Effects of angiotensin II and bladder condition on hydration behavior and water uptake in the toad,Bufo woodhousei

• 1.1. Water absorption response (WR) behavior and water weight gain were examined in hydrated toads, Bufo woodhousei, treated with angiotensin II (All) or with a control Ringer's solution. The effects of urinary bladder condition (ad lib. bladder urine or empty bladder) were examined concurrently.
• 2.2. Toads treated with All (100μg/100g body weight), spent more time in WR posture and absorbed more water than Ringer's-injected toads.
• 3.3. Toads with empty bladders maintained WR posture for longer periods of time and gained more weight than toads whose bladders were not emptied.
• 4.4. The effects of All and bladder urine on water absorption by B. woodhousei appear to be separate and additive.

     

Comparison of dehydration and angiotensin II-stimulated cutaneous drinking in toads, Bufo punctatus

Toads (Bufo punctatus) use a sequence of two postures to place the ventral skin on a moist surface and absorb water osmotically. First, the skin contacts the surface (seat patch down, SPD), and then the hindlimbs are abducted to maximize skin contact area (water absorption response, WR). Toads modulated behavior in response to hydration status and osmotic content of the hydration source. Dehydrated toads placed on water displayed both SPD and WR. Hydrated toads injected with angiotensin II (AII) displayed SPD longer than Ringer-injected controls but did not initiate WR and absorbed less water than dehydrated toads. These results suggest that dehydration has a more robust dipsogenic effect than AII. Dehydrated toads placed on 250 mM NaCl briefly initiated SPD but not WR. The addition of amiloride to the hyperosmotic salt solution resulted in brief display of WR but no water loss. Hydrated toads placed on 250 mM NaCl showed shorter periods of SPD behavior. The combination of AII injection and amiloride addition to the salt solution increased SPD initiation but SPD duration was short and water loss was prevented. Neither AII nor dehydration overrides chemosensory mechanisms in the skin that suppress cutaneous drinking from hypertonic solutions.     

Elevated plasma osmotic concentration stimulates water absorption response in a toad.

The water-seeking behavior (WR) of toads (Bufo viridis) was investigated. Fully hydrated toads that are allowed free choice of wet or dry filter paper voluntarily and spontaneously select to sit on water-soaked paper at a regular frequency during trials. Dehydration of bladder-emptied toads by 14% elicits WR in all animals. Injection of aldosterone or angiotensin-I reduced the dehydration threshold to 7% weight loss. WR frequency increased when plasma osmolality was elevated by injection of NaCl or other solutes (both ionic and non-ionic). Only urea, to which cell membranes are highly permeable, was the exception that did not produce this response. The increase in WR frequency induced by elevated plasma osmolality was augmented by injection of aldosterone or angiotensin-I. In vivo water uptake, measured in a water bath, was increased by an NaCl or oxytocin injection, but not by aldosterone. It is concluded that elevated plasma osmolality induces an increase in WR frequency that is separate and prior to the water uptake process. Different hormones are involved in each step.     

Intra- and extracellular dehydration has no effect on plasma levels of angiotensin II in an amphibian

Previous studies have demonstrated that both dehydration (intra and extracellular) and treatment with angiotensin II (A-II) induce changes in thirst-related behavior in the spadefoot toad, Scaphiopus couchii. One of the steps in determining a causal relationship between a hormone and a behavior is to determine that there is association between an animal's performance of the behavior and changes in endogenous hormonal concentrations. The hypothesis tested that plasma levels of the peptide hormone A-II would change as a result of dehydration known to induce water absorption response (WR) behavior in the spadefoot toad. Plasma samples were taken from toads dehydrated intracellularly by injection of hypertonic solutions of NaCl or sucrose at levels known to induce WR behavior. As an osmotic control, a group of animals was injected with urea, which has been demonstrated to not induce WR behavior. In order to determine the effects of extracellular dehydration on plasma, A-II levels in toads dehydrated by plasma volume depletion via cardiac puncture were compared to sham-punctured controls. None of the treatments in any experiment resulted in significant differences in plasma levels of angiotensin II among groups sampled at the time when WR behavior occurs. These results do not support the hypothesis that dehydration-induced thirst is stimulated by changes in plasma A-II concentrations at the onset of WR behavior. J. Exp. Zool. 286:343-349, 2000.     

Behavioral and neural responses of toads to salt solutions correlate with basolateral membrane potential of epidermal cells of the skin

Dehydrated toads initiated water absorption response (WR) behavior and absorbed water from dilute NaCl solutions. With 200-250 mM NaCl, WR behavior and water absorption were both suppressed. With 200-250 mM Na-gluconate, WR initiation was significantly greater than with NaCl but water loss was greater. Neural recordings from spinal nerve #6 showed a greater integrated response to 250 mM NaCl than to 250 mM Na-gluconate, whereas a larger rinse response was seen with Na-gluconate. Studies with isolated epithelium showed a large increase in conductance (G(t)) when 250 mM NaCl replaced NaCl Ringer's as the apical bathing solution that was accompanied by depolarization of the transepithelial potential (V(t)) and basolateral membrane potential (V(b)). Depolarization of V(b) corresponded with the neural response to 250 mM NaCl. When 250 mM Na-gluconate replaced Ringer's as the apical solution G(t) remained low, V(b) transiently hyperpolarized to values near the equilibrium potential for K(+) and corresponded with the reduced neural response. These results support the hypothesis that chemosensory function of the skin is analogous to that of mammalian taste cells but utilizes paracellular ion transport to a greater degree.     

Water potential receptors in the skin regulate blood perfusion in the ventral pelvic patch of toads

Blood cell flux (BCF) in ventral pelvic skin capillaries was measured in conscious unrestrained Bufo bufo, using a laser Doppler flowcytometer. Hydrated toads responded to water contact with a small but significant increase in BCF. Dehydration alone did not change the BCF in seat patch skin before water contact. However, water contact by dehydrated toads elicited a rapid 600% increase in BCF. The BCF and water uptake of dehydrated toads rehydrating in water declined over 2 h but remained significantly above the low, constant values measured in hydrated toads. Arginine vasotocin injection in hydrated toads did not change skin BCF, but water uptake increased, and urine production decreased. Injection of the beta -adrenergic agonist isoproterenol increased BCF in hydrated toads by 900% and also increased the rate of water uptake. These increases corresponded in magnitude and duration to the response to water contact observed in dehydrated toads. Injection of dehydrated toads with the beta -adrenergic antagonist propranolol significantly reduced both BCF and water uptake. These results are consistent with an autonomic reflex mediated by skin water potential receptors that regulate blood perfusion of ventral pelvic skin.     

Effects of dehydration on plasma osmolality, thirst-related behavior, and plasma and brain angiotensin concentrations in Couch's spadefoot toad, Scaphiopus couchii

Under dehydrating conditions, many terrestrial vertebrates species exhibit increases in plasma osmolality and their drinking behavior. Under some circumstances, this behavioral change is accompanied by changes in plasma and central angiotensin concentrations, and it has been proposed that these changes in angiotensin levels induce the thirst-related behaviors. In response to dehydration, the spadefoot toad, Scaphiopus couchii, exhibits thirst-related behavior in the form of cutaneous drinking. This behavior has been termed water absorption response (WR) behavior. Spadefoot toads live in harsh desert environments and are subject annually to dehydrating conditions that may induce thirst-related behavior. We tested the hypothesis that an increase in WR behavior is associated with both an increase in plasma osmolality and an increase in plasma and brain angiotensin concentrations. First, we determined the degree of dehydration that was necessary to initiate WR behavior. Animals dehydrated to 85% of their standard bladder-empty weight via deprivation of water exhibited WR behavior more frequently than control toads left in home containers with water available. Next, using the same dehydration methods, we determined the plasma osmolality and sodium concentrations of dehydrated toads. Toads dehydrated to 85% standard weight also had a significant increase in plasma osmolality, but exhibited no overall change in plasma sodium concentrations, indicating that while an overall increase in plasma osmolality appears to be associated with WR behavior in S. couchii, changes in sodium concentrations alone are not sufficient to induce the behavior. Finally, plasma and brain angiotensin concentrations were measured in control toads and toads dehydrated to 85% standard weight. Plasma and brain angiotensin concentrations did not increase in dehydrated toads, indicating that dehydration-induced WR behavior that is associated with changes in plasma osmolality may not be induced by changes in endogenous angiotensin concentrations in S. couchii.     

Cutaneous blood flow and water absorption by dehydrated toads

Blood cell flux (BCF) in ventral pelvic skin capillaries was measured in toads, Bufo woodhouseii and Bufo punctatus, using a chamber that allowed hydration behavior and water absorption to be observed concurrently in unrestrained animals. Dehydrated B. woodhouseii and B. punctatus placed on a rehydration solution significantly increased BCF relative to that on a dry surface in less than 2 min. Skin contact with a rehydration solution rather than dehydration alone is the primary stimulus for increased seat patch blood flow. In B. woodhouseii, the water absorption response was initiated after the increase in BCF had started but before maximum BCF was reached. BCF and water uptake across the ventral skin of both species placed on deionized water were not different from those of toads placed on 50 mM NaCl. Similarly, no significant correlation between BCF and rate of water uptake could be observed in dehydrated toads of either species. Angiotensin II (AII) injection in hydrated B. punctatus had no effect on BCF, suggesting that factors other than AII are responsible for the increase in blood flow upon water contact in dehydrated toads.     

Angiotensin II alters blood flow distribution in amphibians.

In toads, angiotensin II (ANG II) induces the water absorption response (WR) during which the seat patch (pelvic+inner-thigh skin) is pressed to a wet substrate from which water flows osmotically into the animal. Since ANG II is a potent vasoconstrictor, it has the potential to redistribute blood flow. To determine the regional circulatory effects of ANG II, we used microsphere methods to measure relative changes in blood flow to several skin regions and other organs before and after ANG II administration in terrestrial toads and aquatic bullfrogs. In toads, after ANG II administration, seat patch and bladder blood flow increased by 264.2%+/-197.6% and 287.2%+/-86.7%, respectively (P<0.05), while dorsal and pectoral skin flow decreased by 48.0%+/-19.4% and 21.3%+/-25.4%, respectively (P<0.05). In bullfrogs, ANG II caused no significant changes in blood flow. Our results support our hypothesis that, in toads, ANG II increases and decreases blood flow to regions of the body associated with water gain and water loss, respectively.     

Behaviour and body temperature of hibernating common toads Bufo bufo

Hibernation behaviour of six common toads Bufo bufo was studied. Transmitters, generating pulses with a temperature dependent repetition rate, implanted in the toad's abdominal cavity, were used to measure the body temperature. Measurements of body temperature and soil temperature at various depths were used to establish the hibernating depth. All toads moved vertically between a few preferred depths and hibernating depth appeared to vary for individual toads. Upward movements mostly were apparent reactions on changes of the soil temperature gradient at the depth of the toad.     

6-Hydroxydopamine produces a weight increase in toads

A large increase in body weight occurred after injection of 6-hydroxydopamine (6-OHDA, 100 mg kg-1) into the iliac lymph sac of toads. The body weight increase was apparently due to retention of fluid and only occurred when toads could not avoid contact of their ventral skin with water. Lower doses of 6-OHDA (25 or 50 mg kg-1) did not cause a weight increase although they were sufficient to eliminate fluorescence from peripheral adrenergic nerves. A peripheral injection of 6-OHDA did not affect the appearance of central aminergic nerves. Removal of the pituitary and median eminence did not affect the weight increase. Although the primary action of 6-OHDA in upsetting water balance in toads has not been revealed by this study, it is clearly not related to the chemical sympathectomy of adrenergic nerves.     

Comparative effects of angiotensin II on osmotic water permeability in the toad (Bufo arenarum)

The possible relationship between the renin-angiotensin system and water balance in the toad Bufo arenarum has been indirectly explored. A positive correlation was found between the hydrosmotic response of ventral pelvic toad skin to angiotensin II (A II) and some age indicators (body weight, snout-urostyle length or head width). A different hydrosmotic response for oxytocin and isoproterenol (but not for A II) was found between four cutaneous regions of toad body. We conclude that A II may not be directly involved in the regulation of water balance mediated by water absorption across the skin of Bufo arenarum toads.     

Spawn site selection and colony size of the frog (Rana temporaria) and the toad (Bufo bufo)

Information has been collected from various sources on spawn site selection and colony size of the frog ( Rana temporaria ) and the toad ( Bufo bufo ). Details are tabulated for 764 frog breeding sites in Britain and in the Republic of Ireland and for 139 toad sites in Britain (the toad does not occur in Ireland). Both species tend to breed in whatever types of site are most available. In Britain the garden pond has steadily increased in importance for both species since 1950. Frogs usually spawned in shallow water, 266 reports out of a total of 588 (45%) referring to water up to six inches (15 cm) deep. Toads tended to spawn in deeper water, 48 reports out of 82 (59%) mentioning water 7–18 inches (18–46 cm) in depth. Median depths were: frog, seven inches (18 cm); toad 13 inches (33 cm). Information on colony size was assimilated for 574 frog colonies and 86 toad colonies. Colonies in gardens were smaller than those in other types of site. Relatively fewer large frog colonies (≡100 animals) were found to occur in London than elsewhere in Britain, and British colonies were generally larger than those in Ireland. Colonies of 100 or more individuals were recorded relatively more frequently for toads than for frogs. By combining information on choice of site and colony size, rough estimates can be derived of the percentage of frogs and toads breeding in gardens and other types of site. For instance, in the London area in the 1960s perhaps 20–25% of the frogs bred in gardens and private grounds. The ecological significance of these observations is discussed.     

Transcellular and paracellular elements of salt chemosensation in toad skin

Dehydrated toads absorb water by pressing a specialized (seat patch) area of the skin to moist surfaces. This behavior, the water absorption response (WR), is preceded by periods of more limited skin contact (seat patch down, SPD) in which the suitability of the rehydration source is evaluated. WR and SPD behaviors were suppressed on 250 mM NaCl and 200 mM KCl solutions. Ten micromolar amiloride partially restored SPD and WR on NaCl solutions. The addition of 5 mM La(3+) also partially restored the initiation of WR and this effect was additive to the effect of amiloride, suggesting transcellular and paracellular pathways exist in parallel. Similarly, 5 mM La(3+) partially restored the initiation of WR on KCl solutions, to levels comparable to those with K(+)gluconate, suggesting a paracellular pathway for detection of K(+). Hyperosmotic (250 mM) NaCl solutions bathing the mucosal surface rapidly and reversibly increased the paracellular conductance of isolated skin and this increase was partially inhibited by 5 mM La(3+). These results suggest that the regulation of tight junctions has a chemosensory role in toad skin.     

The relationship between body length, age and sexual maturity in the common toad, Bufo bufo

An investigation of the relationship between the onset of sexual maturity, age and body size in common toads Bufo bufo was started in 1984 at a pond in southern England. During the toad breeding seasons of 1984 and 1985, 7259 common toad'metamorphs'were captured and permanently marked so that if captured as adults their true age would be known. Although a very small proportion of males and females reached sexual maturity at ages of two and four years respectively, most did so much later and asynchronously. The body size of toads breeding for the first time was relatively constant for each sex and not correlated with age. The rational behind our understanding of the mechanism of female choice is discussed in the light of these results.     

Biochemical and toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin in immature male and female chickens

It has been reported that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced body wasting in mammals is associated with decreased adipose tissue lipoprotein lipase (LPL) and glucose transporting (GT) activity with differential sensitivity between genders. This study extends those findings to chickens as an avian model. A significant decrease in body weight gain was demonstrated in immature male and female chickens 10 days after treatment with a single intraperitoneal (i.p.) dose of 10 and 100 microg TCDD/kg. Body weight gain decrease was associated with hepatomegaly and induction of hepatic CYP1A enzymes in both genders. The increase in liver/body weight ratio (48%) and the decreased LPL activity (28%) were significant only in females at 10 microg TCDD/kg. However, the increase in liver/body weight ratio (31%) and the decrease in LPL activity (26%) were significantly demonstrated in males at 100 microg TCDD/kg. Levels of GT were significantly decreased in females (46%) and in males (48%) following treatment with 10 microg TCDD/kg and 100 microg TCDD/kg, respectively. Therefore, in chickens, as in mammals, the TCDD-induced body wasting is accompanied with decreased LPL activity and decreased GT activity and the magnitude of these changes is gender dependent. In contrast to mammals, this study suggests that female chickens are equally, if not more responsive to TCDD toxicity than males.     

心房钠尿肽以血管紧张素及禁水引起大鼠饮水行为的影响

Atrial natriuretic peptide (ANP) present in the brain has been reported to have profound effects on water and salt metabolism. This study was designed to observe the effect of intracerebroventricular (ICV) injection of ANP on drinking behavior of rats, induced by centrally administered angiotensin II (Ang II) and 24-hours water deprivation, by using a T-maze to measure the speed they ran in a runway for water rewards. In 24-hours water deprived rats ICV injection of ANP resulted in a significant decrease of either running speed or water intake. Drinking behavior induced by ICV injection of Ang II in normally hydrated rats was also significantly inhibited by a prior injection of ANP. These findings suggest that ANP in the brain plays an important role in the central control of drinking behavior.     

Molecular phylogenetic relationship of toads distributed in the Far East and Europe inferred from the nucleotide sequences of mitochondrial DNA genes

The toads of the Bufo bufo species group are widely distributed in the Eurasian continent and Japanese Archipelago. In this study, we analyzed the mtDNA gene sequences of this species group and estimated the divergence time to clarify the evolutionary relationships and biogeography of toads distributed in the Far East and Europe. The phylogenetic tree indicated that this group produced Bufo bufo in Europe, whereas it produced B. japonicus in the Far East. B. japonicus was divided into three major clades corresponding to a group consisting of B. j. gargarizans in China, B. j. bankorensis in Taiwan, and B. j. miyakonis on Miyako Isl. and eastern and western groups of Japanese B. j. japonicus subspecies group. The eastern and western groups were divided into several subclades which tended to reflect the region-specific geographic distribution of all localities except B. j. japonicus from Hakodate. The estimated branching times of these clades suggest that geological events may have influenced the divergence of the toads distributed in the Far East and Europe.     

Adrenal responses to chronic and acute water stress in Japanese quail Coturnix japonica

Water deprivation (WD) resulted in increased serum osmotic pressure (OP) and decreased body weight (WB); adrenal aldosterone content did not change. Adrenal corticosterone content tended to be elevated during early WD, indicating a stress response, but tended to decrease after seven days of WD, suggesting adrenal fatigue. During water restriction (WR), after the period of weight loss, adrenal corticosterone content and serum OP were elevated. As the birds began to gain weight, aldosterone levels did not change but adrenal corticosterone content and serum OP approached control values, suggesting that the birds were beginning to adapt to the WR. Adrenal sensitivity to ACTH was indicated by the elevated adrenal aldosterone and corticosterone content after ACTH injection.     

中药蟾酥的药理作用研究进展

蟾酥是我国传统中药,是蟾蜍科动物中华大蟾蜍或黑眶蟾蜍的干燥分泌物。蟾酥所含化学成分主要有蟾蜍内酯类、蟾毒色胺类等,现代药理学研究表明其具有抗肿瘤、强心、局麻、镇痛、抗炎等多种作用。本文综述近年来国内外学者对蟾酥的化学成分、药理作用的研究概况。