A comprehensive knowledge base of synaptic electrophysiology in the rodent hippocampal formation

The cellular and synaptic architecture of the rodent hippocampus has been described in thousands of peer‐reviewed publications. However, no human‐ or machine‐readable public catalog of synaptic electrophysiology data exists for this or any other neural system. Harnessing state‐of‐the‐art information technology, we have developed a cloud‐based toolset for identifying empirical evidence from the scientific literature pertaining to synaptic electrophysiology, for extracting the experimental data of interest, and for linking each entry to relevant text or figure excerpts. Mining more than 1,200 published journal articles, we have identified eight different signal modalities quantified by 90 different methods to measure synaptic amplitude, kinetics, and plasticity in hippocampal neurons. We have designed a data structure that both reflects the differences and maintains the existing relations among experimental modalities. Moreover, we mapped every annotated experiment to identified potential connections, that is, specific pairs of presynaptic and postsynaptic neuron types. To this aim, we leveraged Hippocampome.org , an open‐access knowledge base of morphologically, electrophysiologically, and molecularly characterized neuron types in the rodent hippocampal formation. Specifically, we have implemented a computational pipeline to systematically translate neuron type properties into formal queries in order to find all compatible potential connections. With this system, we have collected nearly 40,000 synaptic data entities covering 88% of the 3,120 potential connections in Hippocampome.org . Correcting membrane potentials with respect to liquid junction potentials significantly reduced the difference between theoretical and experimental reversal potentials, thereby enabling the accurate conversion of all synaptic amplitudes to conductance. This data set allows for large‐scale hypothesis testing of the general rules governing synaptic signals. To illustrate these applications, we confirmed several expected correlations between synaptic measurements and their covariates while suggesting previously unreported ones. We release all data open‐source at Hippocampome.org in order to further research across disciplines.     

Glutamate transporter 1-expressing glia in the rat substantia nigra—Morphometric analysis and relationships to synapses

Glial cells have a major role in protecting neurons against various forms of stress. Especially, astrocytes mediate the bulk of glutamate clearance in the brain via specific membrane transporters (GLAST and GLT1), thereby preventing the occurrence of excitotoxic events. Although glutamate-mediated mechanisms are thought to contribute to nigral dopaminergic neuron degeneration in Parkinson's disease, detailed information on the organization of glia in the substantia nigra is still lacking. The present study was performed to provide quantitative information on the organization of astroglia and on the relationships between astrocytes and excitatory synapses in the rat substantia nigra. Using immunolabeling of GLT1 and confocal imaging, we found that the substantia nigra was filled with a dense meshwork of immunoreactive astrocyte processes. Stereological analysis performed on electron microscope images revealed that the density of immunoreactive astrocyte plasma membranes was substantial, close to 1 μm²/μm³, in the substantia nigra neuropil, both in the pars compacta and the pars reticulata. Excitatory synapses had on average two thirds of their perimeters free from glia, a disposition that may favor transmitter spillover. The density of glutamatergic synapses, as quantified on confocal images by the simultaneous detection of bassoon and of vesicular glutamate transporter 1 or 2, was very low (0.01 and 0.025 per μm³ in the reticulata and compacta subdivisions, respectively). Thus the ratio of GLT1-expressing glial membrane surface to glutamatergic synapses was very high (40–100 μm²), suggesting an efficient regulation of extracellular glutamate concentrations.     

Target‐ and input‐dependent organization of AMPA and NMDA receptors in synaptic connections of the cochlear nucleus

We examined the synaptic structure, quantity, and distribution of α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionic acid (AMPA)‐ and N‐methyl‐D‐aspartate (NMDA)‐type glutamate receptors (AMPARs and NMDARs, respectively) in rat cochlear nuclei by a highly sensitive freeze‐fracture replica labeling technique. Four excitatory synapses formed by two distinct inputs, auditory nerve (AN) and parallel fibers (PF), on different cell types were analyzed. These excitatory synapse types included AN synapses on bushy cells (AN‐BC synapses) and fusiform cells (AN‐FC synapses) and PF synapses on FC (PF‐FC synapses) and cartwheel cell spines (PF‐CwC synapses). Immunogold labeling revealed differences in synaptic structure as well as AMPAR and NMDAR number and/or density in both AN and PF synapses, indicating a target‐dependent organization. The immunogold receptor labeling also identified differences in the synaptic organization of FCs based on AN or PF connections, indicating an input‐dependent organization in FCs. Among the four excitatory synapse types, the AN‐BC synapses were the smallest and had the most densely packed intramembrane particles (IMPs), whereas the PF‐CwC synapses were the largest and had sparsely packed IMPs. All four synapse types showed positive correlations between the IMP‐cluster area and the AMPAR number, indicating a common intrasynapse‐type relationship for glutamatergic synapses. Immunogold particles for AMPARs were distributed over the entire area of individual AN synapses; PF synapses often showed synaptic areas devoid of labeling. The gold‐labeling for NMDARs occurred in a mosaic fashion, with less positive correlations between the IMP‐cluster area and the NMDAR number. Our observations reveal target‐ and input‐dependent features in the structure, number, and organization of AMPARs and NMDARs in AN and PF synapses. J. Comp. Neurol. 522:4023–4042, 2014. © 2014 Wiley Periodicals, Inc.     

线粒体去乙酰化蛋白SIRT3在噪声诱导隐匿性听力损失中的作用

目的 探讨线粒体去乙酰化蛋白(sirtuin-3,SIRT3)在噪声性隐性听力损失(noise-induced hidden hearing loss, NIHHL)中的作用。方法 将注射过SIRT3抑制剂[3-(1H-1,2,3-triazol-4-yl) pyridine,3-TYP]与未经处理的C57BL/6J小鼠分别于100 dB白噪声下暴露2 h,并在噪声暴露 (noise exposure, NE) 后1 d和14 d进行测听。利用免疫荧光染色观察内毛细胞 (inner hair cell, IHCs)带状突触(ribbon synapses, RS)的变化。冰冻切片下,4-羟基壬烯醛 (4-hydroxynonenal, 4-HNE)染色观察耳蜗内活性氧(reactive oxygen species, ROS) 的累积。结果 与对照组相比,噪声组及3-TYP组小鼠均在噪声后出现短暂性听力阈值的升高,ROS的累积及带状突触的丢失。与噪声组小鼠相比,3-TYP组小鼠在噪声后第14天听力未恢复正常,带状突触丢失明显,且伴有ROS的显著累积。结论 NE诱发了内耳的氧化应激反应,SIRT3可能通过抵抗氧化应激作用来保护耳蜗带状突触及听功能。     

Neuroplastic changes in the sensorimotor cortex associated with orthodontic tooth movement in rats

Orthodontic tooth movement (OTM) causes transient pain and changes in the dental occlusion that may lead to altered somatosensory inputs and patterns of mastication. This study used intracortical microstimulation (ICMS) and electromyographic (EMG) recordings to test whether neuroplastic changes occur in the ICMS‐defined motor representations of left and right anterior digastric (LAD, RAD), masseter, buccinator, and genioglossus (GG) muscles within the rat's face primary motor cortex (face‐M1) and adjacent face primary somatosensory cortex (face‐S1) during OTM. Analyses included any changes in the number of ICMS sites representing these muscles and in the onset latencies of ICMS‐evoked responses in the muscles. Sprague–Dawley rats were divided into experimental (E), sham (S), and naive (N) groups; OTM was induced in the E group. Statistical analyses involved a mixed model repeated‐measures analysis of variance (MMRM ANOVA). OTM resulted in significant neuroplastic changes in the number of positive sites in the E group for LAD, RAD, and GG muscles in face‐M1 and face‐S1 at days 1, 7, and 28 of continuous orthodontic force application, and in the number of sites in face‐M1 from which ICMS could simultaneously evoke EMG responses in different combinations of LAD, RAD, and GG muscles. However, the onset latencies of ICMS‐evoked responses were not significantly different between groups or between face‐M1 and face‐S1. The neuroplastic changes documented in this study may reflect adaptive sensorimotor changes in response to the altered environment in the oral cavity induced by OTM. J. Comp. Neurol. 523:1548–1568, 2015. © 2015 Wiley Periodicals, Inc.     

Alterations in the motor neuron–renshaw cell circuit in the Sod1G93A mouse model

Motor neurons become hyperexcitable during progression of amyotrophic lateral sclerosis (ALS). This abnormal firing behavior has been explained by changes in their membrane properties, but more recently it has been suggested that changes in premotor circuits may also contribute to this abnormal activity. The specific circuits that may be altered during development of ALS have not been investigated. Here we examined the Renshaw cell recurrent circuit that exerts inhibitory feedback control on motor neuron firing. Using two markers for Renshaw cells (calbindin and cholinergic nicotinic receptor subunit alpha2 [Chrna2]), two general markers for motor neurons (NeuN and vesicular acethylcholine transporter [VAChT]), and two markers for fast motor neurons (Chondrolectin and calcitonin‐related polypeptide alpha [Calca]), we analyzed the survival and connectivity of these cells during disease progression in the Sod1^G93A mouse model. Most calbindin‐immunoreactive (IR) Renshaw cells survive to end stage but downregulate postsynaptic Chrna2 in presymptomatic animals. In motor neurons, some markers are downregulated early (NeuN, VAChT, Chondrolectin) and others at end stage (Calca). Early downregulation of presynaptic VAChT and Chrna2 was correlated with disconnection from Renshaw cells as well as major structural abnormalities of motor axon synapses inside the spinal cord. Renshaw cell synapses on motor neurons underwent more complex changes, including transitional sprouting preferentially over remaining NeuN‐IR motor neurons. We conclude that the loss of presynaptic motor axon input on Renshaw cells occurs at early stages of ALS and disconnects the recurrent inhibitory circuit, presumably resulting in diminished control of motor neuron firing. J. Comp. Neurol. 521:1449–1469, 2013. © 2012 Wiley Periodicals, Inc.     

Sublaminar organization of the human subplate: developmental changes in the distribution of neurons,glia, growing axons and extracellular matrix

The objective of this paper was to collect normative data essential for analyzing the subplate (SP) role in pathogenesis of developmental disorders, characterized by abnormal circuitry, such as hypoxic‐ischemic lesions, autism and schizophrenia. The main cytological features of the SP, such as low cell density, early differentiation of neurons and glia, plexiform arrangement of axons and dendrites, presence of synapses and a large amount of extracellular matrix (ECM) distinguish this compartment from the cell‐dense cortical plate (CP; towards pia) and large fiber bundles of external axonal strata of fetal white matter (towards ventricle). For SP delineation from these adjacent layers based on combined cytological criteria, we analyzed the sublaminar distribution of different microstructural elements and the associated maturational gradients throughout development, using immunocytochemical and histological techniques on postmortem brain material (Zagreb Neuroembryological Collection). The analysis revealed that the SP compartment of the lateral neocortex shows changes in laminar organization throughout fetal development: the monolayer in the early fetal period (presubplate) undergoes dramatic bilaminar transformation between 13 and 15 postconceptional weeks (PCW), followed by subtle sublamination in three ‘floors’ (deep, intermediate, superficial) of midgestation (15–21 PCW). During the stationary phase (22–28 PCW), SP persists as a trilaminar compartment, gradually losing its sublaminar organization towards the end of gestation and remains as a single layer of SP remnant in the newborn brain. Based on these sublaminar transformations, we have documented developmental changes in the distribution, maturational gradients and expression of molecular markers in SP synapses, transitional forms of astroglia, neurons and ECM, which occur concomitantly with the ingrowth of thalamo‐cortical, basal forebrain and cortico‐cortical axons in a deep to superficial fashion. The deep SP is the zone of ingrowing axons – ‘entrance (ingrowth) zone’. The process of axonal ingrowth begins with thalamo‐cortical fibers and basal forebrain afferents, indicating an oblique geometry. During the later fetal period, deep SP receives long cortico‐cortical axons exhibiting a tangential geometry. Intermediate SP (‘proper’) is the navigation and ‘nexus’ sublamina consisting of a plexiform arrangement of cellular elements providing guidance and substrate for axonal growth, and also containing transient connectivity of dendrites and axons in a tangential plane without radial boundaries immersed in an ECM‐rich continuum. Superficial SP is the axonal accumulation (‘waiting compartment’) and target selection zone, indicating a dense distribution of synaptic markers, accumulation of thalamo‐cortical axons (around 20 PCW), overlapping with dendrites from layer VI neurons. In the late preterm brain period, superficial SP contains a chondroitin sulfate non‐immunoreactive band. The developmental dynamics for the distribution of neuronal, glial and ECM markers comply with sequential ingrowth of afferents in three levels of SP: ECM and synaptic markers shift from deep to superficial SP, with transient forms of glia following this arrangement, and calretinin neurons are concentrated in the SP during the formation phase. These results indicate developmental and morphogenetic roles in the SP cellular (transient glia, neurons and synapses) and ECM framework, enabling the spatial accommodation, navigation and establishment of numerous connections of cortical pathways in the expanded human brain. The original findings of early developmental dynamics of transitional subtypes of astroglia, calretinin neurons, ECM and synaptic markers presented in the SP are interesting in the light of recent concepts concerning its functional and morphogenetic role and an increasing interest in SP as a prospective substrate of abnormalities in cortical circuitry, leading to a cognitive deficit in different neurodevelopmental disorders.     

重复经颅磁刺激对大鼠缺血脑组织中突触素表达及运动功能康复的影响

目的探讨重复经颅磁刺激(rTMS)对脑可塑性的影响及其作用机制。方法雄性SD大鼠96只,随机分为模型组、假刺激组、急性期rTMS组和慢性期rTMS组,每组24只;采用线栓法制备大鼠右侧大脑中动脉闭塞的脑缺血再灌注模型。分别于建模后6、13、27 d时用免疫组织化学方法检测大鼠梗死侧皮质突触素的表达,同时评价大鼠神经功能。电镜观察大鼠梗死侧皮质突触损害情况。结果与模型组比较,急性期rTMS组梗死侧皮质突触素阳性细胞数在再灌注6、13、27 d明显增多,神经功能障碍评分在再灌注6、13 d明显增高,慢性期rTMS组梗死侧皮质突触素阳性细胞数在再灌注13、27 d明显增多,神经功能障碍评分在再灌注13 d明显增高[(1 3.63±1.06)分vs(11.88±1.25)分,P<0.05,P<0.01]。电镜观察,急性期rTMS组和慢性期rTMS组梗死侧皮质突触损害程度较模型组相对较轻。结论 rTMS使脑缺血再灌注损伤后大鼠梗死侧皮质的突触素阳性细胞数目有不同程度的增加;rTMS对脑缺血再灌注损伤后大鼠的神经功能恢复有一定程度的促进作用。