Terrestrial amphibians take up water by abducting the hind limbs and pressing a specialized portion of the ventral skin to a moist surface, using a characteristic behavior called the water absorption response. An assay of the water absorption response was used to quantify physiological factors associated with thirst and water uptake. Dramatic changes in the water absorption response resulted from subtle changes in hydration state and from altering the reserve water supply in the urinary bladder. The water absorption response could be induced by intraperitoneal and intracerebroventricular injection of angiotensin II, demonstrating that components of the renin-angiotensin system on both sides of the blood-brain barrier have a dipsogenic function in amphibians. These experiments also demonstrated that the water absorption response could be influenced by changes in barometric pressure. Toads avoided the water absorption response on hyperosmotic substrates, and behavioral experiments showed that the amphibian skin served a sensory function similar to that of the lingual epithelium of mammals. The water absorption response assay has enormous potential as a tool for the investigation of physiological processes and sensory capabilities of amphibians.

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Toads obtain water by absorption across their skin. When dehydrated, desert toads exhibit stereotyped hydration behavior in which they press their ventral skin onto a moist surface. However, dehydrated toads avoid surfaces moistened with hyperosmotic NaCl and KCl solutions (Hoff KvS, Hillyard SD. 1993. J. Exp. Biol. 183:347–351). We have studied neural mechanisms for this avoidance with physiologic, behavioral, and morphologic approaches. Spinal nerves innervating the ventral skin could be stimulated by exposure to a hyperosmotic NaCl solution applied to the outer surface of the skin. This neural response occurred with much longer latency than to mechanical stimulation and could be reduced by amiloride, a blocker for Na+ channels known to be responsible for epithelial ion transport and salt taste transduction. In behavioral experiments, avoidance of a NaCl solution was also reduced by adding amiloride to the solution, suggesting involvement of amiloride‐sensitive Na+ channels for detecting the hyperosmotic salt solution. Neural tracing with fluorescent dye revealed spinal nerve endings and connections to putative receptor cells, both located in the deeper layer of the epidermis. Either of these or both may be associated with the transduction of Na+ flowing into the skin. The ability of toads to detect hyperosmotic salt solutions in their environment reveals a previously unknown chemosensory function for spinal nerves in anuran amphibians. J. Comp. Neurol. 408:125–136, 1999. © 1999 Wiley‐Liss, Inc.

In order to estimate metabolic demands of desert pupfish for conservation purposes, we measured oxygen consumption in fish acclimated to the ecologically relevant temperatures of 28 or 33°C. For these experiments, we used fish derived from a refuge population of Devils Hole pupfish (Cyprinodon diabolis). Measurement of routine oxygen consumption (V O2,routine ) revealed some 33°C-acclimated fish (10% of 295 assayed fish) periodically exhibited periods of no measurable oxygen consumption despite available ambient oxygen tensions that were above the critical P O2 . We call this phenomenon paradoxical anaerobism. The longest observed continuous bout with no oxygen consumption was 149 min, although typical bouts were much shorter. Fish maintained normal posture and ventilation rate (>230 ventilations per minute) during paradoxical anaerobism. Fish rarely demonstrated a compensatory increase in oxygen use following a period of paradoxical anaerobism. In contrast, only one out of 262 sampled fish acclimated at 28°C spontaneously demonstrated paradoxical anaerobism. Muscle lactate concentration was not elevated during periods of paradoxical anaerobism. However, the amount of ethanol released by the 33°C-acclimated fish was 7.3 times greater than that released by the 28°C acclimation group, suggesting ethanol may be used as an alternative end product of anaerobic metabolism. Exposure to exogenous ethanol, in concentrations as low as 0.1%, produced periods of paradoxical anaerobism even in 28°C-acclimated fish.

Regions of specialization for water absorption across the skin of Bufonid and Ranid anurans were identified by immunohistochemistry and Western blot analysis, using antibodies raised against arginine vasotocin (AVT)-stimulated aquaporins (AQPs) that are specific to absorbing regions of Hyla japonica. In Bufo marinus, labeling for Hyla urinary bladder-type AQP (AQP-h2), which is also localized in the urinary bladder, occurred in the ventral surface of the hindlimb, pelvic, and pectoral regions. AQP-h2 was not detected in any skin regions of Rana catesbeiana, Rana japonica, or Rana nigromaculata. Hyla ventral skin-type AQP (AQP-h3), which is found in the ventral skin but not the bladder of H. japonica, was localized in the hindlimb, pelvic, and pectoral skins of Bufo marinus, in addition to AQP-h2. AQP-h3 was also localized in ventral skin of the hindlimb of all three Rana species and also in the pelvic region of R. catesbiana. Messenger RNA for AQP-x3, a homolog of AQP-h3, could be identified by RT-PCR from the hindlimb, pectoral, and pelvic regions of the ventral skin of Xenopus laevis, although AVT had no effect on water permeability. In contrast, 10 Ϫ8 M AVT-stimulated water permeability and translocation of AQP-h2 and AQP-h3 into the apical membrane of epithelial cells in regions of the skin of species where they had been localized by immunohistochemistry and Western blot analysis. Finally, water permeability of the hindlimb skin of B. marinus and all the Rana species was stimulated by hydrins 1 and 2 to a similar level as seen for AVT. The present data demonstrate species differences in the occurrence, distribution, and regulation of AQPs in regions of skin specialized for rapid water absorption that can be associated with habitat and also phylogeny. hindlimb skin; arginine vasotocin; immunohistochemistry; water permeability; frogs MANY ADULT ANURAN AMPHIBIANS do not drink through their mouth. Rather, they absorb water across their skin and form dilute urine that is stored in their urinary bladder and can be reabsorbed when foraging away from a hydration source (4, 5). Hillman et al. (14) refer to this water balance strategy as semiterrestrial to distinguish it from terrestrial species that are completely independent of water. The semiterrestrial classification applies to tree frogs in the family Hylidae and toads in the family Bufonidae that have been traditionally classified as arboreal and terrestrial, respectively, in that both have large urinary bladder capacity and specializations for rapidly rehydrating when water is available. Specifically, they utilize an area of skin in the posteroventral region of the body that is specialized for rapid water absorption from shallow water sources or moist substrates. This region, termed the pelvic patch or seat patch, extends laterally to the ventral surface of the hindlimbs and shows a pattern of elevations and grooves termed verrucae hydrophilicae (14). The seat patch is also an area where capillaries form intimate contact with the basement membrane that underli...