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PubMed1995

Ontogeny of arginine vasopressin-like immunoreactivity in the Brazilian opossum brain.

The neuropeptide arginine vasopressin is involved in many centrally mediated functions and brain development. In this study, we have examined the ontogeny of arginine vasopressin-like immunoreactivity (AVP-IR) in the Brazilian opossum (Monodelphis domestica) brain to further understand the involvement of AVP in the forming central nervous system. Monodelphis is a small pouchless marsupial and its pups are born in an extremely immature state before neurogenesis is completed. In the adult brain, cell bodies containing AVP-IR were found in several nuclear groups and areas, and immunoreactive fibers were found to be widely distributed throughout the brain. The distribution of AVP-IR in the adult opossum brain generally resembled that reported for other species including the rat, however, some differences in localization of immunoreactive cells were observed. In the developing opossum brain, AVP-IR was first seen in the mesencephalon and diencephalon between embryonic days 12 and 13. Subsequently, a distinct group of AVP immunoreactive cells was present in the forming supraoptic nucleus on day 1 of postnatal life (1 PN) and at 3 PN in the paraventricular nucleus. Between 1 and 3 PN, a few cells transiently expressed AVP-IR in the forming thalamus and tegmental area. At these ages a few immunoreactive fibers were also detected in the forming cerebellum. These fibers were not seen at later ages in these areas. By 5 PN, an increased expression of AVP-IR was seen in the forming supraoptic and paraventricular hypothalamic nuclei, median eminence, and posterior pituitary. At 7 PN, immunoreactive cells and fibers were seen in several forebrain areas. The distribution pattern of AVP-IR became adult-like by 60 PN. A sex difference in the amount of AVP-IR in the lateral septum was also observed in the opossum brain at 60 PN. This difference persisted in the adult brain. Due to the early presence of AVP-IR in the Monodelphis brain before neurogenesis and morphogenesis is completed, we suggest that AVP may be involved in morphogenesis of the central nervous system. In addition, AVP may have a significant physiological function in regard to homeostasis before the forebrain contributes to these control mechanisms. Further studies, including physiological and developmental manipulations, will define the significance of the early presence of AVP during the differentiation and maturation of the central nervous system in Monodelphis.

Animals↗

PubMed1986

Efferent projections of the basolateral amygdala in the opossum, Didelphis virginiana.

The autoradiographic anterograde axonal transport technique was used to study efferent projections of the opossum basolateral amygdala. All nuclei of the basolateral amygdala send topographically organized fibers to the bed nucleus of the stria terminalis (BST) via the stria terminalis (ST). Injections into rostrolateral portions of the basal nuclei label fibers that surround the commissural bundle of the ST, cross the midline by passing along the outer aspect of the anterior commissure, and terminate primarily in the contralateral BST, anterior subdivision of the basolateral nucleus (BLa), ventral putamen, and olfactory cortex. Each of the basal nuclei project ipsilaterally to the anterior amygdaloid area, substantia innominata and topographically to the ventral part of the striatum and adjacent olfactory tubercle. The posterior subdivision of the basolateral nucleus (BLp), but not the basomedial nucleus (BM), projects to the ventromedial hypothalamic nucleus. BLa and BLp have projections to the nucleus of the lateral olfactory tract and also send fibers to the central nucleus, as does the lateral nucleus (L). The lateral nucleus also has a strong projection to BM and both nuclei project to the amygdalo-hippocampal area. BLa and BLp send axons to the ventral subiculum and ventral lateral entorhinal area whereas L projects only to the latter area. The lateral nucleus and BLp project to the perirhinal cortex and the posterior agranular insular area. The BLa sends efferents to the anterior agranular insular area. Rostrally this projection is continuous with a projection to the entire frontal cortex located rostral and medial to the orbital sulcus. All of the nuclei of the basolateral amygdala project to areas on the medial wall of the frontal lobe that appear to correspond to the prelimbic and infralimbic areas of other mammals. Despite the great phylogenetic distance separating the opossum from placental mammals, the projections of the opossum basolateral amygdala are very similar to those seen in other mammals. The unique frontal projections of the opossum BLa to the dorsolateral prefrontal cortex appear to be related to the distinctive organization of the mediodorsal thalamic nucleus and prefrontal cortex in this species.

Amygdala↗

PubMed1998

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

Sarcocystis neurona was isolated in nude mice and gamma-interferon knockout mice fed sporocysts from faeces of naturally infected opossums (Didelphis virginiana). Mice fed sporocysts became lethargic and developed encephalitis. Protozoa were first found in the brain starting 21 days post-inoculation. Sarcocystis neurona was recovered in cell culture from the homogenate of liver, spleen and brain of a nude mouse 11 days after feeding sporocysts. The protozoa in mouse brain and in cell culture multiplied by schizogony and mature schizonts often had a residual body. Sarcocystis falcatula, which has an avian-opossum cycle, was not infective to nude or knockout mice. Protozoa were not found in tissues of nude mice or knockout mice after subcutaneous injection with culture-derived S. falcatula merozoites and sporocysts from the faeces of opossums presumed to contain only S. falcatula. Results demonstrate that S. neurona is distinct from S. falcatula, and that opossums are hosts for both species.

Animals↗

PubMed1999

New methodology for the obtainment of antibothropic factors from the South American opossum (Didelphis marsupialis) and jararaca snake (Bothrops jararaca).

The antibothropic factor (ABF) from D. marsupialis was collected from perforated hollow plastic golf balls which were surgically implanted subcutaneously in anesthetized opossums, a technique originally described for the production of polyclonal antibodies. Two months after the implantation of the balls, approximately 15 ml of seromatous fluid from D. marsupialis (SFDm-50 mg total protein/ml) could be recovered monthly. Opossum serum as well as SFDm showed similar SDS-PAGE profiles and antihemorrhagic potencies against Bothrops jararaca snake venom (Bjv). The presence of ABF in SFDm was confirmed by immunoblotting, using rabbit polyclonal antibodies raised against ABF isolated from opossum serum. ABF isolated from SFDm or from serum by ion-exchange chromatography showed identical chromatographic and electrophoretic profiles. ABF fromboth sources displayed very similar antihemorrhagic and anticaseinolytic activities against Bjv. In the case of B. jararaca, polyethylene perforated tubes were inserted in the abdominal cavity and two months after implantation, approximately 4 ml of seromatous fluid from B. jararaca (SFBj-23 mg total protein/ml) were recovered. B.jararaca serum and SFBj showed the same native and SDS-PAGE band pattern. Both serum and SFBj inhibited Bjv hemorrhagic activity. We conclude that this new methodology is very suitable for continuously obtaining opossum ABF and SFBj, in large scale and in an easier way, avoiding animal suffering and eventual sacrifice.

Animals↗

PubMed1999

Development of the facial and hypoglossal motor nuclei in the neonatal Brazilian opossum brain.

The development of the facial and hypoglossal motor nuclei were examined in the neonatal Brazilian opossum (Monodelphis domestica), a marsupial in which postnatal central nervous system development has been well characterized. In this study, we utilized postnatal injection of the retrograde tracer cholera toxin subunit B (CtB) to characterize the formation of the facial and hypoglossal motor nuclei in the developing neonatal opossum brainstem. Injections of CtB were made into the cheek/lip region or tongue of opossum pups to retrogradely label the facial or hypoglossal motor nuclei, respectively. Following a 2 h survival time, facial motoneurons in newborn opossum pups (1 PN) exhibited CtB labeling, with their cell bodies localized near the developing cranial abducens nucleus. At 3 and 5 PN, following a 48 h survival time, CtB-labeled facial motoneurons were observed in and migrating to the region of the adult facial motor nucleus in the rostral medulla. Between 7 and 10 PN, almost all facial motoneurons had migrated to their destination within the facial motor nucleus. Hypoglossal motoneurons also exhibited CtB labeling from 1 PN; however, their cell bodies were localized within the hypoglossal motor nucleus at the earliest age examined. Double label studies, to examine guidance of facial motoneurons during migration, demonstrated that CtB-labeled facial motoneurons are in close proximity to vimentin-like immunostained radial glial fibers during migration. These results suggest: (1) migration of facial motoneurons to the facial motor nucleus is a postnatal event, (2) efferent projections from facial and hypoglossal motoneurons project into the peripheral region of their target muscles from the day of birth, and (3) facial motoneurons migrate to their destination in the brainstem thereafter, in close association with radial glial fibers.

Aging↗

PubMed1997

Glucose kinetics of the Virginia opossum: possible implications for predicting glucose turnover in mammals.

Primed continuous infusions of 6-3H-glucose, respirometry and measurements of nitrogen excretion were carried out in adult Virginia opossums (Didelphis virginiana). Our goals were to determine resting glucose turnover rate, to establish the relative importance of this oxidative fuel in the energy budget of this species, to assess whether metabolic rate is a better predictor of glucose turnover rate than body mass and to demonstrate that glucose kinetics can be measured in the opossum for future use of this animal model in studies of mammalian glucoregulation. Results show that the resting glucose turnover rate of opossums averages 17.5 +/- 0.9 mumol/kg.min (n = 9), that 31% of total glucose flux is oxidized and that glucose oxidation represents 9% of their metabolic rate. An allometric equation predicting glucose turnover rate from body mass for placental mammals overestimates the measured turnover rate of this marsupial by 30%, suggesting that turnover rate is better predicted from metabolic rate than from body mass. Finally, this study demonstrates that the Virginia opossum is a convenient model to study glucose metabolism in vivo, and we propose that the very rapid aging characteristics of this species should be exploited to investigate the effects of aging on glucose homeostasis.

Aging↗

PubMed2001

Sarcocystis neurona infections in raccoons (Procyon lotor): evidence for natural infection with sarcocysts, transmission of infection to opossums (Didelphis virginiana), and experimental induction of neurologic disease in raccoons.

Equine protozoal myeloencephalitis (EPM) is a serious neurologic disease of horses in the Americas and Sarcocystis neurona is the most common etiologic agent. The distribution of S. neurona infections follows the geographical distributions of its definitive hosts, opossums (Didelphis virginiana, Didelphis albiventris). Recently, cats and skunks were reported as experimental and armadillos as natural intermediate hosts of S. neurona. In the present report, raccoons (Procyon lotor) were identified as a natural intermediate host of S. neurona. Two laboratory-raised opossums were found to shed S. neurona-like sporocysts after ingesting tongues of naturally-infected raccoons. Interferon-gamma gene knockout (KO) mice fed raccoon-opossum-derived sporocysts developed neurologic signs. S. neurona was identified immunohistochemically in tissues of KO mice fed sporocysts and the parasite was isolated in cell cultures inoculated with infected KO mouse tissues. The DNA obtained from the tongue of a naturally-infected raccoon, brains of KO mice that had neurological signs, and from the organisms recovered in cell cultures inoculated with brains of neurologic KO mice, corresponded to that of S. neurona. Two raccoons fed mature S. neurona sarcocysts did not shed sporocysts in their feces, indicating raccoons are not likely to be its definitive host. Two raccoons fed sporocysts from opossum feces developed clinical illness and S. neurona-associated encephalomyelitis was found in raccoons killed 14 and 22 days after feeding sporocysts; schizonts and merozoites were seen in encephalitic lesions.

Animals↗

PubMed2003

Prevalence and tissue distribution of Besnoitia darlingi cysts in the Virginia opossum (Didelphis virginiana) in Michigan.

Specimens of Virginia opossums (Didelphis virginiana) in Michigan were examined over 1 year to document the presence of Besnoitia darlingi cysts. Cyst morphology, prevalence, seasonal variation, and tissue sites of isolation were studied. Histology and ultrastructural features of the detected cysts and bradyzoites were consistent with B. darlingi. In the opossums, B. darlingi had intracellular tissue cysts. Tissue cysts had a mean diameter of 560 microm and were separated from the host tissue by a thick (5-20 microm) cyst wall. Overall prevalence of B. darlingi cysts in opossums was 10.9% (15/137). Variations in the prevalence were detected during spring (3/17; 17.6%), summer (10/34; 29.4%), and fall (2/60; 3.3%). No cysts were detected in the specimens examined during winter (0/26; 0%). Numerous B. darlingi cysts were detected in ears, conjunctiva, tongue, abdominal muscles, diaphragm, stomach, heart, liver, kidney, lung, and spleen. Cysts were detected mainly in adult female opossums that were debilitated. Ear was the most frequent organ from which the cysts were reported (10/15; 66.7%) when compared individually with other body tissues (P<0.05).

Animals↗

PubMed1994

Spatial behavior in the Brazilian short-tailed opossum (Monodelphis domestica): comparison with the Norway rat (Rattus norvegicus) in the Morris water maze and radial arm maze.

The spatial behavior displayed in a swimming pool and radial arm maze by the Brazilian gray, short-tailed opossum (Monodelphis domestica) was compared with that of the laboratory rat (Rattus norvegicus). The performance of the opossums was clearly different from that of the rats in both tasks. The opossums failed to learn to find a hidden platform in the swimming pool (place task), but they did learn to swim to a visible platform. The opossums did learn working and reference memory components of the radial arm maze but showed different search strategies in this situation. The results are discussed with reference to possible species differences in motor behavior, ecology, brain structure and evolution.

Animals↗

PubMed1988

Evolutionary and developmental aspects of two hemoglobin beta-chain genes (epsilon M and beta M) of opossum.

A series of gene duplications that began in a stem species of Mammalia and led to five developmentally regulated hemoglobin beta-chain loci (epsilon, gamma, eta, delta, and beta) in a common ancestor of eutherian orders Artiodactyla, Rodentia, Lagomorpha, and Primates had important consequences in mammalian evolution. Findings reported here indicate that two progenitors of the five linked genes existed by the time of the eutherian (placental mammal)-metatherian (marsupial mammal) split and that these two genes were already differentiated with respect to their promoter regions and developmental expression. Southern blot and sequence analyses of the hemoglobin beta-chain genes of the opossum (Didelphis virginiana) revealed only two genes, one with coding and promoter sequences similar to eutherian prenatally expressed epsilon, gamma, and eta genes and the other coding for adult opossum hemoglobin beta-chains and having eutherian adult beta-type promoters. The most parsimonious arrangement of greater than 80 beta-globin exon sequences depicts the opossum embryonic-type gene as orthologously related to eutherian epsilon, gamma, and eta genes and the opossum adult-type gene as orthologously related to delta and beta genes. These data further indicate that after the initial beta duplication in the stem of Mammalia, the locus that became developmentally delayed in its expression evolved at a faster rate than the locus that became embryonically expressed.

Amino Acid Sequence↗

PubMedDate not supplied

Seasonal changes in the physiology of male Virginia opossums (Didelphis virginiana): signs of the Dasyurid semelparity syndrome?

Semelparity, which is multiplying once in a lifetime, is a rare reproductive strategy among mammals. Several species of the marsupial family Dasyuridae experience 100% male mortality following an intense mating period. We investigated seasonal physiological changes that may be associated with early mortality in the male Virginia opossum (Didelphis virginiana; Didelphidae) and compared these changes with those of semelparous, male dasyurids. Free-ranging male Virginia opossums (n=36) were collected during 2001 at the Oklahoma State University Cross Timbers Experimental Range. Seasonal data were collected on hematological, morphological, and helminth parameters of these individuals. We used one-way ANOVA to determine whether there were seasonal differences among means for each parameter. It appeared that male Virginia opossums experienced some physiological changes similar to those of male dasyurids exhibiting semelparity. All males collected in summer (August) were juveniles of the year. Lack of adult males in August suggests high mortality of this cohort during the breeding season. Opossum characteristics exhibiting the dasyurid semelparity syndrome included packed cell volume, adrenal mass, and helminth numbers. Minor lymphocytopenia, neutrophilia, and testosterone concentrations also were similar to semelparous dasyurids. However, a lack of change in serum cortisol concentration and body mass and dynamics in immunoglobulin protein, serum protein, and testes mass were not consistent with previous reports of semelparous dasyurid physiology. Evolutionary divergence and differences in breeding behavior between dasyurids and didelphids may be responsible for the lack of consistency between the taxa.

Animals↗

PubMed2005

Immunohistochemical localization of gastrin-releasing peptide, neuronal nitric oxide synthase and neurone-specific enolase in the uterus of the North American opossum, Didelphis virginiana.

The present study has demonstrated the immunohistochemical localization of gastrin-releasing peptide (GRP), neuronal nitric oxide synthase (nNOS) and neurone-specific enolase (NSE) in the uterus of the North American opossum. Although the presence of GRP, nNOS and NSE has been reported recently in the uterus of eutherian species this is the first description of these peptides in a metatherian species. Metatherian mammals are of interest because in these species it is the prolonged lactation phase of development that is the period of primary reproductive investment rather than intrauterine development as is true of eutherian mammals. The opossum, like other marsupial species, has a very abbreviated gestation period which in Didelphis lasts only 12.5 days. GRP was localized in the cytoplasm of cells forming the surface lining epithelium and the glandular epithelium of the opossum endometrium late in pregnancy, at 11.5 days of gestation. Likewise, immunoreactivities of nNOS and NSE were found primarily within the epithelial cells of the endometrium at 11.5 days of gestation. As these peptides and enzymes appear primarily at the time of establishment of the yolk sac placenta (between day 10 and day 12.5 gestation), the present results strongly suggest that these factors may play a fundamental role in the placentation of the opossum.

Animals↗

PubMed1997

Chemosensory and cholinergic stimulation of fictive respiration in isolated CNS of neonatal opossum.

1. The aim of the present experiments was to characterize the central chemical drive of fictive respiration in the isolated CNS of the newborn opossum, Monodelphis domestica. This opossum preparation, in contrast to those of neonatal rats and mice, produces respiratory rhythm of high frequency in vitro. 2. Fictive respiration was recorded from C3-C5 ventral roots of the isolated CNS of 4- to 14-day-old opossums using suction electrodes. At room temperature (21-23 degrees C) the frequency of respiration was 43 +/- 5.3 min-1 (mean +/- S.E.M., n = 50) in basal medium Eagle's medium (BMEM) equilibrated with 5% CO2-95% O2, pH 7.37-7.40. Respiratory discharges remained regular throughout 8 h experiments and continued for more than 20 h in culture. 3. Superfusion of the brainstem confirmed that solutions of pH 6.3-7.2 increased both the amplitude and frequency of respiration. High pH solutions (7.5-7.7) had the opposite effect and abolished the rhythm at pH 7.7. Addition of ACh (50-100 microM) or carbachol (0.01-10 microM) to the brainstem superfusion also increased the amplitude and frequency of respiratory activity, as did physostigmine (50-100 microM) or neostigmine (20-50 microM). Conversely, scopolamine (50-100 microM) reduced the amplitude and frequency of the basal respiratory rhythm by about 30%. 4. H(+)- and cholinergic-sensitive areas on the surface of the isolated CNS were explored with a small micropipette (outer tip diameter, 100 microns) filled with BMEM (pH 6.5) or 1 microM carbachol. Carbachol applied to H(+)- and cholinergic-sensitive areas in the ventral medulla mimicked the changes of respiratory pattern produced by low pH application. Responses to altered pH and carbachol were abolished by scopolamine (50 microM). Histochemistry demonstrated several medullary groups of neurons stained for acetylcholinesterase. The superficial location of one of these groups coincided with a functional and anatomically well-defined pH- and carbachol-sensitive area placed medial to the hypoglossal roots. 5. Exploration of chemosensitive areas revealed that application of drugs or solutions of different pH to a single well-defined spot could have selective and distinctive effects upon amplitude and frequency of respiratory activity. 6. These results show that fictive respiration in the isolated CNS of the newborn opossum is tonically driven by chemical- and cholinergic-sensitive areas located on the ventral medulla, the activity of which regulates frequency and amplitude of respiration. They suggest that a cholinergic relay, although not essential for rhythm generation, is involved in the central pH chemosensory mechanism, or that cholinergic and chemical inputs converge upon the same input pathway to the respiratory pattern generator.

Acetylcholinesterase↗

PubMed1984

A pharmacological study of oesophageal muscularis mucosae from the cat, dog and American opossum (Didelphis virginiana).

Strips of muscularis mucosae from the oesophagi of cat, dog and opossum have been studied to determine their responses to drugs to electrical field stimulation. All tissues were contracted by acetylcholine, histamine and, with the exception of strips of muscularis mucosae from the opossum proximal oesophagus, noradrenaline. The effects of acetylcholine and histamine were competitively antagonized by atropine (50 nM) and mepyramine (50 nM) and were abolished by atropine (1 microM) and mepyramine (1 microM) respectively. Contractile responses to noradrenaline were competitively antagonized by phentolamine (50 nM) but were converted to propranolol (50 nM)-sensitive relaxations by phentolamine (1 microM). Relaxations were abolished by propranolol (1 microM). Cholecystokinin octapeptide, gastrin 1 and vasoactive intestinal polypeptide were ineffective on any of the tissues examined. Substance P caused contractions in tissue from all three species. These effects were atropine and tetrodotoxin insensitive. All tissues gave atropine (50 nM)- and tetrodotoxin (100 nM)-sensitive contractions in response to electrical field stimulation. Contractions were not followed by relaxations and spontaneous mechanical activity was not suppressed between periods of stimulation. No evidence was obtained for the presence of non-adrenergic, non-cholinergic inhibitory innervation of the oesophageal muscularis mucosae in any species. During electrical field stimulation noradrenaline always reduced the amplitude of evoked contractions and, with the exception of tissue from proximal opossum oesophagus, increased resting tension. In opossum distal oesophageal muscularis mucosae, the effects of noradrenaline during electrical field stimulation were abolished by a 30 min pretreatment of the tissue with phentolamine (1 microM) and propranolol (1 microM). To Achieve this in all other tissues, it was also necessary to use yohimbine (1 microM). 7 In all tissues where noradrenaline caused a phentolamine (1 microM)-and propranolol (1 microM)- resistant depression of electrically evoked responses, clonidine produced a yohimbine (1 microM)- sensitive depression. 8 Evidence was obtained for the presence of excitatory alpha 1-and inhibitory alpha 2- and beta-adrenoceptors. Inter-species differences in their distribution are discussed.

Acetylcholine↗

PubMed1998

Proteolytic activity of opossum tooth extracts.

Amelogenins are the main component of the developing enamel matrix. In placental mammals, amelogenins are rapidly cleaved following their secretion. HPLC fractionation of tooth extracts produces a complex chromatographic profile. The fractions are rich in amelogenin cleavage products that generally retain the amino-terminus of the parent protein but have varying lengths of peptide removed from the original carboxyl-terminus. In contrast, HPLC fractionation of opossum tooth extracts produces a simple profile with a single major chromatographic peak. SDS-and Western blot analyses demonstrated that most of the amelogenin consisted of a prominent protein band that migrated at 28 kDa. Mass spectroscopy confirmed the presence of two uncleaved, alternatively spliced forms of opossum amelogenin, Op202 and Op57, but did not detect major amelogenin cleavage products evident in tooth extracts from placental mammals. Amino acid composition analysis supported the conclusion that uncleaved amelogenin is the major component in the developing enamel matrix. Enzymogram analyses using gelatin, casein and recombinant amelogenin as substrates, comparing porcine, rat and opossum tooth extracts, suggested that fewer proteinases are present in opossum. These results identify potentially significant differences in the proteolytic processing of amelogenins between metatherian and eutherian mammals.

Amelogenin↗

PubMed1988

Medullary inspiratory activity during opossum development.

Discharge properties of medullary neurons associated with inspiration were assessed during opossum development. Location of these neurons and projections to the spinal cord were also evaluated. Studies were performed on suckling opossums, from 15-20 days of age to weaning (approximately 100 days), as well as adult animals, anesthetized with Inactin. Most cells either discharged during inspiration (I cells) or were phase-spanning between late expiration and inspiration (EI cells). Number of spikes per breath for I and EI cells significantly increased as the animal matured. For younger opossums (up to approximately 60 days of age), bulbospinal I and EI cells discharged an average of 1.9 spikes per breath, with a minimum interspike interval of 59 ms. Their conduction velocities, determined by use of antidromic stimulation, were in the unmyelinated range (less than 2 m/s). The occurrence of few spikes per breath with long interspike interval, in bulbospinal cells of the younger opossums, suggests that inspiration is not subject to fine feedback control and that rate of high-frequency motor bursts is relatively slow.

Aging↗

PubMed1988

Receptors and cGMP signalling mechanism for E. coli enterotoxin in opossum kidney.

Receptors for the heat-stable enterotoxin produced by Escherichia coli were found in the kidney and intestine of the North American opossum and in cultured renal cell lines. The enterotoxin markedly increased guanosine 3',5'-cyclic monophosphate (cGMP) production in slices of kidney cortex and medulla, in suspensions of intestinal mucosa, and in the opossum kidney (OK) and rat kangaroo kidney (PtK-2) cell lines. In contrast, atrial natriuretic factor elicited much smaller increases in cGMP levels of kidney, intestine, or cultured kidney cell lines. The enterotoxin receptors in OK cells had a molecular mass of approximately 120 kDa when measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of receptors crosslinked with 125I-enterotoxin. The occurrence of receptors for the E. coli peptide in OK implies that these receptors may be involved in the regulation of renal tubular function in the opossum. E. coli enterotoxin caused a much larger increase in urine cGMP excretion (10- to 50-fold over control) than did atrial natriuretic factor when these peptides were injected intravenously into opossums. However, atrial natriuretic factor elicited a marked diuresis, natriuresis, and increased urinary excretion of calcium, phosphate, potassium, and magnesium. In contrast, the enterotoxin did not acutely influence OK fluid and electrolyte excretion. Thus the substantial increase in cGMP synthesis produced by the bacterial peptide in OK cortex and medulla in vitro and the increased renal excretion of cGMP in vivo were not associated with changes in electrolyte or water excretion. Whether cGMP represents a second messenger molecule in the kidney is an interesting question that was raised but not answered in this series of experiments.

Animals↗

PubMed1993

Maximum discharge rates of respiratory neurons during opossum development.

The observed low frequencies of action potentials observed in medullary respiratory neurons of immature opossums (Didelphis virginiana) could occur because these cells are incapable of achieving higher sustained firing rates. Nonsustainability of firing might also help explain why the inspired breath is brief (approximately 0.1 s) in the youngest opossums and rises very slowly during postnatal life. Firing frequencies of medullary respiratory neurons were examined in spontaneously breathing thiobarbiturate-anesthetized opossums before and after stimulation by the glutamate agonists, N-methyl-D-aspartate (NMDA; 20 mM) or kainic acid (KA; 0.5 mM). Drugs were applied using progressively larger pressure injections through a micropipette; animals were tested from the 5th postnatal wk to adulthood. With a sufficient injection volume, stimulation of cell firing would be followed by apparent suppression of action potentials. A maximum "sustained" firing frequency was obtained from the last injection where discharge remained elevated for at least 0.5 s. Inspiratory and expiratory neurons tested with either drug showed the lowest rates of firing in opossums at 5-9 wk of age compared with 10- to 14-wk-old animals and/or adults. Despite higher rates of discharge in 10- to 14-wk-old animals and/or adults, maximum sustained neuronal firing in the youngest animals was at a higher frequency than during spontaneous breathing and, at least in the cell population tested, does not represent a limitation that might affect breathing pattern.

Action Potentials↗