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Saturday, 25 October 2014

PubMed articles: Week 3, October 2014

1. Cold Spring Harb Protoc. 2014 Oct 23. doi: 10.1101/pdb.prot084624. [Epub ahead of print]

In Vivo Recording of Single-Unit Activity during Singing in Zebra Finches.

Okubo TS, Mackevicius EL, Fee MS.
Author information:
Department of Brain and Cognitive Sciences, McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139.


The zebra finch is an important model for investigating the neural mechanisms that underlie vocal production and learning. Previous anatomical and gene expression studies have identified an interconnected set of brain areas in this organism that are important for singing. To advance our understanding of how these various brain areas act together to learn and produce a highly stereotyped song, it is necessary to record the activity of individual neurons during singing. Here, we present a protocol for recording single-unit activity in freely moving zebra finches during singing using a miniature, motorized microdrive. It includes procedures for both the microdrive implant surgery and the electrophysiological recordings. There are several advantages of this technique: (1) high-impedance electrodes can be used in the microdrive to obtain well-isolated single units; (2) a motorized microdrive is used to remotely control the electrode position, allowing neurons to be isolated without handling the bird, and (3) a lateral positioner is used to move electrodes into fresh tissue before each penetration, allowing recordings from well-isolated neurons over the course of several weeks. We also describe the application of the antidromic stimulation and the spike collision test to identify neurons based on the axonal projection patterns.
© 2014 Cold Spring Harbor Laboratory Press.
PMID: 25342072 [PubMed - as supplied by publisher]

2. Cold Spring Harb Protoc. 2014 Oct 23. doi: 10.1101/pdb.prot084582. [Epub ahead of print]

An Optimized Protocol for High-Throughput In Situ Hybridization of Zebra Finch Brain.

Carleton JB, Lovell PV, McHugh A, Marzulla T, Horback KL, Mello CV.
Author information:
Department of Behavioral Neuroscience, Oregon Health & Science University, Portland, Oregon 97239.


In situ hybridization (ISH) is a sensitive technique for documenting the tissue distribution of mRNAs. Advanced nonradioactive ISH methods that are based on the use of digoxigenin (DIG)-labeled probes and chromogenic detection have better spatial resolution than emulsion autoradiography techniques and, when paired with high-resolution digital imaging, allow for large-scale profiling of gene expression at cellular resolution within a histological context. However, technical challenges restrict the number of genes that can be investigated in a small laboratory setting. This protocol describes an optimized, low-cost, small-footprint, high-throughput ISH procedure to detect gene expression patterns in 10-µm brain sections from zebra finches. It uses DIG-labeled riboprobes synthesized from cDNA templates available through the Songbird Neurogenomics Consortium. The method is compatible with high-resolution digital imaging; it produces images with low background and a resolution approaching that of immunohistochemical methods. Approximately 180 slides can be processed each week using this protocol, but it can be scaled to accommodate a broad range of tissues from which cryosections can be obtained.
© 2014 Cold Spring Harbor Laboratory Press.
PMID: 25342071 [PubMed - as supplied by publisher]

3. Cold Spring Harb Protoc. 2014 Oct 23. doi: 10.1101/pdb.emo084574. [Epub ahead of print]

The Zebra Finch, Taeniopygia guttata: An Avian Model for Investigating the Neurobiological Basis of Vocal Learning.

Mello CV.
Author information:
Department of Behavioral Neuroscience, Oregon Health and Science University, Portland, Oregon 97239.


Songbirds are capable of learning their vocalizations by copying a singing adult. This vocal learning ability requires juveniles to hear and memorize the sound of the adult song, and later to imitate it through a process involving sensorimotor integration. Vocal learning is a trait that songbirds share with humans, where it forms the basis of spoken language acquisition, with other avian groups (parrots and hummingbirds), and with a few other mammals (cetaceans, bats). It is however absent in traditional model organisms such as rodents and nonhuman primates. Zebra finches, a songbird species from Australia, are popular pets and are easy to breed. They also sing a relatively simple and stereotyped song that is amenable to quantitative analysis. Zebra finches have thus emerged as a choice model organism for investigating the neurobiological basis of vocal learning. A number of tools and methodologies have been developed to characterize the bioacoustics properties of their song, analyze the degree of accurate copying during vocal learning, map the brain circuits that control singing and song learning, and investigate the physiology of these circuits. Such studies have led to a large base of knowledge on song production and learning, and their underlying neural substrate. Several molecular resources have recently become available, including brain cDNA/EST databases, microarrays, BAC libraries, a molecular brain atlas, a complete genome assembly, and the ability to perform transgenesis. The recent availability of many other avian genomes provides unique opportunities for comparative analysis in the search for features unique to vocal learning organisms.
© 2014 Cold Spring Harbor Laboratory Press.
PMID: 25342070 [PubMed - as supplied by publisher]

4. Cold Spring Harb Protoc. 2014 Oct 23. doi: 10.1101/pdb.prot084590. [Epub ahead of print]

A Method for Exploring Adult Neurogenesis in the Songbird Brain.

Asik K, Rao JL, Kirn JR.
Author information:
Department of Biology, Neuroscience & Behavior Program, Wesleyan University, Middletown, Connecticut 06459.


The avian brain is a valuable model for exploring adult neurogenesis. Here we use immunohistochemical methods to detect cell division and the incorporation of new neurons in the adult zebra finch brain. The nonradioactive, relatively inexpensive thymidine analog bromodeoxyuridine (BrdU) is used to label replicating DNA in dividing cells. The brain is harvested, fixed, and dehydrated before being embedded in polyethylene glycol (PEG), which results in superior histology compared to frozen specimens. After the PEG-embedded brain tissue is sectioned and mounted on slides, standard immunohistochemical procedures are used to detect both BrdU and the neuron-specific marker Hu.
© 2014 Cold Spring Harbor Laboratory Press.
PMID: 25342069 [PubMed - as supplied by publisher]

5. Cold Spring Harb Protoc. 2014 Oct 23. doi: 10.1101/pdb.prot084608. [Epub ahead of print]

Generation of Transgenic Zebra Finches with Replication-Deficient Lentiviruses.

Velho TA, Lois C.
Author information:
Department of Neurobiology, University of Massachusetts Medical School, Worcester, Massachusetts 01655.


Zebra finches have been a rich experimental system for studying neurobiological questions of relevance to human health for decades. In particular, finches are the leading nonhuman model organisms for investigating the biological basis of vocal learning, a critical behavioral substrate for speech acquisition. In addition, zebra finches are an ideal system for the study of brain asymmetry, hormonal control of brain development, physiological function of sleep, sex differences in the brain, behavioral-induced gene expression, and adult neurogenesis, among other questions. Despite their importance for neurobiology, the usefulness of finches as an experimental system has been restricted by a lack of genetic manipulation methods. To overcome this barrier, our laboratory has developed methods for generating transgenic birds, including zebra finches. The successful implementation of this transgenic technology by multiple research laboratories has the potential to dramatically accelerate the progress of our understanding of the genetic basis of complex biological processes such as vocal learning. Moreover, the ability to genetically manipulate zebra finches could also be used to generate novel genetic models for human disorders that cannot be studied elsewhere or that can be more easily studied in this small bird. Here, we describe a protocol to generate transgenic zebra finches using recombinant lentiviruses.
© 2014 Cold Spring Harbor Laboratory Press.
PMID: 25342068 [PubMed - as supplied by publisher]

6. Cold Spring Harb Protoc. 2014 Oct 23. doi: 10.1101/pdb.prot084780. [Epub ahead of print]

Proper Care, Husbandry, and Breeding Guidelines for the Zebra Finch, Taeniopygia guttata.

Olson CR, Wirthlin M, Lovell PV, Mello CV.
Author information:
Department of Behavioral Neuroscience, Oregon Health and Science University, Portland, Oregon 97239.


The zebra finch Taeniopygia guttata castanotis is a songbird commonly used in the laboratory, particularly for studies of vocal learning, neurobiology, and physiology. Within the laboratory, it is important to adopt careful husbandry practices that allow for normal development of the birds. For example, their song is a learned trait, passed culturally from adult males to juveniles, and thus its learning can be influenced by the health and social conditions of the birds present in the laboratory. Here we present guidelines for the successful maintenance and breeding of captive zebra finches.
© 2014 Cold Spring Harbor Laboratory Press.
PMID: 25342067 [PubMed - as supplied by publisher]

7. J R Soc Interface. 2014 Dec 6;11(101). pii: 20140961. doi: 10.1098/rsif.2014.0961.

Nanomechanical properties of bird feather rachises: exploring naturally occurring fibre reinforced laminar composites.

Laurent CM1, Palmer C2, Boardman RP3, Dyke G4, Cook RB5.
Author information:
1Ocean and Earth Science, National Oceanography Centre, University of Southampton, Southampton, UK
2Earth Sciences, University of Bristol, Bristol, UK.
3µ-Vis X-ray Imaging Centre, Faculty of Engineering and the Environment, University of Southampton, Southampton, UK.
4Ocean and Earth Science, National Oceanography Centre, University of Southampton, Southampton, UK Department of Evolutionary Zoology and Human Biology, University of Debrecen, Debrecen, Hungary.
5National Centre for Advanced Tribology at Southampton (nCATS), Faculty of Engineering and the Environment, University of Southampton, Southampton, UK.


Flight feathers have evolved under selective pressures to be sufficiently light and strong enough to cope with the stresses of flight. The feather shaft (rachis) must resist these stresses and is fundamental to this mode of locomotion. Relatively little work has been done on rachis morphology, especially from a mechanical perspective and never at the nanoscale. Nano-indentation is a cornerstone technique in materials testing. Here we use this technique to make use of differentially oriented fibres and their resulting mechanical anisotropy. The rachis is established as a multi-layered fibrous composite material with varying laminar properties in three feathers of birds with markedly different flight styles; the Mute Swan (Cygnus olor), the Bald Eagle (Haliaeetus leucocephalus) and the partridge (Perdix perdix). These birds were chosen not just because they are from different clades and have different flight styles, but because they have feathers large enough to gain meaningful results from nano-indentation. Results from our initial datasets indicate that the proportions and orientation of the laminae are not fixed and may vary either in order to cope with the stresses of flight particular to the bird or with phylogenetic lineage.
© 2014 The Author(s) Published by the Royal Society. All rights reserved.
PMID: 25339689 [PubMed - in process]
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8. Parasitol Res. 2014 Oct 24. [Epub ahead of print]

Culex pipiens s.l. and Culex torrentium (Culicidae) in Wrocław area (Poland): occurrence and breeding site preferences of mosquito vectors.

Weitzel T1, Jawień P, Rydzanicz K, Lonc E, Becker N.
Author information:
1German Mosquito Control Association (GMCA/KABS), Georg-Peter-Süß-Str. 3, 67346, Speyer, Germany,


Both ornithophilic mosquito species, Culex pipiens s.l. (L.) and Culex torrentium (Martini, 1925), occur sympatric in temperate Europe. They are presumed to be primary vectors of West Nile and Sindbis viruses. Differentiation of these morphologically similar Culex species is essential for evaluation of different vector roles, for mosquito surveillance and integrated control strategies. Cx. torrentium has been neglected or erroneously determined as Cx. pipiens s.l. in some previous studies, because only males of both species can be diagnosed reliably by morphology. Thus, knowledge about species abundance, geographical distribution, breeding site preferences and the zoonotic risk assessment is incomplete also in Poland. In Wrocław area (Silesian Lowland), besides typical urban breeding sites, huge sewage irrigation fields provide suitable breeding conditions for Culex species. They are also inhabited by 180 resident and migratory bird species serving as potential virus reservoirs. In this study, morphology of larvae and males as well as species diagnostic enzyme markers, namely adenylate kinase (AK) and 2-hydroxybutyrate dehydrogenase (HBDH), were used to discriminate Cx. pipiens s.l. and Cx. torrentium. In a total of 650 Culex larvae from 24 natural and artificial breeding sites, Cx. pipiens s.l. had a proportion of 94.0 % and Cx. torrentium only 6.0 %. It could be shown that both species are well adapted to various breeding site types like ditches, catch basins, flower pots and buckets with diverse water quality. Cx. torrentium preferred more artificial water containers in urban surrounding (12 % species proportion), whereas in semi-natural breeding sites, Cx. torrentium was rare (3 %). In 12 of 24 breeding sites, larvae of both species have been found associated.
PMID: 25339516 [PubMed - as supplied by publisher]

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9. Mem Inst Oswaldo Cruz. 2014 Oct 14;0:0. [Epub ahead of print]

Phlebotomine fauna, natural infection rate and feeding habits of Lutzomyia cruzi in Jaciara, state of Mato Grosso, Brazil.

Brito VN1, Almeida AD2, Nakazato L2, Duarte R3, Souza CD2, Sousa VR2.
Author information:
1Secretaria Estadual de Saúde de Mato Grosso, Cuiabá, MT, Brasil.
2Departamento de Clínica Médica Veterinária, Faculdade de Agronomia, Medicina Veterinária e Zootecnia, Universidade Federal de Mato Grosso, Cuiabá, MT, Brasil.
3Escola Nacional de Saúde Pública Sérgio Arouca-Fiocruz, Rio de Janeiro, RJ, Brasil.


Visceral leishmaniasis (VL) in Brazil is transmitted by the phlebotomine Lutzomyia longipalpis and in some midwestern regions by Lutzomyia cruzi. Studies of the phlebotomine fauna, feeding habits and natural infection rate by Leishmania contribute to increased understanding of the epidemiological chain of leishmaniases and their vectorial capacity. Collections were performed in Jaciara, state of Mato Grosso from 2010-2013, during which time 2,011 phlebotomines (23 species) were captured (68.70% Lu. cruzi and 20.52% Lutzomyia whitmani). Lu. cruzi females were identified by observing the shapes of the cibarium (a portion of the mouthpart) and spermatheca, from which samples were obtained for polymerase chain reaction to determine the rates of natural infection. Engorged phlebotomines were assessed to identify the blood-meal host by ELISA. A moderate correlation was discovered between the number of Lu. cruzi and the temperature and the minimum rate of infection was 6.10%. Twenty-two females were reactive to the antisera of bird (28%), dog (3.30%) and skunk (1.60%). We conclude that Lu. cruzi and Lu. whitmani have adapted to the urban environment in this region and that Lu. cruzi is the most likely vector of VL in Jaciara. Moreover, maintenance of Leishmania in the environment is likely aided by the presence of birds and domestic and synanthropic animals.
Free Article
PMID: 25338156 [PubMed - as supplied by publisher]
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10. PLoS One. 2014 Oct 22;9(10):e109397. doi: 10.1371/journal.pone.0109397. eCollection 2014.

Relative roles of grey squirrels, supplementary feeding, and habitat in shaping urban bird assemblages.

Bonnington C1, Gaston KJ2, Evans KL1.
Author information:
1Department of Animal and Plant Sciences, University of Sheffield, Sheffield, United Kingdom.
2Environment and Sustainability Institute, University of Exeter, Penryn, Cornwall, United Kingdom.


Non-native species are frequently considered to influence urban assemblages. The grey squirrel Sciurus carolinensis is one such species that is widespread in the UK and is starting to spread across Europe; it predates birds' nests and can compete with birds for supplementary food. Using distance sampling across the urbanisation intensity gradient in Sheffield (UK) we test whether urban grey squirrels influence avian species richness and density through nest predation and competition for supplementary food sources. We also assess how urban bird assemblages respond to supplementary feeding. We find that grey squirrels slightly reduced the abundance of breeding bird species most sensitive to squirrel nest predation by reducing the beneficial impact of woodland cover. There was no evidence that grey squirrel presence altered relationships between supplementary feeding and avian assemblage structure. This may be because, somewhat surprisingly, supplementary feeding was not associated with the richness or density of wintering bird assemblages. These associations were positive during the summer, supporting advocacy to feed birds during the breeding season and not just winter, but explanatory capacity was limited. The amount of green space and its quality, assessed as canopy cover, had a stronger influence on avian species richness and population size than the presence of grey squirrels and supplementary feeding stations. Urban bird populations are thus more likely to benefit from investment in improving the availability of high quality habitats than controlling squirrel populations or increased investment in supplementary feeding.
Free Article
PMID: 25338062 [PubMed - in process]

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