Frozen embryos? Torpor during pregnancy in the Tasmanian short-beaked echidna Tachyglossus aculeatus setosus.

We studied the interaction between torpor and reproduction in free-ranging female Tasmanian echidnas using a combination of techniques including urogenital smears, hormone analysis, ultrasonography, external temperature loggers and camera traps. Male echidnas initiated mating activity by locating hibernating females. All females that mated or were disturbed by males prior to July 27 re-entered hibernation, including many that were pregnant. Pregnant females only entered hibernation in early pregnancy when plasma progesterone concentrations were about twice basal and progesterone then remained constant during torpor. By re-entering hibernation pregnant females extended their gestation period and delayed egg-laying. Progesterone peaked 4-6days before egg-laying, then dropped rapidly.

Distinct development of the trigeminal sensory nuclei in platypus and echidna.

Both lineages of the modern monotremes have been reported to be capable of electroreception using the trigeminal pathways and it has been argued that electroreception arose in an aquatic platypus-like ancestor of both modern monotreme groups. On the other hand, the trigeminal sensory nuclear complex of the platypus is highly modified for processing tactile and electrosensory information from the bill, whereas the trigeminal sensory nuclear complex of the short-beaked echidna (Tachyglossus aculeatus) is not particularly specialized. If the common ancestor for both platypus and echidna were an electroreceptively and trigeminally specialized aquatic feeder, one would expect the early stages of development of the trigeminal sensory nuclei in both species to show evidence of structural specialization from the outset. To determine whether this is the case, we examined the development of the trigeminal sensory nuclei in the platypus and short-beaked echidna using the Hill and Hubrecht embryological collections. We found that the highly specialized features of the platypus trigeminal sensory nuclei (i.e. the large size of the principal nucleus and oral part of the spinal trigeminal nuclear complex, and the presence of a dorsolateral parvicellular segment in the principal nucleus) appear around the time of hatching in the platypus, but are never seen at any stage in the echidna. Our findings support the proposition that the modern echidna and platypus are derived from a common ancestor with only minimal trigeminal specialization and that the peculiar anatomy of the trigeminal sensory nuclei in the modern platypus emerged in the ornithorhynchids after divergence from the tachyglossids.

Development of the cerebellum in the platypus (Ornithorhynchus anatinus) and short-beaked echidna (Tachyglossus aculeatus).

The monotremes are a unique group of mammals whose young are incubated in a leathery-shelled egg and fed with milk from teatless areolae after hatching. As soon as they hatch, monotreme young must be able to maneuver around the nest or maternal pouch to locate the areolae and stimulate milk ejection. In the present study, the embryological collections at the Museum für Naturkunde, Berlin, have been used to follow the development of the monotreme cerebellum through incubation and lactational phases, to determine whether cerebellar circuitry is able to contribute to the coordination of locomotion in the monotreme hatchling, and to correlate cerebellar development with behavioral maturation. The structure of the developing monotreme cerebellum and the arrangement of transitory neuronal populations are similar to those reported for fetal and neonatal eutherians, but the time course of the key events of later cerebellar development is spread over a much longer period. Expansion of the rostral rhombic lip and formation of the nuclear and cortical transitory zones occurs by the time of hatching, but it is not until after the end of the first post-hatching week that deep cerebellar neurons begin to settle in their definitive positions and the Purkinje cell layer can be distinguished. Granule cell formation is also prolonged over many post-hatching months and the external granular layer persists for more than 20 weeks after hatching. The findings indicate that cerebellar circuitry is unlikely to contribute to the coordination of movements in the monotreme peri-hatching period. Those activities are most likely controlled by the spinal cord and medullary reticular formation circuitry.

Development of the hypothalamus and pituitary in platypus (Ornithorhynchus anatinus) and short-beaked echidna (Tachyglossus aculeatus).

The living monotremes (platypus and echidnas) are distinguished by the development of their young in a leathery-shelled egg, a low and variable body temperature and a primitive teat-less mammary gland. Their young are hatched in an immature state and must deal with the external environment, with all its challenges of hypothermia and stress, as well as sourcing nutrients from the maternal mammary gland. The Hill and Hubrecht embryological collections have been used to follow the structural development of the monotreme hypothalamus and its connections with the pituitary gland both in the period leading up to hatching and during the lactational phase of development, and to relate this structural maturation to behavioural development. In the incubation phase, development of the hypothalamus proceeds from closure of the anterior neuropore to formation of the lateral hypothalamic zone and putative medial forebrain bundle. Some medial zone hypothalamic nuclei are emerging at the time of hatching, but these are poorly differentiated and periventricular zone nuclei do not appear until the first week of post-hatching life. Differentiation of the pituitary is also incomplete at hatching, epithelial cords do not develop in the pars anterior until the first week, and the hypothalamo-neurohypophyseal tract does not appear until the second week of post-hatching life. In many respects, the structure of the hypothalamus and pituitary of the newly hatched monotreme is similar to that seen in newborn marsupials, suggesting that both groups rely solely on lateral hypothalamic zone nuclei for whatever homeostatic mechanisms they are capable of at birth/hatching.

Development of the spinal cord and peripheral nervous system in platypus (Ornithorhynchus anatinus) and short-beaked echidna (Tachyglossus aculeatus).

The modern monotremes (platypus and echidnas) are characterized by development of their young in a leathery egg that is laid into a nest or abdominal pouch. At hatching, the young are externally immature, with forelimbs capable of digitopalmar prehension, but hindlimbs little advanced beyond limb buds. The embryological collections at the Museum für Naturkunde in Berlin were used to examine the development of the spinal cord and early peripheral nervous system in developing monotremes and to correlate this with known behavioural development. Ventral root outgrowth to the bases of both the fore- and hindlimbs occurs at 6.0 mm crown-rump length (CRL), but invasion of both limbs does not happen until about 8.0-8.5 mm CRL. Differentiation of the ventral horn precedes the dorsal horn during incubation and separate medial and lateral motor columns can be distinguished before hatching. Rexed's laminae begin to appear in the dorsal horn in the first week after hatching, and gracile and cuneate fasciculi emerge during the first two post-hatching months. Qualitative and quantitative comparisons of the structure of the cervicothoracic junction spinal cord in the two monotremes with that in a diprotodont marsupial (the brush-tailed possum, Trichosurus vulpecula) of similar size at birth, did not reveal any significant structural differences between the monotremes and the marsupial. The precocious development of motor systems in the monotreme spinal cord is consistent with the behavioural requirements of the peri-hatching period, that is, rupture of embryonic membranes and egg, and digitopalmar prehension to grasp maternal hair or nest material.

Development of the dorsal and ventral thalamus in platypus (Ornithorhynchus anatinus) and short-beaked echidna (Tachyglossus aculeatus).

The living monotremes (platypus and echidnas) are distinguished from therians as well as each other in part by the unusual structure of the thalamus in each. In particular, the platypus has an enlarged ventral posterior (VP) nucleus reflecting the great behavioural importance of trigeminosensation and electroreception. The embryological collections of the Museum für Naturkunde in Berlin were used to analyse the development of the dorsal thalamus and ventral thalamus (prethalamus) in both species. Prosomeric organization of the forebrain emerged at 6 mm crown-rump length (CRL), but thalamic neurogenesis did not commence until about 8-9 mm CRL. Distinctive features of the dorsal thalamus in the two species began to emerge after hatching (about 14-15 mm CRL). During the first post-hatching week, dense clusters of granular cells aggregated to form the VP of the platypus, whereas the VP complex of the echidna remained smaller and divided into distinct medial and lateral divisions. At the end of the first post-hatching week, the thalamocortical tract was much larger in the platypus than the echidna. The dorsal thalamus of the platypus is essentially adult-like by the sixth week of post-hatching life. The similar appearance of the dorsal thalamus in the two species until the time of hatching, followed by the rapid expansion of the VP in the platypus, is most consistent with ancestral platypuses having undergone changes in the genetic control of thalamic neurogenesis to produce a large VP for trigeminal electroreception after the divergence of the two lineages of monotreme.

Development of the olfactory pathways in platypus and echidna.

The two groups of living monotremes (platypus and echidnas) have remarkably different olfactory structures in the adult. The layers of the main olfactory bulb of the short-beaked echidna are extensively folded, whereas those of the platypus are not. Similarly, the surface area of the piriform cortex of the echidna is large and its lamination complex, whereas in the platypus it is small and simple. It has been argued that the modern echidnas are derived from a platypus-like ancestor, in which case the extensive olfactory specializations of the modern echidnas would have developed relatively recently in monotreme evolution. In this study, the development of the constituent structures of the olfactory pathway was studied in sectioned platypus and echidna embryos and post-hatchlings at the Museum für Naturkunde, Berlin, Germany. The aim was to determine whether the olfactory structures follow a similar maturational path in the two monotremes during embryonic and early post-hatching ages or whether they show very different developmental paths from the outset. The findings indicate that anatomical differences in the central olfactory system between the short-beaked echidna and the platypus begin to develop immediately before hatching, although details of differences in nasal cavity architecture emerge progressively during late post-hatching life. These findings are most consistent with the proposition that the two modern monotreme lineages have followed independent evolutionary paths from a less olfaction-specialized ancestor. The monotreme olfactory pathway does not appear to be sufficiently structurally mature at birth to allow olfaction-mediated behaviour, because central components of both the main and accessory olfactory system have not differentiated at the time of hatching.

Distinct development of the cerebral cortex in platypus and echidna.

Both lineages of the modern monotremes have distinctive features in the cerebral cortex, but the developmental mechanisms that produce such different adult cortical architecture remain unknown. Similarly, nothing is known about the differences and/or similarities between monotreme and therian cortical development. We have used material from the Hill embryological collection to try to answer key questions concerning cortical development in monotremes. Our findings indicate that gyrencephaly begins to emerge in the echidna brain shortly before birth (crown-rump length 12.5 mm), whereas the cortex of the platypus remains lissencephalic throughout development. The cortices of both monotremes are very immature at the time of hatching, much like that seen in marsupials, and both have a subventricular zone (SubV) within both the striatum and pallium during post-hatching development. It is particularly striking that in the platypus, this region has an extension from the palliostriatal angle beneath the developing trigeminoreceptive part of the somatosensory cortex of the lateral cortex. The putative SubV beneath the trigeminal part of S1 appears to accommodate at least two distinct types of cell and many mitotic figures and (particularly in the platypus) appears to be traversed by large numbers of thalamocortical axons as these grow in. The association with putative thalamocortical fibres suggests that this region may also serve functions similar to the subplate zone of Eutheria. These findings suggest that cortical development in each monotreme follows distinct paths from at least the time of birth, consistent with a long period of independent and divergent cortical evolution.

Distinct development of peripheral trigeminal pathways in the platypus (Ornithorhynchus anatinus) and short-beaked echidna (Tachyglossus aculeatus).

The extant monotremes (platypus and echidnas) are believed to all be capable of electroreception in the trigeminal pathways, although they differ significantly in the number and distribution of electroreceptors. It has been argued by some authors that electroreception was first developed in an aquatic environment and that echidnas are descended from a platypus-like ancestor that invaded an available terrestrial habitat. If this were the case, one would expect the developmental trajectories of the trigeminal pathways to be similar in the early stages of platypus and short-beaked echidna development, with structural divergence occurring later. We examined the development of the peripheral trigeminal pathway from snout skin to trigeminal ganglion in sectioned material in the Hill and Hubrecht collections to test for similarities and differences between the two during the development from egg to adulthood. Each monotreme showed a characteristic and different pattern of distribution of developing epidermal sensory gland specializations (electroreceptor primordia) from the time of hatching. The cross-sectional areas of the trigeminal divisions and the volume of the trigeminal ganglion itself were also very different between the two species at embryonic ages, and remained consistently different throughout post-hatching development. Our findings indicate that the trigeminal pathways in the short-beaked echidna and the platypus follow very different developmental trajectories from the earliest ages. These findings are more consistent with the notion that the platypus and echidna have both diverged from an ancestor with rudimentary electroreception and/or trigeminal specialization, rather than the contention that the echidna is derived from a platypus-like ancestor.