Daniel A. Nicholson

Reduction in Size of Perforated Postsynaptic Densities in Hippocampal Axospinous Synapses and Age-Related Spatial Learning Impairments

A central problem in the neurobiology of normal aging is why learning is preserved in some aged individuals yet impaired in others. To investigate this issue, we examined whether age-related deficits in spatial learning are associated with a reduction in postsynaptic density (PSD) area in hippocampal excitatory synapses (i.e., with a structural modification that is likely to have a deleterious effect on synaptic function). A hippocampus-dependent version of the Morris water maze task was used to separate Long-Evans male rats into young adult, aged learning-unimpaired, and equally aged learning-impaired groups. Axospinous synapses from the CA1 stratum radiatum were analyzed using systematic random sampling and serial section analyses. We report that aged learning-impaired rats exhibit a marked (∼30%) and significant reduction in PSD area, whereas aged learning-unimpaired rats do not. The observed structural alteration involves a substantial proportion of perforated synapses but is not observed in nonperforated synapses. These findings support the notion that many hippocampal perforated synapses become less efficient in aged learning-impaired rats, which may contribute to cognitive decline during normal aging.

Distance-Dependent Differences in Synapse Number and AMPA Receptor Expression in Hippocampal CA1 Pyramidal Neurons

The ability of synapses throughout the dendritic tree to influence neuronal output is crucial for information processing in the brain. Synaptic potentials attenuate dramatically, however, as they propagate along dendrites toward the soma. To examine whether excitatory axospinous synapses on CA1 pyramidal neurons compensate for their distance from the soma to counteract such dendritic filtering, we evaluated axospinous synapse number and receptor expression in three progressively distal regions: proximal and distal stratum radiatum (SR), and stratum lacunosum-moleculare (SLM). We found that the proportion of perforated synapses increases as a function of distance from the soma and that their AMPAR, but not NMDAR, expression is highest in distal SR and lowest in SLM. Computational models of pyramidal neurons derived from these results suggest that they arise from the compartment-specific use of conductance scaling in SR and dendritic spikes in SLM to minimize the influence of distance on synaptic efficacy.

Differences in the expression of AMPA and NMDA receptors between axospinous perforated and nonperforated synapses are related to the configuration and size of postsynaptic densities

Axospinous synapses are traditionally divided according to postsynaptic density (PSD) configuration into a perforated subtype characterized by a complex‐shaped PSD and nonperforated subtype exhibiting a simple‐shaped, disc‐like PSD. It has been hypothesized that perforated synapses are especially important for synaptic plasticity because they have a higher efficacy of impulse transmission. The aim of the present study was to test this hypothesis. The number of postsynaptic AMPA receptors (AMPARs) is widely regarded as the major determinant of synaptic efficacy. Therefore, the expression of AMPARs was evaluated in the two synaptic subtypes and compared with that of NMDA receptors (NMDARs). Postembedding immunogold electron microscopy was used to quantify the immunoreactivity following single labeling of AMPARs or NMDARs in serial sections through the CA1 stratum radiatum of adult rats. The results showed that all perforated synapses examined were immunopositive for AMPARs. In contrast, only a proportion of nonperforated synapses (64% on average) contained immunogold particles for AMPARs. The number of immunogold particles for AMPARs was markedly and significantly higher in perforated synapses than in immunopositive nonperforated synapses. Although all synapses of both subtypes were NMDAR immunopositive perforated synapses contained significantly more immunogold particles for NMDARs than nonperforated ones. Multivariate analysis of variance revealed that the mode of AMPAR and NMDAR expression is related to the complexity of PSD configuration, not only to PSD size. These findings support the notion that perforated synapses may evoke larger postsynaptic responses relative to nonperforated synapses and, hence, contribute to an enhancement of synaptic transmission associated with some forms of synaptic plasticity. J. Comp. Neurol. 468:86–95, 2004. Published 2003 Wiley‐Liss, Inc.

The Alzheimer's β-secretase BACE1 localizes to normal presynaptic terminals and to dystrophic presynaptic terminals surrounding amyloid plaques.

Abstractβ-Site amyloid precursor protein (APP) cleaving enzyme-1 (BACE1) is the β-secretase that initiates Aβ production in Alzheimer’s disease (AD). BACE1 levels are increased in AD, which could contribute to pathogenesis, yet the mechanism of BACE1 elevation is unclear. Furthermore, the normal function of BACE1 is poorly understood. We localized BACE1 in the brain at both the light and electron microscopic levels to gain insight into normal and pathophysiologic roles of BACE1 in health and AD, respectively. Our findings provide the first ultrastructural evidence that BACE1 localizes to vesicles (likely endosomes) in normal hippocampal mossy fiber terminals of both non-transgenic and APP transgenic (5XFAD) mouse brains. In some instances, BACE1-positive vesicles were located near active zones, implying a function for BACE1 at the synapse. In addition, BACE1 accumulated in swollen dystrophic autophagosome-poor presynaptic terminals surrounding amyloid plaques in 5XFAD cortex and hippocampus. Importantly, accumulations of BACE1 and APP co-localized in presynaptic dystrophies, implying increased BACE1 processing of APP in peri-plaque regions. In primary cortical neuron cultures, treatment with the lysosomal protease inhibitor leupeptin caused BACE1 levels to increase; however, exposure of neurons to the autophagy inducer trehalose did not reduce BACE1 levels. This suggests that BACE1 is degraded by lysosomes but not by autophagy. Our results imply that BACE1 elevation in AD could be linked to decreased lysosomal degradation of BACE1 within dystrophic presynaptic terminals. Elevated BACE1 and APP levels in plaque-associated presynaptic dystrophies could increase local peri-plaque Aβ generation and accelerate amyloid plaque growth in AD.

Inactivation of the microRNA-183/96/182 cluster results in syndromic retinal degeneration

The microRNA-183/96/182 cluster is highly expressed in the retina and other sensory organs. To uncover its in vivo functions in the retina, we generated a knockout mouse model, designated “miR-183CGT/GT,” using a gene-trap embryonic stem cell clone. We provide evidence that inactivation of the cluster results in early-onset and progressive synaptic defects of the photoreceptors, leading to abnormalities of scotopic and photopic electroretinograms with decreased b-wave amplitude as the primary defect and progressive retinal degeneration. In addition, inactivation of the miR-183/96/182 cluster resulted in global changes in retinal gene expression, with enrichment of genes important for synaptogenesis, synaptic transmission, photoreceptor morphogenesis, and phototransduction, suggesting that the miR-183/96/182 cluster plays important roles in postnatal functional differentiation and synaptic connectivity of photoreceptors.

Synapse distribution suggests a two-stage model of dendritic integration in CA1 pyramidal neurons.

Competing models have been proposed to explain how neurons integrate the thousands of inputs distributed throughout their dendritic trees. In a simple global integration model, inputs from all locations sum in the axon. In a two-stage integration model, inputs contribute directly to dendritic spikes, and outputs from multiple branches sum in the axon. These two models yield opposite predictions of how synapses at different dendritic locations should be scaled if they are to contribute equally to neuronal output. We used serial-section electron microscopy to reconstruct individual apical oblique dendritic branches of CA1 pyramidal neurons and observe a synapse distribution consistent with the two-stage integration model. Computational modeling suggests that the observed synapse distribution enhances the contribution of each dendritic branch to neuronal output.

Synapses with a segmented, completely partitioned postsynaptic density express more AMPA receptors than other axospinous synaptic junctions

Axospinous perforated synapses of one morphological subtype exhibit multiple transmission zones, each one being formed by an axon terminal protrusion apposing a postsynaptic density (PSD) segment and separated from others by complete spine partitions. Such segmented, completely partitioned (SCP) synapses have been implicated in synaptic plasticity and postulated to be exceptionally efficacious. The present study explored the validity of this supposition. Postembedding immunogold electron microscopy was used for quantifying the postsynaptic AMPA receptor (AMPAR) expression, which is widely regarded as a major determinant of synaptic efficacy. Various subtypes of axospinous synapses were examined in the rat CA1 stratum radiatum. The results showed that the number of immunogold particles for AMPARs in SCP synapses markedly and significantly exceeded that in other perforated subtypes (by 101% on the average) and in nonperforated immunopositive synapses (by 1086%). Moreover, the particle number per single PSD segment, each of which also contained NMDA receptors, was significantly higher than that per nonperforated PSD (by 485%). SCP synapses also exhibited a higher particle density per unit PSD area, as well as a larger overall PSD area as compared with other synaptic subtypes. Analysis of covariance revealed that the high AMPAR expression in SCP synapses was related to the segmented PSD configuration, not only to the PSD size. Moreover, the subpopulations of SCP and other perforated synapses with either overlapping or equal PSD sizes differed in AMPAR content and concentration, with both measures being significantly higher in SCP synapses. Thus, the elevated AMPAR expression in SCP synapses is associated with the presence of separate PSD segments, not only with their large PSD area. These findings are consistent with the idea that SCP synapses have a relatively greater efficacy and may support maximal levels of synaptic enhancement characteristic of certain forms of synaptic plasticity such as the early LTP phase.

Identification of TMEM230 mutations in familial Parkinson's disease

Parkinsons disease is the second most common neurodegenerative disorder without effective treatment. It is generally sporadic with unknown etiology. However, genetic studies of rare familial forms have led to the identification of mutations in several genes, which are linked to typical Parkinsons disease or parkinsonian disorders. The pathogenesis of Parkinsons disease remains largely elusive. Here we report a locus for autosomal dominant, clinically typical and Lewy body–confirmed Parkinsons disease on the short arm of chromosome 20 (20pter-p12) and identify TMEM230 as the disease-causing gene. We show that TMEM230 encodes a transmembrane protein of secretory/recycling vesicles, including synaptic vesicles in neurons. Disease-linked TMEM230 mutants impair synaptic vesicle trafficking. Our data provide genetic evidence that a mutant transmembrane protein of synaptic vesicles in neurons is etiologically linked to Parkinsons disease, with implications for understanding the pathogenic mechanism of Parkinsons disease and for developing rational therapies.

Deletion of the Hyperpolarization-Activated Cyclic Nucleotide-Gated Channel Auxiliary Subunit TRIP8b Impairs Hippocampal Ih Localization and Function and Promotes Antidepressant Behavior in Mice

Output properties of neurons are greatly shaped by voltage-gated ion channels, whose biophysical properties and localization within axodendritic compartments serve to significantly transform the original input. The hyperpolarization-activated current, Ih, is mediated by hyperpolarization-activated cyclic nucleotide-gated (HCN) channels and plays a fundamental role in influencing neuronal excitability by regulating both membrane potential and input resistance. In neurons such as cortical and hippocampal pyramidal neurons, the subcellular localization of HCN channels plays a critical functional role, yet mechanisms controlling HCN channel trafficking are not fully understood. Because ion channel function and localization are often influenced by interacting proteins, we generated a knock-out mouse lacking the HCN channel auxiliary subunit, tetratricopeptide repeat-containing Rab8b-interacting protein (TRIP8b). Eliminating expression of TRIP8b dramatically reduced Ih expression in hippocampal pyramidal neurons. Loss of Ih-dependent membrane voltage properties was attributable to reduction of HCN channels on the neuronal surface, and there was a striking disruption of the normal expression pattern of HCN channels in pyramidal neuron dendrites. In heterologous cells and neurons, absence of TRIP8b increased HCN subunit targeting to and degradation by lysosomes. Mice lacking TRIP8b demonstrated motor learning deficits and enhanced resistance to multiple tasks of behavioral despair with high predictive validity for antidepressant efficacy. We observed similar resistance to behavioral despair in distinct mutant mice lacking HCN1 or HCN2. These data demonstrate that interaction with the auxiliary subunit TRIP8b is a major mechanism underlying proper expression of HCN channels and Ih in vivo, and suggest that targeting Ih may provide a novel approach to treatment of depression.

Impingement syndrome: Temporal outcomes of nonoperative treatment

HYPOTHESIS We prospectively studied patients with impingement syndrome to look at outcomes of nonoperative treatment on a temporal basis. MATERIALS AND METHODS Temporal outcomes of 100 consecutive patients treated for impingement syndrome were prospectively evaluated. All patients began a standardized, nonoperative treatment protocol consisting of a subacromial steroid injection, followed by physical therapy. RESULTS Data were available on 94 patients at the final two-year follow-up assessment. Overall, 74 of 94 patients did not require surgery. In that group, the average American Shoulder and Elbow Surgeons (ASES) outcome score increased from 56 to 95, with an average decrease in the pain score from 4.8 to 0.6. Improvement was demonstrated in patient shoulder outcome scores (ASES score) and visual analog pain scores between treatment initiation and the one-year follow-up assessment (p < .0001); no improvement was identified past one year. Of the non-surgical patients, 22 continued to have some shoulder pain. CONCLUSION Of patients with impingement syndrome treated nonoperatively, 79% did not require surgery after two-year follow-up. Predictors of patients going on to surgical intervention included the total number of subacromial steroid/lidocaine injections and patient response to the initial subacromial injection. Of the patients not undergoing surgery, 30% continued to have some shoulder pain. LEVEL OF EVIDENCE Level 1; Prospective prognosis study, >80% follow-up.