Heart disease in reptiles remains infrequently reported in the veterinary literature, as do successful treatments.1–6 Congestive heart failure has been diagnosed in snakes, chelonians, and lizards.7–10,a Related clinical findings are generally nonspecific, including anorexia, lethargy, peripheral edema, ascites, cyanosis, dyspnea, exophthalmos, blepharedema, and sudden death, and can be attributable to congestive heart failure.2,8,a Many reptiles identified with heart disease are subsequently euthanized or do not survive to treatment.2,a
Diuretics such as furosemide are a mainstay of cardiac disease treatment for humans and other animals, with the aim of reducing fluid overload, edema, and effusion.11 In mammals, furosemide administration reduces reabsorption of sodium and chloride in the renal loops of Henle, thereby leading to increased excretion of water from the kidneys.12 In reptiles, use of furosemide is controversial, given that reptiles have metanephric kidneys that lack the loop-of-Henle structure and, therefore, are unable to produce hypertonic urine.13 However, in chelonians and snakes, furosemide produces diuresis by altering renal tubular electrolyte absorption and renal excretion.14–17 In addition, postrenal diuretic effects of furosemide have been reported for lizards and turtles.18,19 Clinical reports describe the use of furosemide in reptiles (lizards, snakes, and a tortoise) with congestive heart failure at dosages ranging from 1 to 15 mg/kg every 6 to 48 hours, with inconsistent results.2,9,a In loggerhead sea turtles (Caretta caretta) with brevetoxicosis, furosemide (5 mg/kg, IM, q 24 h) has been successfully used to maintain dehydration and, thereby, aid in recovery.20
The purpose of the study reported here was to evaluate the diuretic effects of and associated changes in hematologic and plasma biochemical values following furosemide administration to inland bearded dragons (Pogona vitticeps) by measuring body weight, PCV, plasma biochemical variables, and water consumption before and after furosemide treatment. It was hypothesized that SC furosemide administration would produce a diuretic effect, therefore leading to clinically measurable changes, reflected in body weight, plasma biochemical concentrations, and osmolarity in water-deprived bearded dragons.
Materials and Methods
Animals
The study protocol was approved by the University of Wisconsin-Madison School of Veterinary Medicine Institutional Animal Care and Use Committee. Nine inland bearded dragons (6 males and 3 females), ranging in age from 1 to 2 years, with a mean ± SD body weight of 0.257 ± 0.05 kg were obtained from a commercial breeder. The reptiles were housed as individuals or with conspecifics in glass tanks in a climate-controlled room with a 12-hour light cycle and room temperature maintained between 26° and 29°C. Ultraviolet B light was provided to each enclosure for 12 h/d. Within each enclosure, a temperature range existed, from approximately 34°C under the UV light to 26° to 29°C at the other end of the enclosure. Each reptile was offered gut-loadedb crickets (Acheta domestica), superworms (Zophobas morio), or mixed leafy greens once a day 6 d/wk on a set and consistent schedule. Fresh water was provided to all reptiles in a bowl at all times, and all reptiles were placed in a shallow warm water bath twice weekly.
Reptiles were allowed to acclimate to the housing conditions for at least 6 weeks before experimental procedures began. All were deemed healthy through repeated physical examinations and monitoring of food intake, fecal output, body weight, and bloodwork throughout the study.
Treatment protocol
In a complete crossover design, the 9 animals were assigned to receive furosemidec at 5 or 10 mg/kg or no treatment (control treatment) in a balanced randomized treatment sequence over a 5-week period, with a 1-week washout period provided between treatment sessions. Before each session, food was withheld for 36 hours and no greens were fed in the meal before this withholding period began. No food or water was provided for 48 hours after each treatment session began; water was removed from enclosures at the time of the first furosemide injection. Body weights were measured before each treatment session began, and a 0.4-mL blood sample was obtained from the tail vein at those points for baseline measurements by use of a 25-gauge needle and a commercial preheparinized 1-mL syringe.d Then, furosemide (5 or 10 mg/kg) was administered SC in the axillary region on alternating sides every 12 hours for 4 doses or nothing was administered during the same period.
Reptiles were weighed every 12 hours throughout each treatment session, and a second blood sample was collected 12 hours after the last furosemide injection or 48 hours after the control treatment session began. To measure water consumption, each bearded dragon was then soaked for 5 minutes in warm tap water in the same plastic tub (18 inches long by 8 inches wide by 12 inches high), During this stage, each reptile wassubmerged to the level of the radiohumeral joints when sitting up (as all did initially when being soaked). The period of 5 minutes was chosen because preliminary research had shown that many reptiles would attempt to exit the soaking tub after 5 minutes. During soaking, active drinking behavior was observed and recorded. After soaking, body weight was again measured to estimate water consumption.
Hematologic and plasma biochemical analyses
Plasma total protein, uric acid, sodium, potassium, calcium, phosphorus, and glucose concentrations were measured in blood samples collected before and after each treatment session with a commercial plasma biochemical assay marketed for reptiles.e Packed cell volume was measured with standard laboratory methods, and plasma total solids concentration was measured with a refractometer. Plasma osmolarity was measured via freezing point osmometry.f
Statistical analysis
Statistical softwareg was used to perform the data analysis. Data for each treatment session were analyzed as percentage differences from baseline values, and these values were evaluated for normality with the Shapiro-Wilk test and for equal distribution with the Brown-Forsythe test. The effects of treatment and time on measured values were assessed via repeated-measures 1-way or 2-way ANOVA. The Holm-Sidak method was used for post hoc comparisons. A value of P < 0.05 was considered significant. Data are reported as mean ± SD unless otherwise noted.
Results
Compared with control values, the percentage loss of body weight from baseline (pretreatment) was significantly greater with furosemide administered at 5 mg/kg (5.5 ± 3.2%; P = 0.006) and 10 mg/kg (5.2 ± 4.1%; P = 0.006) by 48 hours after treatment began, but not at earlier measurement points (Figure 1). No significant or clinically relevant differences in weight loss were identified between the 2 furosemide doses at any point.
Mean percentage changes in body weight from baseline (0 hours) at various points for 9 water-deprived inland bearded dragons (Pogona vitticeps) following SC furosemide administration at 5 mg/kg (white circles) or 10 mg/kg (triangles) every 12 hours for 4 doses or no treatment (black circles) for the same period in a complete crossover study design. Treatment sessions were separated by a 1-week washout period. Error bars represent SEM. *At this measurement point, the control value differs significantly (P = 0.006) from the value for furosemide at both 5 and 10 mg/kg.
Citation: American Journal of Veterinary Research 79, 11; 10.2460/ajvr.79.11.1204
Mean percentage changes in body weight from baseline (0 hours) at various points for 9 water-deprived inland bearded dragons (Pogona vitticeps) following SC furosemide administration at 5 mg/kg (white circles) or 10 mg/kg (triangles) every 12 hours for 4 doses or no treatment (black circles) for the same period in a complete crossover study design. Treatment sessions were separated by a 1-week washout period. Error bars represent SEM. *At this measurement point, the control value differs significantly (P = 0.006) from the value for furosemide at both 5 and 10 mg/kg.
Citation: American Journal of Veterinary Research 79, 11; 10.2460/ajvr.79.11.1204
Mean percentage changes in body weight from baseline (0 hours) at various points for 9 water-deprived inland bearded dragons (Pogona vitticeps) following SC furosemide administration at 5 mg/kg (white circles) or 10 mg/kg (triangles) every 12 hours for 4 doses or no treatment (black circles) for the same period in a complete crossover study design. Treatment sessions were separated by a 1-week washout period. Error bars represent SEM. *At this measurement point, the control value differs significantly (P = 0.006) from the value for furosemide at both 5 and 10 mg/kg.
Citation: American Journal of Veterinary Research 79, 11; 10.2460/ajvr.79.11.1204
Compared with percentage differences from baseline in control values, plasma total solids and total protein concentrations increased significantly from baseline with furosemide at both doses (Table 1). Packed cell volume also increased significantly from baseline with furosemide administered at 10 mg/kg (P = 0.006) but not at 5 mg/kg (P = 0.07). No significant or clinically important differences in plasma osmolarity and plasma uric acid and electrolyte concentrations were identified between percentage differences for control values and those for furosemide administered at 5 or 10 mg/kg.
Mean ± SD PCV and plasma biochemical values for 9 water-deprived inland bearded dragons (Pogona vitticeps) before (baseline) and after SC furosemide administration at 5 or 10 mg/kg every 12 hours for 4 doses or no treatment (control treatment) for the same period in a complete crossover study design.
| Measurement point | Comparison between measurement points | |||||
|---|---|---|---|---|---|---|
| Variable | Treatment | Baseline | 48 hours | Absolute difference | Percentage difference | P value* |
| PCV (%) | Control | 27.0 ± 4.0 | 25.9 ± 4.5 | −1.1 ± 1.1 | −4.4 ± 3.7 | - |
| 5 mg/kg | 25.3 ± 5.0 | 26.2 ± 4.6 | 0.9 ± 1.4 | 4.1 ± 5.7 | 0.07 | |
| 10 mg/kg | 27.0 ± 5.0 | 29.6 ± 6.3 | 2.6 ± 4.0 | 9.5 ± 14.0 | 0.006 | |
| Total solids (g/dL) | Control | 5.4 ± 0.7 | 4.9 ± 0.9 | −0.6 ± 0.6 | −10.6 ± 11.3 | - |
| 5 mg/kg | 5.4 ± 0.8 | 5.9 ± 0.8 | 0.5 ± 0.6 | 9.8 ± 11.9 | 0.03 | |
| 10 mg/kg | 5.4 ± 1.1 | 6.7 ± 1.0 | 1.3 ± 1.3 | 27.0 ± 28.5 | 0.001 | |
| Osmolarity (mOsm/L) | Control | 314.0 ± 6.4 | 308.9 ± 7.7 | −5.1 ± 6.8 | −1.6 ± 2.2 | - |
| 5 mg/kg | 312.4 ± 12.6 | 315.6 ± 12.9 | 3.1 ± 10.2 | 1.0 ± 3.2 | 0.17 | |
| 10 mg/kg | 317.8 ± 7.4 | 321.0 ± 16.8 | 3.2 ± 15.8 | 1.0 ± 5.0 | 0.22 | |
| Total protein (g/dL) | Control | 4.9 ± 0.6 | 4.7 ± 0.8 | −0.3 ± 0.4 | −5.4 ± 8.7 | - |
| 5 mg/kg | 5.1 ± 0.8 | 5.4 ± 0.9 | 0.4 ± 0.3 | 7.9 ± 6.7 | 0.02 | |
| 10 mg/kg | 4.9 ± 0.8 | 5.6 ± 0.9 | 0.8 ± 0.8 | 16.5 ± 17.1 | 0.002 | |
| Uric acid (mg/dL) | Control | 4.5 ± 2.8 | 2.8 ± 2.4 | −1.6 ± 0.8 | −39.2 ± 24.0 | - |
| 5 mg/kg | 4.2 ± 3.3 | 4.1 ± 4.6 | 0.0 ± 2.3 | −6.3 ± 57.1 | 0.20 | |
| 10 mg/kg | 4.5 ± 2.0 | 4.1 ± 2.8 | −0.3 ± 2.3 | −0.2 ± 54.8 | 0.22 | |
| Sodium (mmol/L) | Control | 141.9 ± 4.4 | 141.7 ± 5.5 | −0.2 ± 2.7 | −0.2 ± 1.9 | - |
| 5 mg/kg | 141.6 ± 5.5 | 143.8 ± 6.0 | 2.2 ± 1.6 | 1.6 ± 1.2 | 0.51 | |
| 10 mg/kg | 144.6 ± 4.4 | 144.9 ± 8.5 | 0.3 ± 7.3 | 0.2 ± 5.0 | 0.81 | |
| Potassium (mmol/L) | Control | 4.4 ± 1.5 | 4.6 ± 1.2 | 0.2 ± 1.6 | 18.1 ± 53.3 | - |
| 5 mg/kg | 4.0 ± 1.1 | 3.7 ± 0.7 | −0.3 ± 1.3 | −4.0 ± 24.9 | 0.40 | |
| 10 mg/kg | 4.3 ± 0.9 | 3.8 ± 0.5 | −0.5 ± 1.0 | −8.8 ± 18.5 | 0.48 | |
| Calcium (mg/dL) | Control | 12.6 ± 1.8 | 12.2 ± 2.0 | −0.5 ± 0.6 | −3.8 ± 4.9 | - |
| 5 mg/kg | 12.4 ± 1.5 | 12.3 ± 1.2 | −0.1 ± 0.6 | −0.8 ± 4.7 | 0.38 | |
| 10 mg/kg | 12.3 ± 1.6 | 12.5 ± 1.4 | 0.2 ± 0.9 | 2.5 ± 7.9 | 0.13 | |
| Phosphorus (mg/dL) | Control | 7.0 ± 0.9 | 7.7 ± 0.8 | 0.7 ± 1.1 | 12.1 ± 17.2 | - |
| 5 mg/kg | 7.0 ± 0.8 | 8.3 ± 1.2 | 1.3 ± 1.3 | 19.2 ± 19.2 | 0.36 | |
| 10 mg/kg | 6.8 ± 1.0 | 8.3 ± 1.1 | 1.5 ± 1.4 | 24.3 ± 20.5 | 0.33 | |
| Glucose (mg/dL) | Control | 189.1 ± 31.3 | 178.8 ± 23.7 | −10.3 ± 25.3 | −4.6 ± 12.1 | - |
| 5 mg/kg | 194.8 ± 35.3 | 199.6 ± 21.3 | 4.8 ± 26 | 4.6 ± 15 | 0.22 | |
| 10 mg/kg | 192.6 ± 15.1 | 208.6 ± 23.7 | 16.0 ± 27.5 | 8.9 ± 14.6 | 0.08 | |
Negative values for differences represent decreases from baseline, and positive values represent increases.
P values represent comparisons between furosemide treatments and the control treatment; values < 0.05 were considered significant.
— = Not applicable.
Soaking in a warm water bath for 5 minutes, 12 hours after each treatment session ended, resulted in active drinking behavior by 1 lizard when no furosemide was administered, 8 lizards when furosemide was administered at 5 mg/kg, and 7 lizards when furosemide was administered at 10 mg/kg. Soaking and the observed active drinking behavior resulted in a significant increase in body weight following furosemide treatment at 5 mg/kg (2.9 ± 1.8%; P = 0.03) and 10 mg/kg (5.6 ± 2.5%; P < 0.001), compared with results for the control treatment (0.7 ± 0.7%). Weight gain was significantly (P = 0.03) greater for the 10 mg/kg treatment, compared with results for the 5 mg/kg treatment.
Discussion
Repeated SC administration of furosemide at 5 or 10 mg/kg to the bearded dragons deprived of a water source in the present study resulted in diuresis and dehydration. The unique anatomic characteristics of reptile kidneys, wherein the nephrons contain no loops of Henle nor a macula densa or structure with similar function, have called into question the usefulness of furosemide in reptiles.14 Nevertheless, furosemide has been shown to induce diuresis in turtles, lizards, and snakes.14–19
In a study14 involving freshwater turtles, administration of a single dose of furosemide (2 or 5 mg/kg, IV) resulted in marked reduction in renal sodium and chloride reabsorption as well as an increase in potassium excretion and urine production, with no change in plasma electrolyte concentrations, Hct, or plasma renin concentrations. In other studies,16,17 repeated furosemide administration to freshwater turtles (10 mg/kg, IM, q 24 h for 3 doses) as well as tortoises (10 mg/kg and 20 mg/kg, IP, q 24 h for 2 doses) resulted in a significant increase in plasma renin concentration and decrease in plasma sodium and potassium concentrations. The difference in plasma potassium concentration between furosemide-treated and control tortoises was −0.5 mmol/L.16 A decrease in plasma potassium concentration, compared with that for no furosemide, was observed with repeated SC administration of furosemide at both 5 and 10 mg/kg in the present study, but this difference was not significant. Without information regarding the mechanism of action furosemide has in bearded dragons, it is unclear whether a significant change might be expected with higher or additional doses.
In the bearded dragons in the study reported here, the changes in plasma total protein concentration and PCV associated with furosemide administration and dehydration were consistent with findings for other species. For example, water restriction of chickens and pigeons leads to dose-dependent increases in PCV and blood uric acid, total protein, and globulin concentrations.21,22 However, no significant increase in plasma uric acid concentration or plasma osmolarity was identified in the present study. Characterization of hydration status in reptiles through physical examination can be challenging, so it is unfortunate that no consistent plasma biochemical changes were identified that could be used for this purpose.
Plasma osmolarity was expected to be an indicator of the hydration status of the bearded dragons in the present study, but it was not. Most vertebrates maintain plasma osmolarity between 250 and 300 mOsm.23 However, in some reptile species, periods of hyperosmolarity are tolerated to deal with periods of restricted environmental water sources.23 Mean ± SD plasma osmolarity in inland bearded dragons was 295 ± 9.35 mOsm/kg in a previous study.24 In the bearded dragons of the present report, plasma osmolarity was substantially higher than this value (mean ± SD of all baseline values, 314.0 ± 9.1 mOsm/kg; range, 294 to 331 mOsm/kg). The reason for this difference in findings between the 2 studies is unclear. One possibility is that the bearded dragons in the present study were under greater osmotic stress prior to baseline sample collection, although we consider this unlikely given the amount of water offered to them on a regular basis and the regular feeding schedule that included fresh greens. Clinicians should be aware that despite a degree of dehydration that would be considered clinically important in mammals, the bearded dragons in our study had no increase in plasma osmolarity. A greater degree of dehydration may be necessary to induce an increase in plasma osmolarity in this species.
Despite no change in plasma osmolarity from baseline, the bearded dragons in the study reported here took in water to compensate for the furosemide-induced dehydration. Drinking behavior in response to dehydration has been observed in many species, including green iguanas.25,26 Findings suggested that furosemide administration had a dose-dependent effect on the amount of drinking when bearded dragons were soaked after a concurrent 48 hours of water and food withholding. On average, they took up nearly twice as much water (when expressed as a percentage of body weight) after furosemide treatment at 10 mg/kg, compared with after the 5 mg/kg treatment.
In green iguanas, a nondesert species, 48 hours of food and water withholding was sufficient to prompt consumption of a similar percentage of body weight via drinking (approx 5.4%).26 In contrast, following a 48-hour period of food and water withholding alone, inland bearded dragons, a desert-adapted species, undergoing the control treatment consumed < 1% of their body weight in water once given access. This finding suggested that inland bearded dragons were less prone to environmentally induced dehydration than other reptile species not adapted to arid environments.
Overall, the results of the study reported here indicated that furosemide administration at 5 and 10 mg/kg, SC, had diuretic effects in bearded dragons. Therefore, furosemide should be considered for treatment of congestive heart failure and other conditions requiring diuresis in this species. Additional research is needed to determine the safety of the long-term furosemide administration to this species as well as the doses of furosemide required to achieve favorable outcomes in ill bearded dragons requiring diuresis.
Acknowledgments
Supported by the University of Wisconsin School of Veterinary Medicine Companion Animal Fund, the Association of Reptile and Amphibian Veterinarians, and Abaxis Global Diagnostics Inc.
Footnotes
Simone-Freilicher E, Sullivan P, Quinn R, et al. Two cases of congestive heart failure in lizards, in Proceedings. 1st ExoticsCon 2015;505–509.
Orange Cube cricket diet, Hi-Calcium cricket diet, and Cricket Quencher with calcium, Fluker Farms, Port Allen, La.
Salix, Merck Animal Health, Kenilworth, NJ.
Pro-Vent, Smiths Medical Inc, Keene, NH.
VetScan Avian/Reptile Profile Plus, Abaxis Inc, Union City, Calif.
Advanced micro-osmometer model No. 3300, Advanced Instruments Inc, Norwood, Mass.
SigmaPlot, version 13, Systat Software, San Jose, Calif.
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