Presynaptic Bouton Density Research Articles

Mice deficient in synaptic protease neurotrypsin show impaired spaced long-term potentiation and blunted learning-induced modulation of dendritic spines

Neurotrypsin (NT) is a neuronal trypsin-like serine protease whose mutations cause severe mental retardation in humans. NT is activated in vitro by Hebbian-like conjunction of pre- and postsynaptic activities, which promotes the formation of dendritic filopodia via proteolytic cleavage of the proteoglycan agrin. Here, we investigated the functional importance of this mechanism for synaptic plasticity, learning, and extinction of memory. We report that juvenile neurotrypsin-deficient (NT−/−) mice exhibit impaired long-term potentiation induced by a spaced stimulation protocol designed to probe the generation of new filopodia and their conversion into functional synapses. Behaviorally, juvenile NT−/− mice show impaired contextual fear memory and have a sociability deficit. The latter persists in aged NT−/− mice, which, unlike juvenile mice, show normal recall but impaired extinction of contextual fear memories. Structurally, juvenile mutants exhibit reduced spine density in the CA1 region, fewer thin spines, and no modulation in the density of dendritic spines following fear conditioning and extinction in contrast to wild-type littermates. The head width of thin spines is reduced in both juvenile and aged NT−/− mice. In vivo delivery of adeno-associated virus expressing an NT-generated fragment of agrin, agrin-22, but not a shorter agrin-15, elevates the spine density in NT−/− mice. Moreover, agrin-22 co-aggregates with pre- and postsynaptic markers and increases the density and size of presynaptic boutons and presynaptic puncta, corroborating the view that agrin-22 supports the synaptic growth.

Alzheimer's Disease-Like Neurodegeneration in Porphyromonas gingivalis Infected Neurons with Persistent Expression of Active Gingipains.

Background:Porphyromonas gingivalis (P. gingivalis) and its gingipain virulence factors have been identified as pathogenic effectors in Alzheimer’s disease (AD). In a recent study we demonstrated the presence of gingipains in over 90% of postmortem AD brains, with gingipains localizing to the cytoplasm of neurons. However, infection of neurons by P. gingivalis has not been previously reported.Objective:To demonstrate intraneuronal P. gingivalis and gingipain expression in vitro after infecting neurons derived from human inducible pluripotent stem cells (iPSC) with P. gingivalis for 24, 48, and 72 h.Methods:Infection was characterized by transmission electron microscopy, confocal microscopy, and bacterial colony forming unit assays. Gingipain expression was monitored by immunofluorescence and RT-qPCR, and protease activity monitored with activity-based probes. Neurodegenerative endpoints were assessed by immunofluorescence, western blot, and ELISA.Results:Neurons survived the initial infection and showed time dependent, infection induced cell death. P. gingivalis was found free in the cytoplasm or in lysosomes. Infected neurons displayed an accumulation of autophagic vacuoles and multivesicular bodies. Tau protein was strongly degraded, and phosphorylation increased at T231. Over time, the density of presynaptic boutons was decreased.Conclusion:P. gingivalis can invade and persist in mature neurons. Infected neurons display signs of AD-like neuropathology including the accumulation of autophagic vacuoles and multivesicular bodies, cytoskeleton disruption, an increase in phospho-tau/tau ratio, and synapse loss. Infection of iPSC-derived mature neurons by P. gingivalis provides a novel model system to study the cellular mechanisms leading to AD and to investigate the potential of new therapeutic approaches.

Multiple Morphological Factors Underlie Experience-Dependent Cross-Modal Plasticity in the Developing Sensory Cortices.

Sensory experience regulates the structural and functional wiring of sensory cortices. In previous work, we showed that whisker deprivation (WD) from birth not only reduced excitatory synaptic transmission of layer (L) 2/3 pyramidal neurons of the correspondent barrel cortex in mice, but also cross-modally reduced synaptic transmission of L2/3 pyramidal neurons in other sensory cortices. Here, we used in utero electroporation, in combination with optical clearing, to examine the main morphological components regulating neural circuit wiring, namely presynaptic bouton density, spine density, as well as dendrite and axon arbor lengths. We found that WD from P0 to P14 reduced presynaptic bouton density in both L4 and L2/3 inputs to L2/3 pyramidal neurons, as well as spine density across the dendritic tree of L2/3 pyramidal neurons, in the barrel field of the primary somatosensory cortex. The cross-modal effects in the primary auditory cortex were manifested mostly as reduced dendrite and axon arbor size, as well as reduced bouton density of L2/3 inputs. Increasing sensory experience by rearing mice in an enriched environment rescued the effects of WD. Together, these results demonstrate that multiple morphological factors contribute to experience-dependent structural plasticity during early wiring of the sensory cortices.

Fornix deep brain stimulation induces reduction of hippocampal synaptophysin levels

Fornix deep brain stimulation (DBS) has the ability to refurbish memory functions in animal models with experimental dementia. One of the possible underlying mechanisms is the acute increase of acetylcholine in the hippocampus. Another suggested hypothesis is neuroplasticity. Recent work in rats has shown that acute fornix DBS can modulate neurotrophic factors as well as synaptic plasticity markers on the short-term. Here, we want to test the hypothesis that acute fornix DBS can also lead to long-term effects on neuroplasticity. Rats received DBS at 100 Hz, 100 μA and 100 μs pulse width for 4 h with electrodes placed bilaterally in the fornix. Seven weeks after stimulation, rats were sacrificed. BDNF, p-CREB, SV2 and synaptophysin immunohistochemistry was performed for their brains. No differences were found in the number of BDNF, p-CREB or SV2 positive cells for fornix DBS rats when compared to sham. Surprisingly, the density of synaptophysin immunoreactive presynaptic boutons was significantly decreased in the CA1 and CA3 subregion of the hippocampus for DBS rats. Therefore, fornix DBS might induce long-term depression related mechanisms.

Semaphorin 4D promotes inhibitory synapse formation and suppresses seizures in vivo.

We previously discovered a role for the extracellular domain of the transmembrane protein semaphorin 4D (Sema4D) as a fast-acting, selective, and positive regulator of functional γ-aminobutyric acid (GABA)ergic synapse formation in hippocampal neuronal culture. We also demonstrated that Sema4D treatment increases inhibitory tone and suppresses hyperexcitability in an organotypic hippocampal slice culture model of epilepsy. Here, we investigate the ability of Sema4D to promote GABAergic synapse formation and suppress seizure activity in vivo in adult mice. We performed a 3-hour, intrahippocampal infusion of Sema4D or control protein into the CA1 region of adult mice. To quantify GABAergic presynaptic bouton density, we performed immunohistochemistry on hippocampal tissue sections isolated from these animals using an antibody that specifically recognizes the glutamic acid decarboxylase isoform 65 protein (GAD65), which is localized to presynaptic GABAergic boutons. To assess seizure activity, we employed 2 in vivo mouse models of epilepsy, intravenous (iv) pentylenetetrazol (PTZ) and hippocampal electrical kindling, in the presence or absence of Sema4D treatment. We monitored seizure activity by behavioral observation or electroencephalography (EEG). To assay the persistence of the Sema4D effect, we monitored seizure activity and measured the density of GAD65-positive presynaptic boutons 3 or 48 hours after Sema4D infusion. Sema4D-treated mice displayed an elevated density of GABAergic presynaptic boutons juxtaposed to hippocampal pyramidal neuron cell bodies, consistent with the hypothesis that Sema4D promotes the formation of new inhibitory synapses in vivo. In addition, Sema4D acutely suppressed seizures in both the PTZ and electrical kindling models. When we introduced a 48-hour gap between Sema4D treatment and the seizure stimulus, seizure activity was indistinguishable from controls. Moreover, immunohistochemistry on brain sections or hippocampal slices isolated 3 hours, but not 48 hours, after Sema4D treatment displayed an increase in GABAergic bouton density, demonstrating temporal correlation between the effects of Sema4D on seizures and GABAergic synaptic components. Our findings suggest a novel approach to treating acute seizures: harnessing synaptogenic molecules to enhance connectivity in the inhibitory network.

Effects of early nerve repair on experimental brachial plexus injury in neonatal rats.

Obstetrical brachial plexus injury refers to injury observed at the time of delivery, which may lead to major functional impairment in the upper limb. In this study, the neuroprotective effect of early nerve repair following complete brachial plexus injury in neonatal rats was examined. Brachial plexus injury induced 90% loss of spinal motoneurons and 70% decrease in biceps muscle weight at 28 days after injury. Retrograde degeneration in spinal cord was associated with decreased density of dendritic branches and presynaptic boutons and increased density of astrocytes and macrophages/microglial cells. Early repair of the injured brachial plexus significantly delayed retrograde degeneration of spinal motoneurons and reduced the degree of macrophage/microglial reaction but had no effect on muscle atrophy. The results demonstrate that early nerve repair of neonatal brachial plexus injury could promote survival of injured motoneurons and attenuate neuroinflammation in spinal cord.

Presynaptic mitochondrial morphology in monkey prefrontal cortex correlates with working memory and is improved with estrogen treatment.

Humans and nonhuman primates are vulnerable to age- and menopause-related decline in working memory, a cognitive function reliant on the energy-demanding recurrent excitation of neurons within Brodmann's Area 46 of the dorsolateral prefrontal cortex (dlPFC). Here, we tested the hypothesis that the number and morphology (straight, curved, or donut-shaped) of mitochondria in dlPFC presynaptic boutons are altered with aging and menopause in rhesus monkeys (Macaca mulatta) and that these metrics correlate with delayed response (DR) accuracy, a well-characterized measure of dlPFC-dependent working memory. Although presynaptic bouton density or size was not significantly different across groups distinguished by age or menses status, DR accuracy correlated positively with the number of total and straight mitochondria per dlPFC bouton. In contrast, DR accuracy correlated inversely with the frequency of boutons containing donut-shaped mitochondria, which exhibited smaller active zone areas and fewer docked synaptic vesicles than those with straight or curved mitochondria. We then examined the effects of estrogen administration to test whether a treatment known to improve working memory influences mitochondrial morphology. Aged ovariectomized monkeys treated with vehicle displayed significant working memory impairment and a concomitant 44% increase in presynaptic donut-shaped mitochondria, both of which were reversed with cyclic estradiol treatment. Together, our data suggest that hormone replacement therapy may benefit cognitive aging, in part by promoting mitochondrial and synaptic health in the dlPFC.

Altered synapses and gliotransmission in Alzheimer's disease and AD model mice

Amyloid-β (Aβ) plaque accumulation in Alzheimer's disease (AD) is associated with glutamatergic synapse loss, but less is known about its effect on inhibitory synapses. Here, we demonstrate that vesicular γ-aminobutyric acid (GABA) transporter (VGAT) presynaptic bouton density is unaffected in human preclinical and end-stage AD and in APP/PS1 transgenic (TG) mice. Conversely, excitatory vesicular glutamate transporter 1 (VGlut1) boutons are significantly reduced in end-stage AD cases and less reduced in preclinical AD cases and TGs. Aged TGs also show reduced protein levels of VGlut1 and synaptophysin but not VGAT or glutamate decarboxylase (GAD). These findings indicate that GABAergic synapses are preserved in human AD and mouse TGs. Synaptosomes isolated from plaque-rich TG cortex had significantly higher GAD activity than those from plaque-free cerebellum or the cortex of wild-type littermates. Using tissue fractionation, this increased activity was localized to glial synaptosomes, suggesting that Aβ plaques stimulate increased astrocyte GABA synthesis.

Peripheral Nerve Injury Leads to Working Memory Deficits and Dysfunction of the Hippocampus by Upregulation of TNF-α in Rodents

Patients with chronic pain usually suffer from working memory deficits, which may decrease their intellectual ability significantly. Despite intensive clinical studies, the mechanism underlying this form of memory impairment remains elusive. In this study, we investigated this issue in the spared nerve injury (SNI) model of neuropathic pain, a most common form of chronic pain. We found that SNI impaired working memory and short-term memory in rats and mice. To explore the potential mechanisms, we studied synaptic transmission/plasticity in hippocampus, a brain region critically involved in memory function. We found that frequency facilitation, a presynaptic form of short-term plasticity, and long-term potentiation at CA3-CA1 synapses were impaired after SNI. Structurally, density of presynaptic boutons in hippocampal CA1 synapses was reduced significantly. At the molecular level, we found that tumor necrosis factor-α (TNF-α) increased in cerebrospinal fluid, in hippocampal tissue and in plasma after SNI. Intracerebroventricular or intrahippocampal injection of recombinant rat TNF mimicked the effects of SNI in naive rats, whereas inhibition of TNF-α or genetic deletion of TNF receptor 1 prevented both memory deficits and synaptic dysfunction induced by SNI. As TNF-α is critical for development of neuropathic pain, we suggested that the over-production of TNF-α following peripheral nerve injury might lead to neuropathic pain and memory deficits, simultaneously.

Quantal Analysis Reveals a Functional Correlation between Presynaptic and Postsynaptic Efficacy in Excitatory Connections from Rat Neocortex

At many central synapses, the presynaptic bouton and postsynaptic density are structurally correlated. However, it is unknown whether this correlation extends to the functional properties of the synapses. To investigate this, we made recordings from synaptically coupled pairs of pyramidal neurons in rat visual cortex. The mean peak amplitude of EPSPs recorded from pairs of L2/3 neurons ranged between 40 microV and 2.9 mV. EPSP rise times were consistent with the majority of the synapses being located on basal dendrites; this was confirmed by full anatomical reconstructions of a subset of connected pairs. Over a third of the connections could be described using a quantal model that assumed simple binomial statistics. Release probability (P(r)) and quantal size (Q), as measured at the somatic recording site, showed considerable heterogeneity between connections. However, across the population of connections, values of P(r) and Q for individual connections were positively correlated with one another. This correlation also held for inputs to layer 5 pyramidal neurons from both layer 2/3 and neighboring layer 5 pyramidal neurons, suggesting that during development of cortical connections presynaptic and postsynaptic strengths are dependently scaled. For 2/3 to 2/3 connections, mean EPSP amplitude was correlated with both Q and P(r) values but uncorrelated with N, the number of functional release sites mediating the connection. The efficacy of a cortical connection is thus set by coordinated presynaptic and postsynaptic strength.

Fetal asphyxia leads to the loss of striatal presynaptic boutons in adult rats

Fetal asphyxic insults in the brain are known to be associated with developmental and neurological problems like neuromotor disorders and cognitive deficits. Little is known, however, about the long-term consequences of fetal asphyxia contributing to the development of different neurological diseases common in the adult or the aging brain. For that reason the present study aimed to investigate the long-term effects of fetal asphyxia on synaptic organization within the adult rat brain. Fetal asphyxia was induced at embryonic day 17 by 75-min clamping of the uterine and ovarian arteries. Presynaptic bouton densities and numbers were analyzed in the striatum and prefrontal cortex at the age of 19 months. A substantial decrease in presynaptic bouton density and number was observed in the striatum of fetal asphyxia rats compared to control rats, while an increase was found in the fifth layer of the prefrontal cortex. These results suggest that fetal asphyxia can have long-lasting effects on synaptic organization that might contribute to a developmental etiology of different neurological disorders and aging.

Cognitive impairment and transmitter‐specific pre‐ and postsynaptic changes in the rat cerebral cortex during ageing

Recent studies suggest that age-related cognitive decline is correlated with an excitatory-inhibitory imbalance in synaptic discharges on pyramidal neurons. This study focuses on whether ageing and cognitive status correlates with relative numbers of excitatory and inhibitory presynaptic boutons. We investigated the density of excitatory and inhibitory presynaptic inputs across several areas of the rat cerebral cortex in young and aged male Fischer 344 rats. Aged animals were segregated into aged cognitively impaired (AI) and aged cognitively unimpaired (AU) groups using the Morris water maze. We applied immunohistochemistry to reveal the majority of excitatory and inhibitory presynaptic boutons captured with confocal microscopy and quantitative image analysis. A gradual decline in the density of excitatory and inhibitory presynaptic boutons occurred from young to AU to AI animals; however, the ratios of excitatory to inhibitory presynaptic bouton densities were not significantly altered. We further investigated the density of receptor scaffolding proteins representing key excitatory and inhibitory receptor postsynaptic sites, using antibodies against specific markers of excitatory and inhibitory postsynaptic densities, respectively. Significant changes in the ratios of excitatory to inhibitory postsynaptic densities were observed only in AI compared to young rats.

Paradoxical Upregulation of Glutamatergic Presynaptic Boutons during Mild Cognitive Impairment

Synaptic integrity is now recognized as a central component of Alzheimer's disease. Surprisingly, however, the structural status of glutamatergic synapses in Alzheimer's disease is unclear, despite the fact that glutamate is the major excitatory transmitter of the CNS and has key roles in excitotoxicity and long-term potentiation. The identification of specific markers of glutamatergic neurons now allows an assessment of the structural involvement of the glutamatergic system across progressive stages of the Alzheimer's pathology, an opportunity not afforded by previously used neurochemical approaches. Glutamatergic presynaptic bouton density and dystrophic neurite abundance were quantified in midfrontal gyrus brain tissue from subjects with no cognitive impairment, mild cognitive impairment, or mild- or severe-stage Alzheimer's disease. Our study demonstrates a striking pathology-dependent pattern of glutamatergic synaptic remodeling with disease progression. Subjects with mild cognitive impairment display a paradoxical elevation in glutamatergic presynaptic bouton density, a situation akin to that observed in the cholinergic system, which then depletes and drops with disease progression. This pattern of synaptic remodeling mirrors our previous findings in transgenic animal models and is of major relevance to current transmitter-based therapeutics.

LDL receptor deficiency results in decreased cell proliferation and presynaptic bouton density in the murine hippocampus

An aberrant cholesterol metabolism in the brain may contribute to the pathogenesis of Alzheimer's disease (AD). The LDL receptor (LDLR) regulates plasma cholesterol levels and recently we and others obtained evidence that it is also involved in regulating brain cholesterol homeostasis. Moreover, we found that LDLR-deficient mice display impaired spatial memory. Because cholesterol, in part derived from cellular uptake via LDLR, is required for peripheral cell proliferation and growth, we examined the effect of absence of the LDLR on hippocampal proliferation and the density of synaptic connections. Mice deficient for the LDLR displayed a reduced number of proliferating (BrdU-labeled) cells in the hippocampus as compared to wild type control mice. In addition, the number of synaptophysin-immunoreactive presynaptic boutons in the hippocampal CA1 and the dentate gyrus (DG) areas, but not in cortical areas, was lower in the LDLR-knockout mice than in the control mice. In vitro experiments showed that LDLR activity is increased when cell growth is enhanced by the addition of N2 supplement. This further supports a role for the LDLR in the outgrowth of neurites. These findings support the notion that, similar to its role in the periphery, the LDLR is important for the cellular uptake of cholesterol in the brain and that disturbance of this process affects neuronal plasticity.

ADAM-10 over-expression increases cortical synaptogenesis

Cortical cholinergic, glutamatergic and GABAergic terminals become upregulated during early stages of the transgenic amyloid pathology. Abundant evidence suggests that sAPPα, the product of the non-amyloidogenic α-secretase pathway, is neurotrophic both in vitro and when exogenously applied in vivo. The disintegrin metalloprotease ADAM-10 has been shown to have α-secretase activity in vivo. To determine whether sAPPα has an endogenous biological influence on cortical presynaptic boutons in vivo, we quantified cortical cholinergic, glutamatergic and GABAergic presynaptic bouton densities in either ADAM-10 moderate expressing (ADAM-10 mo) transgenic mice, which moderately overexpress ADAM-10, or age-matched non-transgenic controls. Both early and late ontogenic time points were investigated. ADAM-10 mo transgenic mice display significantly elevated cortical cholinergic, glutamatergic and GABAergic presynaptic bouton densities at the early time point (8 months). Only the cholinergic presynaptic bouton density remains significantly elevated in late-staged ADAM-10 mo transgenic animals (18 months). To confirm that the observed elevations were due to increased levels of endogenous murine sAPPα, exogenous human sAPPα was infused into the cortex of non-transgenic control animals for 1 week. Exogenous infusion of sAPPα led to significant elevations in the cholinergic, glutamatergic and GABAergic cortical presynaptic bouton populations. These results are the first to demonstrate an in vivo influence of ADAM-10 on neurotransmitter-specific cortical synaptic plasticity and further confirm the neurotrophic influence of sAPPα on cortical synaptogenesis.

P3-443: Glutamatergic presynaptic bouton density is elevated in MCI, lowered in late ad and correlates to MMSE

P3-443: Glutamatergic presynaptic bouton density is elevated in MCI, lowered in late ad and correlates to MMSE

Circulating insulin‐like growth factor I and functional recovery from spinal cord injury under enriched housing conditions

Voluntary locomotor training as induced by enriched housing of rats stimulates recovery of locomotion after spinal cord injury (SCI). Generally it is thought that spinal neural networks of motor- and interneurons located in the ventral and intermediate laminae within the lumbar intumescence of the spinal cord, also referred to as central pattern generators (CPGs), are the 'producers of locomotion' and play a pivotal role in the amelioration of locomotor deficits after SCI. It has been suggested that locomotor training provides locomotor-specific sensory feedback into the CPGs, which stimulates remodeling of central nervous system pathways, including motor systems. Several molecules have been proposed to potentiate this process but the underlying mechanisms are not yet known. To understand these mechanisms, we studied the role of insulin-like growth factor (IGF) I in functional recovery from SCI under normal and enriched environment (EE) housing conditions. In a first experiment, we discovered that subcutaneous administration of IGF-I resulted in better locomotor recovery following SCI. In a second experiment, detailed analysis of the observed functional recovery induced by EE revealed full recovery of hindlimb coordination and stability of gait. This EE-dependent functional recovery was attenuated by alterations in the pre-synaptic bouton density within the ventral gray matter of the lumbar intumescence or CPG area. Neutralization of circulating IGF-I significantly blocked the effectiveness of EE housing on functional recovery and diminished the EE-induced alterations in pre-synaptic bouton density within the CPG area. These results support the use of IGF-I as a possible therapeutic aid in early rehabilitation after SCI.

Structural involvement of the glutamatergic presynaptic boutons in a transgenic mouse model expressing early onset amyloid pathology

While the cholinergic depletion in Alzheimer's disease (AD) has been known for some time, a definitive involvement of other neurotransmitter systems has been somewhat more elusive. Our study demonstrates a clear involvement of both glutamatergic and, to a lesser extent, GABAergic neurons in an early onset transgenic mouse model of AD-like amyloid pathology. Immunohistochemical staining and subsequent quantification has revealed a statistically significant increased density of glutamatergic and GABAergic presynaptic boutons in both the plaque free and plaque adjacent cortical neuropile areas of transgenic mice as compared to non-transgenic controls. Furthermore, amyloid plaque size was shown to have a statistically significant effect on the relative area occupied by dystrophic glutamatergic neurites in the peri-plaque neuropile. These findings support our hypothesis that the amyloid pathology progresses in a time and neurotransmitter specific manner, first in the cholinergic system which appears to be most vulnerable, followed by the glutamatergic presynaptic boutons and finally the somewhat more resilient GABAergic terminals.

The impact of Aβ-plaques on cortical cholinergic and non-cholinergic presynaptic boutons in alzheimer's disease-like transgenic mice

A previous study in our laboratory, involving early stage, amyloid pathology in 8-month-old transgenic mice, demonstrated a selective loss of cholinergic terminals in the cerebral and hippocampal cortices of doubly transgenic (APP K670N,M671L+PSl M146L) mice, an up-regulation in the single mutant APP K670N,M671L mice and no detectable change in the PSl M146L transgenics [J Neurosci 19 (1999) 2706]. The present study investigates the impact of amyloid plaques on synaptophysin and vesicular acetylcholine transporter (VAChT) immunoreactive bouton numbers in the frontal cortex of the three transgenic mouse models previously described. When compared as a whole, the frontal cortices of transgenic and control mice show no observable differences in the densities of synaptophysin-immunoreactive boutons. An individual comparison of layer V of the frontal cortex, however, shows a significant increase in density in transgenic models. Analysis of the cholinergic system alone shows significant alterations in the VAChT-immunoreactive bouton densities as evidenced by an increased density in the single (APP K670N,M671L) transgenics and a decreased density in the doubly transgenics (APP K670N,M671L+PSl M146L). In investigating the impact of plaque proximity on bouton density at early stages of the amyloid pathology in our doubly (APP K670N,M671L+PSl M146L) transgenic mouse line, we observed that plaque proximity reduced cholinergic pre-synaptic bouton density by 40%, and yet increased synaptophysin-immunoreactive pre-synaptic bouton density by 9.5%. Distance from plaques (up to 60 μm) seemed to have no effect on bouton density; however a significant inverse relationship was visible between plaque size and cholinergic pre-synaptic bouton density. Finally, the number of cholinergic dystrophic neurites surrounding the truly amyloid, Thioflavin-S + plaque core, was disproportionately large with respect to the incidence of cholinergic boutons within the total pre-synaptic bouton population. Confocal and electron microscopic observations confirmed the preferential infiltration of dystrophic cholinergic boutons into fibrillar amyloid aggregates. We therefore hypothesize that extracellular Aβ aggregation preferentially affects cholinergic terminations prior to progression onto other neurotransmitter systems. This is supported by the observable presence of non-cholinergic sprouting, which may be representative of impending neuritic degeneration.

Perinatal asphyxia results in changes in presynaptic bouton number in striatum and cerebral cortex — a stereological and behavioral analysis

Deficits in cognitive function have been related to quantitative changes in synaptic population, particularly in the cerebral cortex. Here, we used an established model of perinatal asphyxia that induces morphological changes, i.e. neuron loss in the cerebral cortex and striatum, as well as behavioural deficits. We hypothesized that perinatal asphyxia may lead to a neurodegenerative process resulting in cognitive impairment and altered presynaptic bouton numbers in adult rats. We studied cognitive performance at 18 months and presynaptic bouton numbers at 22 months following perinatal asphyxia. Data of the spatial Morris water escape task did not reveal clear memory or learning deficits in aged asphyctic rats compared to aged control rats. However, a memory impairment in aged rats versus young rats was observed, which was more pronounced in asphyctic rats. We found an increase in presynaptic bouton density in the parietal cortex, whereas no changes were found in striatum and frontal cortex in asphyctic rats. An increase of striatal volume was observed in asphyctic rats, leading to an increase in presynaptic bouton numbers in this area. These findings stress the issue that volume measurements have to be taken into account when determining presynaptic bouton density. Furthermore, perinatal asphyxia led to region-specific changes in presynaptic bouton numbers and it worsened the age-related cognitive impairment. These results suggest that perinatal asphyxia induced neuronal loss, which is compensated for by an increase in presynaptic bouton numbers.