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PubMed1984

Gene mapping in marsupials and monotremes. I. The chromosomes of rodent-marsupial (Macropus) cell hybrids, and gene assignments to the X chromosome of the grey kangaroo.

Somatic cell genetic mapping of marsupial and monotreme species will greatly extend the power of comparative gene mapping to detect ancient mammalian gene arrangements. The use of eutherian-marsupial cell hybrids for such mapping is complicated by the frequent retention of deleted and rearranged marsupial chromosomes. We used staining techniques, involving the fluorochromes Hoechst 33258 and chromomycin A3, to facilitate rapid and unequivocal identification of marsupial chromosomes and chromosome segments and to make chromosome assignment and regional localization of marsupial genes possible. Chromosome segregation in rodent-macropod hybrids was consistent with preferential loss of the marsupial complement. The extent of loss was very variable. Some hybrids retained 30% of the marsupial complement; some retained small centric fragments; and some, no cytologically identifiable marsupial material. We examined the chromosomes and gene products of a number of rodent-grey kangaroo Macropus giganteus hybrids, and have assigned the genes Pgk-A (phosphoglycerate kinase-A), Hpt (Hypoxanthine-phosphoribosyl transferase), and Gpd (Glucose-6-phosphate dehydrogenase) to the long arm of the kangaroo X chromosome, and provisionally established the gene order Pgk-A--Hpt--Gpd.

Animals↗
PubMed1981

Hoechst 33258 induced uncondensed sites in marsupial chromosomes.

The fluorochrome Hoechst 33258 induces pronounced uncondensed regions at mitosis at one or more specific sites on the X chromosomes of all eighteen species of marsupials belonging to the family Macropodidae which have been examined. The Y chromosomes of nearly all of these species also show sensitive sites. Autosomal regions which respond to this chemical were observed in only five species and there is evidence of polymorphism for two of these. The regions which respond usually show C-banding, but not all C-banding regions are affected. No specific effect was found in the chromosomes of eleven other species examined which are representative of 5 different Australian marsupial families. The implications of the apparent restriction of sex chromosome sensitive sites to macropods are discussed.

Animals↗
PubMed1984

Transformation of the Hprt gene with DNA from spermatogenic cells. Implications for the evolution of X chromosome inactivation.

DNA-mediated transformation of hypoxanthine guanine phosphoribosyl transferase (HPRT)-deficient cells was used to assess the state of the chromosome Hprt gene in spermatogenic cells. It had been shown previously that DNA from the inactive X chromosome of somatic cells functions poorly or not at all in HPRT transformation, indicating that DNA modification is involved in somatic cell X chromosome inactivation (XCI). In contrast, DNA from mature sperm does function in HPRT transformation suggesting that DNA modification may not be the basis of XCI in mature sperm. In this paper, transformation of HPRT- mouse and hamster cells has been performed to test the nature of XCI during earlier stages of spermatogenesis. DNA from these developing murine germ cells was shown to be capable of HPRT transformation, extending the observation that XCI in sperm does not appear to involve a DNA modification. We also show here that DNA from mature sperm of marsupials functions in HPRT transformation, a result consistent with a role for sperm XCI in the evolution of somatic X inactivation.

Animals↗
PubMed1995

Full-length cDNA sequence of X-linked G6PD of an Australian marsupial, the wallaroo.

In order to study the mechanism of X Chromosome (Chr) inactivation in marsupials, the cDNA for glucose-6-phosphate dehydrogenase (G6PD) from an Australian marsupial, the wallaroo (Macropus robustus), was cloned. A partial clone containing the 3' half of the cDNA was obtained by screening a liver cDNA library. The majority of the coding region was obtained by polymerase chain reaction of cDNA with primers designed from regions of conservation between human and opossum G6PD. The 5' end was obtained by rapid amplification of cDNA ends. High homology was observed between mammalian species in the coding region. The 5' untranslated region is highly G+C rich, and appears to be part of a CpG island, as is the case in the human and mouse genes. This is the first report of the full sequence of the cDNA for any marsupial X-linked gene.

Amino Acid Sequence↗
PubMed1976

Satellite DNA in the kangaroo Macropus rufogriseus.

Two distinct satellite DNAs, amounting to 25% of the total DNA, were isolated from the nuclei of the red-necked wallaby, Macropus rufogriseus. The physical properties of native, single-stranded and reassociated molecules were studied in buoyant-density gradient centrifugation. The homogeneity of each satellite fraction was examined using melting characteristics of native and reassociated DNA, and renaturation kinetics. These data suggest that sequence heterogeneity exists in both fractions. Each satellite fraction was found by in situ hybridization to be localized in heterochromatin of interphase nuclei and in the centromeric regions of metaphase chromosomes. The chromosomal distributions of the two satellite DNAs differentiate the sex chromosomes, which have sequences of only one satellite, from the autosomes which have sequences of both satellites in the centromeric heterochromatin. Giemsa C-banding techniques also showed a differentiation of the centromeric regions of sex chromosomes from those of the autosomes.

Animals↗
PubMed1981

The structure of the cold-stable kinetochore fiber in metaphase PtK1 cells.

When metaphase PtK1 cells are cooled to 6-8 degrees C for 4-6 h the free, polar, and astral spindle microtubules (MTs) disassemble while the MTs of each kinetochore fiber cluster together and persist as bundles of cold-stable MTs. These cold-stable kinetochore fibers are similar to untreated kinetochore fibers in both their length (i.e., 5-6 micrometer) and in the number of kinetochore-associated MTs (i.e., 20-45) of which they are comprised. Quantitative information concerning the lengths of MTs within these fibers was obtained by tracking individual MTs between serial transverse sections. Approximately 1/2 of the kinetochore MTs in each fiber were found to run uninterrupted into the polar region of the spindle. It can be inferred from this and other data that a substantial number of MTs run uninterrupted between the kinetochore and the polar region in untreated metaphase PtK1 cells.

Animals↗
PubMed1976

Nucleoli and ploidy in Potorous cells (PTK2) in vitro.

Most cells of the male PTK2 cell line contain one nucleolus, and they are diploid. However, a proportion of cells have more than one nucleolus. Cells with two and three nucleoli were isolated and cloned into viable populations. Greater than 90% of the cells in these clonal populations maintained the abnormal nucleolar number of the originally isolated cell. Karyotype analysis of cells with two and three nucleoli demonstrated that the cells were respectively tetraploid and hexaploid. It was concluded that in PTK2 cells, nucleolarity is a good index of ploidy even if the ploidy level is abnormal. Furthermore, long term monitoring of the tetraploid cells demonstrated virtually no tendency towards reversion to the diploid condition as suggested by other studies in Potorous.

Animals↗
PubMed1992

Relationships between plasma composition and parotid salivary composition and secretion rates in the potoroine marsupials, Aepyprymnus rufescens and Potorous tridactylus.

Parotid salivation was investigated in two species of potoroine marsupial, Aepyprymnus rufescens and Potorous tridactylus to ascertain flow rates and composition, the buffer capacity of the saliva with respect to possible dependence of these animals on foregut fermentation, and the similarity of anion excretion patterns to those of the kangaroo parotid. Under anaesthesia neither species secreted spontaneously and secretion was stimulated by intravenous infusion of carbachol, bethanechol and isoprenaline. Under cholinergic stimulation in Aepyprymnus, the concentrations of Na, Cl, HCO3 and osmolality were positively correlated with flow rate, whereas K, Mg, PO4, H+ and urea were negatively correlated with flow. Amylase activity and the concentrations of protein and Ca showed no consistent relation to flow. Relative to Aepyprymnus, saliva of Potorous had much lower amylase activity and amylase activity per gram protein, lower concentrations of urea and Ca, and higher Na. Protein, K and HCO3 concentrations were similar in both species. The plasma of both species had similar electrolyte concentrations, but Potorous had lower protein, urea, osmolality and amylase activity. Plasma amylase activity in Aepyprymnus rose during cholinergic stimulation to levels in excess of rodent plasma. Isoprenaline infusion in Aepyprymnus increased salivary amylase activity and concentrations of protein, Ca, HCO3 and PO4, and reduced the concentrations of Cl and H+. The patterns of anion excretion in the two potoroine marsupials were dissimilar to those of the kangaroo parotid suggesting that parotid fluid secretion is not HCO3 driven to the same extent as that of Kangaroos. Buffer anion concentrations and secretion rates were similar to koalas and low relative to kangaroos, indicating that these potoroines do not rely on foregut fermentation.

Amylases↗
PubMed1988

The ultrastructure of the pituitary and the adrenal gland of three newborn marsupials (Dasyurus hallucatus, Trichosurus vulpecula and Isoodon macrourus).

The ultrastructure of the pituitary glands of the newborn northern native cat, brushtail possum and the northern brown bandicoot and the adrenal glands of the former two marsupials were examined to determine whether these endocrine glands were functional at birth. The anterior pituitary of all three species was well vascularised and many cells contained electron-dense, membrane bound granules. The adrenal glands were composed of two distinct cell populations. One group of cells possessed dark staining granules, normally observed in catecholamine-secreting cells, and the second group contained large amounts of smooth endoplasmic reticulum and mitochondria with tubulovesicular cristae, indicative of steroid hormone secreting cells. The ultrastructure of the cells of the fetal pituitary and adrenal of all three species would suggest that these glands are functional at birth and, as with many eutherians, the marsupial pituitary and adrenal gland may play an important role in determining the length of gestation.

Adrenal Glands↗
PubMed1988

Ultrastructural identification of Merkel cells around the mouth of the newborn marsupial.

The ultrastructure of the epidermal cells surrounding the mouth of three newborn marsupial species, the Northern native cat Dasyurus hallucatus, the brush tail possum Trichosurus vulpecula and the Northern brown bandicoot Isoodon macrourus were examined. The presence of Merkel cells, highly sensitive touch receptors, would suggest that the sense of touch aids the relatively underdeveloped newborn marsupial to move from the urinogenital sinus to the pouch and to locate the teat.

Animals↗
PubMed1987

Topography and ultrastructure of sensory nerve endings in the joint capsules of the Kowari (Dasyuroides byrnei), an Australian marsupial.

The present investigation in concerned with the topography and ultrastructure of sensory nerve endings in the joint capsules of the Kowari (Dasyuroides byrnei), an Australian marsupial. Material for light and electron microscopy was obtained from shoulder, elbow and knee joint capsules. On the basis of differences in the organization of the connective tissue belonging to the fibrous layer, 3 variants of capsule structure have been distinguished: a rigid, a flaccid and an intermediate type. Whilst the rigid type is characterized by dense connective tissue in the clearly demarcated fibrous layer, the flaccid type shows loose, irregularly arranged connective tissue in the fibrous layer which merges into the synovial layer of the joint capsule. The morphology of the intermediate type corresponds to an intermediate stage between the former two types. In the fibrous layer of the joint capsules three different types of sensory nerve endings were observed: free nerve endings, Ruffini corpuscles and lamellated corpuscles. The free nerve endings are supplied by myelinated afferent axons (1-2 microns in diameter); the terminal thickenings of which are incompletely surrounded by a terminal Schwann cell. Ruffini corpuscles are present in three different varieties: small corpuscles without a perineural capsule predominantly within the flaccid part of the capsule; slightly larger corpuscles with an incomplete perineural capsule and large corpuscles resembling Golgi tendon organs which predominantly occur in the rigid parts of the capsule. The afferent myelinated axons measure 2-4 microns in diameter. The lamellated corpuscles show two variants: small corpuscles with a 2 to 4-layered perineural capsule in the rigid parts of the joint capsules and large corpuscles with two longitudinal clefts of the inner core in the flaccid parts. Both types are supplied by myelinated axons of 3-5 microns in diameter. Thus, in the fibrous layer of the rigid type of joint capsules large Ruffini and small lamellated corpuscles predominate, whereas the fibrous layer of the flaccid type coincides with small Ruffini and large lamellated corpuscles. The present data, therefore, corroborate the concept that the morphology of mechanoreceptors depends upon the texture of the surrounding connective tissue.

Animals↗
PubMed1987

The early development of the eye of the pouch-young of the marsupial Dasyurus hallucatus.

The development of the eye of the pouch-young of the marsupial native cat Dasyurus hallucatus from birth to day 30 is described. Following the staging system used by other authors, the stage of development of the eye at birth is comparable to that of a six-week human embryo, a 33 day rhesus embryo or a 14 day rat embryo. In contrast to eutherian mammals where development progresses from one stage to another at approximately the same rate, in the native cat the rate tends to progressively decrease. All of the neural development of the native cat eye occurs while the young is in the pouch.

Animals↗
PubMed1987

Ultrastructure and possible function of giant crystalloids in the Sertoli cell of the juvenile and adult koala (Phascolarctos cinereus).

Sertoli cells of the juvenile and adult koala testis exhibit a unique morphology due to their large nuclei and in particular, a remarkable abundance of large cytoplasmic crystalloid inclusions. Numerous crystalloid subunits in immature Sertoli cells are aggregated into distinct clusters where by assembly and union, they form large slender crystalloids consisting of an ordered substructure of filaments and tubules. Adult Sertoli cells contain large numbers of basally-positioned crystalloids up to 60 micron in length and the observations suggest a possible mechanism for their growth from collections of tubules assembled together within membrane-bound inclusions. The trunk and adluminal cytoplasm of the adult Sertoli cell also contains crystalloids, usually single, positioned between germ cells or their excess residual cytoplasm. Following sperm release, crystalloids are not shed from the seminiferous epithelium but are retained within the apical Sertoli cell cytoplasm. Although their subsequent fate could not be determined crystalloids did not show evidence of breakdown or phagocytosis by the Sertoli cell, suggesting that they may be reutilized and possibly function to stabilize the association between Sertoli cell cytoplasm and the developing germ cells.

Animals↗
PubMed1989

Histological and electron microscopic milestones in the development of the retina of a marsupial wallaby, Macropus eugenii.

Retinal differentiation in the pouch young of the wallaby Macropus eugenii was characterized microscopically and morphometrically. Mitosis occurs until early in the second month in the central retina, and until early in the fourth month, peripherally. Separation of the neuroblast layer by the outer plexiform layer did not immediately halt cell division. The retinal surface continued to expand well past the time of cessation of proliferation. Cell death in the ganglion cell layer continued through the fourth month centrally and to nearly five months in the periphery. The major period of cell death was coincident with the segregation of retinal afferents and the refinement of topography in the superior colliculus and dorsal lateral geniculate nucleus. Beginning in the third month retinal thickness, measured between the outer limiting membrane and nerve fiber layer, declined equally in peripheral and central regions. At all stages the combined thicknesses of the outer and inner nuclear layer in the retinal periphery was greater than that in the center. Together with a late thickening of the inner plexiform layer, the data are consistent with the suggestion that expansion of peripheral non-ganglion cell elements may play a role in development of center to periphery differences in ganglion cell distributions. Retinal differentiation of the wallaby follows the pattern of most mammals. The onset of development of key milestones for the acquisition of retinal function occurred in the sequence: conventional synapse formation prior to ribbon synapse formation in the inner plexiform layer, and photoreceptor outer segment differentiation prior to terminal triad synapse formation.

Animals↗
PubMed1985

The embryological development and cytodifferentiation of the anterior pituitary in the marsupial, Isoodon macrourus.

Light and electron microscopy were used to study the development of the anterior pituitary gland from fetal stages to the end of pouch life in the marsupial I. macrourus. The early morphological development of the anterior pituitary in I. macrourus follows a similar pattern of events to that described for eutherians. Rathke's and Seessel's pouches were present in 10 1/2 day old embryos. At birth these pouches had formed a multi-chambered vesicle which was still connected to the stomodeum by a thin cord of tissue. A small number of granules (200-400 nm dia.) were found in cells at birth. These cells could not be classified on ultrastructural features but alcian blue-periodic acid Schiffs-orange G staining suggested one cell type was possibly a presumptive thyrotroph. There were no capillaries in the pars distalis at birth. The cords connecting Rathke's and Seessel's pouches to the stomodeum were located at the site of the periosteal bud of the developing basisphenoid which commenced to ossify at 7 days. At this stage presumptive thyrotrophs, gonadotrophs, and somatotrophs could be distinguished using alcian blue-periodic acid Schiffs-orange G staining. However, five cell types could be categorised at the same age using ultrastructural characteristics alone. Precise names for these cells are unavailable but two closely resemble presumptive mammotrophs and thyrotrophs described for another marsupial M. eugenii. By 13 days after birth the anterior pituitary of I. macrourus had become vascular and acidophils were concentrated in a posterior zone. There was little gross morphological change from 13 to 66 days after birth by which the time weaning has occurred. Cilia were seen in cells of the anterior pituitary and mitosis of granulated cells was observed from birth onwards. There is a considerable range of variation in pituitary cytogenesis amongst marsupials, and its functional significance awaits further investigation.

Animals↗