Developmental plasticity of ascending spinal axons studies using the North American opossum, Didelphis virginiana

The polymerase chain reaction was used to characterize the trypanosomes infecting Glossina morsitans submorsitans and G. tachinoides in the game ranch of Nazinga, Burkina Faso, situated near an agropastoral zone. Dissection of 435 tsetse flies, and PCR analysis of 166 infected flies were conducted to assess the epidemiological situation. Trypanosomes of the Nannomonas subgenus were the most abundant in the two tsetse species (80.4% and 73.7% of identified infections in G. m. submorsitans and G. tachinoides respectively). T. vivax and T. brucei infection rates were comparable between the two tsetse species. Mature infection pattern identified by PCR differed from overall infections, mainly because T. simiae infections did not mature, whereas T. vivax represented the predominant taxon. Parasitological and PCR results showed some discrepancies; possibly some typical Duttonella strains could not be recognized by the sets of primers used. The technologies used in this work helped to determine the high trypanosomosis risk in this area.     

Evidence for GAP-43 within descending spinal axons in the North American opossum, Didelphis virginiana

We have shown previously that GAP-43, a growth associated protein characteristically present in growing and regenerating axons, is relatively abundant in the spinal cord of adult opossums. In the present study, we combined the orthograde transport of the fluorescent marker Fluoro-Ruby with immunofluorescence for GAP-43 to determine if any of it is present within descending spinal axons. When Fluoro-Ruby was injected into the red nucleus and midbrain tegmentum, the medial pontine or medullary reticular formation, the medullary raphe or the lateral vestibular nucleus, axons were labeled in the expected areas of the spinal cord, but in most cases none showed evidence for GAP-43. In two of the four cases with rubral injections, however, a few labeled axons within the rubrospinal tract showed GAP-43 immunofluorescence, and in one case with an injection of the gigantocellular reticular nucleus and adjacent raphe, labeled axons within lamina IX immunostained for the protein. Since serotoninergic neurons are present within the gigantocellular reticular nucleus and adjacent raphe, and axons of the same phenotype are abundant within lamina IX, we asked whether serotoninergic axons contain GAP-43. When sections of the spinal cord were immunostained for both serotonin and GAP-43, many axons within lamina IX showed evidence for both substances. Such axons appeared to contact presumptive motoneurons. In cases with Fluoro-Ruby injections of the forelimb motor cortex, labeled axons were present within the pyramidal tract, and some of them showed evidence for GAP-43.     

Origin, course, and laterality of spinocerebellar axons in the North American opossum, Didelphis virginiana

Inhibition of neurons containing gamma-aminobutyric acid (GABA) may underlie some of the excitatory effects of opioids in the central nervous system (CNS). In the present study, we examined the relationship of the cloned mu- and delta-opioid receptors (MOR1 and DOR1, respectively) to GABAergic neurons in brain and spinal cord. This was done by combining immunofluorescent staining for MOR1 or DOR1 with that for GABA or glutamic acid decarboxylase (GAD); fluorescent retrograde tract-tracing was used in some cases to identify neurons with particular projections. In rats, cells double labeled for GABA and MOR1 were observed in layers II-VI of the parietal cortex and in layers II-IV of the piriform cortex. In the hippocampus, double labeling was observed in the dentate gyrus and in regions CA1 and CA3. Double labeling was very prominent in the striatum and in the reticular nucleus of the thalamus; it was also observed in other portions of the diencephalon. However, double labeling for GABA and MOR1 was never observed in the cerebellar cortex. Cells double labeled for GABA and MOR1 were common in the periaqueductal gray (PAG) and the medial rostral ventral medulla (RVM) of both rats and monkeys, suggesting that involvement of GABAergic neurons with supraspinal opioid antinociception may extend to primates. In the RVM of rats, many of those double-labeled neurons were retrogradely labeled from the dorsal spinal cord. In contrast, double-labeled neurons in the PAG were almost never retrogradely labeled from the RVM. No unequivocal examples of double labeling for DOR1 and GAD were found in any region of the CNS that we examined in either rats or monkeys. However, GABAergic neurons were often apposed by DOR1 immunoreactive varicosities. Our findings suggest that activation of mu-opioid receptors directly modulates the activity of GABAergic neurons throughout the CNS, including neurons involved in the supraspinal component of opioid analgesia. In contrast, delta-opioid receptors appear to be positioned to modulate the activity of GABAergic neurons indirectly.     

North American opossum Didelphis virginiana as a fetal nephrotoxicity model: histologic and ultrastructural assessment of uranyl nitrate (UN)-induced damage

Individuals report a variety of reasons for having sex. Understanding these reasons can improve HIV and STD prevention efforts because they may constitute an important component in the aetiology of sexual risk-taking behaviours. Relationships between self-reported reasons for having sex and frequency of participation in sexual practices among 146 heterosexual men recruited from public STD clinics in Southern California were examined. Using a self-administered questionnaire, respondents reported how often they engaged in sex for each of 16 reasons and how frequently they participated in high, moderate, and low-risk sexual practices. A principal components analysis identified five factors used to construct scales: love; compliance; pleasure; altered states; and potency. Higher-risk sexual practices were positively associated with the pleasure and potency scales, whereas lower-risk practices were positively associated with the love scale. These findings suggest that some reasons men report for having sex may influence sexual risk-taking. Interventions to reduce unsafe sex should explicitly address how men can practise safer sex and still experience pleasure and potency.     

The detection of hemorrhagic proteins in snake venoms using monoclonal antibodies against Virginia opossum (Didelphis virginiana) serum

Most snakes and a few warm-blooded animals have a resistance to snake venoms because of naturally occurring antihemorrhagins found in their sera. The antihemorrhagins in serum of Virginia opossum (Didelphis virginiana) neutralize hemorrhagic activity by binding to hemorrhagins in snake venoms. The binding characteristic of antihemorrhagins in D. virginiana serum was used to develop a five-step western blot. The detection of hemorrhagic proteins were measured indirectly with antihemorrhagins in Virginia opossum serum and with DV-2LD#2, a monoclonal antibody specific for Virginia opossum antihemorrhagins. Snake venoms were separated by native-PAGE, transferred to a Millipore Immobilon-P membrane and then incubated with crude Virginia opossum serum. The hemorrhagins in snake venom bind to antihemorrhagins in Virginia opossum serum which react with DV-2LD#2 a monoclonal antibody that is specific for Virginia opossum antihemorrhagins. DV-2LD#2 monoclonal antibody inhibits antihemorrhagic activity in Virginia opossum serum when mixed in equal amounts. The inhibition of antihemorrhagins by DV-2LD#2 monoclonal antibody suggests specificity. DV-2LD#2 monoclonal antibody does not recognize antihemorrhagins in gray woodrat (Neotoma micropus) serum. The five-step western blot reveals two well-defined bands which represent hemorrhagins found in Western diamondback rattlesnake (Crotalus atrox) venom. Venoms from 15 different snake species were examined to determine the usefulness of the five-step western blot. Other hemorrhagic venoms (Great Basin rattlesnake (C. viridis lutosus), Prairie rattlesnake (C. viridis viridis), Tancitaran dusky rattlesnake (C. pusillus), Northern Mojave rattlesnake (C. scutulatus scutulatus type B) and Northern Pacific rattlesnake (C. v. oreganus)) had one single band in the five-step western blot. DV-2LD#2 did not bind to the non-hemorrhagic venoms and reacted with 50% of the hemorrhagic venoms used in this study. The monoclonal antibody, CAH, reacted with all the hemorrhagic venoms except for the venom of the King cobra (Ophiophagus hannah) and did not react with the non-hemorrhagic venoms. The hemorrhagic binding site of CAH monoclonal antibody and the antihemorrhagin in Virginia opossum are different binding sites. The five-step western blot will be a very useful assay for determining hemorrhagic activity without using live animals.     

Inhibitory properties of the antibothropic complex from the South American opossum (Didelphis marsupialis) serum

The South American opossum Didelphis marsupialis is known to be highly resistant to snake envenomation. In this paper it is shown that the opossum serum inhibits haemorrhage induced by both Crotalinae and Viperinae venoms. Tested against Bothrops jararaca (jararaca) venom, the antibothropic complex (ABC) isolated from the opossum serum was at least six times more antihaemorrhagic than the commercial antivenom. ABC showed no proteolytic activity by itself and was not hydrolysed by the venom. It inhibited the hydrolysis of casein by B. jararaca venom, but did not inhibit its hydrolytic activities upon N alpha-benzoyl-L-arginine ethyl ester (BAEE) and N alpha-benzoyl-DL-arginine p-nitroanilide (BAPNA). The inhibitor did not interfere with trypsin and bacterial collagenase activities on BAPNA and N-(3-[2-furyl]acryloyl)-Leu-Gly-Pro-Ala (FALGPA), respectively. It reduced chymotrypsin hydrolysis of N-acetyl-L-tyrosine ethyl ester (ATEE) because ABC is also a substrate for this enzyme. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis, B. jararaca venom preferentially degraded fibrinogen A alpha-chain and fibrin alpha-chain. Tested on extracellular matrix proteins, the venom hydrolysed collagen IV, gelatins I and V, laminin and fibronectin, besides depolimerizing collagen I alpha-chain dimers. Fibrillar collagen V was not digested. These hydrolyses were inhibited by ABC and by EDTA. Our results show that the antibothropic complex is a venom metalloproteinase inhibitor, which could, at least partially, account for its antihaemorrhagic activity. Electrophoretic evidence indicated non-covalent complex formation between the antihaemorrhagic factor and component(s) of B. jararaca venom.     

Ultrastructural pattern of glucagon producing-cells in the gastric mucosa of the developing opossum Didelphis albiventris (Marsupialia)

We present the case of a 59-year-old woman with an International Federation of Gynecology and Obstetrics stage IIIB serous psammocarcinoma. Only 12 cases of this rare ovarian neoplasm have been reported previously. These tumors act like borderline tumors and, therefore, do not usually require chemotherapy or radiation therapy after appropriate surgical debulking and staging.     

Isolation in immunodeficient mice of Sarcocystis neurona from opossum (Didelphis virginiana) faeces, and its differentiation from Sarcocystis falcatula

BACKGROUND: There has been conflicting evidence of the effect of angiotensin-converting enzyme (ACE) inhibitors on exercise tolerance. Meta-analysis of published results has suggested that a beneficial effect of ACE inhibitors is demonstrated if a trial design is adequate. SETTING: Multicentre International Trial. METHODS: In a double-blind, randomized, multicentre trial, 292 patients with moderate (New York Heart Association Grades II and III) heart failure were treated with trandolapril or placebo in addition to diuretics, and followed for 16 weeks. Exercise tolerance on a treadmill was assessed at baseline and after 4, 8, 12 and 16 weeks of treatment. Both a modified Bruce and a modified Naughton protocol were used. RESULTS: Exercise tolerance improved in both treatment groups, with no significant benefit from trandolapril treatment. CONCLUSION: Trandolapril does not improve exercise tolerance as measured by treadmill testing.     

The effect of enzyme replacement on red cell adenine deoxyribonucleotides in adenosine deaminase-deficient erythrocytes of the opossum, Didelphis virginiana

1. Polyethyleneglycol-modified bovine adenosine deaminase was administered (10-20 U/kg/week) intramuscularly to two opossums for 15 weeks and changes in red cell adenine ribo- and deoxyribonucleotides quantitated by HPLC. 2. Only a moderate decline of erythrocyte dAXP was observed at the end of the study when compared to results of enzyme replacement seen in human adenosine deaminase deficient patients. 3. Opossum red cells salvage substantial amounts of deoxyadenosine provided in physiologic (50 nM) concentration from plasma having either low or high adenosine deaminase activity.     

Features of myoneural junctions in the extraocular muscles of the opossum (Didelphis albiventris)

The myoneural junctions present in the opossum (Didelphis albiventris) extraocular muscles were studied through histochemical and ultrastructural techniques. Three different types of junction were shown: two single types, 'en grappe' and 'en plaque', present in the middle third of the muscle fibers; and one multiple type, more rare and observed in the distal third of the muscle.     

Adult opossums (Didelphis virginiana) demonstrate near normal locomotion after spinal cord transection as neonates

When the thoracic spinal cord of the North American opossum (Didelphis virginiana) is transected on postnatal day (PD) 5, the site of injury becomes bridged by histologically recognizable spinal cord and axons which form major long tracts grow through the lesion. In the present study we asked whether opossums lesioned on PD5 have normal use of the hindlimbs as adults and, if so, whether that use is dependent upon axons which grow through the lesion site. The thoracic spinal cord was transected on PD5 and 6 months later, hindlimb function was evaluated using the Basso, Beattie, and Bresnahan (BBB) locomotor scale. All animals supported their weight with the hindlimbs and used their hindlimbs normally during overground locomotion. In some cases, the spinal cord was retransected at the original lesion site or just caudal to it 6 months after the original transection and paralysis of the hindlimbs ensued. Surprisingly, however, these animals gradually recovered some ability to support their weight and to step with the hindlimbs. Similar recovery was not seen in animals transected only as adults. In order to verify that descending axons which grew through the lesion during development were still present in the adult animal, opossums subjected to transection of the thoracic cord on PD5 were reoperated and Fast blue was injected several segments caudal to the lesion. In all cases, neurons were labeled rostral to the lesion in each of the spinal and supraspinal nuclei labeled by comparable injections in unlesioned, age-matched controls. The results of orthograde tracing studies indicated that axons which grew through the lesion innervated areas that were appropriate for them.     

Photoreceptors in a primitive mammal, the South American opossum, Didelphis marsupialis aurita: characterization with anti-opsin immunolabeling

The retinas of placental mammals appear to lack the large number and morphological diversity of cone subtypes found in diurnal reptiles. We have now studied the photoreceptor layer of a South American marsupial (Didelphis marsupialis aurita) by peanut agglutinin labeling of the cone sheath and by labeling of cone outer segments with monoclonal anti-visual pigment antibodies that have been proven to consistently label middle-to-long wavelength (COS-1) and short-wavelength (OS-2) cone subpopulations in placental mammals. Besides a dominant rod population (max. = 400,000/mm2) four subtypes of cones (max. = 3000/mm2) were identified. The outer segments of three cone subtypes were labeled by COS-1: a double cone with a principal cone containing a colorless oil droplet, a single cone with oil droplet, and another single cone. A second group of single cones lacking oil droplets was labeled by OS-2 antibody. The topography of these cone subtypes showed striking anisotropies. The COS-1 labeled single cones without oil droplets were found all over the retina and constituted the dominant population in the area centralis located in the temporal quadrant of the upper, tapetal hemisphere. The population of OS-2 labeled cones was also ubiquitous although slightly higher in the upper hemisphere (200/mm2). The COS-1 labeled cones bearing an oil droplet, including the principal member of double cones, were concentrated (800/mm2) in the inferior, non-tapetal half of the retina. The two spectral types of single cones resemble those of dichromatic photopic systems in most placental mammals. The additional set of COS-1 labeled cones is a distinct marsupial feature. The presence of oil droplets in this cone subpopulation, its absence in the area centralis, and the correlation with the non-tapetal inferior hemisphere suggest a functional specialization, possibly for mesopic conditions. Thus, sauropsid features have been retained but probably with a modified function.     

Isolation of protein factors from opossum (Didelphis albiventris) serum which protect against Bothrops jararaca venom

The fractionation of Didelphis albiventris serum by DEAE-Sephadex A50 yields a fraction (DA2) which protects the opossum against Bothrops venom. One polypeptide (DA2-II) responsible for this protection was isolated from fraction DA2 by ion exchange chromatography and biochemically characterized. DA2-II is a 43,000 mol. wt glycoprotein with the following N-terminal sequence: LKAMDTTPPLKIKKEPVK. Pairwise comparison of the amino acid sequence with four anti-hemorrhagic factors isolated from other opossum species indicated that DA2-II possesses high similarity (60-80%) with these proteins.     

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.