Fins to Feet

Coming of the Amniotes

Steamy, water-logged swamps, primitive amphibians trawling the undergrowth for food and gigantic insects buzzing overhead – this was the image of the Carboniferous period (which lasted from about 360 to 300 million years ago) that every illustrated natural history book I ever read as a child drilled into my mind’s eye. Great rainforests of clubmosses, horse-tails and ferns ranged throughout the tropics. Oxygen levels were much higher than they are today – allowing invertebrates to grow to extraordinary sizes. Amphibians were the dominant land vertebrates – and ran the gamut from small newt-like creatures to large predators similar in appearance to modern crocodiles. It was in this lush, wet world that the very first Amniotes – the rather diminutive ancestors of modern reptiles, mammals and birds – made their debut. This section of Fins to Feet, will deal with the appearance of this animal group – a critical event in evolutionary history that completed the vertebrate conquest of land.

The amniote kinship of mammals and reptiles

In contrast to Amphibians, both Mammals and Reptiles are fully terrestrially adapted organisms. The exceptions to this rule include a relatively small number of reptile and mammal lineages that became secondarily adapted to living in the water – like, for example, whales and ichthyosaurs – though they still retain a number of features, common to all ‘amniotes’, that betray their terrestrial ancestry.

Both Mammals and Reptiles trace their origins to a common amniote ancestor – in other words, to creatures that laid waterproof eggs on dry land.

The first Amniotes were not the first backboned animals to walk on land – though they were more practiced land-lubbers than any vertebrate group that preceded them. Tetrapods, or four limbed vertebrates, made the first tentative forays onto land in the swamps of the late Devonian (374 – 359 million years ago). The epic vertebrate transition from water to land (the event referenced in the very title of this blog) will be the subject of a future post – and it can be regarded as having ended with the coming of the Amniotes. Early Amniotes appeared some 350 million years ago (in the first half of the Carboniferous period) in terrestrial environments already overrun with plant-life, insects and land-walking amphibians.

So what, then, are these “amniotes”? What distinguishes them from other tetrapods?

In younger years, I would have seen the anamniote-amniote transition as simply the transition from “amphibian” to “reptile” – and while this is not an altogether terrible way to think about it, some caveats are necessary. Modern amphibians, or Lissamphibians – that is, frogs, salamanders, newts, toads and the less familiar caecilians – are our only existing examples of non-amniote tetrapods. But we must not make the mistake of drawing a neat equivalence between modern Lissamphibia and the ‘amphibious’ tetrapod ancestors of amniotes. Lissamphibia lack ribs, have smooth, wet skins highly-adapted for cutaneous respiration and, for the most part, lead arboreal, aquatic or fossorial lifestyles. The ancient “amphibians” from which amniotes are derived were, by contrast, stout ribbed ground-dwelling creatures with bodies covered in thick dermal plates.

The earliest uncovered examples of Amniotes are animals that resemble – at least superficially – insectivorous terrestrial lizards. These include fossil species like Hylonomus and Paleothyris. They have slender bodies that run a length of about 20 centimeters from nose to tail-tip. They shared their world with a wide range of small to giant primitive amphibians that together made up the bulk of vertebrate terrestrial life on earth, including – for example – temnospondylls and anthracosaurs. These animals lived under various degrees of dependence on bodies of water for reproduction and egg-development. The amniotes achieved emancipation from the aquatic realm by a number of important biological innovations. Foremost among these is the Amniote egg.

The Amniote egg is characterized by a hard semi-permeable shell membrane and a suite of “extra-embryonic” tissues that surround the developing embryo  – these include the amnion, the allantois and the chorion. These eggs could be laid on relatively dry land, away from the hydrating embrace of a pond or river. Terrestrial egg-laying probably reduced the risk of predation and also enabled eggs to weather seasonal dry periods better than aquatic amphibian eggs.

Later in evolutionary history, various amniote groups would make massive modifications to this basic design – mammals, for example, did away with the fibrous shell and allowed embryonic development to proceed entirely within the uterus – but for now, let us consider what makes the primitive amniote egg a “terrestrial” egg. To do this, we now turn to the amphibian egg and consider some of its limitations in comparison to the Amniote egg.

Most modern amphibians deposit their eggs in water. The eggs hatch and undergo an aquatic larval period – a “tadpole” phase during which the organism is equipped with gills and fins and lacks limbs. Eventually the tadpole metamorphoses into the adult form, growing limbs and discarding its gills. The amniote egg is distinguished by the fact that it gives rise to developed hatchlings (which are, essentially, miniature versions of the adult form) without an intervening larval phase. This lack of a free-swimming tadpole stage is also seen in certain amphibian species– largely occupants of wet and humid tropical rainforests – which lay their eggs away from free-standing water (instead, the embryo undergoes its tadpole stage within the confines of the egg). Such a “direct developing” amphibian egg might have served as the precursor to the amniote egg.

The typical amphibian egg is bound by a vitelline membrane and one or more layers of egg jelly (together constituting the “egg capsule”). These structures protect the egg and provide support, but they do not help prevent water-loss – and thus, the egg must be laid in water or in a wet environment to avoid desiccation. Amniote eggs are more resistant to water-loss and can be laid in drier environments.

The egg capsule of amphibians is more of a barrier to the diffusion of respiratory gases (namely carbon dioxide and oxygen) than is the fibrous egg shell membrane of modern reptiles. This imposes stricter limits on the size and metabolic activity of the growing embryo in amphibians. To illustrate: As embryo size increases, the radius of the egg increases. As the radius of a spherical egg increases, the volume (4/3 pi r^3) increases much faster than does the surface area available for gas exchange (4 pi r^2 ). Given that the egg capsule is a poor agent of respiratory exchange to begin with, this relationship means that oxygen uptake through the capsule is insufficient to support embryos beyond a certain size.  Furthermore, an increase in size also means an exponential increase in the weight of the supporting tissues around it. The egg capsule would have to get thicker to support the mass, thereby reducing effective diffusion. A gelatinous egg-capsule simply does not provide enough physical support, beyond a certain egg-size and particularly on land, to prevent the deformation of the egg or the embryo by gravity or surface tension, without seriously compromising the rate of gaseous exchange.

Thus, the range of egg sizes that can be realized under the amphibian egg-plan is limited.

So how might early Amniotes have overcome this size/metabolic constraint? And how did they ultimately make the transition to egg-laying on dry land? 

1)      By enveloping the egg in a fibrous air-permeable shell membrane that [a] allowed for easier exchange of gases but kept fluids in and [b] provided enough mechanical support to resist the force of gravity on land. This proteinacious shell membrane is secreted onto the egg within the oviduct. This may have antecedents in certain amphibian species, where a layer of glycoproteins is secreted onto the egg-capsule in the oviduct before deposition.

2)      By moving water from the egg-jelly to the yolk – drying up the outer portions of the egg – effectively eliminating the egg-jelly as a barrier to gas diffusion. A process similar to this is observed during egg development in reptiles, when water is withdrawn from the albumen and transported to the yolk. Whether or not the albumen and the egg jelly are structures that are “homologous” is not known for certain.

3)      By the evolution of an extra-embryonic membrane specialized for respiratory exchange called the chorioallantois. We shall discuss this briefly in my section on extra-embryonic membranes.

All reptile eggs measured to date are larger than those observed in amphibians and generally appear to display a higher rate of metabolic activity and oxygen consumption. Amniotes on the whole display a vast range of egg sizes. Freedom from the mentioned size constraints allowed for bigger offspring and, by extension, bigger adults. This newly available range of body-sizes was crucial to the evolutionary success of the amniotes in the long run.

Some researchers suggest that elevated oxygen levels in the late Carboniferous may have eased the various size constraints (related to gas diffusion) discussed earlier and helped along with the evolution of large eggs.

Why are Amniote eggs “yolkier” than the typical amphibian egg?

Since development within the egg among Amniotes is geared towards the production of a fully-formed miniature adult rather than a free-living aquatic larva (as in most amphibians), it takes a large store of nutrients (in the form of “yolk”) to carry the embryo through the whole extent of development. In other words, amniote eggs are stocked up for the long haul – and have large yolk sacs richly provided with blood vessels to convey nutrients to the growing embryo. And so, the increased nutritional requirement imposed by “direct development” is the reason Amniotes needed bigger eggs, or larger amounts of yolk at any rate. Because more has to be invested in each egg, reptiles tend to lay fewer eggs than amphibians or fish.

What about those extra-embryonic membranes?

The position and function of the various extra-embryonic membranes has been a source of confusion for me throughout the course of my schooling, but hopefully a simple diagram will help us suss out the complicated attendant tissues and boundaries that compartmentalize the egg and regulate material exchange both between compartments and between the egg and the external world. Although there are amphibians that lay eggs on land, none of them show the complex arrangement of extra-embryonic tissues depicted below.

Let us use the chicken egg, a familiar household food-item, as our model for understanding the essential structure of the amniote egg. The embryo and supporting tissues arise from a small disc of cells, called the blastoderm, settled on surface of the yolk of the egg. The movements of the various embryonic tissue layers need not concern us here, but sufficeth to say that the growth and rearrangement of the tissues of this disc gives rise to three extra-embryonic layers: [1] the chorion is the outer-most extra-embryonic layer and, apart from providing overall enclosure for the embryo, plays a role in respiration along with the allantois (mentioned below) [2] the yolk sac encloses the yolk and is supplied with blood vessels that convey food from the yolk to the embryo [3] during development, the chorion folds over the embryo (as diagrammed below) to enclose it in the amniotic cavity. The fluid in this cavity buffers shocks and acts as a protective mechanical barrier. Finally, a fourth extra-embryonic membrane, called the allantois, develops as an outpocketing of the hindgut. It functions to store nitrogenous wastes (i.e. uric acid) produced by the metabolic activity of the chick. It expands to make contact with the undersurface of the chorion (forming the chorioallantois mentioned earlier) and serves as the principal respiratory organ for the embryo. It is richly supplied with blood vessels for gaseous exchange. As development proceeds, the yolk sac diminishes in size while the allantois grows.

The Albumen surrounds the yolk and provides additional support and nutrition.

Although mammals do not lay external eggs (with the exception of Platypi and echidnae), similar extra-embryonic membranes are seen around the developing fetus in the maternal womb.

So what really distinguishes the Amniote egg from the eggs of other tetrapods is the fibrous shell membrane and a host of extra-embryonic membranes. What else marked the first amniotes apart from their amphibious cousins?

The first Amniotes appeared to have smaller, narrower and deeper skulls than other tetrapods. A diagnostic feature that is often used to differentiate amphibians from amniotes is the absence in reptiles of an invagination called the “otic notch” which is present behind the eye-orbits in anamniote tetrapods. The anamniote-amniote transition also appears to have involved alterations in the musculature of the jaw, elaboration of the tongue, a reduction in the number of movable elements in the skull, strengthening of the ankles and the appearance of more slender bones. All in all, however, the skeletal differences between known basal amniotes and closely related tetrapods are rather minimal.

A large number of ancient amphibians, very probably including the carboniferous ancestors of amniotes, were armored in heavy dermal plates – the first amniotes appear to have traded these for horny keratinous epidermal scales that cover most of the body (with dermal gastralia on the underside). This loss of massy dermal bone lightened the body and made speedier locomotion on land possible.

What did the first amniotes eat?

Amniotes today exhibit a far wider range of diets than do amphibians – and this was key to their evolutionary success on land. Early fossil amniotes like Holonymus and Paleothyris have sharp teeth designed to pierce through the tough carapaces of invertebrates to reach the flesh within. It has been suggested that the radiation of amniotes may have to do with rising levels of insect diversity.

The development of terrestrial herbivory was a key event in the evolution of the amniotes and we shall spend a few moments trying to make sense of it.

Diadectes, a reptile-like amphibian closely related to basal amniotes, is one of the earliest terrestrial herbivores known to science (its diet is inferred from its dental and skeletal anatomy). The evolution of terrestrial herbivory (a trait unseen in modern amphibians, but common among amniotes) probably involved some sort of progression through the following stages: [a] an omnivorous stage where a diet of invertebrates was supplemented with low-fiber, high-nutrition, cellulose-poor plants or plant-parts (buds, shoots, young leaves etc.) [b] a stage in which the primary diet was high-quality, low-fiber plants and [c] finally, a stage of obligate herbivory where the diet consisted of abundant low-quality, high-fiber vegetation – [c] is an ecological role that has been continuously occupied by a succession of large land-walking animals, from Diadectes to Apatosaurus to modern grazing mammals, for the last 300 million years. High-fiber mature stems and leaves, though widely distributed, are not very digestible. They are rich in cellulose, a carbohydrate which cannot be broken down by the vertebrate digestive system without the aid of certain kinds of bacteria. It is likely that animals foraging in the leaf-litter picked up bacterial species that survived in the gut and, over evolutionary time, became members of the microbial gut flora of a species. What may have begun as incidental commensalism – where neither party benefited particularly from interaction – may have eventually evolved into symbiosis, where the bacteria broke down cellulose in ingested plant matter, making nutrients available to the host, while the host provided shelter and food to the bacteria. The disproportionately large, barrel shaped body in animals like Diadectes is designed to help bacterially ferment ingested plant material (bacterial fermentation is slow and a large space to store plant material for extended periods of time is supremely useful for an obligate herbivore). The evolution of terrestrial herbivory in Amniotes (and in closely related groups) allowed them to attain truly enormous sizes on land.

What divisions do we see among early amniotes?

We now turn, briefly, to consider some of the taxonomic divisions that can be established among early fossil amniotes by studying the structure of the skull. Temporal fenestrae are openings seen in the skull behind the eye-orbits. The establishment of these holes lightens the skull and provides additional edges for the attachment of muscles. The presence, number and position of these holes can be used to categorize amniotes. We observe four different conditions:

1)      Anapsid: No temporal fenestra is seen. Examples: Turtles and possibly Hylonomus and Paleothyris.

2)      Synapsid: A lower temporal fenestra is seen. Examples: Mammals and their extinct fossil relatives and ancestors

3)      Diapsid: Two temporal fenestrae are seen. Examples: Dinosaurs, birds, mosasaurs, lizards

4)      Euryapsid: A temporal fenestra is seen in the upper skull. This condition is probably derived from the Diapsid condition by the loss of the lower fenestra. Example: Icthyosaurs

The crucial bifurcation in the tree of Amniote life is between the Synapsida (mammals and their ancestors) and the Sauropsida (including animals with the Diapsid, Anapsid and Euryapsid conditions – that is, birds and reptiles).

The story of the Sauropsids and the Synapsids is matter for future posts. The sizes and shapes that amniotes would attain over the course of their evolution is truly astonishing. Once vertebrate life solidified its grip on land, sky really was the limit.

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