Phase I Cultural Resources Survey Of The Proposed Texas Eastern Transmission, Lp, Dot 2015 - Mexi-Stfe Project, In Hidalgo County, Texas

This document describes the results of Phase I cultural resources survey of the proposed Texas Eastern Transmission, LP, DOT 2015 – MEXI-STFE Project, in Hidalgo County, Texas (Figures 1.1 and 1.2). The project entailed the examination of a 275 m (2378.6 ft) long pipeline replacement, as well as 2.1 ha (5.2 ac) of additional temporary workspace and 826.0 m (2710 ft) of project access roads. Together these project items totaled 4.4 ha (10.9 ac) in area. This investigation was completed on behalf of Texas Eastern Transmission, LP, by R. Christopher Goodwin & Associates, Inc. in May of 2015. The goal of fieldwork was to identify and to evaluate all historic properties, archeological sites, cultural resources loci, standing structures, and/or cemeteries that may be impacted adversely by the proposed project. All work was performed in accordance with the procedures outlined in the National Historic Preservation Act of 1966, as amended; the Archaeological and Historic Preservation Act of 1974; the Archaeological Resources Protection Act of 1979, as amended; and Title 36 of the Code of Federal Regulations, Parts 60-66 and 800, as appropriate. Additionally, this survey effort abided by the standards set forth in the Federal Energy Regulatory Commission, Office of Pipeline Regulation’s Guidelines for Reporting on Cultural Resources Investigations (1994:11-13), the Archeology and Historic Preservation: The Secretary of the Interior’s Guidelines, and with guidelines contained within the Texas Historical Commission’s Preserving Our Heritage: A Statewide Plan for Texas. Information provided by the Council of Texas Archeologists, the Texas Archaeological Research Laboratory, the Antiquities Code of Texas, and the Texas Historical Commission’s Rules of Practice and Procedure for the Antiquities Code of Texas, also was utilized.. All of the proposed project items fell within the bounds of the Louisiana-Rio Grande Canal Company Irrigation System National Register District. The historic property consists of two pumping stations and a network of canals, irrigation pipes, and drainage ditches that extend approximately 500 miles total in length (Meyers and Weitze 1995). The two pumping stations are located outside the project area, but the project area is in the vicinity of drainage ditches likely part of the former Louisiana – Rio Grande Canal Company system, now under the jurisdiction of the Hidalgo County Irrigation District No. 2. Despite the proximity of the district, the proposed projects, currently defined as replacement of an existing pipeline and hydrostatic testing, will not pose an adverse effect to, or have any additional visual impacts on, the district. If the project scope changes, further investigation of this national register property may be warranted. Furthermore, no cultural resources were identified within the limits of the current survey areas. No additional investigation or recordation of the project items associated with the Texas Eastern Transmission, LP, Project Name Project is recommended

Transcriptome Analysis of Single Cells

Many gene expression analysis techniques rely on material isolated from heterogeneous populations of cells from tissue homogenates or cells in culture.1,2,3 In the case of the brain, regions such as the hippocampus contain a complex arrangement of different cell types, each with distinct mRNA profiles. The ability to harvest single cells allows for a more in depth investigation into the molecular differences between and within cell populations. We describe a simple and rapid method for harvesting cells for further processing. Pipettes often used in electrophysiology are utilized to isolate (using aspiration) a cell of interest and conveniently deposit it into an Eppendorf tube for further processing with any number of molecular biology techniques. Our protocol can be modified for the harvest of dendrites from cell culture or even individual cells from acute slices

A Neurotoxic Phosphoform of Elk-1 Associates with Inclusions from Multiple Neurodegenerative Diseases

Neurodegenerative diseases are characterized by a number of features including the formation of inclusions, early synaptic degeneration and the selective loss of neurons. Molecules serving as links between these shared features have yet to be identified. Identifying candidates within the diseased microenvironment will open up novel avenues for therapeutic intervention. The transcription factor Elk-1 resides within multiple brain areas both in nuclear and extranuclear neuronal compartments. Interestingly, its de novo expression within a single dendrite initiates neuronal death. Given this novel regionalized function, we assessed whether extranuclear Elk-1 and/or phospho-Elk-1 (pElk-1) protein might be associated with a spectrum of human neurodegenerative disease cases including Lewy body Disease (e.g. Parkinson's), Alzheimer's disease, and Huntington's Disease. We first determined the importance of Elk-1 post-translational modifications on its ability to initiate regionalized cell death. We next screened human cases from three major neurodegenerative diseases to look for remarkable levels of Elk-1 and/or pElk-1 protein as well as their association with inclusions characteristic of these diseases. We compared our findings to age-matched control cases. We find that the ability of Elk-1 to initiate regionalized neuronal death depends on a specific phosphosite, T417. Furthermore, we find that T417+ Elk-1 uniquely associates with several types of inclusions present in cases of human Lewy body Disease, Alzheimer's disease, and Huntington's Disease. These results suggest a molecular link between the presence of inclusions and neuronal loss that is shared across a spectrum of neurodegenerative disease

Neuroethics guiding principles for the NIH Brain Initiative

Neuroscience presents important neuroethical considerations. Human neuroscience demands focused application of the core research ethics guidelines set out in documents such as the Belmont Report. Various mechanisms, including institutional review boards (IRBs), privacy rules, and the Food and Drug Administration, regulate many aspects of neuroscience research and many articles, books, workshops, and conferences address neuroethics. (Farah, 2010; https://bioethicsarchive.georgetown.edu/pcsbi/studies.html; http://www.neuroethicssociety.org/annual-meeting). However, responsible neuroscience research requires continual dialogue among neuroscience researchers, ethicists, philosophers, lawyers, and other stakeholders to help assess its ethical, legal, and societal implications. The Neuroethics Working Group of the National Institutes of Health (NIH) Brain Research through Advancing Innovative Neurotechnologies (BRAIN) Initiative, a group of experts providing neuroethics input to the NIH BRAIN Initiative Multi-Council Working Group, seeks to promote this dialogue by proposing the following Neuroethics Guiding Principles (Table 1)

Cytoplasmic Intron Sequence-Retaining Transcripts Can Be Dendritically Targeted via ID Element Retrotransposons

SummaryRNA precursors give rise to mRNA after splicing of intronic sequences traditionally thought to occur in the nucleus. Here, we show that intron sequences are retained in a number of dendritically-targeted mRNAs, by using microarray and Illumina sequencing of isolated dendritic mRNA as well as in situ hybridization. Many of the retained introns contain ID elements, a class of SINE retrotransposon. A portion of these SINEs confers dendritic targeting to exogenous and endogenous transcripts showing the necessity of ID-mediated mechanisms for the targeting of different transcripts to dendrites. ID elements are capable of selectively altering the distribution of endogenous proteins, providing a link between intronic SINEs and protein function. As such, the ID element represents a common dendritic targeting element found across multiple RNAs. Retention of intronic sequence is a more general phenomenon than previously thought and plays a functional role in the biology of the neuron, partly mediated by co-opted repetitive sequences

Cell Surface Protein mRNAs Show Differential Transcription in Pyramidal and Fast-Spiking Cells as Revealed by Single-Cell Sequencing

The prefrontal cortex (PFC) plays a key role in higher order cognitive functions and psychiatric disorders such as autism, schizophrenia, and depression. In the PFC, the two major classes of neurons are the glutamatergic pyramidal (Pyr) cells and the GABAergic interneurons such as fast-spiking (FS) cells. Despite extensive electrophysiological, morphological, and pharmacological studies of the PFC, the therapeutically utilized drug targets are restricted to dopaminergic, glutamatergic, and GABAergic receptors. To expand the pharmacological possibilities as well as to better understand the cellular and network effects of clinically used drugs, it is important to identify cell-type-selective, druggable cell surface proteins and to link developed drug candidates to Pyr or FS cell targets. To identify the mRNAs of such cell-specific/enriched proteins, we performed ultra-deep single-cell mRNA sequencing (19 685 transcripts in total) on electrophysiologically characterized intact PFC neurons harvested from acute brain slices of mice. Several selectively expressed transcripts were identified with some of the genes that have already been associated with cellular mechanisms of psychiatric diseases, which we can now assign to Pyr (e.g., Kcnn2, Gria3) or FS (e.g., Kcnk2, Kcnmb1) cells. The earlier classification of PFC neurons was also confirmed at mRNA level, and additional markers have been provided

Avian Primordial Germ Cells Contribute to and Interact With the Extracellular Matrix During Early Migration

During early avian development, primordial germ cells (PGC) are highly migratory, moving from the central area pellucida of the blastoderm to the anterior extra-embryonic germinal crescent. The PGCs soon move into the forming blood vessels by intravasation and travel in the circulatory system to the genital ridges where they participate in the organogenesis of the gonads. This complex cellular migration takes place in close association with a nascent extracellular matrix that matures in a precise spatio-temporal pattern. We first compiled a list of quail matrisome genes by bioinformatic screening of human matrisome orthologs. Next, we used single cell RNA-seq analysis (scRNAseq) to determine that PGCs express numerous ECM and ECM-associated genes in early embryos. The expression of select ECM transcripts and proteins in PGCs were verified by fluorescent in situ hybridization (FISH) and immunofluorescence (IF). Live imaging of transgenic quail embryos injected with fluorescent antibodies against fibronectin and laminin, showed that germinal crescent PGCs display rapid shape changes and morphological properties such as blebbing and filopodia while surrounded by, or in close contact with, an ECM fibril meshwork that is itself in constant motion. Injection of anti-β1 integrin CSAT antibodies resulted in a reduction of mature fibronectin and laminin fibril meshwork in the germinal crescent at HH4-5 but did not alter the active motility of the PGCs or their ability to populate the germinal crescent. These results suggest that integrin β1 receptors are important, but not required, for PGCs to successfully migrate during embryonic development, but instead play a vital role in ECM fibrillogenesis and assembly

Cytoplasmic BK\u3csub\u3eCa\u3c/sub\u3e channel intron-containing mRNAs contribute to the intrinsic excitability of hippocampal neurons

High single-channel conductance K+ channels, which respond jointly to membrane depolarization and micromolar concentrations of intracellular Ca2+ ions, arise from extensive cell-specific alternative splicing of pore-forming α-subunit mRNAs. Here, we report the discovery of an endogenous BKCa channel α-subunit intron-containing mRNA in the cytoplasm of hippocampal neurons. This partially processed mRNA, which comprises ≈10% of the total BKCa channel α-subunit mRNAs, is distributed in a gradient throughout the somatodendritic space. We selectively reduced endogenous cytoplasmic levels of this intron-containing transcript by RNA interference without altering levels of the mature splice forms of the BKCa channel mRNAs. In doing so, we could demonstrate that changes in a unique BKCa channel α-subunit introncontaining splice variant mRNA can greatly impact the distribution of the BKCa channel protein to dendritic spines and intrinsic firing properties of hippocampal neurons. These data suggest a new regulatory mechanism for modulating the membrane properties and ion channel gradients of hippocampal neurons

Single-Cell Transcriptional Analysis Reveals Novel Neuronal Phenotypes and Interaction Networks Involved in the Central Circadian Clock

Single-cell heterogeneity confounds efforts to understand how a population of cells organizes into cellular networks that underlie tissue-level function. This complexity is prominent in the mammalian suprachiasmatic nucleus (SCN). Here, individual neurons exhibit a remarkable amount of asynchronous behavior and transcriptional heterogeneity. However, SCN neurons are able to generate precisely coordinated synaptic and molecular outputs that synchronize the body to a common circadian cycle by organizing into cellular networks. To understand this emergent cellular network property, it is important to reconcile single-neuron heterogeneity with network organization. In light of recent studies suggesting that transcriptionally heterogeneous cells organize into distinct cellular phenotypes, we characterized the transcriptional, spatial, and functional organization of 352 SCN neurons from mice experiencing phase-shifts in their circadian cycle. Using the community structure detection method and multivariate analytical techniques, we identified previously undescribed neuronal phenotypes that are likely to participate in regulatory networks with known SCN cell types. Based on the newly discovered neuronal phenotypes, we developed a data-driven neuronal network structure in which multiple cell types interact through known synaptic and paracrine signaling mechanisms. These results provide a basis from which to interpret the functional variability of SCN neurons and describe methodologies toward understanding how a population of heterogeneous single cells organizes into cellular networks that underlie tissue-level function

Mammalian cell transfection: the present and the future

Transfection is a powerful analytical tool enabling study of the function of genes and gene products in cells. The transfection methods are broadly classified into three groups; biological, chemical, and physical. These methods have advanced to make it possible to deliver nucleic acids to specific subcellular regions of cells by use of a precisely controlled laser-microcope system. The combination of point-directed transfection and mRNA transfection is a new way of studying the function of genes and gene products. However, each method has its own advantages and disadvantages so the optimum method depends on experimental design and objective