Chapter 5 The Reptiles Reptiles are considered ph yletically to represent th e first truly terrestrial vertebrates. They originated in the early Mesozoic period from an amphibian-like ancestor. In those times the y became the predominant tetrapod vertebrates, living not only on the dry land, but also in the fresh water of lakes and rivers, and in the sea. Four principal groups of reptiles have persisted to the present day. The Chelonia (turtles and tortoises) have changed little since their origin in early Triassic times and today are represented by about fifty genera. These reptiles have a world wide distribution; they are usuall y aquatic (or more strictly amphibious) in their habits. Five species live in the sea although they must return to dry land in order to lay their eggs. A number of Chelonians have adopted a life in arid desert regions and these include the North American desert tortoise, Gopherus agassiz ii, and the Mediterranean tortoise, Testudo graeca. The Crocodilia (9 genera) have existed in a relatively unchanged form since the y first appeared in the late Triassic period. They are mostly aquatic, living in the vicinity of fresh water, but at least one species, Crocodilus porosus, ventures into the sea for periods of uncertain duration. The Squamata, numerically the principal contemporary reptiles, consist of two main groups; the Lacertilia (lizards) and Ophidia (snakes), which originated in the Jurassic and Cretaceous periods respectively. Today the y are each represented by about 300 genera. The lizards have the widest geographic distribution of the reptiles and are even found on man y oceanic islands. The marine iguana of the Galapagos islands, Amblyrbyncbus cristatus, spends much of its time feeding on the algae in the sea. The snakes also have a wide distribution and one famil y, the H ydrophiidae (15 genera) lives principally in the sea. Terrestrial snakes and lizards live in habitats ranging from tropical rain forests to dry desert regions. The remaining major group is the Rh ynchocephalia, a relict branch of the reptiles with one surviving species, Sphenodon (the tuatara), which is now confined to a wet temperate environment on a few small islands situated off the coast of New Zealand. Compared with th eir amphibian ancestors, reptiles are considered to be relatively independent of th e proximity of fresh water. Many species, however, continue to live in the vicinity of Jakes and rivers, though, unlike almost all amphibians they produce an egg that, by virtue of its shell and amnionic membrane, can be laid on land. Some of the sea snakes, Pelamis, however, do not even need to return to land for this purpose, as the y give birth to live young in the sea. As compared to the amphibians the reptiles have a relatively impermeable skin which limits exchanges of their water and salts with the environment. Many reptiles, especially those living in arid areas, can form uric acid (instead of urea or ammonia) as the principal end-product of nitrogen metabolism, so that they require little water for the renal excretion of catabolic nitrogen. The three aforementioned factors, amniotic egg, relatively impermeable inte gument and uricotelism, distinguish the reptiles osmotically from their ancestral amphibian forms and must cont ribute substantially to the cosmopolitan radiation of this group into regions of potential osmotic hostility. 135 P. J. Bentley, Endocrines and Osmoregulation Springer-Verlag Berlin Heidelberg 1970
The geographic dispersion of many terrestrial species across large expanses of the oceans is often diffigult to envisage, but this has undoubtedly occurred among the reptiles (DARLINGTON, 1957), a feat that must largely reflect their relative osmotic independence from their environment. Reptiles are thought to have originated in the old world and migrated to the new world, with a subsequent pilgrimage from South America to Australia. DARLINGTON states that these movements can be explained ' without resorting to special land bridges or continental drift' presumably by making transoceanic voyages. Fossil representatives of an extinct group of land tortoises (Meiolaniidae) from South America have been found on Lord Howe Island and Walpole Island in the eastern Pacific Ocean. It seems likely that these reptiles floated across the many hundreds of kilometers of ocean to occupy these remote islands. Turtles have a remarkable propensity to survive such conditions. When at Duke University I placed three turtles, Pseudemys scripta, in a tank of sea-water, two of these animals died within 14 days but the third was still alive after 31 days. I replaced it in fresh water after this time, and it proceeded to carryon with its normal existence. Among the snakes, the genus Natrix has a remarkably wide geographic distribution and is found in Europe, Asia, Africa, Australia, America and islands such as Cuba and Madagascar. These snakes are primarily aquatic, but have probably attained their cosmopolitan status by crossing the seas. PETTUS (1958) has.cornpared the fluid metabolism of two races of Natrix sipedon living in the southern United States. One of these groups normally lives in fresh water and the other in brackish water; but the former cannot usually survive transfer to a saline medium. The success of such adaptation seems to reside solely in a predisposition to refrain from drinking salt solutions. Providing that reptiles maintain the relative impermeability of their integument, dispersal of terrestrial species through the seas is physiologically conceivable. Reptiles have a water content equivalent to about 70% of their body weight, an amount similar to that of birds and mammals, though less than that of the Amphibia. The concentration of the body fluids conforms to the usual tetrapod pattern, being about 250 to 300 m-osmole/l. Variations in the concentration and electrolyte content of the body fluids do, however, occur; reptiles in a marine environment have a slightly higher plasma concentration than those living on land (Table 5.1) though they remain hypoosmotic to sea-water. Reptiles can tolerate quite large changes in the concentrations of their body fluids; the sodium concentration in the plasma of the lizard, Trachysaurus rugosus, may rise from 150 to nearly 200 m-equiv/l during the dry Australian summer (Table 5.1) and comparable increases have also been observed in the lizard, Amphibolurus ornatus, and the desert tortoise (BRADSHAW and SHOEMAKER, 1967; DANTZLER and SCHMIDT-NIELSEN, 1966). Decreased plasma electrolyte levels can also be readily tolerated; the softshell turtle, Trionyx spinifer, normally has a plasma sodium concentration of about 150 m equiv/l, but in healthy hibernating individuals this may decrease to 80 m-equiv/l (Table 5.1). Birds and mammals do not seem to be able to withstand such variations in the concentrations of their body fluids, but in some reptiles such tolerance may be an important factor in their ability to survive adverse conditions when the possibility of osmotic regulation is limited. Reptiles are poikilotherms and so, unlike birds and mammals, do not utilize evaporative water loss for thermal cooling. Nevertheless, they do attempt to 136
Table 5.1 Concentrations of sodium and potassium in the plasma of reptiles from various habitats Habitat Plasma concentration (m-equiv/l) Sodium Potassium Chelonia Chelonia mydas l (Green turtle) a) Kept in sea-water b) Kept in fresh water Trionyx spinifer 2 (Soflshell turtle) a) Normal b) Hibernating Marine Fresh water 158 1.5 130 2.6 144 90 Lacertilia Trachysaurus rugosus 3 (Bobtail lizard) a) Winter Semi-desert 152 3.5 b) Summer Uromastyx acantbinurust ('Dob') Amblyrhynchus crlstatusr (Marine iguana) 196 4.6 Desert 150 4.4 (Sahara) Marine 178 11 Ophidia N atrix cyclopiont (Water snake) Fresh water 134 5.9 Pelamis platurus' (Yellow-bellied sea snake) Marine 264 11 References: IHOLMES and McBEAN (1964); 2DuNSON and WEYMOUTH (1965); 3BENTLEY (1959b); 4GRENOT (1968); 5D unso N (1969) ; 6ELIZONDO and LEBRJE (1969); 7DuNsoN (1968). maintain their body temperature at levels that are optimal for their activity and discreetly utilize external heat sources for this purpose. This is performed principally by conforming to a certain pattern of behaviour; warming by periodic basking in the sun and opposing the ventral body surface to warm surfaces, and cooling by seeking shade and refuges, such as rock crannies and burrows, where the temperature is more moderate. Modification of the body temperature may be assisted physiologically by changing the conductivity of the body by means of circulatory adjustments. Some species can change the colour of their skin so as to alter the rate of absorption of solar radiation (see SCHMIDT-NIELSEN, 1964a). Such adjust- 137
ments are probably mediated principally by the nervous system, though in some reptiles melanocyte stimulating hormone mediates colour change (WARING, 1963). Evaporation of water from the respiratory tract and skin increases when the temperature of the body and the environment rises, but this is physically obligatory and cannot be considered as part of a regulatory response to facilitate cooling. The principal avenues for the exchange of water and solutes between the reptile and its environment are basically the same as in mammals and birds. The physiological significance and magnitude of the exchanges through the different channels however, differ. The main osmoregulatory organs are the kidneys, as in all vertebrates, but in certain reptiles the action of these organs may be augmented.by cephalic 'salt' glands, and the cloaca and urinary bladder. 1. Water Exchanges a) Skin and Respiratory Tract In a terrestrial environment reptiles lose water by evaporation from the external integument, and the lungs and pulmonary passages. This loss increases when body temperature and environmental temperature increase; thus the bobtail goanna, Trachysaurus rugosus, when kept in dry air at 25 0, loses water by evaporation at the rate of 0.7 g/100 g day while at 37.5 0 it is 2.9 g/100 g day (WARBURG, 1965a). This water loss is small, relative to that in birds, mammals and the Amphibia, and makes little contribution to thermal cooling; the temperature of the goannas when equilibrated to such-conditions only being one or two degrees less than that of the environment. The increased evaporation of water at high environmental temperatures is due to the decrease in the saturation deficit for water in the surrounding air, and an increase in the bod y temperature of the animal. The metabolic rate of reptiles increases two to three times for every 10 0 rise in body temp~rature(benedict, 1932), so that there is an increased rate of gas exchange in the lungs that further facilitates water loss. Different species of reptiles have been observed to lose water by evaporation at various rates (BOGERT and COWLES, 1947). Thereasonsfor suchdifferenceswere not initially clear as they could involve several factors such as: the rate of oxygen consumption, the ratio of surface area to body weight or the particular properties of the skin of the different species. This was not further investigated until SCHMIDT NIELSEN in 1964 (a) suggested, after examining the fragmentary information that was then available, that cutaneous evaporation in reptiles may be greater and more variable than was hitherto considered likely. When cutaneous and pulmonary water losses were measured separately in a variety of species of reptiles (at 23 0) the movement through the integument was indeed found to contribute from 66 to 87% of the total evaporative loss (see Table 5.2; BENTLEY and SCHMIDT-NIELSEN, 1966; SCHMIDT-NIELSEN and BENTLEY, 1966). At higher temperatures (35 0 and 40 0 ) pulmonary water loss was increased relative to that from the skin but was still about 50% of the total. Similar results have been described in a variety of lizards (see CLAUSSEN, 1967). The rates of water loss across the skin vary considerably in different reptile species, in a manner that suggests that animals normally occupying 138
... VJ '-D T abl e 5.2 Eva por ative w ater loss from the respiratory tract and skin of v arious rept iles in dry ai r at 23 R espiratory H 20 Loss Cu tan eous H 20 Loss O2 Consum ption Habitat total mg / g day mg/rnl O j rng/cm" day as % to tal mllg day Crocodilia Caiman sclerops 9.6 4.9 33 87 1.8 Aquatic (Caima n) - Chelonia Pseud emy s scripta 4.3 4.7 12 78 0.9 Aquatic (Slider turtle) Tcrrapene carolina 2.6 4.2 5.3 76 0.6 Temp erat e for est (Box turtle ) Go pherus agassizii 0.4 1.5 1.5 76 0.26 Desert (Desert tortoi se) -- Lacertilia Iguana iguan a 3.4 0.9 4.8 72 2.6 Tropical for est Sauroma lus obesus 1.1 0.5 1.3 66 1.2 De sert (C huckawalla) -- Ophidia' Natrix taxispilot a 3.6 1.6 16.7 88 2.1 Aquati c (Brown wat er sna ke) Pitu oph is catenifer 3.1 1.3 3.7 64 2.3 Desert (Sonora gophe r snake) References: lfro m PRANGE and SCHMIDT-NIELSEN (1969) others from BENTLEY and SCHMIDT- N IELSEN (1966) and SCHMIDT-NIELSEN and BENTLEY (1966)
T abl e 5.3 Cu taneous water exchange in reptiles k ept in fr esh water or 3.3% sodiu m chloride solut ion (= sea-water) mgt em» day fresh water 3.3 % NaCI (gain) (loss) Crocodilia Caiman scleropss (C aiman) 26 14 Chelonia Trionyx spinifer 2 (Softshell turtle) Pseudemys scripta 2 (Slider turtle) 26 6 38 10 Lacertilia Sauromalus obesus 2 (C huckawalla ) Not detectable IBENTLEY an d SCHMIDT-NIELSEN (1965) 2BENTLEY and SCHMIDT-NIELSEN (1970) dr y habitats suffer a smaller loss in this way than those from wetter situations. Thus, the aquatic crocodilian, Caiman sclerops, loses water by evaporation through the skin 25 times as rapidly as the desert lizard, Sauromalus obesus. The skins of various species of reptiles also exhibit differ ences in their osmotic perm eability to water. In fresh water, whi ch is hypoosmotic to its body fluids, the crocodilian, Caim an sclerops, takes up water acro ss its inte gum ent at the rate of 26 mg/crrr' day (Table 5.3) which is 20 to 50% the rate of osmotic water uptake across the skin of amphibians. The softshell turtle, Trionyx spinifer, takes up water osmotically at a similar rate, but the slider turtle, Pseudemys scripta, takes up water onl y about 20% as rapidl y. The softshell turtle is covered with a soft pliable skin whil e about two-thirds of the surface of Pseudemys consists of hard keratin plates which probably have a more restricted permeability to water. When these reptiles were kept in hyperosmotic sodium chloride solution, similar in concentration to sea-water, the y lost water (Table 5.3). The desert lizard, Sauromalus, has a very restricted permeability with respect to evaporative loss and we could not detect any mo vement across the skin of these animals when they were bathed for a day in fresh water or 3.3 % sodium chloride solution. TERcAFs (1963) has, ho wever, shown that the skin of the lizard, Urom astyx, is osmotically permeable to water in vitro, an observation that is not comparable with ou r in v ivo experiments. The skin of man y Amphibia exhibits an increas ed osmotic permeability to water in the pres ence of neu rohypophysial hormones but there is no evidence that suggests that this occurs in reptiles. 140
Water loss from the respiratory tract depends on the rate of oxygen con sumption and the ability to extract this gas from the inspired air. The oxygen consumption of reptiles depends on their activity and bod y temperature. In addition, the basal metabolic rate seems to vary considerably in diffe rent reptiles. An inspection of the rate s of oxygen consumption of the different reptiles in Table 5.2 indicates a range from 0.26 mllg day in the desert tortoise to 2.6 mllg day in the iguana. Such differences are partly reflected in the respective rates of water loss from the respiratory tract of such animals. The desert tortoise thus loses water through this channel at the rate of 0.4 mg/g day while the iguana loses 3.4 mg/ g day. The relationship between oxygen consumption and respiratory water loss is, however, not an exact one as there is considerable variation in the ability of the different animals to extract oxygen from the inspired air. Usually this corresponds to a reduction of oxygen from the atmospheric volume of 20.9 % to only 17 to 20%. The chuckawalla, Sauromalus obesus, can on occasion breathe sporadically at the rate of only 2 or 3 breaths an hour and reduce the oxygen concentration in its lungs to 5% of the volume in the atmosphere (SCHMIDT-NIELSEN, CRAWFORD, and BENTLEY, 1966). Such behaviour would reduce respiratory water loss, but whether this occurs normally as a response to promote water conservation is uncertain. The thyroid hormones control the rate of oxygen consumption in birds and mammals but this does not normally seem to be so in reptiles, though in certain circumstances, as when the temperature is elevated, the y may be involved (LYNN, MCCORMICK, and GREGOREK, 1965). Evaporative water losses in reptiles are probably temporally regulated, in conjunction with the body temperature, by changes in behaviour. Reptiles may be diurnal or nocturnal in their habits and avoid extreme temperatures by seeking the shelter of cool refuge s. WARBURG (1965a) found goannas, Trachysaurus rugosus, in burrows 2 to 3 metres deep. The temperature in these holes never exceeded 28 0, even though the external air temperature was as high as 40 0 and that of the surface soil 50 0 b) Urinary and Faecal Water Loss Like in birds, it is difficult to distinguish clearly between urinary and faecal water losses in reptiles, as both products pass through a cloaca prior to their expulsion. The proportion of the animals' total water loss that is lost through such channels varies, and in terrestrial situations this is partly dependent on the temperature, which influences the rate of evaporation more than other water losses. At a temperature of 23 0 (in dry air) the crocodilian, Caiman sclerops, loses about 20% of its daily total water deficit through the cloaca (BENTLEY and SCHMIDT-NIELSEN, 1965 and 1966). The lizard, Trachysaurus, which lives in semi-arid areas, loses 40% of its overall water loss in this way at 23 0 but only about 25 % at 35 0 At 30 0 Uta besperis, a lacertilian from southern California, onl y loses 5% of its total daily water decrement in the urine and faeces (CLAUSSEN, 1967). While in aquatic situations evaporation is expected to be small, a net water loss may still occur osmotically across the skin if the environmental fluid is hyperosmotic, like the sea. Additional water may also be excreted through the cephalic 'salt' glands in such species. When the estuarine turtle, Malaclemys centrata, is kept in sea-water, it loses about equal 141
amounts of water through cloacal and extrarenal channels (BENTLEY, BRETZ, and ScHMIDT-NIELSEN, 1967a). The normal rates of urine flow differ considerably in different reptiles (Table 5.4) and can vary from amounts equivalent to about 0.5% of the body weight in Malaclemys kept in sea-water, to 6% in the gecko lizard and 9% in the caiman when kept in air. Water loss from the kidney and gut can constitute an avenue of water loss, which in circumstances when such supplies are restricted, may be physiologically critical. Reptiles, unlike mammals and birds, cannot form a urine that is hyperosmotic to their body fluids and so cannot conserve water by increasing urine concentration in this way. However, some do have the ability to convert increased amounts of theirwaste nitrogen into uric acid, which requires less water for its renal excretion than the alternative products, ammonia and urea. The desert tortoise, Table 5.4 Variations in the urine {low of reptiles in different states of hydration mllkg hr Habitat Normal Dehydrated Water loaded Saline loaded Crocodilia Caiman scleropss a) In fresh water b) In air 3.5 1.1 Aquatic Chelonia Pseudemys scripta 2 (Slider turtle) Malacl emys centratas (Diamondback terrapin) a) In fresh water b) In sea-water Gopherus agassizii» (Desert tortoise) 1.3 Aprox. Anuric 1.0 0.2 2.0 Aprox. Anuric 3.6 8.3 0.4 to Anuric Aquatic Estuarine 0.9 to Desert Anuric Lacertilia Hemidactylus Sp.4 (Gecko) Phrynosoma cornutumi (Horned toad) Trachysaurus rugosus 5 (Bobtail lizard) 2.6 2.0 0.24 1.3 0.8 Anuric 12.1 1.8 11.3 2.4 Desert 0.6 Wet tropical Anuric Semidessert Ophidia N atrix sipedon': (Water snake) Varies with vasotocin injections Aquatic IBENTLEY and SCHMIDT-NIELSEN (1965); 2DANTZLER and SCHMIDT-NIELSEN (1966) ; 3BENTLEY et at. (1967) ; 4RoBERTS and SCHMIDT-NIELSEN (1966); 5BENTLEY (1959b); 6DANTZLER (1967a). 142
... ~ V.J T abl e 5.5 Glomerular filtration rate of va rious rept iles in different states of hydration GFR mllkg h Normal Saline loaded Wa ter load ed D ehyd rated H abi tat Crocodilia Cr ocodilus acut us! 9.6 6.1 13.0 7.3 Fresh water Crocodilus poros us': 1.5-18.8 2.8 Sometimes marine (Sal t water crocodi le) - - Chelonia Pseudemys scrip ta 3 4.7-10.3 2.8 Fresh water (Slider turtle) Chelonia mydas 2 14.3 - - - Marine (Green turtle) Gopherus agassizii 3 4.7-15.1 2.9 D esert (Dese rt tortoise) -- Lacertilia H emidactylus Sp.4 loa 3.3 24.3 11.0 Wet tropical (Gecko) Phr yn osoma cornu tum 3.5 2.1 5.5 1.7 Desert (Horned toad ) Trachysaurus rugosus 5 1 1 3 Zero Semi-Desert (Bobta il lizard ) ' B. SCHMIDT-NI ELSEN and SKADHAUGE (1967); 2B. SCHMIDT-NI ELSEN and DAVIS (1968) ; 3DANTZLER and SCHMIDT-NI ELSE:-l (1966); 4RoBERTS and SCHMIDT-NIELSEN (1966); 5Calculated from SHOEMAKER et al. (1966) and BENTLEY (1959b).
Gopherus agassizii, normally excretes relatively more waste nitrogen as uric acid than the aquatic turtle, Pseudemys scripta (DANTZLER and SCHMIDT-NIELSEN, 1966). Some tortoises also may alter the relative quantities of these nitrogenous end-products in response to the state of the reptiles' hydration (KHALIL and HAG GAG, 1955), uric acid predominating when water is restricted. The urine flow of reptiles exhibits considerable lability (Table 5.4). When water is administered to the lizard, Trachysaurus rugosus, its urine flow may increase nearly 50 times as compared to that normally observed, while if the water intake is restricted, the urine volume may be too small to measure. In reptiles, variations in urine flow are related to changes in both the GFR (Table 5.5) and reabsorption of water from the renal tubules. This contrasts with mammals, in which the latterprocess normally predominates. Thepossibleamplitudeof the changes in GFRdiffers in various species and may be only 2 or 3-fold in a lizard such as the horned toad, and more than la-fold in the crocodile (Table 5.5). As indicated by differences in the inulin or creatine urine/plasma concentration ratios, renal tubular water reabsorption may also vary. In mammals this normally amounts to more than 99% of the water that is filtered across the glomerulus, but in the horned toad this is only 45% in normally hydrated animals though it may increase to 60% during dehydration (ROBERTS and SCHMIDT-NIELSEN, 1966). In the Australian lizard, Trachysaurus, tubular water reabsorption is only 40% in hydrated individuals, but it may increase to more than 95% when the urine flow is low (SHOEMAKER, LICHT, and DAWSON, 1966). Thus, the relative importance of the changes in GFR and renal tubular water reabsorption varies in different species of reptiles and is also dependent on the physiological circumstances. At high rates of urine flow, increases in the GFR may playa predominant role in controlling urine volume, but when the urine volume is small, tubular water reabsorption may be relatively more important. Ideally the two processes probably work in conjunction with each other. Changes in GFR may be due to an increased rate in filtration across individual glomeruli, or can also result from changes in the numbers of active units (glomerular intermittency). The latter process has been shown to occur in the snake, Natrix sipedon (LEBRIE and SUTHERLAND, 1962; DANTZLER, 1967a) and the turtle, Pseudemys scripta (DANTZLER and SCHMIDT-NIELSEN, 1966). Thesite of the tubular water reabsorption has been examined by 'stop-flow' procedures in Natrix sipedon (DANTZLER, 1967b) and the results suggest that during antidiuresis, water is reabsorbed from the distal parts of the nephron. While it is clear that both changes in the GFR and reabsorption of water by the renal tubule mediate alteration in the urine flow of reptiles, the immediate physiological reasons for such changes are, at the best, speculative. Changes in the GFR can be envisaged as resulting to some extent from dilution or concentration of plasma protein, which could, respectively, increase or decrease the forces for ultrafiltration across the glomerulus. Haemodynamic factors, resulting from changes in the blood pressure, can also conceivably alter glomerular activity. Tubular water reabsorption can be influenced by the rate at which the filtrate is delivered to the absorption sites. Water transfer across the renal tubule may occur as an osmotic accompaniment of solute movement or, as in mammals, result from a change in the properties of the tubular epithelium, allowing osmotic equilibration between the two sides to occur more rapidly. We do not know which of these factors occurs 144
physiologically in reptiles, but injections of peptides from the neurohypophysis can initiate decreases in glomerular activity and increase fluid absorption from the renal tubules. a) R ole of Neurohypophysis. Three peptides with such actions on the kidney have been identified in the neurohypophyses of different reptile species. H E L LE R (1942) showed that the pituitary of the European grass snake, Tropidonotus natrix, con tained a biological activity similar to that in mammals in so far as it had an antidiu- Table 5.6 Distribution of neurohypophysial pep tides in reptiles Vasotocin Mesotocin Oxytocin Crocodilia Caima n scleropss (Caiman) Chelonia Testudo graeca 2 (Tortoise) Chelonia mydas 1 3 (Gr een turtle) Pseudem ys scripta' (Slider turtle) Lepidochelys kempi 3 (Ridle y turtle) Caretta Sp. 3 (Loggerhead turtle) Oxytocin - like (?) (?) Oxytocin - like (?) (?) (?) (?) Lacer tilia Iguana iguanas (Iguana) Oxytocin - like Ophidia Tropidonotus natrix" 3 (Grass snake) Crota lus atroxi (Ra ttle r) Naja naja 6 8 (Cobra) Vipera aspis" 8 (Viper) Elaphe quadrivergata 8 (Elaph) (?) (?) ± ISAWYER et al. (1961) ; 2HELLER and PICKERING (1961); 3FoLLETT (1967); 4SAWYER (1968); 5MuNSICK (1966); 6PICKERING (1967); 7ACHER, CHAUVET, and CHAUVET (1968); BAcHER, CHAUVET, and CHAUVET (1969 a). 145
retic effect when injected into rats. However, its properties were not identical with those of mammalian neurohypophysial peptides, as it exerted a far more potent 'water balance effect' (water retention) when injected into frogs. The activity in the neurohypophysis of the grass snake was later (HELLER and PICKERING, 1961) shown to be pharmacologically identical with 8-arg101Oe-oxytocin (vasotocin). A second peptide, similar in its properties to mammalian oxytocin, was also found in the grass snake. The caiman neurohypophysis was found to contain vasotocin, but a second activity could not be identified in this species, though both were found in the green turtle, Chelonia mydas (SAWYER, MUNSICK, and VAN DYKE, 1961). Vasotocins have since been identified in othergroups (Table5.6), includingthesnakes, in which their structure has been confirmed by amino acid analysis (PICKERING, 1967; ACHER, CHAUVET, and CHAUVET, 1968). The second peptide activity that was found initially in most of the reptiles examined has been pharmacologically identified as 8-ileu-oxytocin (mesotocin) in the rattlesnake (MUNSICK, 1966) and found chemically to be such in the viper, cobra and elaph (ACHER et al., 1968; ACHER, CHAUVET, and CHAUVET, 1969a). PICKERING (1967) found that amino acid analysis, indicated both the presence of mesotocin and oxytocin in the neurohypophysis of the cobra and FOLLETT (1967) has presented tentative pharmacological evidence to suggest that this may also be so in other reptiles. ACHER et al. (1969 a) emphasize that they are unable chemically to detect oxytocin in the viper, cobra or elaph. PICK ERING also found an additional peptide, similar to vasotocin, in the cobra, but this has not yet been exhaustively characterized. Few results indicating concentrations of such peptides 10reptiles are available, but in the grass snake the level of vasotocin is about 4-9 X 10. 9 M kg body weight (calc. from HELLER and PICKERING, 1961; FOLLETT, 1967), a value comparable with those in other tetrapods. There is usually 2-4 times as much vasotocin as the other active peptides. Mammalian neurohypophysial peptide preparations have been shown to exert an antidiuretic effect when injected into the alligator and into Trachysaurus rugosus (Table 5.7). Extracts of reptilian neurohypophyses also exert such an action when injected back into the contributing species. DANTZLER (1967 a) has made a careful analysis of the actions of vasotocin, mesotocin and oxytocin on the renal function of the water snake Natrix sipedon. All of these neurohypophysialpeptides can produce an antidiuresis, but mesotocin and oxytocin are only effective in large doses that are always associated with a reduced GFR and a decrease in blood pressure. Vasotocin reduces urine flow in small doses that have no action on blood pressure and acts both by reducing the GFR and by increasing tubular water reabsorption. The renal tubular action of vasotocin has a lower dose-response threshold than its glomerular effects. The results suggested that the decreased GFR was due to a reduction in the number of active glomeruli. It is uncertain whether vasotocin has this pattern of action throughout the Reptilia for, as we have seen, there is a considerable diversity in the renal processes of this group. Injection of large amounts of Pitressin into the alligator thus was found to reduce the GFR without affecting tubular absorption but such experiments should be repeated using small doses of vasotocin. The evidence for a physiological role of neurohypophysial peptides in controlling urine flow in reptiles remains circumstantial, and less complete than in any other tetrapod group. Unfortunately the effects of neurohypophysectomy on urine 146
Table 5.7 The effects 0/ neurohypophysial pep tides on urine flow, the GFR and renal tubular water reabsorption in various reptiles Mechanism Antidiuresis GFR Tubular H20 deer. Reabsorption Pep tides Used : Crocodilia Alligator mlssissippiensis'v: (Alligator) Chelonia Pseudemys scripta 3 (Slider turtle) a) Small dose b) Large dose Lacertilia Trachysaurus rugosus" (Bobtail goanna) Pit ressin, Pitocin Neural lobe extract from Pseudemys Pitocin, Pitressin, extract from lizard Ophidia Natrix sipedon 5, 6 (Water snake) Vasotocin High dose mesotocin or oxytocin = effect - = no effect IBURGESS et al. (1933) 2SAWYER and SAWYER (1952) 3DANTZLER and SCHMIDT-NIELSEN (1966) 4BENTLEY (1959 b) 5LEBRIE and SUTHERLAND (1962) 6DANTZLER (1967 a) flow have not been reported, nor has the presence of such pep tides been demonstrated in the circulation. It is noteworthy that neurohypophysial peptides exert other actions in reptiles. They may reduce the blood pressure as in Trachysaurus rugosus (WOOLLEY, 1959) and Natrix sipedon (DANTZLER, 1967 a) but high doses are required, so that these effects are probably only of pharmacological significance. Vasotocin (in vitro) contracts the oviduct of the turtle, Pseudemys scripta, when it is in an ovulating condition (M UNSICK et al., 1960). LAPOINTE (1969) has shown that vasotocin also contracts the oviduct of the viviparous island night lizard, Klauberina riversiana, in v itro. Oxytocin and mesotocin were very much less effective than vasotocin. It is possible that this action of vasotocin reflects a physiological role for this peptide in reptilian oviposition and parturition just as oxytocin assists the latter process in mammals. 147
fl) Role of Cloaca and Urinary Bladder. Urine does not pass from the reptiles kidney directly to the outside of the animal, but is first accumulated in a cloaca and in some species may pass from here into a urinary bladder. A urinary bladder is absent in Crocodilia, Ophidia and some Lacertilia, but is present in many of the latter, as well as in the Chelonia. As the urinary and faecal pellets in many reptiles contain little water, its reabsorption presumably occurs across the cloaca and large intestine, as in birds. The Brazilian snake, Xenodon sp., suffers an increased rate of water loss when the openings of its ureters are cannulated so as to allow water to by-pass the cloaca (JUNQUEIRA et al., 1966). The increased fluid loss was equivalent to 2.6% of the body weight in a day, and this may represent the amount which is normally reabsorbed in the region of the cloaca and large intestine of these snakes. Direct measurements of fluid (water and sodium) absorption from the cloaca of a large Australian lizard, Varanus gouldii, have been made (BRAYSHER and GREEN, 1970). The coprodaeum of this lizard was isolated in vivo from the rest of the cloaca with the aid of rubber bulbs. When isotonic saline was introduced into this segment it was absorbed at the rate of 8 mllkg of the lizards body weight each hour. This was increased to 21 mllkg h following the injection of vasotocin. Sodium absorption was facilitated, but the relative roles of this and differences in colloid osmotic pressure (MURRISH and SCHMIDT-NIELSEN, 1970) in promoting the water movement are not clear. The large intestine of the tortoise, Testudo graeca, is osmotically permeable to water in vitro, but high concentrations of vasotocin have no effect on water movement (BENTLEY, 1962 b). The urinary bladder of chelonians is also permeable to water and in Pseudemys scripta water movement may take place both along an osmotic gradient and as an accompaniment to active sodium transport from the mucosal to serosal surface (BRODSKY and SCHILB, 1965). The permeability of the bladder to water is, in contrast to frogs and toads, unaffected by vasotocin (CANDIA, GONZALES, GENTILE, and BENTLEY, unpublished observations). The urinary bladder of the terrestrial tortoise, Testudo graeca, is also osmotically permeable in vitro, and is also unaffected by vasotocin (BENTLEY, 1962 b). DANTZLER and SCHMIDT-NIELSEN (1966) examined the permeability of the bladder in the desert tortoise, Gopherus agassizii, to water (in vivo), and found that when water was introduced into the bladder, it was reabsorbed at the rate of about 20 mllh in either hydrated or non-hydrated animals. The urinary bladder of turtles may hold large amounts of fluid ; I have observed volumes equivalent to 20% of the body weight in the bladder of the diamondback terrapin, Malaclemys centrata. Such stores of water could facilitate survival in circumstances in which water supplies are restricted. As reptiles onl y lose water at a slow rate. rapid increases in the osmotic permeability of the bladder, like those mediated by vasotocin in frogs and toads, are probably unnecessary, so that the absence of such an action of this hormone is not surprising. Many lizards also store fluid in a urinary bladder and an investigation of the functioning of this organ, particularly in species from desert areas, would be interesting. 148
2. Salt Exchange a) Skin The relative impermeability of the integument considerably restricts the movements of salts across the body surface of reptiles. However, precise measurements of the movements of ions like sodium and potassium are lacking. When the caiman is placed in distilled water it loses sodium across its skin, the net loss amounting to about 1,u-equiv/cm 2h (BENTLEY and SCHMIDT-NIELSEN, 1965). This is about half the rate of loss in frogs and salamanders kept under similar conditions. Unlike the skin of the amphibians, no electrical p. d. could be recorded across the caiman integument in vitro when it was bathed on both sides with physiological saline, though a small p.d.(4 my, outside negative) was seen when the outside media was diluted. The latter can be accounted for by different rates of diffusion of sodium and chloride, an observation that at least confirms the permeability of the skin to ions. When the caiman is placed in 3.3 % sodium chloride solution (= to sea-water) it gains large amounts of sodium chloride but this occurs as a result of drinking, little measurable uptake occurring across the skin. I have also measured cutaneous sodium loss in the softshell turtle when it is bathed in distilled water and it loses this ion at about the same rate as the caiman. When the diamondback terrapin, Malaclemys centrata, is kept in sea-water, circumstantial evidence obtained from sodium levels in the blood and urine indicates that a small accumulation of sodium is taking place through the skin (BENTLEY, et al., 1967a). In normal physiological circumstances cutaneous exchanges of solutes are probably only of minor significance, but as apparent from the above experiments, the information is fragmentary. Species like the caiman and softshell turtle have a more permeable integument than most reptiles so they must be considered atypical, while the conditions (bathed in distilled water) for such measurements are hardly physiological. By utilizing labelled isotopes of sodium and potassium it should be possible to measure movements of electrolytes across the skin of a wider range of reptiles, under conditions that can be more properly considered physiological. b) Kidney Excretion and conservation of electrolytes occur during the process of urine formation. If excess water is accumulated, as a result of feeding, drinking or cutaneous uptake, it will be excreted by the kidney with a simultaneous, and unavoidable, loss of some salt. The ability of reptiles to form a hypoosmotic urine by reabsorption of such salts from the glomerular filtrate minimizes such losses. When the diamondback terrapin is fasted and maintained in fresh water, the urine concentration is about 60 m-osmole/l, while that of the sodium present is less than 1 m-equiv/i (BENTLEY et al., 1967a). I have found the concentration of the urine of the softshell turtle, kept under the same conditions, to be similar, while Pseudemys scripta forms a urine with a higher sodium (about 5 m-equiv/l) level. Terrestrial reptiles do not appear to perform as well as these aquatic species. I have observed sodium concentrations as low as 15 m-equiv/l in the lizard, Trachysaurus rugosus, but ROBERTS and SCHMIDT-NIELSEN (1965) found that in three species of lizards 149
(gecko, horned toad and Galapagos lizard), the urine sodium level was always about 100 m-equiv/l, even when the animals had been hydrated. Solute reabsorption from the renal tubule varied from less than 50% in the Galapagos lizards to 85% in the geckos. The ability of many reptiles to restrict urinary sodium loss is not impressive, and may be subject to some adaptation (genetic or physiological?) commensurate with the environmental conditions to which the animals are normally subjected. By analogy with other vertebrates, it appears likely that in the reptiles adrenocortical steroids may increase the reabsorption of sodium across the renal tubule and facilitate tubular secretion of potassium. However, there is little evidence available to indicate that this is so. Some experiments to elucidate the role of such steroids on urinary excretion of these ions in reptiles are clearly needed. Injections of vasotocinnot only decrease urinary waterloss in reptiles, but in the snake, Natrix sipedon, they also increase renal tubular absorption of sodium while at the same time facilitating potassium secretion (DANTZLER, 1967 a). This peptide thus mimics the actions of corticosteroids on the mammalian renal tubule. The action of vasotocin seems to be too rapid for it to be acting by releasing such steroids in the snake, so it would appear to have a direct effect. The physiological significance, if any, of this interesting response is not known. Many reptiles, especially species that live in the sea, may gain excessive quantities of solutes. The reptile kidney has a limited ability to regulate the concentrations of electrolytes, especially sodium, in the body fluids, as the concentration of the urine never exceeds that of the plasma. Indeed, when hyperosmotic solutions are administered to reptiles like the lizard, Trachysaurus rugosus, or the desert tortoise, Gopherus agassizii, they may become anuric and completely fail to excrete any of the solute (BENTLEY, 1959 b; DANTZLER and SCHMIDT-NIEL SEN, 1966). Even hypoosmotic solutions of sodium chloride are poorly excreted; when 100 mm NaCl solutions are given to Trachysaurus rugosus they excrete less than 20% of this through the kidney in 24 hours. The renal responses to potassium chloride are slightly more rapid and this may be due to the ability to secrete potassium across the renal tubule (SHOEMAKER et al., 1966). The latter response may be important in herbivorous reptiles that gain excess potassium from their diet. The relative inability of the reptilian kidney to excrete salts, may be compensated for in two ways. As will be described shortly, many reptiles have the ability to excrete sodium and potassium extrarenally through cephalic 'salt' glands. In other reptiles, which lack such accessory excretory glands, an ability to withstand considerable increases in the solute concentrations of the body fluids may exist. As we have seen, this has been shown in natural conditions in the Australian lizards, Trachysaurus rugosus and Amphibolurus ornatus. Such animals retain electrolytes and await the arrival of more adequate supplies of water which they then utilize for the renal excretion of such solutes. BRADSHAW and SHOEMAKER (1967) found that the plasma sodium levels of Amphibolurus could rise to as much as 300 m-equiv/l during periods of summer drought. During a summer rainstorm these lizards were observed to dart about and drink the water rapidly as it fell; ten hours later plasma samples were collected and the electrolyte concentrations were found to have returned to normal. Diamondback terrapins kept in sea-water accumulate sodium, but when they are placed in fresh water they drink and rapidly excrete 150
the excess solute through extrarenal channels, presumably the orbital 'salt' gland (BENTLEY et al., 1967a). Some reptiles lack extrarenal channels for salt excretion and have a poor tolerance to the presence of such solutes in their body fluids; we have found that young caiman die within 24 h after being placed in 3.3% sodium chloride solutions (BENTLEY and SCHMIDT-NIELSEN, 1965). The reptile kidney, in the absence of adequate osmotically-free water has a very poor ability to excrete sodium. c) Cloaca and Urinary Bladder The urine of reptiles, like that of birds, passes into a cloaca where it may be accumulated prior to periodic expulsion to the outside. In certain species it may pass from the cloaca to be stored in a urinary bladder. The composition of the urine, including its electrolyte content, may be altered by both of these organs. Urine is stored for about 4 h in the cloaca of the caiman, during which time the sodium concentration decreases by 50% (BENTLEY and SCHMIDT-NIELSEN, 1965). This probably affords these crocodilians a substantial reduction in renal sodium loss. Such an economy has also been observed in the crocodile, Crocodilus acutus (B. SCHMIDT-NIELSEN and SKADHAUGE, 1967). The cloaca of the caiman, in vitro, has an electrical p.d.of about 10 mv (mucosa negative) across its wall which is consistent with the process of active sodium transport. The cloaca and large intestine of the Ophidia and the Chelonia cannot be readily separated for in vitro observations, but active sodium transport also appears to occur in this region of the gut of various snakes and in Testudo graeca (JUNQUEIRA et al., 1966; BAILLIEN and SCHOEFFENIELS, 1961; BENTLEY, 1962 b). In Testudo this process is not affected in vitro by the presence of vasotocin or aldosterone. As we shall see in the next chapter sodium transport across the large intestine of toads can be facilitated by such hormones, suggesting that the actions of these substances on the cloaca and large intestine of reptiles may be worth further investigation. The urinary bladder of the chelonians, Pseudemys scripta and Testudo graeca, also has the ability to transport sodium actively from the mucosal to the serosal surfaces (BRODSKY and SCHILB, 1960; BENTLEY, 1962 b) thus further aiding conservation of sodium from the urine. In Testudo graeca this process is unaffected, in vitro, by the presence of vasotocin, an observation that contrasts with the effects of this peptide hormone in the Amphibia. However, like in frogs and toads, aldosterone increases the rate of sodium transport across the bladder of the Greek tortoise, while if these reptiles are pretreated with an antialdosterone drug (spirolactone), sodium transport is reduced. These observations need to be extended, especially to other species of reptiles, as they suggest that endogenous aldosterone may physiologically facilitate sodium transport across the urinary bladder of reptiles. The urinary bladder and cloaca of turtles may also be involved in the accumulation of sodium from the external environment. Pseudemys scripta exchange sodium with bathing fluids at the rate of 0.04 to 10 fl-moles/100g h (DUNSON, 1967). When the cloaca is blocked, this rate of exchange is reduced by 70%, indicating that the processes of sodium transfer in the cloaca and urinary bladder may be involved. Turtles are known to irrigate their cloacal regions with the fluid 151
that bathes them, so that an active transport of sodium in such regions could mediate a net accumulation of this ion by turtles. In the softshell turtle DUNSON considers that active sodium transport across the pharynx may be responsible for such solute collection. d) 'Salt' Glands "'So they went up to the Mock turtle who looked at them with large eyes full of tears'; 'for I am the crocodile' and he wept crocodile-tears to show it was quite true'''. These two quotations from LEWIS CARROLL and RUDYARD KIPLING introduce a delightful account by SCHMIDT-NIELSEN and FANGE (1958) of their discovery of the salt secreting supraorbital ('tear') glands in reptiles. The diamondback terrapin, Malaclemys centrata, and the loggerhead turtle, Caretta caretta, secretehighly concentrated salt solutions from such tear (Harderian) glands, situated in the orbit. The marine iguana, Amblyrhynchus cristatus, of the Galapagos islands utilizes a nasal gland for this purpose, just as in the birds. Crocodiles, however, have not been shown to secrete such salty 'tears'. Other marine reptiles that utilize 'salt' glands for electrolyte excretion are: the yellow-banded sea snake, Laticauda semzfasciata, and the yellow-bellied sea snake, Pelamis platurus, in which the particular tissue concerned appears to be an Harderian gland (DUNSON and T AUB, 1967; DUNSON, 1968). SCHMIDT-NIELSEN and his collaborators (1963) also found functioning nasal salt glands in a number of terrestrial lizards; the tropical iguana, Iguana iguana, the desert iguana, Dipsosauras dorsalis, and the North African desert lizard Uromastyx aegyptus. Such secretory glands have also been demonstrated in the North American lizards: the chuckawalla, Sauromalus obesus, the false iguana, Ctenosaura pectinata, and the blue spmy lizard, Sceloporus dorsalis, (TEMPLETON, 1964 and 1966), as well as a lizard, Uromastyx acanthinurus, that lives in the Sahara (GRENOT, 1967). Such functioning secretory glands are not, however, present in all terrestrial reptiles, nor when present, do they all function with similar efficiency'. The fluid produced by such salt glands in reptiles is many times more concentrated than the blood plasma while the electrolyte levels in the secretions from marine species are even higher than those in sea-water (Table 5.8). Reptiles with such accessory salt secreting organs can thus excrete the salts from a varied marine diet, and probably even drink some sea-water without incurring a deficit in their body water. While in marine reptiles the sodium concentrations present in the salt gland secretions are much higher than those of potassium (Na/K as much as 20/1), the terrestriallacertilians usually secrete a relatively potassium-rich fluid, Na/K< 0.1. Such an arrangement is consistent with the relative abundance of these ions in the diets of marine and terrestrial species. The stimuli that inititiate secretion by the salt glands of reptiles appear to be similar to those seen in birds. Injections of hyperosmotic solutions of sodium chloride, potassium chloride or sucrose and the cholinergic drug methacholine can all bring about secretion. It is interesting that even in terrestrial lizards, in which potassium is the predominant ion present, injections of sodium chloride still have a kalitropic action (TEMPLETON, 1964 and 1966). However, after such administra- 152
Table 5.8 Sodium and potassium secretion from the 'salt ' glands of reptiles Concentration Rate of secretion m-equiull u-equio! kg h Na K Na K Chelonia Malaclemys cerurata' 784 70 (Diamondback terrapin) Caretta caretta! 878 31 (Loggerhead turtle) Chelonia mydas 2 685 645 1340 50 (Green turtle) Lacertilia Ctenosaura pectinatas 21 21 0.6 19 (False iguana) Sauro malus obe sus» 44 430 3 27 (Chuckawalla) Dipsosaurus dorsalis', 5 494 1387 22 51 (Desert iguana) Uromastyx aegyptus" 639 1398 (Desert lizard) Amblyrhynchus cristatus': 969 123 2550 510 (Marine iguana) Ophidia Laticauda semijasciatal 730 33 (Yellow-banded sea snake) Pelamis platurus 8 2180 92 (Yellow-bellied sea snake)!schmldt-nielsen and FANGE (1958); 2HoLMES and McBEAN (1964) ; 3T EMPLETON (1964); 4SCHMlDT-NIELSEN et al. (1963) ; 5TEMPLETON (1966); 6DuNsoN (1969); 7D u Nso N and TAUB (1967); 8DuNsoN (1968). tion of sodium chloride a slow adjustment may occur, through the course of several days and, as shown in the false iguana, the KINa, ratio can drop from 8/1 to 1/1 (TEMPLETON, 1967). Such an effect could be mediated by hormones. In marine reptiles the quantities of salt excreted by the salt glands far exceeds those excreted in the urine; it makes up more than 90% of the total loss in the green turtle and sea snakes (HOLMES and McBEAN, 1964; DUNSON, 1968) and 75% in the marine iguana (DUNSON, 1969). Measurement of the rates of solute excretion in terrestrial lizards indicate that they are less efficient, only 20% of an administered dose of sodium chloride being excreted through the nasal glands of the desert iguana and less than 1% in the blue spiny lizard (TEMPLETON, 1966). Potassium is excreted relatively more efficiently in these lizards, the respective proportions being 40% and 10% of the given dose. Marine reptiles also appear to be able to secrete salts 153