Histological and immunohistological investigation of lymphoid tissue in the platypus (Ornithorhynchus anatinus)

The gross and histological appearance and the distribution of T and B lymphocytes and plasma cells are described for lymphoid tissues obtained from 15 platypuses. The spleen was bilobed and surrounded by a thick capsule of collagen, elastic fibres and little smooth muscle. White pulp was prominent and included germinal centres and periarterial lymphoid sheaths. Red pulp contained haematopoietic tissue. A thin lobulated thymus was located within the mediastinum overlying the heart. The cortex of lobules consisted of dense aggregates of small and medium lymphocytes, scattered macrophages and few reticular epithelial cells. In the medulla, Hassall's corpuscles were numerous, lymphocytes were small and less abundant, and reticular cells were more abundant than in the cortex. Lymphoid nodules scattered throughout loose connective tissue in cervical, pharyngeal, thoracic, mesenteric and pelvic sites measured 790±370 μm (mean±S.D. , n = 39) in diameter, the larger of which could be observed macroscopically. These consisted of single primary or secondary follicles supported by a framework of reticular fibres. Macrophages were common in the germinal centres. The platypus had a full range of gut-associated lymphoid tissue. No tonsils were observed macroscopically but histologically they consisted of submucosal follicles and intraepithelial lymphocytes. Peyer's patches were not observed macroscopically but histologically they consisted of several prominent submucosal secondary follicles in the antimesenteric wall of the intestine. Caecal lymphoid tissue consisted of numerous secondary follicles in the submucosa and densely packed lymphocytes in the lamina propria. Bronchus-associated lymphoid tissue was not observed macroscopically but was identified in 7 of 11 platypus lungs assessed histologically. Lymphoid cells were present as primary follicles associated with bronchi, as aggregates adjacent to blood vessels and as intraepithelial lymphocytes. The distribution of T lymphocytes, identified with antihuman CD3 and CD5, and B lymphocytes and plasma cells, identified with antihuman CD79a and CD79b and antiplatypus immunoglobulin, within lymphoid tissues in the platypus was similar to that described in therian mammals except for an apparent relative paucity of B lymphocytes. This study establishes that the platypus has a well-developed lymphoid system which is comparable in histological structure to that in therian mammals. It also confirms the distinctiveness of its peripheral lymphoid tissue, namely lymphoid nodules. Platypus lymphoid tissue has all the essential cell types, namely T and B lymphocytes and plasma cells, to mount an effective immune response against foreign antigens.     

On the morphology of the brachial plexus of the platypus (Ornithorhynchus anatinus) and the echidna (Tachyglossus aculeatus)

Four forelimbs of 3 platypuses and 3 forelimbs of 2 echidnas were examined to study the precise form of the brachial plexus and to clarify the structural characteristics of the brachial plexus in phylogeny. The spinal components contributing to the plexus (C4–T2) and the formation patterns of the 3 trunks of the plexus were the same as those generally observed in mammals. In the cranial half of the brachial plexus from C4, 5 and 6 in monotremes, division into the ventral bundle (lateral cord) and dorsal bundle (axillary nerve) is clear, as in other mammals. However, for monotremes, in the caudal half of the plexus from C7 and T1 (+T2) and the nerves arising from the caudal plexus there is no definite division into the ventral and dorsal bundles, which distribute to the flexor and extensor parts of the forelimbs, respectively. The lower trunk of the monotreme brachial plexus forms a cord which contains both ventral and dorsal components. This characteristic diverges from the generally accepted idea that the tetrapod limb plexus is divided clearly into 2 layers: a dorsal layer for extensors and a ventral layer for flexors of the limb. Considering the incomplete dorsoventral division of forelimb nerves in some reptiles and urodeles, the caudal half of the monotreme brachial plexus has characteristics in common with those of lower tetrapods.     

Cellular distribution of the calcium-binding proteins parvalbumin, calbindin, and calretinin in the neocortex of mammals: phylogenetic and developmental patterns

The three calcium-binding proteins parvalbumin, calbindin, and calretinin are found in morphologically distinct classes of inhibitory interneurons as well as in some pyramidal neurons in the mammalian neocortex. Although there is a wide variability in the qualitative and quantitative characteristics of the neocortical subpopulations of calcium-binding protein-immunoreactive neurons in mammals, most of the available data show that there is a fundamental similarity among the mammalian species investigated so far, in terms of the distribution of parvalbumin, calbindin, and calretinin across the depth of the neocortex. Thus, calbindin- and calretinin-immunoreactive neurons are predominant in layers II and III, but are present across all cortical layers, whereas parvalbumin-immunoreactive neurons are more prevalent in the middle and lower cortical layers. These different neuronal populations have well defined regional and laminar distribution, neurochemical characteristics and synaptic connections, and each of these cell types displays a particular developmental sequence. Most of the available data on the development, distribution and morphological characteristics of these calcium-binding proteins are from studies in common laboratory animals such as the rat, mouse, cat, macaque monkey, as well as from postmortem analyses in humans, but there are virtually no data on other species aside of a few incidental reports. In the context of the evolution of mammalian neocortex, the distribution and morphological characteristics of calcium-binding protein-immunoreactive neurons may help defining taxon-specific patterns that may be used as reliable phylogenetic traits. It would be interesting to extend such neurochemical analyses of neuronal subpopulations to other species to assess the degree to which neurochemical specialization of particular neuronal subtypes, as well as their regional and laminar distribution in the cerebral cortex, may represent sets of derived features in any given mammalian order. This could be particularly interesting in view of the consistent differences in neurochemical typology observed in considerably divergent orders such as cetaceans and certain families of insectivores and metatherians, as well as in monotremes. The present article provides an overview of calcium-binding protein distribution across a large number of representative mammalian species and a review of their developmental patterns in the species where data are available. This analysis demonstrates that while it is likely that the developmental patterns are quite consistent across species, at least based on the limited number of species for which ontogenetic data exist, the distribution and morphology of calcium-binding protein-containing neurons varies substantially among mammalian orders and that certain species show highly divergent patterns compared to closely related taxa. Interestingly, primates, carnivores, rodents and tree shrews appear closely related on the basis of the observed patterns, marsupials show some affinities with that group, whereas prototherians have unique patterns. Our findings also support the relationships of cetaceans and ungulates, and demonstrates possible affinities between carnivores and ungulates, as well as the existence of common, probably primitive, traits in cetaceans and insectivores.     

Generation of reactive oxygen responses by monotreme and marsupial granulocytes

The granules of circulating leukocytes contain reactive oxygen species that are important components of host defence against bacterial invasion. We report the capacity of marsupials and monotremes to mount such a defence in a manner similar to their eutherian relatives. Using the nitroblue tetrazolium (NBT) test, reactive oxygen species were detected in the peripheral blood cells of five captive marsupial species (the tammar wallaby, Macropus eugenii, the Rufous hare wallaby, Lagorchestes hirsutus, the Brush-tailed bettong, Bettongia penicillata, the Long-footed potoroo, Potorous longipes, and the Long-nosed potoroo, Potorous tridactylus). The study included animals that were clinically healthy and those that were affected by mycobacterial disease. Animals in poor health elicited weak responses, consistent with the NBT test being used as a diagnostic assay for immunodeficiency. The NBT slide assay was also applied to platypus (Ornithorhyncus anatinus) and short-beaked echidna (Tachyglossus aculeatus) granulocytes to confirm the applicability of the test to this unique group of mammals. Presented in part in poster form at the 8th Congress of the International Society of Developmental and Comparative Immunology (Cairns, Australia, 2000).