuated hippocampal theta power. Artificial theta stimulation produced continuous theta oscillations that were almost similar to endogenous theta rhythm in amplitude and frequency. However, contrary to our predictions, artificial theta stimulation impaired conditioned fear response in both sham and fimbria-fornix lesioned animals. These data suggest that restoration of theta oscillation per se is not sufficient to support memory encoding after fimbria-fornix lesion and that universal theta oscillation in the hippocampus with a fixed frequency may actually impair memory.

Spontaneous epileptiform discharges in a mouse model of Alzheimer's disease are suppressed by antiepileptic drugs that block sodium channels.

Previous studies have demonstrated an increased risk of epilepsy in patients with Alzheimer's disease (AD). Also, in many mouse models of AD, animals have spontaneous seizures and frequent epileptiform discharges (EDs). Abnormal function of sodium channels has been proposed to contribute to hyperexcitability in a manner suggesting that drugs that block sodium channels might exacerbate the condition. Here we addressed this question by investigating whether common antiepileptic drugs (AEDs) that block sodium channels, including carbamazepine (CBZ), phenytoin (DPH), or valproic acid (VPA) have any effect on spontaneous seizures or EDs in APdE9 mice. Mice were successively treated with vehicle, followed by CBZ (10mg/kg, t.i.d.), DPH (10mg/kg, t.i.d.), or VPA (260 mg/kg, b.i.d.) for 3d. After wash-out and new vehicle treatment, higher doses of CBZ (40 mg/kg, t.i.d.), DPH (40 mg/kg, t.i.d.), or VPA (400mg/kg, b.i.d.) were administered for 3d (DPH) or 5d (CBZ, VPA). During the entire experiment, mice were under continuous (24/7) video-EEG monitoring. Our data show that each treatment reduced the number of spontaneous electrographic EDs. VPA was the most effective by reducing the ED frequency below 50% of that at baseline in 75% of mice. Western blot analysis of the Na(v)1.1 protein levels in the ventral temporal cortex and the hippocampus did not reveal any differences between the genotypes. Under the conditions tested, sodium channel blocking AEDs suppressed epileptiform activity in APdE9 mice with increased amyloid pathology. Whether this applies to other mouse models of AD with different APP mutations and/or genetic background remains to be explored.

Aging and alpha-synuclein affect synaptic plasticity in the dentate gyrus.

Although intracellular accumulation of alpha-synuclein (alpha-syn) is a characteristic pathological change in Parkinson's disease, Lewy body dementia and Alzheimer's disease, the normal function of this presynaptic protein is still unknown. To assess the contribution of alpha-syn to synaptic plasticity as well as to age-related synaptic degeneration in mice, we compared adult and aged mice overexpressing mutated (A30P) human alpha-syn with their nontransgenic littermates using behavioral tests and electrophysiological measures in the dentate gyrus. We found decreased basal synaptic transmission and paired-pulse facilitation in the perforant path-dentate granule cell synapses of aged mice. In addition, alpha-syn accumulation in aged A30P mice but not in aged wild-type mice led to long-term depression of synaptic transmission after a stimulation protocol that normally induces long-term potentiation. These findings suggest that overexpression of mutated alpha-syn exacerbates the aging process and leads to impaired synaptic plasticity.

Role of alpha-synuclein in synaptic glutamate release.

Defective mobilization of dopamine from the reserve pool has been reported in both alpha-synuclein knockout mice (KO) and pPrp-A30P transgenic mice. The present study extends these findings to glutamate release. Standard hippocampal slices were prepared from KO, pPrp-A30P, and C57BL/6J wild type (WT1) mice, as well as from mice with transgenic overexpression of wild type human alpha-synuclein (pSyn-hASY) and their negative littermates (WT2), and field responses were measured in CA3 in response to mossy fiber stimulation. The input/output curves indicated no differences in basal synaptic transmission between groups. Paired-pulse facilitation was significantly weaker in both transgenic alpha-synuclein lines and KO mice compared to their controls. High-frequency stimulation induced LTP only in transgenic mice. Frequency-facilitation was absent in KO mice and different from other mouse lines. These findings support the idea that lack of alpha-synuclein impairs mobilization of glutamate from the reserve pool. However, transgenic expression of A30P mutated or wild type alpha-synuclein does not appear to prevent endogenous mouse alpha-synuclein to carry out this function.

Increased hippocampal and cortical beta oscillations in mice deficient for the HNK-1 sulfotransferase.

The HNK-1 carbohydrate is detectable in perineuronal nets around inhibitory neurons in the hippocampus and neocortex. To address the functional contribution of HNK-1 to interneuron function in the adult brain, we recorded EEG and auditory-evoked potential in freely moving mice deficient for HNK-1 sulfotransferase (ST-/- mice) and in wild-type littermates. While ST-/- mice displayed normal theta oscillations, both cortical and hippocampal oscillations within the beta range were enhanced, and gamma oscillations showed an opposite trend. ST-/- mice had amplitudes of auditory-evoked potentials similar to control mice, but the latencies of their hippocampal responses were shorter. Morphological analysis revealed a decreased density of parvalbumin-positive interneurons in the hippocampal CA3 subfield of ST-/- mice, which may contribute to the observed changes in networks oscillations. These findings reveal alterations in ST-/- mice that differ from EEG abnormalities of mice deficient in the HNK-1 carrier molecule tenascin-R.