Loss of TDP-43 induces synaptic dysfunction that is rescued by UNC13A splice-switching ASOs.

TDP-43 loss of function induces multiple splicing changes, including a cryptic exon in the amyotrophic lateral sclerosis and fronto-temporal lobar degeneration risk gene UNC13A, leading to nonsense-mediated decay of UNC13A transcripts and loss of protein. UNC13A is an active zone protein with an integral role in coordinating pre-synaptic function. Here, we show TDP-43 depletion induces a severe reduction in synaptic transmission, leading to an asynchronous pattern of network activity. We demonstrate that these deficits are largely driven by a single cryptic exon in UNC13A. Antisense oligonucleotides targeting the UNC13A cryptic exon robustly rescue UNC13A protein levels and restore normal synaptic function, providing a potential new therapeutic approach for ALS and other TDP-43-related disorders.

Excitatory and inhibitory synapses show a tight subcellular correlation that weakens over development.

Neurons receive correlated levels of excitation and inhibition, a feature that is important for proper brain function. However, how this relationship between excitatory and inhibitory inputs is established during the dynamic period of circuit wiring remains unexplored. Using multiple techniques, including in utero electroporation, electron microscopy, and electrophysiology, we reveal a tight correlation in the distribution of excitatory and inhibitory synapses along the dendrites of developing CA1 hippocampal neurons. This correlation was present within short dendritic stretches (<20 µm) and, surprisingly, was most pronounced during early development, sharply declining with maturity. The tight matching between excitation and inhibition was unexpected, as inhibitory synapses lacked an active zone when formed and exhibited compromised evoked release. We propose that inhibitory synapses form as a stabilizing scaffold to counterbalance growing excitation levels. This relationship diminishes over time, suggesting a critical role for a subcellular balance in early neuronal function and circuit formation.

Corrigendum: Correlative Live-Cell and Super-Resolution Imaging to Link Presynaptic Molecular Organisation With Function.

[This corrects the article DOI: 10.3389/fnsyn.2022.830583.].

Correlative Live-Cell and Super-Resolution Imaging to Link Presynaptic Molecular Organisation With Function.

Information transfer at synapses occurs when vesicles fuse with the plasma membrane to release neurotransmitters, which then bind to receptors at the postsynaptic membrane. The process of neurotransmitter release varies dramatically between different synapses, but little is known about how this heterogeneity emerges. The development of super-resolution microscopy has revealed that synaptic proteins are precisely organised within and between the two parts of the synapse and that this precise spatiotemporal organisation fine-tunes neurotransmission. However, it remains unclear if variability in release probability could be attributed to the nanoscale organisation of one or several proteins of the release machinery. To begin to address this question, we have developed a pipeline for correlative functional and super-resolution microscopy, taking advantage of recent technological advancements enabling multicolour imaging. Here we demonstrate the combination of live imaging of SypHy-RGECO, a unique dual reporter that simultaneously measures presynaptic calcium influx and neurotransmitter release, with post hoc immunolabelling and multicolour single molecule localisation microscopy, to investigate the structure-function relationship at individual presynaptic boutons.

HIV-1 Rev interacts with HERV-K RcREs present in the human genome and promotes export of unspliced HERV-K proviral RNA.

BACKGROUND: The HERV-K (HML-2) viruses are the youngest of the human endogenous retroviruses. They are present as several almost complete proviral copies and numerous fragments in the human genome. Many HERV-K proviruses express a regulatory protein Rec, which binds to an element present in HERV-K mRNAs called the RcRE. This interaction is necessary for the nucleo-cytoplasmic export and expression of HERV-K mRNAs that retain introns and plays a role analogous to that of Rev and the RRE in HIV replication. There are over 900 HERV-K RcREs distributed throughout the human genome. Thus, it was of interest to determine if Rev could functionally interact with selected RcRE elements that map either to HERV-K proviruses or human gene regions. This interaction would have the potential to alter the expression of both HERV-K mRNAs and cellular mRNAs during HIV-1 infection. RESULTS: In this study we employed a combination of RNAseq, bioinformatics and cell-based functional assays. Potential RcREs were identified through a number of bioinformatic approaches. They were then tested for their ability to promote export and translation of a reporter mRNA with a retained intron in conjunction with Rev or Rec. Some of the selected elements functioned well with either Rev, Rec or both, whereas some showed little or no function. Rev function on individual RcREs varied and was also dependent on the Rev sequence. We also performed RNAseq on total and cytoplasmic RNA isolated from SupT1 cells expressing HIV Rev, with or without Tat, or HERV-K Rec. Proviral mRNA from three HERV-K loci (4p16.1b, 22q11.23 and most significantly 3q12.3) accumulated in the cytoplasm in the presence of Rev or Tat and Rev, but not Rec. Consistent with this, the 3' RcRE from 3q12.3 functioned well with HIV-Rev in our reporter assay. In contrast, this RcRE showed little or no function with Rec. CONCLUSIONS: The HIV Rev protein can functionally interact with many RcREs present in the human genome, depending on the RcRE sequence, as well as the Rev sequence. This leads to export of some of the HERV-K proviral mRNAs and also has the potential to change the expression of non-viral genes.

Nanoscale Structural Plasticity of the Active Zone Matrix Modulates Presynaptic Function.

The active zone (AZ) matrix of presynaptic terminals coordinates the recruitment of voltage-gated calcium channels (VGCCs) and synaptic vesicles to orchestrate neurotransmitter release. However, the spatial organization of the AZ and how it controls vesicle fusion remain poorly understood. Here, we employ super-resolution microscopy and ratiometric imaging to visualize the AZ structure on the nanoscale, revealing segregation between the AZ matrix, VGCCs, and putative release sites. Long-term blockade of neuronal activity leads to reversible AZ matrix unclustering and presynaptic actin depolymerization, allowing for enrichment of AZ machinery. Conversely, patterned optogenetic stimulation of postsynaptic neurons retrogradely enhanced AZ clustering. In individual synapses, AZ clustering was inversely correlated with local VGCC recruitment and vesicle cycling. Acute actin depolymerization led to rapid (5 min) nanoscale AZ matrix unclustering. We propose a model whereby neuronal activity modulates presynaptic function in a homeostatic manner by altering the clustering state of the AZ matrix.

Distance-dependent gradient in NMDAR-driven spine calcium signals along tapering dendrites.

Neurons receive a multitude of synaptic inputs along their dendritic arbor, but how this highly heterogeneous population of synaptic compartments is spatially organized remains unclear. By measuring N-methyl-d-aspartic acid receptor (NMDAR)-driven calcium responses in single spines, we provide a spatial map of synaptic calcium signals along dendritic arbors of hippocampal neurons and relate this to measures of synapse structure. We find that quantal NMDAR calcium signals increase in amplitude as they approach a thinning dendritic tip end. Based on a compartmental model of spine calcium dynamics, we propose that this biased distribution in calcium signals is governed by a gradual, distance-dependent decline in spine size, which we visualized using serial block-face scanning electron microscopy. Our data describe a cell-autonomous feature of principal neurons, where tapering dendrites show an inverse distribution of spine size and NMDAR-driven calcium signals along dendritic trees, with important implications for synaptic plasticity rules and spine function.

An annotated bibliography of verbal behavior articles published outside of The Analysis of Verbal Behavior: 2016.

An annotated bibliography is provided that summarizes journal articles on verbal behavior published outside of The Analysis of Verbal Behavior in 2016, the primary journal for scholarship in this area. Thirty-seven such articles were identified and annotated as a resource for practitioners, researchers, and educators.

Visualizing Presynaptic Calcium Dynamics and Vesicle Fusion with a Single Genetically Encoded Reporter at Individual Synapses.

Synaptic transmission depends on the influx of calcium into the presynaptic compartment, which drives neurotransmitter release. Genetically encoded reporters are widely used tools to understand these processes, particularly pHluorin-based reporters that report vesicle exocytosis and endocytosis through pH dependent changes in fluorescence, and genetically encoded calcium indicators (GECIs) that exhibit changes in fluorescence upon binding to calcium. The recent expansion of the color palette of available indicators has made it possible to image multiple probes simultaneously within a cell. We have constructed a single molecule reporter capable of concurrent imaging of both presynaptic calcium influx and exocytosis, by fusion of sypHy, the vesicle associated protein synaptophysin containing a GFP-based pHluorin sensor, with the red-shifted GECI R-GECO1. Due to the fixed stoichiometry of the two probes, the ratio of the two responses can also be measured, providing an all optical correlate of the calcium dependence of release. Here, we have characterized stimulus-evoked sypHy-RGECO responses of hippocampal synapses in vitro, exploring the effects of different stimulus strengths and frequencies as well as variations in external calcium concentrations. By combining live sypHy-RGECO imaging with post hoc fixation and immunofluorescence, we have also investigated correlations between structural and functional properties of synapses.

Engineering FKBP-Based Destabilizing Domains to Build Sophisticated Protein Regulation Systems.

Targeting protein stability with small molecules has emerged as an effective tool to control protein abundance in a fast, scalable and reversible manner. The technique involves tagging a protein of interest (POI) with a destabilizing domain (DD) specifically controlled by a small molecule. The successful construction of such fusion proteins may, however, be limited by functional interference of the DD epitope with electrostatic interactions required for full biological function of proteins. Another drawback of this approach is the remaining endogenous protein. Here, we combined the Cre-LoxP system with an advanced DD and generated a protein regulation system in which the loss of an endogenous protein, in our case the tumor suppressor PTEN, can be coupled directly with a conditionally fine-tunable DD-PTEN. This new system will consolidate and extend the use of DD-technology to control protein function precisely in living cells and animal models.

Research Rankings of Behavior Analytic Graduate Training Programs and Their Faculty.

We evaluated research productivity of several graduate programs that provide Behavior Analyst Certification Board (BACB)-approved course sequences in behavior analysis. Considering the faculty of BACB training programs as a unit, in only about 50 %, programs have faculty combined to publish ten or more total articles in our field's primary empirical journals. Among individual faculty members, a sizeable number have not published an article in any of the field's top journals. To recognize major scholarly contributors, we provide top 10 lists of training programs and individual faculty members. We conclude by discussing the importance of research in an increasingly practice-driven marketplace.

Top 10 Responses to the Commentaries on Dixon, Reed, Smith et al. (2015).

In a previous article (Dixon et al. Behavior Analysis in Practice, 8(1), 7-15, 2015), we put forward data suggesting that most behavior analytic faculty do not publish in major behavior analytic journals, and in only about 50 % of behavior analysis programs have faculty combined to produce ten or more empirical articles. Several commentaries followed the release of our article, with content that ranged from supporting our endeavors and confirming the dangerous position our field may be in to highlighting the need for further refinement in procedures used to rank the quality of behavior analysis graduate training programs. Presented in the present article are our "top 10" responses to these commentaries.