Effect of metoclopramide, ranitidine, and droperidol on the lower sphincter of the esophagus. Experimental study on the opossum (Didelphis albiventris)

Electromanometric measures of the gastroesophageal junction were performed in 20 adult, male and female, anesthetised opossums. The electromanometric examinations were performed according the intermitent pull through technique. The research was divided in four groups, according to the drug to be analysed: group 1 (20 animals) IM injection of physiological solution (control group); group 2 (20 animals) IM injection of metoclopramide; group 3 (20 animals) IM injection of ranitidine; group 4 (20 animals) IM injection of droperidol. Electromanometry was done 15 minutes before the drug injection, just after the injection and 15, 30, 45 and 60 minutes after the injection. In each one of the moments the pressure of the lower esophageal sphincter (LES-mmHg) was analysed. Considering LES pressure the results observed were: in group 1 it was not observed any significative alteration after IM injection of physiologic solution; in group 2 it was observed significative pressure increase, 15 minutes after metoclopramide IM injection; in group 3 it was observed pressure increase, being significative at 15 and 30 minutes after IM injection; in group 4 it was observed significative increase in LES pressure in every moment, 15 minutes after droperidol IM injection.

Morphological characteristics of neuromuscular junctions of the opossum (Didelphis albiventris) extraocular muscles: a scanning-electron-microscopic study

Three types of neuromuscular junctions were described in the extraocular muscles of the opossum. The present study demonstrates the three-dimensional characteristics of these neuromuscular junctions after HCl connective tissue digestion. Adult opossum of both sexes were used and the neuromuscular junctions of the extraocular muscles were examined after removal of the intramuscular connective tissue and basal layer. This material was examined with a scanning electron microscope. Two types of 'en plaque' neuromuscular junction were described: the continuous type revealed elongated and branched primary synaptic grooves separated from each other by sarcolemma protuberances with different sizes, and the discontinuous or punctiform type which presents very shallow and discontinuous grooves when compared with the former. The multiple neuromuscular junctions were observed as two or three junctions associated with the same muscular fiber. The multiple junctions were present in thin fibers (around 11 microm caliber); the en plaque junctions were associated with large diameter fibers (around 21 microm). This study confirms and reveals the detailed morphological characteristics of the three neuromuscular junction types previously described by transmission electron microscope in the extraocular muscles of opossum.

Macroscopic and microscopic analysis of the tongue of the common opossum (Didelphis marsupialis)

We performed a macroscopic and microscopic study of the tongues of common opossums, Didelphis marsupialis, from South America. We studied two males and two females. We collected morphometric data on the tongue with precision calipers. For the light microscopy and scanning electron microscopy analyses, we fixed tissue fragments in 10% formaldehyde and 2.5% glutaraldehyde, respectively. The opossum tongues averaged 5.87 +/- 0.20 cm in length, 3.27 +/- 0.15 cm in width at the lingual body, and 3.82 +/- 0.15 cm in width at the root. The mean thickness of the lingual body was 1.8 +/- 0.1 cm, and the thickness of the root was 3.82 +/- 0.15 cm. Sharp filiform papillae were scattered across the entire tongue; conical filiform papillae occurred on the lingual body and tongue tip; fungiform papillae were scattered among the filiform papillae on the lingual body and tongue tip; and there were three vallate papillae at the root of the tongue. We found two strands of papillary projections in the tongue root. Despite the low variability observed in the lingual papillae, the morphological data obtained in this study may be related to the opossum's diverse food habits and the extensive geographic distribution of the species throughout America. Microsc. Res. Tech. 2012. (c) 2012 Wiley Periodicals, Inc.

Senators from Pennsylvania. Memorial addresses delivered in the Senate and House of Representatives of the United States in memory of Philander C. Knox, Boies Penrose, William E. Crow, late senators from Pennsylvania. Sixty-seventh Congress. Proceedings in the Senate, January 28, 1923. Proceedings in the House, February 18, 1923.

Running title: Memorial addresses.

The lover's week: or, The six days adventures of Philander and Amaryllis.

Signatures: A-H⁴.

Speeches incident to the visit of Philander Chase Knox, Secretary of State of the United States of America, to the countries of the Caribbean. February 23 to April 17, 1912.

Mode of access: Internet.

Nedd4-2functionally interacts with ClC-5: involvement in constitutive albumin endocytosis in proximal tubule cells

Constitutive albumin uptake by the proximal tubule is achieved by a receptor-mediated process in which the Cl– channel, ClC-5, plays an obligate role. Here we investigated the functional interaction between ClC-5 and ubiquitin ligases Nedd4 and Nedd4-2 and their role in albumin uptake in opossum kidney proximal tubule (OK) cells. In vivo immunoprecipitation using an anti-HECT antibody demonstrated that ClC-5 bound to ubiquitin ligases, whereas glutathione S-transferase pull-downs confirmed that the C terminus of ClC-5 bound both Nedd4 and Nedd4-2. Nedd4-2 alone was able to alter ClC-5 currents in Xenopus oocytes by decreasing cell surface expression of ClC-5. In OK cells, a physiological concentration of albumin (10 μg/ml) rapidly increased cell surface expression of ClC-5, which was also accompanied by the ubiquitination of ClC-5. Albumin uptake was reduced by inhibiting either the lysosome or proteasome. Total levels of Nedd4-2 and proteasome activity also increased rapidly in response to albumin. Overexpression of ligase defective Nedd4-2 or knockdown of endogenous Nedd4-2 with small interfering RNA resulted in significant decreases in albumin uptake. In contrast, pathophysiological concentrations of albumin (100 and 1000 μg/ml) reduced the levels of ClC-5 and Nedd4-2 and the activity of the proteasome to the levels seen in the absence of albumin. These data demonstrate that normal constitutive uptake of albumin by the proximal tubule requires Nedd4-2, which may act via ubiquitination to shunt ClC-5 into the endocytic pathway.

Reconstruction of the ancestral marsupial karyotype from comparative gene maps

BACKGROUND: The increasing number of assembled mammalian genomes makes it possible to compare genome organisation across mammalian lineages and reconstruct chromosomes of the ancestral marsupial and therian (marsupial and eutherian) mammals. However, the reconstruction of ancestral genomes requires genome assemblies to be anchored to chromosomes. The recently sequenced tammar wallaby (Macropus eugenii) genome was assembled into over 300,000 contigs. We previously devised an efficient strategy for mapping large evolutionarily conserved blocks in non-model mammals, and applied this to determine the arrangement of conserved blocks on all wallaby chromosomes, thereby permitting comparative maps to be constructed and resolve the long debated issue between a 2n=14 and 2n=22 ancestral marsupial karyotype. RESULTS: We identified large blocks of genes conserved between human and opossum, and mapped genes corresponding to the ends of these blocks by fluorescence in situ hybridization (FISH). A total of 242 genes was assigned to wallaby chromosomes in the present study, bringing the total number of genes mapped to 554 and making it the most densely cytogenetically mapped marsupial genome. We used these gene assignments to construct comparative maps between wallaby and opossum, which uncovered many intrachromosomal rearrangements, particularly for genes found on wallaby chromosomes X and 3. Expanding comparisons to include chicken and human permitted the putative ancestral marsupial (2n=14) and therian mammal (2n=19) karyotypes to be reconstructed. CONCLUSIONS: Our physical mapping data for the tammar wallaby has uncovered the events shaping marsupial genomes and enabled us to predict the ancestral marsupial karyotype, supporting a 2n=14 ancestor. Futhermore, our predicted therian ancestral karyotype has helped to understand the evolution of the ancestral eutherian genome.

A transcriptome resource for the koala (Phascolarctos cinereus) : insights into koala retrovirus transcription and sequence diversity

Background The koala, Phascolarctos cinereus, is a biologically unique and evolutionarily distinct Australian arboreal marsupial. The goal of this study was to sequence the transcriptome from several tissues of two geographically separate koalas, and to create the first comprehensive catalog of annotated transcripts for this species, enabling detailed analysis of the unique attributes of this threatened native marsupial, including infection by the koala retrovirus. Results RNA-Seq data was generated from a range of tissues from one male and one female koala and assembled de novo into transcripts using Velvet-Oases. Transcript abundance in each tissue was estimated. Transcripts were searched for likely protein-coding regions and a non-redundant set of 117,563 putative protein sequences was produced. In similarity searches there were 84,907 (72%) sequences that aligned to at least one sequence in the NCBI nr protein database. The best alignments were to sequences from other marsupials. After applying a reciprocal best hit requirement of koala sequences to those from tammar wallaby, Tasmanian devil and the gray short-tailed opossum, we estimate that our transcriptome dataset represents approximately 15,000 koala genes. The marsupial alignment information was used to look for potential gene duplications and we report evidence for copy number expansion of the alpha amylase gene, and of an aldehyde reductase gene. Koala retrovirus (KoRV) transcripts were detected in the transcriptomes. These were analysed in detail and the structure of the spliced envelope gene transcript was determined. There was appreciable sequence diversity within KoRV, with 233 sites in the KoRV genome showing small insertions/deletions or single nucleotide polymorphisms. Both koalas had sequences from the KoRV-A subtype, but the male koala transcriptome has, in addition, sequences more closely related to the KoRV-B subtype. This is the first report of a KoRV-B-like sequence in a wild population. Conclusions This transcriptomic dataset is a useful resource for molecular genetic studies of the koala, for evolutionary genetic studies of marsupials, for validation and annotation of the koala genome sequence, and for investigation of koala retrovirus. Annotated transcripts can be browsed and queried at http://koalagenome.org

Molecular Tuning of Rhodopsins for High Visual Sensitivity in Different Light Environments : Variation in Absorbance Spectrum and Opsin Sequence within and between Species

Visual pigments of different animal species must have evolved at some stage to match the prevailing light environments, since all visual functions depend on their ability to absorb available photons and transduce the event into a reliable neural signal. There is a large literature on correlation between the light environment and spectral sensitivity between different fish species. However, little work has been done on evolutionary adaptation between separated populations within species. More generally, little is known about the rate of evolutionary adaptation to changing spectral environments. The objective of this thesis is to illuminate the constraints under which the evolutionary tuning of visual pigments works as evident in: scope, tempo, available molecular routes, and signal/noise trade-offs. Aquatic environments offer Nature s own laboratories for research on visual pigment properties, as naturally occurring light environments offer an enormous range of variation in both spectral composition and intensity. The present thesis focuses on the visual pigments that serve dim-light vision in two groups of model species, teleost fishes and mysid crustaceans. The geographical emphasis is in the brackish Baltic Sea area with its well-known postglacial isolation history and its aquatic fauna of both marine and fresh-water origin. The absorbance spectrum of the (single) dim-light visual pigment were recorded by microspectrophotometry (MSP) in single rods of 26 fish species and single rhabdoms of 8 opossum shrimp populations of the genus Mysis inhabiting marine, brackish or freshwater environments. Additionally, spectral sensitivity was determined from six Mysis populations by electroretinogram (ERG) recording. The rod opsin gene was sequenced in individuals of four allopatric populations of the sand goby (Pomatoschistus minutus). Rod opsins of two other goby species were investigated as outgroups for comparison. Rod absorbance spectra of the Baltic subspecies or populations of the primarily marine species herring (Clupea harengus membras), sand goby (P. minutus), and flounder (Platichthys flesus) were long-wavelength-shifted compared to their marine populations. The spectral shifts are consistent with adaptation for improved quantum catch (QC) as well as improved signal-to-noise ratio (SNR) of vision in the Baltic light environment. Since the chromophore of the pigment was pure A1 in all cases, this has apparently been achieved by evolutionary tuning of the opsin visual pigment. By contrast, no opsin-based differences were evident between lake and sea populations of species of fresh-water origin, which can tune their pigment by varying chromophore ratios. A more detailed analysis of differences in absorbance spectra and opsin sequence between and within populations was conducted using the sand goby as model species. Four allopatric populations from the Baltic Sea (B), Swedish west coast (S), English Channel (E), and Adriatic Sea (A) were examined. Rod absorbance spectra, characterized by the wavelength of maximum absorbance (λmax), differed between populations and correlated with differences in the spectral light transmission of the respective water bodies. The greatest λmax shift as well as the greatest opsin sequence difference was between the Baltic and the Adriatic populations. The significant within-population variation of the Baltic λmax values (506-511 nm) was analyzed on the level of individuals and was shown to correlate well with opsin sequence substitutions. The sequences of individuals with λmax at shorter wavelengths were identical to that of the Swedish population, whereas those with λmax at longer wavelengths additionally had substitution F261F/Y in the sixth transmembrane helix of the protein. This substitution (Y261) was also present in the Baltic common gobies and is known to redshift spectra. The tuning mechanism of the long-wavelength type Baltic sand gobies is assumed to be the co-expression of F261 and Y261 in all rods to produce ≈ 5 nm redshift. The polymorphism of the Baltic sand goby population possibly indicates ambiguous selection pressures in the Baltic Sea. The visual pigments of all lake populations of the opossum shrimp (Mysis relicta) were red-shifted by 25 nm compared with all Baltic Sea populations. This is calculated to confer a significant advantage in both QC and SNR in many humus-rich lakes with reddish water. Since only A2 chromophore was present, the differences obviously reflect evolutionary tuning of the visual protein, the opsin. The changes have occurred within the ca. 9000 years that the lakes have been isolated from the Sea after the most recent glaciation. At present, it seems that the mechanism explaining the spectral differences between lake and sea populations is not an amino acid substitution at any other conventional tuning site, but the mechanism is yet to be found.

Structure and activity cyclase activated of OK-GC: A kidney receptor-guanylate cyclase activated by guanylin peptides

Uroguanylin, guanylin, and lymphoguanylin are small peptides that activate renal and intestinal receptor guanylate cyclases (GC). They are structurally similar to bacterial heat-stable enterotoxins (ST) that cause secretory diarrhea. Uroguanylin, guanylin, and ST elicit natriuresis, kaliuresis, and diuresis by direct actions on kidney GC receptors. A 3,762-bp cDNA characterizing a uroguanylin/guanylin/ST receptor was isolated from opossum kidney (OK) cell RNA/cDNA. This kidney cDNA (OK-GC) encodes a mature protein containing 1,049 residues sharing 72.4�75.8% identity with rat, human, and porcine forms of intestinal GC-C receptors. COS or HEK-293 cells expressing OK-GC receptor protein were activated by uroguanylin, guanylin, or ST13 peptides. The 3.8-kb OK-GC mRNA transcript is most abundant in the kidney cortex and intestinal mucosa, with lower mRNA levels observed in urinary bladder, adrenal gland, and myocardium and with no detectable transcripts in skin or stomach mucosa. We propose that OK-GC receptor GC participates in a renal mechanism of action for uroguanylin and/or guanylin in the physiological regulation of urinary sodium, potassium, and water excretion. This renal tubular receptor GC may be a target for circulating uroguanylin in an endocrine link between the intestine and kidney and/or participate in an intrarenal paracrine mechanism for regulation of kidney function via the intracellular second messenger, cGMP.

Structure and activity of OK-GC: a kidney receptor guanylate cyclase activated by guanylin peptides

Uroguanylin, guanylin, and lymphoguanylin are small peptides that activate renal and intestinal receptor guanylate cyclases (GC). They are structurally similar to bacterial heat-stable enterotoxins (ST) that cause secretory diarrhea. Uroguanylin, guanylin, and ST elicit natriuresis, kaliuresis, and diuresis by direct actions on kidney GC receptors. A 3,762-bp cDNA characterizing a uroguanylin/guanylin/ST receptor was isolated from opossum kidney (OK) cell RNA/cDNA. This kidney cDNA (OK-GC) encodes a mature protein containing 1,049 residues sharing 72.4-75.8% identity with rat, human, and porcine forms of intestinal GC-C receptors. COS or HEK-293 cells expressing OK-GC receptor protein were activated by uroguanylin, guanylin, or ST13 peptides. The 3.8-kb OK-GC mRNA transcript is most abundant in the kidney cortex and intestinal mucosa, with lower mRNA levels observed in urinary bladder, adrenal gland, and myocardium and with no detectable transcripts in skin or stomach mucosa. We propose that OK-GC receptor GC participates in a renal mechanism of action for uroguanylin and/or guanylin in the physiological regulation of urinary sodium, potassium, and water excretion. This renal tubular receptor GC may be a target for circulating uroguanylin in an endocrine link between the intestine and kidney and/or participate in an intrarenal paracrine mechanism for regulation of kidney function via the intracellular second messenger, cGMP.

Efeitos do controle da jaqueira, Artocarpus heterophyllus L., sobre a comunidade de pequenos mamíferos e a rede de dispersão de sementes na Ilha Grande, RJ

Atualmente observa-se uma expressiva perda de biodiversidade global resultante de atividades antrópicas, sendo a introdução de espécies exóticas uma das mais impactantes. A jaqueira Artocarpus heterophyllus é uma espécie exótica introduzida no Brasil durante o período colonial, sendo considerada invasora em diversas localidades. Na Mata Atlântica invade áreas de mata aberta e de borda, habitualmente associadas a ambientes antrópicos. Na Ilha Grande é encontrada em grande abundância em decorrência do histórico de ocupação humana. Para compreender como a mastofauna responde a presença da jaqueira, o Laboratório de Ecologia de Mamíferos da Universidade do Estado do Rio de Janeiro (UERJ) vem desenvolvendo um estudo ao longo de seis anos nos arredores da Vila Dois Rios, localizada na face oceânica da Ilha Grande. A partir dos resultados prévios iniciou-se uma segunda etapa do estudo no mesmo local que buscou avaliar diferentes métodos de controle das jaqueiras. O presente estudo é uma continuação direta desses dois trabalhos anteriores e teve como objetivo acompanhar as respostas da comunidade de pequenos mamíferos no período imediatamente posterior ao controle. Durante 18 meses foram amostradas bimestralmente 18 grades, 10 aonde foi efetuado o controle das jaqueiras e 8 aonde não foi constatada a presença desta árvore. Em cada grade foram colocadas 11 armadilhas de captura viva sendo banana a isca utilizada. Os mamíferos capturados foram medidos e suas fezes coletadas. A quantidade de jacas em cada área também foi anotada bimensalmente. As fezes foram analisadas em laboratório e as sementes encontradas identificadas. Os resultados obtidos indicam que a influência de A. heterophyllus sobre a estrutura da comunidade de pequenos mamíferos foi menor após o tratamento de controle. A única espécie que parece ainda responder a abundância de jaqueiras é o roedor Trinomys dimidiatus, que apresentou densidades mais elevadas nas áreas em tratamento, porém mais próximas a resultados obtidos para espécies congêneres em áreas pouco antropizadas. Utilizando uma abordagem de redes complexas observamos que, embora T. dimidiatus seja a espécie mais abundante em termos de número de indivíduos, o gambá Didelphis aurita parece ser a espécie de mamífero mais importante para dispersão de sementes nativas, aparecendo como espécie com maior número de conexões com espécies de sementes nas redes contruídas para as áreas sem jaqueiras e com jaqueiras antes e após o tratamento. Finalmente, a partir dos dados obtidos criamos um modelo matemático para a população de T. dimidiatus dos arredores da Vila Dois Rios, baseado em um crescimento logístico. Os resultados do modelo proposto se mostraram correlacionados com os dados de abundância reais, de modo que ele parece ser um simulador adequado da população local.