A Mixed Methods Approach to Exploring the Relationship between Norway Rat (Rattus norvegicus) Abundance and Features of the Urban Environment in an Inner-City Neighborhood of Vancouver,Canada

Urban rats (Rattus spp.) are among the most ubiquitous pest species in the world. Previous research has shown that rat abundance is largely determined by features of the environment; however, the specific urban environmental factors that influence rat population density within cities have yet to be clearly identified. Additionally, there are no well described tools or methodologies for conducting an in-depth evaluation of the relationship between urban rat abundance and the environment. In this study, we developed a systematic environmental observation tool using methods borrowed from the field of systematic social observation. This tool, which employed a combination of quantitative and qualitative methodologies, was then used to identify environmental factors associated with the relative abundance of Norway rats (Rattus norvegicus) in an inner-city neighborhood of Vancouver, Canada. Using a multivariate zero-inflated negative binomial model, we found that a variety of factors, including specific land use, building condition, and amount of refuse, were related to rat presence and abundance. Qualitative data largely supported and further clarified observed statistical relationships, but also identified conflicting and unique situations not easily captured through quantitative methods. Overall, the tool helped us to better understand the relationship between features of the urban environment and relative rat abundance within our study area and may useful for studying environmental determinants of zoonotic disease prevalence/distribution among urban rat populations in the future.     

The Mouse MC13 Mutant Is a Novel ENU Mutation in Collagen Type II,Alpha 1

Phenotype-driven mutagenesis experiments are a powerful approach to identifying novel alleles in a variety of contexts. The traditional disadvantage of this approach has been the subsequent task of identifying the affected locus in the mutants of interest. Recent advances in bioinformatics and sequencing have reduced the burden of cloning these ENU mutants. Here we report our experience with an ENU mutagenesis experiment and the rapid identification of a mutation in a previously known gene. A combination of mapping the mutation with a high-density SNP panel and a candidate gene approach has identified a mutation in collagen type II, alpha I (Col2a1). Col2a1 has previously been studied in the mouse and our mutant phenotype closely resembles mutations made in the Col2a1 locus.     

Cloning, localization, and functional expression of sodium channel beta1A subunits

Auxiliary beta1 subunits of voltage-gated sodium channels have been shown to be cell adhesion molecules of the Ig superfamily. Co-expression of alpha and beta1 subunits modulates channel gating as well as plasma membrane expression levels. We have cloned, sequenced, and expressed a splice variant of beta1, termed beta1A, that results from an apparent intron retention event. beta1 and beta1A are structurally homologous proteins with type I membrane topology; however, they contain little to no amino acid homology beyond the shared Ig loop region. beta1A mRNA expression is developmentally regulated in rat brain such that it is complementary to beta1. beta1A mRNA is expressed during embryonic development, and then its expression becomes undetectable after birth, concomitant with the onset of beta1 expression. In contrast, beta1A mRNA is expressed in adult adrenal gland and heart. Western blot analysis revealed beta1A protein expression in heart, skeletal muscle, and adrenal gland but not in adult brain or spinal cord. Immunocytochemical analysis of beta1A expression revealed selective expression in brain and spinal cord neurons, with high expression in heart and all dorsal root ganglia neurons. Co-expression of alphaIIA and beta1A subunits in Chinese hamster lung 1610 cells results in a 2.5-fold increase in sodium current density compared with cells expressing alphaIIA alone. This increase in current density reflected two effects of beta1A: 1) an increase in the proportion of cells expressing detectable sodium currents and 2) an increase in the level of functional sodium channels in expressing cells. [(3)H]Saxitoxin binding analysis revealed a 4-fold increase in B(max) with no change in K(D) in cells coexpressing alphaIIA and beta1A compared with cells expressing alphaIIA alone. beta1A-expressing cell lines also revealed subtle differences in sodium channel activation and inactivation. These effects of beta1A subunits on sodium channel function may be physiologically important events in the development of excitable cells.     

Carriage of Clostridium difficile by Wild Urban Norway Rats (Rattus norvegicus) and Black Rats (Rattus rattus)

Clostridium difficile is an important cause of enteric infections in humans. Recently, concerns have been raised regarding whether animals could be a source of C. difficile spores. Although colonization has been identified in a number of domestic species, the ability of commensal pests to serve as a reservoir for C. difficile has not been well investigated. The objective of this study was to determine whether urban rats (Rattus spp.) from Vancouver, Canada, carry C. difficile. Clostridium difficile was isolated from the colon contents of trapped rats and was characterized using ribotyping, toxinotyping, and toxin gene identification. Generalized linear mixed models and spatial analysis were used to characterize the ecology of C. difficile in rats. Clostridium difficile was isolated from 95 of 724 (13.1%) rats, although prevalence differed from 0% to 46.7% among city blocks. The odds of being C. difficile positive decreased with increasing weight (odds ratio [OR], 0.67; 95% confidence interval [CI], 0.53 to 0.87), suggesting that carriage is more common in younger animals. The strains isolated included 9 ribotypes that matched recognized international designations, 5 identified by our laboratory in previous studies, and 21 “novel” ribotypes. Some strains were clustered geographically; however, the majority were dispersed throughout the study area, supporting environmental sources of exposure and widespread environmental contamination with a variety of C. difficile strains. Given that urban rats are the source of a number of other pathogens responsible for human morbidity and mortality, the potential for rats to be a source of C. difficile for humans deserves further consideration.     

Manduca sexta prothoracicotropic hormone: evidence for a role beyond steroidogenesis

Prothoracicotropic hormone (PTTH) is a homodimeric brain peptide hormone that positively regulates the production of ecdysteroids by the prothoracic gland of Lepidoptera and probably other insects. PTTH was first purified from heads of adult domestic silkworms, Bombyx mori. Prothoracic glands of Bombyx and Manduca sexta undergo apoptosis well before the adult stage is reached, raising the recurring question of PTTH function at these later stages. Because Bombyx has been domesticated for thousands of years, the possibility exists that the presence of PTTH in adult animals is an accidental result of domestication for silk production. In contrast, Manduca has been raised in the laboratory for only five or six decades. The present study found that Manduca brains contain PTTH at all stages examined post‐prothoracic gland apoptosis, i.e., pharate adult and adult life, and that PTTH‐dependent changes in protein phosphorylation and protein synthesis were observed in several reproductive and reproduction‐associated organs. The data indicate that PTTH indeed plays a role in non‐steroidogenic tissues and suggest possible future avenues for determining which cellular processes are being so regulated. © 2009 Wiley Periodicals, Inc.     

Radiation induces acute alterations in neuronal function

Every year, nearly 200,000 patients undergo radiation for brain tumors. For both patients and caregivers the most distressing adverse effect is impaired cognition. Efforts to protect against this debilitating effect have suffered from inadequate understanding of the cellular mechanisms of radiation damage. In the past it was accepted that radiation-induced normal tissue injury resulted from a progressive reduction in the survival of clonogenic cells. Moreover, because radiation-induced brain dysfunction is believed to evolve over months to years, most studies have focused on late changes in brain parenchyma. However, clinically, acute changes in cognition are also observed. Because neurons are fully differentiated post-mitotic cells, little information exists on the acute effects of radiation on synaptic function. The purpose of our study was to assess the potential acute effects of radiation on neuronal function utilizing ex vivo hippocampal brain slices. The cellular localization and functional status of excitatory and inhibitory neurotransmitter receptors was identified by immunoblotting. Electrophysiological recordings were obtained both for populations of neuronal cells and individual neurons. In the dentate gyrus region of isolated ex vivo slices, radiation led to early decreases in tyrosine phosphorylation and removal of excitatory N-methyl-D-aspartate receptors (NMDARs) from the cell surface while simultaneously increasing the surface expression of inhibitory gamma-aminobutyric acid receptors (GABA(A)Rs). These alterations in cellular localization corresponded with altered synaptic responses and inhibition of long-term potentiation. The non-competitive NMDAR antagonist memantine blocked these radiation-induced alterations in cellular distribution. These findings demonstrate acute effects of radiation on neuronal cells within isolated brain slices and open new avenues for study.     

Creating Rigidly Stabilized Fractures for Assessing Intramembranous Ossification,Distraction Osteogenesis,or Healing of Critical Sized Defects

Assessing modes of skeletal repair is essential for developing therapies to be used clinically to treat fractures. Mechanical stability plays a large role in healing of bone injuries. In the worst-case scenario mechanical instability can lead to delayed or non-union in humans. However, motion can also stimulate the healing process. In fractures that have motion cartilage forms to stabilize the fracture bone ends, and this cartilage is gradually replaced by bone through recapitulation of the developmental process of endochondral ossification. In contrast, if a bone fracture is rigidly stabilized bone forms directly via intramembranous ossification. Clinically, both endochondral and intramembranous ossification occur simultaneously. To effectively replicate this process investigators insert a pin into the medullary canal of the fractured bone as described by Bonnarens⁴. This experimental method provides excellent lateral stability while allowing rotational instability to persist. However, our understanding of the mechanisms that regulate these two distinct processes can also be enhanced by experimentally isolating each of these processes. We have developed a stabilization protocol that provides rotational and lateral stabilization. In this model, intramembranous ossification is the only mode of healing that is observed, and healing parameters can be compared among different strains of genetically modified mice ^5-7, after application of bioactive molecules ^8,9, after altering physiological parameters of healing ¹⁰, after modifying the amount or time of stabilization ¹¹, after distraction osteogenesis ¹², after creation of a non-union ¹³, or after creation of a critical sized defect. Here, we illustrate how to apply the modified Ilizarov fixators for studying tibial fracture healing and distraction osteogenesis in mice.