Key morphological features of human pyramidal neurons.

The basic building block of the cerebral cortex, the pyramidal cell, has been shown to be characterized by a markedly different dendritic structure among layers, cortical areas, and species. Functionally, differences in the structure of their dendrites and axons are critical in determining how neurons integrate information. However, within the human cortex, these neurons have not been quantified in detail. In the present work, we performed intracellular injections of Lucifer Yellow and 3D reconstructed over 200 pyramidal neurons, including apical and basal dendritic and local axonal arbors and dendritic spines, from human occipital primary visual area and associative temporal cortex. We found that human pyramidal neurons from temporal cortex were larger, displayed more complex apical and basal structural organization, and had more spines compared to those in primary sensory cortex. Moreover, these human neocortical neurons displayed specific shared and distinct characteristics in comparison to previously published human hippocampal pyramidal neurons. Additionally, we identified distinct morphological features in human neurons that set them apart from mouse neurons. Lastly, we observed certain consistent organizational patterns shared across species. This study emphasizes the existing diversity within pyramidal cell structures across different cortical areas and species, suggesting substantial species-specific variations in their computational properties.

Three-dimensional analysis of synaptic organization in the hippocampal CA1 field in Alzheimer's disease.

Alzheimer's disease is the most common form of dementia, characterized by a persistent and progressive impairment of cognitive functions. Alzheimer's disease is typically associated with extracellular deposits of amyloid-ß peptide and accumulation of abnormally phosphorylated tau protein inside neurons (amyloid-ß and neurofibrillary pathologies). It has been proposed that these pathologies cause neuronal degeneration and synaptic alterations, which are thought to constitute the major neurobiological basis of cognitive dysfunction in Alzheimer's disease. The hippocampal formation is especially vulnerable in the early stages of Alzheimer's disease. However, the vast majority of electron microscopy studies have been performed in animal models. In the present study, we performed an extensive 3D study of the neuropil to investigate the synaptic organization in the stratum pyramidale and radiatum in the CA1 field of Alzheimer's disease cases with different stages of the disease, using focused ion beam/scanning electron microscopy (FIB/SEM). In cases with early stages of Alzheimer's disease, the synapse morphology looks normal and we observed no significant differences between control and Alzheimer's disease cases regarding the synaptic density, the ratio of excitatory and inhibitory synapses, or the spatial distribution of synapses. However, differences in the distribution of postsynaptic targets and synaptic shapes were found. Furthermore, a lower proportion of larger excitatory synapses in both strata were found in Alzheimer's disease cases. Individuals in late stages of the disease suffered the most severe synaptic alterations, including a decrease in synaptic density and morphological alterations of the remaining synapses. Since Alzheimer's disease cases show cortical atrophy, our data indicate a reduction in the total number (but not the density) of synapses at early stages of the disease, with this reduction being much more accentuated in subjects with late stages of Alzheimer's disease. The observed synaptic alterations may represent a structural basis for the progressive learning and memory dysfunctions seen in Alzheimer's disease cases.

Volume Electron Microscopy Study of the Relationship Between Synapses and Astrocytes in the Developing Rat Somatosensory Cortex.

In recent years, numerous studies have shown that astrocytes play an important role in neuronal processing of information. One of the most interesting findings is the existence of bidirectional interactions between neurons and astrocytes at synapses, which has given rise to the concept of "tripartite synapses" from a functional point of view. We used focused ion beam milling and scanning electron microscopy (FIB/SEM) to examine in 3D the relationship of synapses with astrocytes that were previously labeled by intracellular injections in the rat somatosensory cortex. We observed that a large number of synapses (32%) had no contact with astrocytic processes. The remaining synapses (68%) were in contact with astrocytic processes, either at the level of the synaptic cleft (44%) or with the pre- and/or post-synaptic elements (24%). Regarding synaptic morphology, larger synapses with more complex shapes were most frequently found within the population that had the synaptic cleft in contact with astrocytic processes. Furthermore, we observed that although synapses were randomly distributed in space, synapses that were free of astrocytic processes tended to form clusters. Overall, at least in the developing rat neocortex, the concept of tripartite synapse only seems to be applicable to a subset of synapses.

Ambient Glutamate Promotes Paroxysmal Hyperactivity in Cortical Pyramidal Neurons at Amyloid Plaques via Presynaptic mGluR1 Receptors.

Synaptic dysfunctions and altered neuronal activity play major role in the pathophysiology of Alzheimer's disease (AD), with underlying mechanisms largely unknown. We report that in the prefrontal cortex of amyloid precursor protein-presenilin 1 and APP23 AD mice, baseline activity of pyramidal cells is disrupted by episodes of paroxysmal hyperactivity. Induced by spontaneous EPSC bursts, these incidents are prevalent in neurons proximal to amyloid plaques and involve enhanced activity of glutamate with metabotropic effects. Abolition of EPSC bursts by tetrodotoxin and SERCA ATPase blockers thapsigargin or cyclopiasonic acid suggests their presynaptic origin and sensitized store-released calcium. Accordingly, the rate of EPSC bursts activated by single axon stimulation is enhanced. Aggravation of the hyperactivity by blockers of excitatory amino acid transporter (±)-HIP-A and DL-TBOA together with histochemical and ultrastructural evidence for enrichment of plaque-related dystrophies with synaptic vesicles and SNARE protein SNAP-25 infer the later as hot-spots for ectopic release of glutamate. Inhibition of EPSC bursts by I/II mGluR1 blocker MCPG or selective mGluR1 antagonist LY367385 implicate metabotropic glutamatergic effects in generation of paroxysmal bursts. These findings demonstrate for the first time that at amyloid plaques, enhanced activity of nonsynaptic glutamate can promote irregular EPSC bursts with hyperactivity of pyramidal cells via mGluR1 receptors.

Analyzing dendritic spine pathology in Alzheimer's disease: problems and opportunities.

Synaptic failure is an immediate cause of cognitive decline and memory dysfunction in Alzheimer's disease. Dendritic spines are specialized structures on neuronal processes, on which excitatory synaptic contacts take place and the loss of dendritic spines directly correlates with the loss of synaptic function. Dendritic spines are readily accessible for both in vitro and in vivo experiments and have, therefore, been studied in great detail in Alzheimer's disease mouse models. To date, a large number of different mechanisms have been proposed to cause dendritic spine dysfunction and loss in Alzheimer's disease. For instance, amyloid beta fibrils, diffusible oligomers or the intracellular accumulation of amyloid beta have been found to alter the function and structure of dendritic spines by distinct mechanisms. Furthermore, tau hyperphosphorylation and microglia activation, which are thought to be consequences of amyloidosis in Alzheimer's disease, may also contribute to spine loss. Lastly, genetic and therapeutic interventions employed to model the disease and elucidate its pathogenetic mechanisms in experimental animals may cause alterations of dendritic spines on their own. However, to date none of these mechanisms have been translated into successful therapeutic approaches for the human disease. Here, we critically review the most intensely studied mechanisms of spine loss in Alzheimer's disease as well as the possible pitfalls inherent in the animal models of such a complex neurodegenerative disorder.

Intraneuronal APP and extracellular Aß independently cause dendritic spine pathology in transgenic mouse models of Alzheimer's disease.

Alzheimer's disease (AD) is thought to be caused by accumulation of amyloid-ß protein (Aß), which is a cleavage product of amyloid precursor protein (APP). Transgenic mice overexpressing APP have been used to recapitulate amyloid-ß pathology. Among them, APP23 and APPswe/PS1deltaE9 (deltaE9) mice are extensively studied. APP23 mice express APP with Swedish mutation and develop amyloid plaques late in their life, while cognitive deficits are observed in young age. In contrast, deltaE9 mice with mutant APP and mutant presenilin-1 develop amyloid plaques early but show typical cognitive deficits in old age. To unveil the reasons for different progressions of cognitive decline in these commonly used mouse models, we analyzed the number and turnover of dendritic spines as important structural correlates for learning and memory. Chronic in vivo two-photon imaging in apical tufts of layer V pyramidal neurons revealed a decreased spine density in 4-5-month-old APP23 mice. In age-matched deltaE9 mice, in contrast, spine loss was only observed on cortical dendrites that were in close proximity to amyloid plaques. In both cases, the reduced spine density was caused by decreased spine formation. Interestingly, the patterns of alterations in spine morphology differed between these two transgenic mouse models. Moreover, in APP23 mice, APP was found to accumulate intracellularly and its content was inversely correlated with the absolute spine density and the relative number of mushroom spines. Collectively, our results suggest that different pathological mechanisms, namely an intracellular accumulation of APP or extracellular amyloid plaques, may lead to spine abnormalities in young adult APP23 and deltaE9 mice, respectively. These distinct features, which may represent very different mechanisms of synaptic failure in AD, have to be taken into consideration when translating results from animal studies to the human disease.

Dense and overlapping innervation of pyramidal neurons by chandelier cells.

Chandelier (or axo-axonic) cells are a distinct group of GABAergic interneurons that innervate the axon initial segments of pyramidal cells and thus could have an important role controlling the activity of cortical circuits. To understand their connectivity, we labeled upper layers chandelier cells (ChCs) from mouse neocortex with a genetic strategy and studied how their axons contact local populations of pyramidal neurons, using immunohistochemical detection of axon initial segments. We studied ChCs located in the border of layers 1 and 2 from primary somatosensory cortex and found that practically all ChC axon terminals contact axon initial segments, with an average of three to five boutons per cartridge. By measuring the number of putative GABAergic synapses in initial segments, we estimate that each pyramidal neuron is innervated, on average, by four ChCs. Additionally, each individual ChC contacts 35-50% of pyramidal neurons within the areas traversed by its axonal arbor, with pockets of very high innervation density. Finally, ChCs have similar innervation patterns at different postnatal ages (P18-P90), with only relatively small lateral expansions of their arbor and increases in the total number of their cartridges during the developmental period analyzed. We conclude that ChCs innervate neighboring pyramidal neurons in a dense and overlapping manner, a connectivity pattern that could enable ChCs to exert a widespread influence on their local circuits.

The influence of phospho-τ on dendritic spines of cortical pyramidal neurons in patients with Alzheimer's disease.

The dendritic spines on pyramidal cells represent the main postsynaptic elements of cortical excitatory synapses and they are fundamental structures in memory, learning and cognition. In the present study, we used intracellular injections of Lucifer yellow in fixed tissue to analyse over 19 500 dendritic spines that were completely reconstructed in three dimensions along the length of the basal dendrites of pyramidal neurons in the parahippocampal cortex and CA1 of patients with Alzheimer's disease. Following intracellular injection, sections were immunostained for anti-Lucifer yellow and with tau monoclonal antibodies AT8 and PHF-1, which recognize tau phosphorylated at Ser202/Thr205 and at Ser396/404, respectively. We observed that the diffuse accumulation of phospho-tau in a putative pre-tangle state did not induce changes in the dendrites of pyramidal neurons, whereas the presence of tau aggregates forming intraneuronal neurofibrillary tangles was associated with progressive alteration of dendritic spines (loss of dendritic spines and changes in their morphology) and dendrite atrophy, depending on the degree of tangle development. Thus, the presence of phospho-tau in neurons does not necessarily mean that they suffer severe and irreversible effects as thought previously but rather, the characteristic cognitive impairment in Alzheimer's disease is likely to depend on the relative number of neurons that have well developed tangles.

Colocalization of α-actinin and synaptopodin in the pyramidal cell axon initial segment.

The cisternal organelle that resides in the axon initial segment (AIS) of neocortical and hippocampal pyramidal cells is thought to be involved in regulating the Ca(2+) available to maintain AIS scaffolding proteins, thereby preserving normal AIS structure and function. Through immunocytochemistry and correlative light and electron microscopy, we show here that the actin-binding protein α-actinin is present in the typical cistenal organelle of rodent pyramidal neurons as well as in a large structure in the AIS of a subpopulation of layer V pyramidal cells that we have called the "giant saccular organelle." Indeed, this localization of α-actinin in the AIS is dependent on the integrity of the actin cytoskeleton. Moreover, in the cisternal organelle of cultured hippocampal neurons, α-actinin colocalizes extensively with synaptopodin, a protein that interacts with both actin and α-actinin, and they appear concomitantly during the development of these neurons. Together, these results indicate that α-actinin and the actin cytoskeleton are important components of the cisternal organelle that are probably required to stabilize the AIS.

The Application of 3D Anatomy for Teaching Veterinary Clinical Neurology.

Neuroanatomy is always a challenging topic for veterinary students. It is widely accepted that understanding the anatomy of the central nervous system (CNS) is essential to explain many of the pathological processes that affect the brain. Although its study has varied over time to achieve this goal, in human and veterinary medicine it is difficult to find a teaching method that associates normal anatomy with pathological alterations of the brain. For the first time, we have created an educational tool that combines neuroanatomy and neuropathology, using different magnetic resonance (MR) images as a basis and EspINA software as analyzer, to obtain segmented structures and 3D reconstructions of the dog brain. We demonstrate that this combination is an optimal tool to help anatomists to understand the encephalon, and additionally to help clinicians to recognize illness including a multitude of neurological problems. In addition, we have tried to see whether photogrammetry, which is a common technique in other sciences, for example geology, could be useful to teach veterinary neuroanatomy. Although we still need further investigations, we have been able to generate 3D reconstructions of the whole brain, with very promising results to date.

Neuroanatomical and psychological considerations in temporal lobe epilepsy.

Temporal lobe epilepsy (TLE) is the most common form of focal epilepsy and is associated with a variety of structural and psychological alterations. Recently, there has been renewed interest in using brain tissue resected during epilepsy surgery, in particular 'non-epileptic' brain samples with normal histology that can be found alongside epileptic tissue in the same epileptic patients - with the aim being to study the normal human brain organization using a variety of methods. An important limitation is that different medical characteristics of the patients may modify the brain tissue. Thus, to better determine how 'normal' the resected tissue is, it is fundamental to know certain clinical, anatomical and psychological characteristics of the patients. Unfortunately, this information is frequently not fully available for the patient from which the resected tissue has been obtained - or is not fully appreciated by the neuroscientists analyzing the brain samples, who are not necessarily experts in epilepsy. In order to present the full picture of TLE in a way that would be accessible to multiple communities (e.g., basic researchers in neuroscience, neurologists, neurosurgeons and psychologists), we have reviewed 34 TLE patients, who were selected due to the availability of detailed clinical, anatomical, and psychological information for each of the patients. Our aim was to convey the full complexity of the disorder, its putative anatomical substrates, and the wide range of individual variability, with a view toward: (1) emphasizing the importance of considering critical patient information when using brain samples for basic research and (2) gaining a better understanding of normal and abnormal brain functioning. In agreement with a large number of previous reports, this study (1) reinforces the notion of substantial individual variability among epileptic patients, and (2) highlights the common but overlooked psychopathological alterations that occur even in patients who become "seizure-free" after surgery. The first point is based on pre- and post-surgical comparisons of patients with hippocampal sclerosis and patients with normal-looking hippocampus in neuropsychological evaluations. The second emerges from our extensive battery of personality and projective tests, in a two-way comparison of these two types of patients with regard to pre- and post-surgical performance.

Three-dimensional synaptic organization of the human hippocampal CA1 field.

The hippocampal CA1 field integrates a wide variety of subcortical and cortical inputs, but its synaptic organization in humans is still unknown due to the difficulties involved studying the human brain via electron microscope techniques. However, we have shown that the 3D reconstruction method using Focused Ion Beam/Scanning Electron Microscopy (FIB/SEM) can be applied to study in detail the synaptic organization of the human brain obtained from autopsies, yielding excellent results. Using this technology, 24,752 synapses were fully reconstructed in CA1, revealing that most of them were excitatory, targeting dendritic spines and displaying a macular shape, regardless of the layer examined. However, remarkable differences were observed between layers. These data constitute the first extensive description of the synaptic organization of the neuropil of the human CA1 region.

There are billions of nerve cells or neurons in the human brain, and each one can form thousands of connections, also called synapses, with other neurons. That means there are trillions of synapses in the brain that keep information flowing. Studying the arrangement of individual neurons in the human brain, and the connections between them, is incredibly difficult because of its complexity. Scientists have tools that can image the whole brain and can measure the activity in different regions, but these tools only visualize brain structures that are large enough to be seen with human eyes. Synapses are much smaller (in the range of nanometers), and can only be seen using thin slices of preserved brain tissue through a technique called electron microscopy. The hippocampus is a part of the human brain that is critical for memory, learning and spatial orientation, and is affected in epilepsy and Alzheimer's disease. Although numerous studies of the hippocampus have been performed in laboratory animals, such as mice, the question remains as to how much of the information gained from these studies applies to humans. Thus, studying the human brain directly is a major goal in neuroscience. However, the scarcity of human brain tissue suitable for the study of synapses is one of the most important issues to overcome. Fortunately, healthy human brain tissue that can be studied using electron microscopy is sometimes donated after death. Using these donations could improve the understanding of the synapses in normal brains and possible changes associated with disease. Now, Montero-Crespo et al. have mapped synapses in the normal human hippocampus in three dimensions ­ providing the first detailed description of synaptic structure in this part of the brain. Using high-powered electron microscopes and donated brain tissue samples collected after death, Montero-Crespo et al. imaged almost 25,000 connections between neurons. The analysis showed that synapses were more densely packed in some layers of the hippocampus than in others. Most synapses were found to be connected to tiny dendritic 'spines' that sprout from dendritic branches of the neuron, and they activated (not suppressed) the next neuron. Beyond its implications for better understanding of brain health and disease, this work could also advance computer modelling attempts to mimic the structure of the brain and its activity.

Alterations of the microvascular network in sclerotic hippocampi from patients with epilepsy.

The main hallmarks of human hippocampal sclerosis are neuronal loss and gliosis; reductions in microvasculature labeling in the cornu Ammonis 1 in this condition have been detected using alkaline phosphatase histochemistry. To determine whether the reduction in alkaline phosphatase activity is coupled with a loss of blood vessels,we examined the volume fraction occupied by blood vessels in toluidine blue-stained hippocampal sections from 24 epilepsy patient resections (19 with hippocampal sclerosis, 5 without hippocampal sclerosis) and 5 normal autopsy controls. Light and electron microscopy and immunohistochemistry were used to determine the distribution of collagen Type IV in relation to the fine structure of the hippocampal microvascular network. We found a consistent and highly significant loss of microvessels in the sclerotic hippocampal cornu Ammonis 1 field; a variety of vascular alterations including spinelike protrusions, disruptions, and atrophic branching, were observed in the remaining blood vessels. We suggest that blood vessel alterations are an additional pathological hallmark of hippocampal sclerosis associated with temporal lobe epilepsy and that they may relate to the pathogenesis of this condition.

Hippocampal sclerosis: histopathology substrate and magnetic resonance imaging.

The term hippocampal sclerosis was originally used to describe a shrunken and hardened hippocampus, which histologically displayed neuronal loss and glial proliferation. These alterations are mainly located in the hilus of the dentate gyrus and in the CA1 and CA3 pyramidal cell layers but all hippocampal regions may show neuronal cell loss to varying degrees. A number of morphologic and cytochemical findings are associated with mesial temporal sclerosis, especially within the dentate gyrus. These changes include selective loss of inhibitory interneurons, abnormal sprouting of axons, reorganization of neural transmitter receptors, alterations in second messenger systems, and hyperexcitability of the granule cells. Extrahippocampal pathology is also found at other temporal lobe structures. Frequent extrahippocampal pathology affects the amygdala, first seen with neuronal cell loss and gliosis in the laterobasal complex. Surgical removal of this epileptogenic area can be curative or provide significant reduction in seizure frequency in the majority of individuals. Magnetic resonance imaging (MRI) is highly sensitive in detecting and locating mesial temporal sclerosis when a correct MRI temporal lobe protocol is used. The most important MRI findings, atrophy and abnormal T2 signal, allow us to detect mesial temporal sclerosis in the majority of the cases. Secondary MRI findings help in the diagnosis and lateralization of mesial temporal sclerosis in patients with subtle primary findings and in cases of bilateral hippocampal abnormalities. The development of advanced magnetic resonance (MR) techniques, such as functional MR, diffusion, or transference of magnetization, will lead to greater understanding of this pathology and will improve our diagnostic capacity.

Seeding and transgenic overexpression of alpha-synuclein triggers dendritic spine pathology in the neocortex.

Although misfolded and aggregated α-synuclein (α-syn) is recognized in the disease progression of synucleinopathies, its role in the impairment of cortical circuitries and synaptic plasticity remains incompletely understood. We investigated how α-synuclein accumulation affects synaptic plasticity in the mouse somatosensory cortex using two distinct approaches. Long-term in vivo imaging of apical dendrites was performed in mice overexpressing wild-type human α-synuclein. Additionally, intracranial injection of preformed α-synuclein fibrils was performed to induce cortical α-syn pathology. We find that α-synuclein overexpressing mice show decreased spine density and abnormalities in spine dynamics in an age-dependent manner. We also provide evidence for the detrimental effects of seeded α-synuclein aggregates on dendritic architecture. We observed spine loss as well as dystrophic deformation of dendritic shafts in layer V pyramidal neurons. Our results provide a link to the pathophysiology underlying dementia associated with synucleinopathies and may enable the evaluation of potential drug candidates on dendritic spine pathology in vivo.

High plasticity of axonal pathology in Alzheimer's disease mouse models.

Axonal dystrophies (AxDs) are swollen and tortuous neuronal processes that are associated with extracellular depositions of amyloid ß (Aß) and have been observed to contribute to synaptic alterations occurring in Alzheimer's disease. Understanding the temporal course of this axonal pathology is of high relevance to comprehend the progression of the disease over time. We performed a long-term in vivo study (up to 210 days of two-photon imaging) with two transgenic mouse models (dE9xGFP-M and APP-PS1xGFP-M). Interestingly, AxDs were formed only in a quarter of GFP-expressing axons near Aß-plaques, which indicates a selective vulnerability. AxDs, especially those reaching larger sizes, had long lifetimes and appeared as highly plastic structures with large variations in size and shape and axonal sprouting over time. In the case of the APP-PS1 mouse only, the formation of new long axonal segments in dystrophic axons (re-growth phenomenon) was observed. Moreover, new AxDs could appear at the same point of the axon where a previous AxD had been located before disappearance (re-formation phenomenon). In addition, we observed that most AxDs were formed and developed during the imaging period, and numerous AxDs had already disappeared by the end of this time. This work is the first in vivo study analyzing quantitatively the high plasticity of the axonal pathology around Aß plaques. We hypothesized that a therapeutically early prevention of Aß plaque formation or their growth might halt disease progression and promote functional axon regeneration and the recovery of neural circuits.

Fibrillar amyloid plaque formation precedes microglial activation.

In Alzheimer's disease (AD), hallmark ß-amyloid deposits are characterized by the presence of activated microglia around them. Despite an extensive characterization of the relation of amyloid plaques with microglia, little is known about the initiation of this interaction. In this study, the detailed investigation of very small plaques in brain slices in AD transgenic mice of the line APP-PS1(dE9) revealed different levels of microglia recruitment. Analysing plaques with a diameter of up to 10 µm we find that only the half are associated with clear morphologically activated microglia. Utilizing in vivo imaging of new appearing amyloid plaques in double-transgenic APP-PS1(dE9)xCX3CR1+/- mice further characterized the dynamic of morphological microglia activation. We observed no correlation of morphological microglia activation and plaque volume or plaque lifetime. Taken together, our results demonstrate a very prominent variation in size as well as in lifetime of new plaques relative to the state of microglia reaction. These observations might question the existing view that amyloid deposits by themselves are sufficient to attract and activate microglia in vivo.

Spatial distribution of neurons innervated by chandelier cells.

Chandelier (or axo-axonic) cells are a distinct group of GABAergic interneurons that innervate the axon initial segments of pyramidal cells and are thus thought to have an important role in controlling the activity of cortical circuits. To examine the circuit connectivity of chandelier cells (ChCs), we made use of a genetic targeting strategy to label neocortical ChCs in upper layers of juvenile mouse neocortex. We filled individual ChCs with biocytin in living brain slices and reconstructed their axonal arbors from serial semi-thin sections. We also reconstructed the cell somata of pyramidal neurons that were located inside the ChC axonal trees and determined the percentage of pyramidal neurons whose axon initial segments were innervated by ChC terminals. We found that the total percentage of pyramidal neurons that were innervated by a single labeled ChC was 18-22 %. Sholl analysis showed that this percentage peaked at 22-35 % for distances between 30 and 60 µm from the ChC soma, decreasing to lower percentages with increasing distances. We also studied the three-dimensional spatial distribution of the innervated neurons inside the ChC axonal arbor using spatial statistical analysis tools. We found that innervated pyramidal neurons are not distributed at random, but show a clustered distribution, with pockets where almost all cells are innervated and other regions within the ChC axonal tree that receive little or no innervation. Thus, individual ChCs may exert a strong, widespread influence on their local pyramidal neighbors in a spatially heterogeneous fashion.

Pharmacological inhibition of BACE1 impairs synaptic plasticity and cognitive functions.

BACKGROUND: BACE1 (beta site amyloid precursor protein cleaving enzyme 1) is the rate limiting protease in amyloid ß production, hence a promising drug target for the treatment of Alzheimer's disease. Inhibition of BACE1, as the major ß-secretase in vivo with multiple substrates, however is likely to have mechanism-based adverse effects. We explored the impact of long-term pharmacological inhibition of BACE1 on dendritic spine dynamics, synaptic functions, and cognitive performance of adult mice. METHODS: Sandwich enzyme-linked immunosorbent assay was used to assess Aß40 levels in brain and plasma after oral administration of BACE1 inhibitors SCH1682496 or LY2811376. In vivo two-photon microscopy of the somatosensory cortex was performed to monitor structural dynamics of dendritic spines while synaptic functions and plasticity were measured via electrophysiological recordings of excitatory postsynaptic currents and hippocampal long-term potentiation in brain slices. Finally, behavioral tests were performed to analyze the impact of pharmacological inhibition of BACE1 on cognitive performance. RESULTS: Dose-dependent decrease of Aß40 levels in vivo confirmed suppression of BACE1 activity by both inhibitors. Prolonged treatment caused a reduction in spine formation of layer V pyramidal neurons, which recovered after withdrawal of inhibitors. Congruently, the rate of spontaneous and miniature excitatory postsynaptic currents in pyramidal neurons and hippocampal long-term potentiation were reduced in animals treated with BACE1 inhibitors. These effects were not detected in Bace1(-/-) mice treated with SCH1682496, confirming BACE1 as the pharmacological target. Described structural and functional changes were associated with cognitive deficits as revealed in behavioral tests. CONCLUSIONS: Our findings indicate important functions to BACE1 in structural and functional synaptic plasticity in the mature brain, with implications for cognition.