Decreased glucose tolerance but normal blood glucose levels in the field in the caviomorph rodent Ctenomys talarum: the role of stress and physical activity

Hystricomorph rodents have a divergent insulin molecule with only 1-10% of the biological activity in comparison to other mammalian species. In this study, we used the subterranean rodent Ctenomys talarum as a model and performed blood glucose tolerance tests (GTTs) with trained and untrained individuals to evaluate blood glucose regulation and the possible role of physical activity as a compensatory mechanism. Additionally, we evaluated the variations in blood glucose during acute and chronic stress and gathered data in the field to evaluate natural-occurring variations in blood glucose levels. The GTTs showed that C. talarum have a diminished capacity of regulating blood glucose levels in comparison to other mammals and suggest that unexplored differences in the compensatory mechanisms, insulin structure and/or glucose transporters exist within species of hystricomorph rodents. However, blood glucose levels in the field stayed within the normal mammalian range. Physical activity did not prove to be a compensatory mechanism for blood glucose regulation. The individuals did not display important increases in blood glucose after acute stressors and managed to adequately regulate blood glucose during chronic stress. We suggest that the species may not face a selective pressure favoring a more tightly, mammalian like, capacity of regulating blood glucose levels.     

Phylogenetics of the speciose and chromosomally variable rodent genus Ctenomys (Ctenomyidae, Octodontoidea), based on mitochondrial cytochrome b sequences

The tuco-tucos [Ctenomys) are subterranean rodents that are widespread in southern South America. On the basis of its 60 living species, Ctenomys is one of the most speciose mammalian genera and displays great chromosomal variation. In order to study the mode of speciation in Ctenomys and to evaluate the role of chromosomes in cladogenesis, it is essential to generate an accurate phylogeny of the genus. From such a phylogeny it should be possible to identify particular species lineages worthy of further study. Following the success in 1998 of Lessa & Cook in generating a phylogeny of 10, mostly Bolivian, species from mitochondrial cytochrome b sequences, we sequenced a further 18, mostly Argentinian, species. By combining our datasi't with that of Lcssa & Cook's we were able to confirm six species lineages within Ctenomys. These lineages correspond well to taxonomic groups suggested on the basis of morphology and biogeographv. At least two of the species groups (the Boliviano-Matogrossense and Chaco) arc very variable chromosomally, and are worthwhile systems to examine the possibility of chromosomal spec iation. While, in general, multiple specimens of the named species did form monophyletic groups on the basis of the cytochrome b analysis, there were some exceptions. Thus, C. bergi and C yolandae did not form separate clades in the phylogenetic analysis, nor did C. roigi and C. dorbignyi. The molecular analysis was also very informative with regards species designations for C. boliviensis and related forms.     

A study of genetic distances and variability in several species pf the gemis Ctenomys (Rodentia: Octodontidae) with special references to a probable causal role of chromosomes in speciation

Genetic variability measured by allozymic electrophoresis has been studied in several species of the subterranean rodent genus Ctenomys (Octodontidae). The study was carried out with the main purpose of analysing a special group known as the 'Corrientes group' which inhabits the province of Corrientes in Argentina. The members of the group are, with high probability, isolated reproductively due to their karyotypic differences diploid numbers are between 42 and 70). To evaluate whether the chromosomal differences were the causes of speciation or if they arose a posteriori, we compared the level of the genetic distance among taxa within the orrientes group with all those measured between intraspecific populations in Ctenomys . The results indicate that the distances among the populations of the Corrientes group are at or below the level of those measured among traspecific populations in Ctenomys . Thus, it is unlikely that genetic differentiation triggered speciation. It is shown that this low level of genetic differentiation is not in contradiction with the high levels expected for species originating from one of the possible mechanisms of chromosomal speciation named, chromosomal transilience. Although for geographical reasons it seems obviousthat gene flow is precluded among the members of the Corrientes group, the Slatkin method for estimating Nm values was also used. Because high values of Nm exist but no isolation by distance could be detected, it is suggested that reproductive and geographical isolation are very ecent. Finally, analysis of population variability suggests that the high levels of heterozygosity observed (a) can be explained by the population structure, and (b) are within the range of expected values if bottlenecks have occurred in the recent history of the Corrientes group. As a general conclusion, the results indicate that in the Corrientes group the genetic data support a putative causal role for chromosomes in speciation.     

Molecular analysis of populations of Ctenomys (Caviomorpha,Rodentia) with high karyotypic variability

The tuco-tucos (Ctenomys) are South American subterranean rodents that are some of the most chromosomally variable of all mammals. In this study we focus on Ctenomys of the "Corrientes species group" from that Argentine province and consisting of C. dorbignyi, C. perrensi, C. roigi and unnamed populations (Ctenomys sp.). A diploid range of 41-70 has been demonstrated for these taxa with multiple Robertsonian changes, pericentric inversions, heterochromatic modifications and other chromosomal rearrangements. To analyse the molecular variation in the Corrientes group, a 402 base pair fragment of mitochondrial cytochrome b was sequenced in 75 individuals from 26 populations. This generated 15 different haplotypes which were subjected to phylogenetic analysis. The different species within the Corrientes group failed to form monophyletic groups in the phylogenetic trees we generated and the divergences between haplotypes were low. Therefore, these molecular data demonstrate the recent subdivision of the Corrientes group with a substantial accumulation of chromosomal differences. Through our new chromosomal data and cytochrome b studies, we are able to subdivide the Ctenomys sp. populations into two groups and hypothesize on their relationships with the described Corrientes group species. Further studies are needed to establish the basis of the phenomenal chromosomal variation in this group.     

Polymorphism of eimerian oocysts can be a problem in naturally infected hosts: an example from subterranean rodents in Bolivia

Since 1986, 364 tuco-tucos (Ctenomys spp.) representing 7 species were collected from 16 major collecting areas representing at least 4 distinct ecological habitats in Bolivia, South America. All were examined for coccidia, and 125 (34%) had oocysts in their feces including 84 of 236 (36%) Ctenomys boliviensis from tropical palm/savanna habitats; 1 of 3 (33%) Ctenomys conoveri from a chaco thorn forest; 3 of 7 (33%) Ctenomys frater from medium altitude grass habitats; and 6 of 8 (75%) Ctenomys lewisi and 31 of 35 (88%) Ctenomys opimus from high altitude/puna habitats. None of 3 Ctenomys leucodon (high altitude/puna) or 72 Ctenomys steinbachi (tropical palm/savanna) were passing oocysts when examined. The 5 infected host species all had oocysts of Eimeria opimi Lambert, Gardner, and Duszynski, 1988, in their feces. These oocysts and their sporocysts varied greatly in size, both within and between host species, but qualitative characters (e.g., residua and wall texture) remained constant. Our conclusion, that all oocysts seen were E. opimi, was supported by multigroup discriminant analysis of 256 individual oocysts, 30-67 selected randomly from each Ctenomys sp. Minimum polygons enclosing the centroid (= multivariate mean) and the spread of individuals for each species group (OTU) showed significant overlap in discriminant space, and Geisser classification showed a 55% miss rate of individuals being classified into the wrong OTUs. Thus, oocyst and sporocyst lengths and widths cannot be used to separate morphotypes of E. opimi from different Ctenomys spp. from different geographic regions of Bolivia.     

Cytogenetics Status of Four Ctenomys Species in the South of Brazil

This report is a review of 20 years of cytogenetics studies in tuco-tucos from the south of Brazil and the implications for the evolution of these species as well as the relation of these data with the geological history of the Coastal Plain of southern Brazil. Two forms of Ctenomys torquatus, 2n = 44 and 46, are separated by a geographic barrier. Ctenomys flamarioni shows a constant karyotype (2n = 48) and presents high variability in FN due to constitutive heterochromatin variation. Ctenomys minutus presents the highest chromosomic variation among the species in the south of Brazil (2n = 42–50) with three chromosomic hybrid zones. Ctenomys lami, like C. minutus, presents a high chromosomic variation due to fusions and fissions of chromosome pairs 1 and 2. Both species present a close evolutionary relationship, including a chromosomic hybrid zone.     

Energetics and thermoregulation during digging in the rodent tuco-tuco (Ctenomys talarum)

For subterranean rodents, searching for food by extension of the tunnel system and maintenance of body temperature are two of the most important factors affecting their life underground. In this study we assess the effect of ambient temperature on energetics and thermoregulation during digging in Ctenomys talarum. We measured VO2 during digging and resting at ambient temperature (Ta) below, within, and above thermoneutrality. Digging metabolic rate was lowest at Ta within the thermoneutral zone and increased at both lower and higher temperatures, but body temperature (Tb) remained constant at all Tas. Below thermoneutrality, the cost of digging and thermoregulation are additive. Heat production for thermoregulation would be compensated by heat produced as a by-product of muscular activity during digging. Above thermoneutrality, conduction would be an important mechanism to maintain a constant Tb during digging.     

Relationship between helminth parasites and demographic attributes of a population of the subterranean rodent Ctenomys talarum (Rodentla: Octodontidae)

The helminth parasite fauna of a natural population of the octodontid, Ctenomys talarum, was studied. Parasites that were found included the nematodes Heligmostrongylus sp. and Trichuris sp. Total prevalence of parasitism was 92.3%, mean intensity of infection was 22.7 worms, and mean abundance was 21 worms. Prevalence and mean abundance of infection with Heligmostrongylus sp. were higher in C. talarum males relative to females. Ecological and physiological causes, as well as the mating system of the host species, influence the likelihood of sex differences in parasite infection. The low parasite burden and diversity of C. talarum are associated with restrictions imposed by the subterranean habitat and with life-history traits of these rodents. Whether these findings apply to other Ctenomys spp. is unknown.     

Surface-mound formation by the Tuco-tuco, Ctenomys fulvus (Rodentia: Ctenomyidae), with comments on earth-pushing in other fossorial mammals

Surface-mound formation by Ctenomys fulvus (and evidently most Ctenomyidae, an autochthonous South American family) differs from most other fossorial mammals in that soil is transported to the surface by kicking with the hind feet (a habit shared with most Bathyergidae. the African mole-rats). The greatly enlarged hind feet of Ctenomys are the largest for any terrestrial fossorial group. The eyes of Ctenomys , also different from many mole types, are large and appear better protected by hind foot soil-pushing. The manner in which the stocky tail of Ctenomys supports the body above the substratum while kicking synchronously with the hind feet is unusual for any mammal.
The body facing forward while pushing soil, the position favoured by most mole-types, allows for the scanning of the surface for predators and defence through the use of teeth, claws and direct vocalizing, while hind foot soil-pushing conserves energy by negating the need to turn around in the tunnel after digging, and again after depositing soil on the surface. Studies are required to determine the efficiency of various predators on the capture of mole-types as soil is being pushed on to the surface, perhaps the most significant selective pressure on the manner of pushing soil.     

Karyotype evolution in South American subterranean rodents Ctenomys magellanicus (Rodentia: Octodontidae): chromosome rearrangements and (TTAGGG)n telomeric sequence localization in 2n=34 and 2n=36 chromosomal forms

Ctenomys is the most numerous genus of South American subterranean rodents and one of the most karyotypically diverse clades of mammals known. Ctenomys magellanicus is the southernmost species of the group and the only one living in Isla Grande de Tierra del Fuego (Argentina). This species presents two chromosomal forms, i.e. 2n=34, and 2n=36 (FN=68). Recent studies suggest that genetic divergence between both karyotypic forms resulted from a chromosomal speciation process. In order to identify the chromosomal rearrangement involved in the process of karyotype evolution in this species, we used chromosome banding techniques and fluorescence in situ hybridization with a telomeric probe to metaphase chromosomes of the two chromosomal forms of Ctenomys magellanicus. Chromosome analysis of Giemsa-stained and G-banding preparations showed that Cm34 and Cm36 karyotypes differ in one rearrangement involving chromosomes A9 from Cm34 and B12 and B17 from Cm36. In addition FISH analysis showed that all of the chromosomes from both chromosomal forms exhibit a telomeric-only distribution pattern of the (TTAGGG)n sequence, indicating that none of the chromosomal forms of Ctenomys magellanicus has true telocentric chromosomes. Our results suggest that a chromosome fission event would have occurred during the process of karyotype evolution in this species.     

Evidence for rolling-circle replication in a major satellite DNA from the South American rodents of the genus Ctenomys

A major PvuII satellite DNA has been cloned from a South American octodontid rodent of the genus Ctenomys (C. porteousi). The satellite monomer, termed RPCS, is 337 bp in size and 42% G + C. Analysis of the nucleotide sequence demonstrates that RPCS is not composed of a series of shorter repeats. RPCS-related sequences were found in 11 of 12 Ctenomys species analyzed by hybridization under high-stringency conditions. The only negative species, C. opimus, was reactive under low-stringency conditions. RPCS-related sequences were not found under high- or low-stringency conditions in Calomys musculinus and Mus musculus. However, under low-stringency conditions, RPCS-related sequences were found in the octodontid Octodontomys gliroides, which is thought to have diverged from the genus Ctenomys more than 10 Myr ago. The pattern of periodicities observed, by restriction analysis, between Ctenomys species in the satellite array can be mainly accounted for by a rolling-circle amplification mechanism but cannot be solely accounted for by unequal crossing-over.     

Chicken ceruloplasmin. Evidence in support of a trinuclear cluster involving type 2 and 3 copper centers

Ceruloplasmin was isolated to purity from chicken plasma by a single-step chromatography on amino-ethyl-derivatized Sepharose. Molecular mass, as estimated by nonreducing sodium dodecyl sulfate-electrophoresis, was approximately 140 kDa, slightly higher than that found for ceruloplasmins from other sources. Specific activity as p-phenylenediamine oxidase was five times higher than that reported for mammalian ceruloplasmins. The copper content was estimated to be 5.01 +/- 0.35 atoms per protein molecule, 50% of which was EPR-detectable. The EPR spectrum was completely devoid of any signal typical of the type 2 copper as seen in the other blue multicopper oxidases and in ceruloplasmin from mammalian species. Anaerobic reduction of chicken ceruloplasmin resulted in the disappearance of the 330 nm optical band typical of type 3 copper, which was followed by the appearance of an EPR signal typical of type 2 copper. Subsequently, the type 1 copper and finally the newly formed type 2 copper were reduced. The original optical and EPR spectra were recovered within few minutes upon exposure of reduced ceruloplasmin to air. It is concluded that in oxidized chicken ceruloplasmin type 2 copper interacts with the diamagnetic pair responsible for the 330 nm absorption in such a way as to become EPR-undetectable and that the interaction is relieved by reduction of the pair. Whether this interaction is intrinsically weaker in other blue oxidases and ceruloplasmins studied or is lost with standard preparation procedures remains to be established.     

Energy and distribution in subterranean rodents: sympatry between two species of the genus Ctenomys

The low basal metabolic rate (BMR) observed in subterranean rodents, compared to that of surface-dwelling species of comparable size, has been proposed to be an adaptation to underground life. Two main hypotheses have been proposed to explain this finding, the cost of burrowing and the thermal stress. The former states that the low BMR is due to the high cost of extending the tunnel system whereas the other relates it to the possibility of overheating in burrows where evaporative and convective heat exchange are restricted. Additionally, both hypotheses related the energetics of subterranean rodent with spatial distribution. The genus Ctenomys is an excellent model to evaluate the cost of burrowing or thermal stress, since they are widely distributed, with members differing markedly in body mass. The aim of this study was to assess digging and basal energetics in two Ctenomys species that live in sympatry in a coastal grassland, but differ in their microspatial distribution by soil preference. We used the obtained energetic data to test both energy-distribution hypotheses. We measured BMR and digging metabolic rate (DMR) through open flow respirometry in two species exposed to soft and hard soils. In brief, DMR in Ctenomys talarum (100-170 g), as in Ctenomys australis (250-600 g), was unaffected by soil hardness. Within thermoneutral zone of each species, DMR/RMR quotient was lower in the smaller species. Our data did not support the thermal stress hypothesis, but the cost of burrowing hypothesis was not rejected. Other alternative hypotheses are proposed to explain the distribution of C. talarum and C. australis.     

Number and Nuclear Localisation of Nucleoli in Mammalian Spermatocytes

In seven mammalian species, including man, the position and number of nucleoli in pachytene spermatocyte nuclei were studied from electron microscope (EM) nuclear sections or bivalent microspreads. The number and position of the nucleolar organiser regions (NORs) in mitotic and meiotic chromosomes were also analysed, using silver staining techniques and in situ hybridisation protocols. The general organisation of pachytene spermatocyte nucleoli was almost the same, with only minor morphological differences between species. The terminal NORs of Thylamys elegans (Didelphoidea, Marsupialia), Dromiciops gliroides (Microbiotheridae, Marsupialia), Phyllotys osgoodi (Rodentia, Muridae) and man, always gave rise to peripheral nucleoli in the spermatocyte nucleus. In turn, the intercalated NORs from Octodon degus, Ctenomys opimus (Rodentia, Octodontidae) and Chinchilla lanigera (Rodentia, Cavidae), gave rise to central nucleoli. In species with a single nucleolar bivalent, just one nucleolus is formed, while in those with multiple nucleolar bivalents a variable number of nucleoli are formed by association of different nucleolar bivalents or NORs that occupy the same nuclear peripheral space (Phyllotis and man). It can be concluded that the position of each nucleolus within the spermatocyte nucleus is mainly dependent upon: (1) the position of the NOR in the nucleolar bivalent synaptonemal complex (SC), (2) the nuclear pathway of the nucleolar bivalent SC, being both telomeric ends attached to the nuclear envelope, and (3) the association between nucleolar bivalents by means of their NOR-nucleolar domains that occupy the same nuclear space. Thus, the distribution of nucleoli within the nuclear space of spermatocytes is non-random and it is consistent with the existence of a species-specific meiotic nuclear architecture.     

Identification of duplicated fourth alpha2-adrenergic receptor subtype by cloning and mapping of five receptor genes in zebrafish

The alpha(2)-adrenergic receptors (alpha(2)-ARs) belong to the large family of rhodopsinlike G-protein-coupled receptors that share a common structure of seven transmembrane (TM) alpha-helices. The aims of this study were (1) to determine the number of alpha(2)-AR genes in a teleost fish, the zebrafish (Danio rerio), (2) to study the gene duplication events that generated the alpha(2)-AR subtypes, and (3) to study changes in receptor structure that have occurred since the divergence of the mammalian and fish lineages. Here, we report the cloning and chromosomal mapping of fish orthologs for all three mammalian alpha(2)-ARs. In addition, we identified a fourth alpha(2)-AR subtype with two duplicates in zebrafish. Chromosomal mapping showed that the zebrafish alpha(2)-AR genes are located within conserved chromosomal segments, consistent with the origin of the four alpha(2)-AR subtypes by two rounds of chromosome or block duplication before the divergence of the ray fin fish and tetrapod lineages. Thus, the fourth subtype has apparently been present in the common ancestor of vertebrates but has been deleted or is yet to be identified in mammals. The overall percentage identity between the fish and mammalian orthologs is 53% to 67%, and in the TM regions 80% to 87%. These values are clearly lower than what is observed between mammalian orthologs. Still, all of the residues thought to be important for alpha(2)-adrenergic ligand binding are conserved across species and subtypes, and even the most divergent regions of the fish receptors show clear "molecular fingerprints" typical for orthologs of a given subtype.     

Changes induced by parturition in serum cholesterol and triglycerides in obese and normal weight women

It is shown that total cholesterol levels in the blood before and immediately after delivery in obese and normal weighing women remain within normal values. Conversely blood triglycerides levels, higher than normal prior to delivery, are shown to drop back to normal values within three days (p 0.001). This is not thought to be attributable to labor stress.     

Observations on the cytochemistry of the hemocytes of an insect, Galleria mellonella

A number of cytochemical parameters of the hemocytes of larval Galleria mellonella, an insect frequently used as a model by comparative cellular immunologists, are described. Cytochemical methods were used to quantify hemocyte granule-associated components, the results are compared to those obtained for leukocytes from higher animals. Granulocytes contained a population of nonlysosomal granules rich in mucopolysaccharide not seen in plasmatocytes. The numbers and dimensions of these granules showed a positive correlation to cell size, probably reflecting a developmental sequence in granulocyte maturation. Both granulocytes and plasmatocytes had other granules containing the typical lysosomal enzymes, acid phosphatase, beta-glucuronidase, esterase, and lysozyme. The nonlysosomal enzyme alkaline phosphatase was not found in Galleria hemocytes; it is also absent from vertebrate monocytes, macrophages, and immature polymorphonuclear leukocytes. Insect hemocytes appear to lack certain components of antibacterial systems typical of mammalian blood cells, such as H2O2-generating systems, cationic proteins, and myeloperoxidase. The bactericidal mechanisms of hemocytes probably involve lysozyme, as well as other biologically active cellular and humoral factors unique to insects.     

Noninvasive, automatic 24-h ambulatory blood pressure monitoring in normotensive subjects

The reliability of noninvasive, automatic blood pressure monitoring is not yet clearly established. A 24-h ambulatory blood pressure profile was obtained in 9 healthy, normotensive subjects with an automatic, noninvasive device. The blood pressure profile showed the typical circadian pattern with lower systolic and diastolic values during sleep, although pulse pressure was fairly constant (about 40 mm Hg). The systolic blood pressure rose steeply in the early morning hours--before waking up. The results were compared with simultaneous hourly readings using the auscultatory method. There were no statistically significant differences between the automatic and auscultatory readings, 13 of the 18 mean values at different time points being within 2 mm Hg of each other. All the auscultatory means fell within the 95% confidence limits of those measured hourly by the automatic method. Although the automatic method seemed to be reliable compared with the auscultatory method, its sensitivity to motion artifacts is a disadvantage in a truly ambulatory setting.     

Dolphin ceruloplasmin: the first proteolytically stable mammalian ceruloplasmin.

1. Ceruloplasmin, the blue protein of the plasma of vertebrates, was isolated from dolphin, a marine mammal. The protein showed overall physico-chemical parameters very similar to those of all other mammalian ceruloplasmins. The spectroscopic properties indicated a conservation of the copper binding sites. 2. Non-denaturing electrophoresis revealed a conformation similar to that of other mammalian ceruloplasmins. EPR spectroscopy and calorimetric analyses indicated a three-domain arrangement of the protein typical of "aged" ceruloplasmin. 3. Dolphin ceruloplasmin is the only mammalian ceruloplasmin insensitive to trypsin, plasmin or chymotrypsin. This property, however, does not result in a higher conformational stability of the molecule. Thus, susceptibility of ceruloplasmin to aging is not directly related to the lability to proteases, which is typical of all other mammalian ceruloplasmins so far studied.     

Recurrent amplifications and deletions of satellite DNA accompanied chromosomal diversification in South American tuco-tucos (genus Ctenomys, Rodentia: Octodontidae): a phylogenetic approach

We investigated the relationship between satellite copy number and chromosomal evolution in tuco-tucos (genus Ctenomys), a karyotypically diverse clade of rodents. To explore phylogenetic relationships among 23 species and 5 undescribed forms, we sequenced the complete mitochondrial cytochrome b genes of 27 specimens and incorporated 27 previously published sequences. We then used quantitative dot-blot techniques to assess changes in the copy number of the major Ctenomys satellite DNA (satDNA), named RPCS. Our analysis of the relationship between variation in copy number of RPCS and chromosomal changes employed a maximum-likelihood approach to infer the copy number of the satellite RPCS in the ancestors of each clade. We found that amplifications and deletions of RPCS were associated with extensive chromosomal rearrangements even among closely related species. In contrast, RPCS copy number stability was observed within clades characterized by chromosomal stability. This example reinforces the suspected role of amplification, deletion, and intragenomic movement of satDNA in promoting extensive chromosomal evolution.