Therapeutic Effects of Tolvaptan, a Potent, Selective Nonpeptide Vasopressin V2 Receptor Antagonist, in Rats with Acute and Chronic Severe H
http://www.100md.com
内分泌学杂志 2005年第7期
Research Institute of Pharmacological and Therapeutical Development, Otsuka Pharmaceutical Co., Ltd., Tokushima 771-0192, Japan
Address all correspondence and requests for reprints to: Toshiki Miyazaki, Research Institute of Pharmacological and Therapeutical Development, Otsuka Pharmaceutical Co., Ltd., 463-10 Kagasuno Kawauchi-cho Tokushima 771-0192, Japan. E-mail: t_miyazaki@research.otsuka.co.jp.
Abstract
The therapeutic efficacy of tolvaptan (OPC-41061), a potent, selective nonpeptide vasopressin V2 receptor antagonist, on acute and chronic severe hyponatremia was assessed in rats. Experiments were designed to demonstrate the efficacy of tolvaptan reducing mortality in an acute model, and controlling the extent of serum sodium elevation without causing abnormal animal behavior suggesting neurological symptoms in a chronic model. In the acute model, rats developed rapidly progressive, severe hyponatremia by continuous sc infusion of [deamino-Cys1, D-Arg8]-vasopressin (10 ng/h) and forced water-loading (additional 10% initial body weight per day). By d 6, untreated rats had a 47% mortality rate. However, rats treated with repeated oral administrations of tolvaptan (1, 3, and 10 mg/kg) produced dose-dependent aquaresis (i.e. urine volume increased and urine osmolality decreased) that resulted in a gradual increase in plasma sodium concentration. Consequently, tolvaptan treatment reduced mortality and, at higher doses, resulted in no observed deaths. In the gradual model, rats receiving a continuous sc infusion of [deamino-Cys1, D-Arg8]-vasopressin (1 ng/h) combined with a liquid diet were induced to stable, severe hyponatremia (110 mEq/liter), which lead to increased organ weight and water content. Rats receiving dose titrations of tolvaptan (0.25, 0.5, 1, 2, 4, and 8 mg/kg) increased plasma sodium to healthy levels without causing abnormal animal behavior suggesting neurological symptoms or death, improved hyponatremia-driven increases in wet weight and water content in the organs. Thus, in animal models, analogous to the hyponatremia forms seen in humans, tolvaptan presents exciting therapeutic implications in the management of patients with severe hyponatremia.
Introduction
HYPONATREMIA, THE MOST common disorder of body fluid and electrolyte balance, is associated with a variety of diseases in which water retention, edema, and fluid shifts produce clinically relevant symptoms (1). These incidences range from 15–22% for serum sodium less than 135 mEq/liter and around 1–5% for serum sodium less than 130 mEq/liter in hospitalized patients (2, 3).
Previous studies suggest that severe hyponatremia itself produces brain edema, which can lead to neurological symptoms or even death if produced acutely (4, 5, 6). In this situation, the physiological responses to a water load, ions, and organic osmoles shifting across the cell membrane, are relatively slow and cannot keep up with harmful tissue swelling. Therefore, acutely hyponatremic patients need to be treated rapidly to decrease the hyponatremic levels to a mild level so symptomatic recovery can be realized (7).
On the other hand, in chronic severe hyponatremia, which is observed most frequently in the syndrome of inappropriate antidiuretic hormone secretion, a rapid correction of serum sodium after the completion of physiological compensation may promote disturbances in the blood-brain barrier and produce central nervous system injuries such as central pontine myelinolysis (5, 8, 9, 10, 11, 12). Therefore, when treating both acute and chronic severe hyponatremia, it is critical to control the rate and extent of the plasma sodium rise.
In many cases of hyponatremia, plasma arginine vasopressin (AVP), which plays a critical role in body fluid regulation, is increased inappropriately. The increased AVP accelerates renal free water reabsorption via an AVP V2 receptor pathway, causing water retention in the body. According to the condition described above, animal models of hyponatremia induced by the combination of water loading and AVP V2 receptor agonist administration are established (13).
In recent years, clinical (14, 15) and experimental (16, 17, 18) studies suggest that AVP V2 receptor antagonists are effective for treating hyponatremia. However, it has not been established whether these antagonists reduce mortality associated with acute severe hyponatremia and how they should be used to manage the magnitude of increase in serum sodium concentration in chronic severe hyponatremia.
Tolvaptan [(±)-7-chloro-5-hydroxy-1-[2-methyl-4-(2-methylbenzoylamino) benzoyl]-2,3,4,5-tetrahydro-1H-benzazepine], is characterized as a highly potent and selective nonpeptide vasopressin V2 receptor antagonist. Tolvaptan antagonized [3H]AVP binding to human and rat V2 receptors in a competitive fashion and showed aquaresis in rats by oral administration (19).
In the present study, we performed two experiments to evaluate the following: 1) Can tolvaptan reduce mortality in acute severe hyponatremia by elevating serum sodium concentrations rapidly via its aquaretic action? 2) Can a paced dose titration of tolvaptan control the extent of increasing serum sodium elevation and gradually eliminate water retention in organs without neurological symptoms in chronic severe hyponatremia?
Materials and Methods
Animals
The care and handling of animals were in accordance with the Guidelines for Animal Care and Use (Otsuka Pharmaceutical Co., Ltd., Tokushima, Japan; October 1, 1994). In all experiments, male Sprague Dawley rats were purchased from Charles River Japan, Inc. (Yokohama, Japan). They were kept in the following conditions: humidity, 60 ± 10%; temperature, 23 ± 2 C; 12-h light and 12-h dark cycle. They had free access to food and tap water before the experiments.
Drugs
[Deamino-Cys1, D-Arg8]-vasopressin (DDAVP) was purchased from Sigma-Aldrich Japan Co., Ltd. (Tokyo, Japan). Hydroxypropylmethylcellulose (HPMC) was purchased from Shin-Etsu Chemical Co., Ltd. (Tokyo, Japan). Tolvaptan [(±)-7-chloro-5-hydroxy-1-[2-methyl-4-(2-methylbenzoylamino) benzoyl]-2,3,4,5-tetrahydro-1H-benzazepine] was prepared at the Formulation Research Institute of Otsuka Pharmaceutical Co., Ltd., and was suspended in 1% HPMC solution.
Experimental schedule of acute hyponatremia
Acute severe hyponatremia was induced by a continuous sc infusion of 10 ng/h of DDAVP per body using osmotic minipumps (infusion rate; 0.5 μl/h, ALZET model 2002; ALZA Corp., Mountain View, CA) and intragastric water administration that was 5% of initial body weight twice a day.
As shown in Fig. 1A, male rats (9 wk) weighing 260–320 g were sc implanted with an osmotic minipump containing DDAVP (n = 133) or saline (n = 10). Rats were housed individually in metabolic cages to collect urine, where they had free access to food (regular rat chow) and water. Blood (0.3 ml) was collected from the tail vein using a capillary tube to measure plasma sodium concentrations. On d 3, the rats, whose plasma sodium concentrations were in the range of 90–120 mEq/liter (n = 68), were divided into four groups of vehicle (1% HPMC) and tolvaptan 1-, 3-, and 10-mg/kg treatments by matching their individual plasma sodium concentrations. Tolvaptan or the vehicle was administrated by gavage from d 4–6. Gross behavioral changes of animals were observed and recorded to evaluate neurological symptoms. At the end of d 6, mortality was evaluated, and surviving animals were killed to measure brain sodium and water content.
FIG. 1. Experimental schedules of acute and chronic hyponatremia models. A, Acute model; B, chronic model.
Experimental schedule of chronic hyponatremia
As shown in Fig. 1B, at 8 wk of age, 21 rats weighing 250–270 g were divided into two groups and an osmotic minipump containing DDAVP (1 ng/h per body sc, n = 14) or saline (n = 7) was implanted. From d –3 to d 1, they received 70 ml/d of standard liquid food (Oriental Yeast Co., Ltd., Osaka, Japan). On all subsequent days, they received 40 ml/d of modified liquid food, which was prepared by exchanging water with a 14% dextrose solution (13). On d 4, the rats in the DDAVP-treated group were further divided into the vehicle (n = 7) and tolvaptan (n = 7) groups by matching their individual body weight and plasma sodium concentrations measured on d 3. Blood and urine were collected every day. Tolvaptan or the vehicle (1% HPMC) was administrated by gavage from d 4. Gross behavioral changes of animals were observed and recorded to evaluate neurological symptoms. On d 14, animals were killed, and the brain, heart, and kidney were isolated to measure organ weight and water content.
General procedures
Urinary sodium, potassium, and chloride and plasma sodium concentrations were measured using ion-specific electrodes (CX-3; Beckman Instruments, Fullerton, CA). Osmolality was measured by freezing-point depression (Model 3400; Fiske Associates, Needham Heights, MA). To measure water content, organs were dried for 48 h at 110 C on heating blocks. Brain sodium was extracted by 0.75 M nitric acid, and its concentration was analyzed by flame photometry (Hitachi 750, Tokyo, Japan).
Statistical analysis
Values were expressed as the means ± SEM. Differences were considered statistically significant at P < 0.05.
Acute hyponatremia
Differences between the DDAVP-infused groups were statistically tested using repeated measures ANOVA, followed by two-tailed Dunnett’s multiple comparison tests between the vehicle- and tolvaptan-treated groups. When comparing the saline-infused and vehicle-treated groups, two-tailed t tests were performed. A two-tailed Fisher’s exact test was used with Bonferroni’s correction to compare mortality between the groups.
Chronic hyponatremia
Differences between the DDAVP-infused vehicle and tolvaptan treatment groups were statistically analyzed using repeated measures ANOVA, followed by two-tailed t test.
Differences between the saline-infused and DDAVP-infused vehicle treatment group were statistically analyzed by a two-tailed t test.
Results
Effects of tolvaptan on acute severe hyponatremia
Mortality and plasma sodium.
DDAVP infusion and excess water loading markedly decreased plasma sodium levels in rats, and severe progressive hyponatremia developed (105.2 ± 1.0 mEq/liter, on d 3). Some of the animals consecutively decreased plasma sodium on subsequent days, then reached plasma sodium less than 90 mEq/liter and died. As shown in Table 1, of the 17 rats treated with the vehicle, seven rats died on d 5 and one died on d 6; total mortality was 47%. When tolvaptan (1, 3, and 10 mg/kg) was orally administrated on d 4, a dramatic improvement in mortality was seen. In the 1-mg/kg tolvaptan-treated group, two rats died on d 5 and four died on d 6 (total mortality = 38%), showing slightly prolonged survival. However, in the other tolvaptan-treated groups (3 and 10 mg/kg), plasma sodium levels gradually elevated after repeated tolvaptan administration (Fig. 2), and all rats survived d 6. In addition, statistically significant differences in mortality were seen between the vehicle- and tolvaptan-treated groups.
TABLE 1. Effects of tolvaptan on mortality in acute hyponatremia
FIG. 2. Time courses of plasma sodium concentrations in the acute model. Values are expressed as the mean ± SEM. Values are taken from the end of each experimental day. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed Dunnett’s multiple comparison test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. x, Saline-infused; , vehicle; , tolvaptan 1 mg/kg; , tolvaptan 3 mg/kg; and , tolvaptan 10 mg/kg.
Four hours after the first oral administration of tolvaptan (1, 3, and 10 mg/kg), mean plasma sodium concentrations increased by 0.5, 2.0, and 12.9 mEq/liter, respectively. For rats in the group given the highest dose of tolvaptan (10 mg/kg), the maximum change in individual plasma sodium concentration after 4 h was 21.6 mEq/liter, indicating that the concentration was rapidly corrected but remained within mild hyponatremic levels. Moreover, the maximum daily change was 24.6 mEq/liter and no abnormal animal behavior was observed, suggesting the correction rate was adequate. Furthermore, the plasma sodium concentration increased after repeated tolvaptan administrations (119.3 ± 2.3, 120.6 ± 2.7, 128.5 ± 1.4 mEq/liter on d 6), resulting in statistically significant differences (Fig. 2).
Urine parameters.
At 3 and 10 mg/kg, significant changes in urine volume and urine osmolality were seen 4 h after drug administration: Urine volume increased and osmolality decreased. The changes were also statistically significant at 24 h, except for urine volume in the 3-mg/kg tolvaptan group on d 6. Even at the lowest dose (1 mg/kg), diluted urine was excreted, and statistically significant differences were seen at some points (Fig. 3).
FIG. 3. Effects of tolvaptan on initial (0–4 h) and total daily urine volume and osmolality in the acute model. Values are expressed as the mean ± SEM. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed Dunnett’s multiple comparison test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. , Vehicle; , tolvaptan 1 mg/kg; , tolvaptan 3 mg/kg; and , tolvaptan 10 mg/kg.
Brain sodium and water content.
In the vehicle-treated group, significant changes in brain sodium and water content were observed after water retention: sodium content decreased and water content increased (Fig. 4). After 3 d of repeated tolvaptan dosing, the decreased brain sodium content increased even at the lower doses, and a statistically significant increment was observed in the highest dosed (10 mg/kg) group. However, despite significant changes in the vehicle-treated group, we observed no effects to brain water content.
FIG. 4. Effects of tolvaptan on brain sodium and water content in the acute model. Values are expressed as the mean ± SEM. Comparing the saline-infused and vehicle-treated groups for sodium content in the brain, a two-tailed t test was performed. Then, a two-tailed Dunnett’s multiple comparison test was performed to compare brain sodium contents. $$, P < 0.01 vs. saline-infused. *, P < 0.05 vs. vehicle.
Effects of tolvaptan on chronic hyponatremia
Plasma sodium control in chronic hyponatremia.
Figure 5 shows plasma sodium changes in chronic hyponatremia. Subcutaneous DDAVP infusion and liquid food consumption decreased plasma sodium concentrations by d 2 to about 110 mEq/liter, which was maintained thereafter, and severe, chronic hyponatremia was induced. Beginning on d 4, dose titrations of tolvaptan were administered. Plasma sodium concentrations gradually and significantly elevated with increasing tolvaptan doses (0.25, 0.5, 1, 2, 4, and 8 mg/kg). Mean elevated plasma sodium concentrations were as follows: 7.4 mEq/liter (0.25 mg/kg, d 4–5), 6.3 mEq/liter (0.5 mg/kg, d 5–6), 5.5 mEq/liter (1 mg/kg, d 6–7), 6.9 mEq/liter (2 mg/kg, d 7–8), and 4.4 mEq/liter (4 mg/kg, d 8–9). After d 9 in the tolvaptan-treated group, plasma sodium concentrations were within healthy levels. Therefore, elevated serum sodium levels were not observed when the tolvaptan dose was increased to 8 mg/kg. Finally, the plasma sodium concentration in the tolvaptan-treated group increased to 142.8 ± 0.4 mEq/liter (d 14). It contrasts with that of the vehicle-treated group, which was still low (110.6 ± 0.9 mEq/liter). Individual plasma sodium concentrations in this group had a maximum daily change of only 11.8 mEq/liter and no abnormal animal behavior (hyperactivity and hyperirritability, jumping, spasticity of the extremities, paralysis of the limbs, occasional seizure activity, and unresponsiveness), as previously described (20), was seen. These results suggest that the rate of correction was properly controlled.
FIG. 5. Effects of tolvaptan on plasma sodium levels in the chronic model. Values are expressed as the mean ± SEM. Seven animals comprise each group. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed t test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. x, Saline-infused; , vehicle; and , tolvaptan.
Aquaretic action of tolvaptan on chronic hyponatremia.
Oral administration of tolvaptan produced aquaresis. In the tolvaptan-treated group, the 0- to 4-h urine volume was 6–37 times greater, and urine osmolality was significantly lower than in the vehicle-treated group. Despite similar water intake for both groups, in the tolvaptan-treated group, the urine volume measured from 0–24 h was 1.1–1.5 times greater than that in the vehicle-treated group (except d 5 and 10), and significant differences were observed on d 6, 7, 8, and 13. At 0–24 h, urine osmolality was significantly lower in the tolvaptan-treated group than in the vehicle-treated group throughout the dosing period (Fig. 6). Figure 7 shows urine sodium excretion 0–24 h after oral administration of tolvaptan or the vehicle. In the tolvaptan-treated group, urine sodium excretion was 1.1–2.0 times less than in the vehicle-treated group (except on d 6), and the differences between the two groups were statistically significant on d 5, 7, 10, 11, and 13.
FIG. 6. Effects of tolvaptan on initial (0–4 h) and total daily urine volume and osmolality in the chronic model. Values are expressed as the mean ± SEM. Seven animals comprise each group. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed t test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. , Vehicle; and , tolvaptan.
FIG. 7. Effects of tolvaptan on urine sodium excretion in the chronic model. Values are expressed as the mean ± SEM. Seven animals comprise each group. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed t test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. x, Saline-infused; , vehicle; and , tolvaptan.
Effects of tolvaptan on organ changes in chronic hyponatremia.
Organ weight and water content are shown in Fig. 8. Significant increases in wet weight of the heart and left kidney were observed in the vehicle-treated group, as compared with the saline-infused group. After dose titration of tolvaptan, the increment increases in wet weight of the heart and kidney were blunted, although the difference was not statistically significant in the heart. Brain, heart, and kidney water content also significantly increased in the vehicle-treated group as opposed to the saline-infused group. Tolvaptan completely reversed the vital water content increases of brain and heart seen in the vehicle-treated animals, with statistical significance. In the kidney, tolvaptan inhibited the water content increase by 43%, although statistical significance was not observed. No significant difference in body weight on d 14 was observed between the vehicle- (265.9 ± 4.1 g) and tolvaptan-treated (266.4 ± 2.6 g) groups. These data suggest that tolvaptan treatment inhibited organ water retention.
FIG. 8. Effects of tolvaptan on organ wet weight and water content in the chronic model. Values are expressed as the mean ± SEM. Seven animals comprise each group. At first, the differences between the saline-infused and vehicle groups were analyzed by a two-tailed t test, then a two-tailed t test was performed. $, P < 0.05; $$, P < 0.01 vs. saline-infused. **, P < 0.01 vs. vehicle.
Discussion
Brain adaptation to hypoosmolality is well known; when plasma sodium decreases, brain cells rapidly export electrolytes and organic osmolytes to prevent excess swelling (21). However, these adaptive mechanisms do not prevent the development of brain edema when serum sodium concentrations fall rapidly, so severe neurological symptoms or death may occur (22).
Therefore, acute severe hyponatremia needs to be rapidly corrected to less critical hyponatremia, then progressively corrected to normonatremic level. It has been reported that with rapid and controlled correction (estimated threshold of brain injury, <25 mEq/liter·d), neurological events and mortality may be avoided in acute severe hyponatremia (7). In chronic severe hyponatremia, however, the brain generally adapts itself to reduced concentrations of electrolytes and organic osmolytes. These adapted brain cells are unable to rapidly equilibrate themselves with extracellular osmolyte levels when hyponatremia is rapidly corrected (23). This situation leads to the contraction of cells, disturbance of the blood-brain barrier, and potentially irreversible neurological damage, which is manifested in extreme cases by paralysis, coma, or death.
In many cases of hyponatremia, plasma AVP levels are increased. When circulating AVP binds to V2 receptors existing in renal collecting ducts, renal free water reabsorption is stimulated via increasing intracellular cAMP and translocating aquaporin-2 from intracellular vesicles to the apical plasma (24). Tolvaptan is a specific AVP V2 receptor antagonist that blocks the binding of AVP at V2 receptors, resulting in aquaresis without depletion of electrolytes, and possibly improves diluted hyponatremia (19, 25).
We found that tolvaptan improved mortality in fluid-overloaded acute severe hyponatremia. In the vehicle-treated group, the rats failed to adapt themselves to hypoosmolality, which resulted in 47% mortality due to severe hyponatremia. In fact, a significant decrease in brain sodium and an increase of brain water content were observed during 6 d of the experiment. However, tolvaptan treatment reduced total mortality even at the lowest dose given, presumably because plasma sodium concentrations improved above some critical threshold. In the higher dose groups, death was not observed because plasma sodium concentrations rapidly elevated to mild hyponatremia levels. Furthermore, significant improvement in brain sodium content was observed although plasma sodium level was still low.
In the chronic hyponatremia study, the rate of increase of plasma sodium concentration was carefully controlled by using paced dose titration of tolvaptan. With this method, the severe state improved gradually to normonatremia without abnormal animal behavior suggesting neurological symptoms or death. In the chronic model, tolvaptan was effective at a lower dose (0.25 mg/kg) than in the acute model of hyponatremia (1 mg/kg). One of the reasons is the doses of DDAVP infused, 10 ng/h in the acute model and 1 ng/h in the chronic model.
After the normalization of plasma sodium, the dosing was extended for several days to confirm whether plasma sodium increased to hypernatremia or reverted back to hyponatremia. As a result, plasma sodium did not change any more and remained within the normal range during the extended dosing period. Similar results were seen in clinical studies (26, 27). In patients with hyponatremia secondary to congestive heart failure, plasma sodium concentrations increased to the normal range after administration of tolvaptan and remained within a normal range during treatment with tolvaptan. Interestingly, a different response of plasma sodium was observed in patients with normonatremia. After treatment, normonatremic patients had a transient increase in sodium levels, with the values returning to baseline within 3 wk of therapy. On the other hand, it has been reported that normalized plasma sodium reverted back to the state of hyponatremia, once the dosing ceased after normalization in similar animal models (16). These suggest that dose titration is a successful strategy for controlling the rate of increase of plasma sodium concentration, and that plasma sodium remains within a normal range while the dosing is extended.
It has been reported that tolvaptan produces aquaresis after single and multiple dose regimens (19) through elevated electrolyte-free water clearance in conscious rats (25). Through repeated tolvaptan administration, we were able to maintain aquaresis. Our results suggest that tolvaptan also increased free-water excretion, resulting in gradual increases in plasma sodium concentrations.
During tolvaptan treatment, 24-h urinary sodium excretion also decreased in both models, which might have partially contributed to the improvement of plasma sodium concentrations. Although the mechanisms are not clear, one possible explanation is the recovery from the situation of "solute depletion" by tolvaptan. It has been reported that the volume expansion state in syndrome of inappropriate antidiuretic hormone secretion is associated with decreased proximal tubular sodium reabsorption (28), so correction of volume expansion after tolvaptan treatment may contribute to the improvement of renal sodium reabsorption.
In the chronic model, the wet weight of the heart and kidney, and water content in the brain, heart, and kidney also significantly increased. In the vehicle-treated group, incremental increases in the heart and kidney weight almost entirely resulted from increased water content because the mean dry weight of these organs was the same as that of the saline-infused group (data not shown). After tolvaptan treatment, those changes significantly reduced organ water content, suggesting that tolvaptan improved water retention in these organs.
In conclusion, tolvaptan treatment showed aquaresis, and controlled the magnitude of plasma sodium elevation both in acute and chronic severe hyponatremia. Furthermore, tolvaptan improved mortality in acute severe hyponatremia and organ water retention in chronic severe hyponatremia. Thus, tolvaptan presents exciting therapeutic implications in the management of clinical hyponatremia.
Acknowledgments
We express sincere thanks to Dr. F. Czerwiec for his critical reading of this manuscript. We also thank Dr. J. Garfield and Mr. K. Komuro for their excellent assistance in the preparation of this manuscript.
References
Anderson RJ, Chung H, Kluge R, Schrier RW 1985 Hyponatremia: a prospective analysis of its epidemiology and the pathogenetic role of vasopressin. Ann Intern Med 102:164–168
Flear CTG, Gill GV, Burn J 1981 Hyponatremia: mechanisms and management. Lancet 2:26–81
Kleinfeld M, Casimir M, Borra S 1979 Hyponatremia as observed in a chronic disease facility. J Am Geriatrics Soc 27:156–161
Arieff AI 1986 Hyponatremia, convulsions, respiratory arrest, and permanent brain damage after elective surgery in healthy women. N Engl J Med 314:1529–1534
Ayus JC, Krothapalli RK, Armstrong DL, Norton HJ 1989 Symptomatic hyponatremia in rats: effect of treatment on mortality and brain lesions. Am J Physiol 257:F18–F22
Ayus JC, Arieff AI 1999 Chronic hyponatremic encephalopathy in postmenopausal women. Association of therapies with mobidity and mortality. JAMA 281:2299–2304
Ayus JC, Krothapalli RK, Arieff AI 1985 Changing concepts in treatment of severe symptomatic hyponatremia. Am J Med 78:897–901
Kleinschmitt-Demasters BK, Norenberg MD 1981 Rapid correction of hyponatremia causes demyelination: relation to central pontine myelinolysis. Science 211:1068–1070
Ayus JC, Krothapalli RK, Armstrong DL 1985 Rapid correction of severe hyponatremia in the rat: histopathological change in the brain. Am J Physiol 248:F711–F719
Verbalis JG, Martinez AJ, Drutarosky MD 1991 Neurological and neuropathological sequelae of correction of chronic hyponatremia. Kidney Int 39:1274–1282
Soupart A, Stenuit A, Perier O, Decaux G 1991 Limits of brain tolerance to daily increments in serum sodium in chronically hyponatremic rats treated with hypertonic saline or urea: advantages of urea. Clin Sci 80:77–84
Soupart A, Penninckx R, Stenuit A, Perier O, Decaux G 1992 Treatment of chronic hyponatremia in rats by intravenous saline: comparison of rate versus magnitude of correction. Kidney Int 41:1662–1667
Verbalis JG, Drutarosky MD 1988 Adaptation to chronic hypoosmolality in rats. Kidney Int 34:351–360
Saito T, Ishikawa S, Abe K, Kamoi K, Yamada K, Shimizu K, Saruta T, Yoshida S 1997 Acute aquaresis by the nonpeptide arginine vasopressin (AVP) antagonist OPC-31260 improves hyponatremia in patient with syndrome of inappropriate secretion of antidiuretic hormone (SIADH). J Clin Endocrinol Metab 82:1054–1057
Decaux G 2001 Long-term treatment of patient with inappropriate secretion of antidiuretic hormone by the vasopressin receptor antagonist conivaptan, urea, or furosemide. Am J Med 110:582–584
Fujisawa G, Ishikawa S, Tsuboi Y, Okada K, Saito T 1993 Therapeutic efficacy of non-peptide ADH antagonist OPC-31260 in SIADH rats. Kidney Int 44:19–23
Naito A, Hasegawa H, Kurasawa T, Ohtake Y, Matsukawa H, Ezure Y, Tsurita Y, Koike K, Shigenobu K 2000 The therapeutic efficacy of VP-343, a selective vasopressin V2 receptor antagonist, in the experimental SIADH rat model. Biol Pharm Bull 23:1323–1327
Naito A, Hasegawa H, Kurasawa T, Ohtake Y, Matsukawa H, Ezure Y, Koike K, Shigenobu K 2001 Histopathological study of kidney abnormalities in an experimental SIADH rat model and its application to the evaluation of the pharmacologic profile of VP-343, a selective vasopressin V2 receptor antagonist. Biol Pharm Bull 24:897–901
Yamamura Y, Nakamura S, Itoh S, Hirano T, Onogawa T, Yamashita T, Yamada Y, Tsujimae K, Aoyama M, Kotosai K, Ogawa H, Yamashita H, Kondo K, Tominaga M, Tsujimoto G, Mori T 1998 OPC-41061, a highly potent human vasopressin V2-receptor antagonist: pharmacological profile and aquaretic effect by single and multiple oral dosing in rats. J Pharmacol Exp Ther 287:860–867
Soupart A, Penninckx R, Crenier L, Stenuit A, Perier O, Decaux G 1994 Prevention of brain demyelination in rats after excessive correction of chronic hyponatremia by serum sodium lowering. Kidney Int 45:193–200
Verbalis JG, Gullans SR 1991 Hyponatremia causes large sustained reductions in brain content of multiple organic osmolytes in rats. Brain Res 567:274–282
Soupart A, Decaux G 1996 Therapeutic recommendations for management of severe hyponatremia: current concepts on pathogenesis and prevention of neurologic complications. Clin Nephrol 46:149–169
Verbalis JG, Gullans SR 1993 Rapid correction of hyponatremia produces differential effects on brain osmolyte reaccumulation in rats. Brain Res 606:19–27
Verbalis JG 2002 Vasopressin V2 receptor antagonists. J Mol Endocrinol 29:1–9
Hirano T, Yamamura Y, Nakamura S, Toshiyuki O, Mori T 2000 Effects of the V2-receptor antagonist OPC-41061 and the loop diuretic furosemide alone and in combination in rats. J Pharmacol Exp Ther 292:288–294
Gheorghiade M, Niazi I, Ouyang J, Czerwiec F, Kambayashi J, Zampino M, Orlandi C 2003 Vasopressin V2-receptor blockade with tolvaptan in patients with chronic heart failure: results from a double-blind, randomized trial. Circulation 107:2690–2696
Gheorghiade M, Gattis WA, O’Connor CM, Adams Jr KF, Elkayam U, Barbagelata A, Ghali JK, Benza RL, McGrew FA, Klapholz M, Ouyang J, Orlandi C 2004 Effects of tolvaptan, a vasopressin antagonist, in patients hospitalized with worsening heart failure: a randomized controlled trial. JAMA 291:1963–1971
Decaux G 2001 Difference in solute excretion during correction of hyponatremic patients with cirrhosis or syndrome of inappropriate secretion of antidiuretic hormone by oral vasopressin V2 receptor antagonist VPA-985. J Lab Clin Med 138:18–21(Toshiki Miyazaki, Yoshita)
Address all correspondence and requests for reprints to: Toshiki Miyazaki, Research Institute of Pharmacological and Therapeutical Development, Otsuka Pharmaceutical Co., Ltd., 463-10 Kagasuno Kawauchi-cho Tokushima 771-0192, Japan. E-mail: t_miyazaki@research.otsuka.co.jp.
Abstract
The therapeutic efficacy of tolvaptan (OPC-41061), a potent, selective nonpeptide vasopressin V2 receptor antagonist, on acute and chronic severe hyponatremia was assessed in rats. Experiments were designed to demonstrate the efficacy of tolvaptan reducing mortality in an acute model, and controlling the extent of serum sodium elevation without causing abnormal animal behavior suggesting neurological symptoms in a chronic model. In the acute model, rats developed rapidly progressive, severe hyponatremia by continuous sc infusion of [deamino-Cys1, D-Arg8]-vasopressin (10 ng/h) and forced water-loading (additional 10% initial body weight per day). By d 6, untreated rats had a 47% mortality rate. However, rats treated with repeated oral administrations of tolvaptan (1, 3, and 10 mg/kg) produced dose-dependent aquaresis (i.e. urine volume increased and urine osmolality decreased) that resulted in a gradual increase in plasma sodium concentration. Consequently, tolvaptan treatment reduced mortality and, at higher doses, resulted in no observed deaths. In the gradual model, rats receiving a continuous sc infusion of [deamino-Cys1, D-Arg8]-vasopressin (1 ng/h) combined with a liquid diet were induced to stable, severe hyponatremia (110 mEq/liter), which lead to increased organ weight and water content. Rats receiving dose titrations of tolvaptan (0.25, 0.5, 1, 2, 4, and 8 mg/kg) increased plasma sodium to healthy levels without causing abnormal animal behavior suggesting neurological symptoms or death, improved hyponatremia-driven increases in wet weight and water content in the organs. Thus, in animal models, analogous to the hyponatremia forms seen in humans, tolvaptan presents exciting therapeutic implications in the management of patients with severe hyponatremia.
Introduction
HYPONATREMIA, THE MOST common disorder of body fluid and electrolyte balance, is associated with a variety of diseases in which water retention, edema, and fluid shifts produce clinically relevant symptoms (1). These incidences range from 15–22% for serum sodium less than 135 mEq/liter and around 1–5% for serum sodium less than 130 mEq/liter in hospitalized patients (2, 3).
Previous studies suggest that severe hyponatremia itself produces brain edema, which can lead to neurological symptoms or even death if produced acutely (4, 5, 6). In this situation, the physiological responses to a water load, ions, and organic osmoles shifting across the cell membrane, are relatively slow and cannot keep up with harmful tissue swelling. Therefore, acutely hyponatremic patients need to be treated rapidly to decrease the hyponatremic levels to a mild level so symptomatic recovery can be realized (7).
On the other hand, in chronic severe hyponatremia, which is observed most frequently in the syndrome of inappropriate antidiuretic hormone secretion, a rapid correction of serum sodium after the completion of physiological compensation may promote disturbances in the blood-brain barrier and produce central nervous system injuries such as central pontine myelinolysis (5, 8, 9, 10, 11, 12). Therefore, when treating both acute and chronic severe hyponatremia, it is critical to control the rate and extent of the plasma sodium rise.
In many cases of hyponatremia, plasma arginine vasopressin (AVP), which plays a critical role in body fluid regulation, is increased inappropriately. The increased AVP accelerates renal free water reabsorption via an AVP V2 receptor pathway, causing water retention in the body. According to the condition described above, animal models of hyponatremia induced by the combination of water loading and AVP V2 receptor agonist administration are established (13).
In recent years, clinical (14, 15) and experimental (16, 17, 18) studies suggest that AVP V2 receptor antagonists are effective for treating hyponatremia. However, it has not been established whether these antagonists reduce mortality associated with acute severe hyponatremia and how they should be used to manage the magnitude of increase in serum sodium concentration in chronic severe hyponatremia.
Tolvaptan [(±)-7-chloro-5-hydroxy-1-[2-methyl-4-(2-methylbenzoylamino) benzoyl]-2,3,4,5-tetrahydro-1H-benzazepine], is characterized as a highly potent and selective nonpeptide vasopressin V2 receptor antagonist. Tolvaptan antagonized [3H]AVP binding to human and rat V2 receptors in a competitive fashion and showed aquaresis in rats by oral administration (19).
In the present study, we performed two experiments to evaluate the following: 1) Can tolvaptan reduce mortality in acute severe hyponatremia by elevating serum sodium concentrations rapidly via its aquaretic action? 2) Can a paced dose titration of tolvaptan control the extent of increasing serum sodium elevation and gradually eliminate water retention in organs without neurological symptoms in chronic severe hyponatremia?
Materials and Methods
Animals
The care and handling of animals were in accordance with the Guidelines for Animal Care and Use (Otsuka Pharmaceutical Co., Ltd., Tokushima, Japan; October 1, 1994). In all experiments, male Sprague Dawley rats were purchased from Charles River Japan, Inc. (Yokohama, Japan). They were kept in the following conditions: humidity, 60 ± 10%; temperature, 23 ± 2 C; 12-h light and 12-h dark cycle. They had free access to food and tap water before the experiments.
Drugs
[Deamino-Cys1, D-Arg8]-vasopressin (DDAVP) was purchased from Sigma-Aldrich Japan Co., Ltd. (Tokyo, Japan). Hydroxypropylmethylcellulose (HPMC) was purchased from Shin-Etsu Chemical Co., Ltd. (Tokyo, Japan). Tolvaptan [(±)-7-chloro-5-hydroxy-1-[2-methyl-4-(2-methylbenzoylamino) benzoyl]-2,3,4,5-tetrahydro-1H-benzazepine] was prepared at the Formulation Research Institute of Otsuka Pharmaceutical Co., Ltd., and was suspended in 1% HPMC solution.
Experimental schedule of acute hyponatremia
Acute severe hyponatremia was induced by a continuous sc infusion of 10 ng/h of DDAVP per body using osmotic minipumps (infusion rate; 0.5 μl/h, ALZET model 2002; ALZA Corp., Mountain View, CA) and intragastric water administration that was 5% of initial body weight twice a day.
As shown in Fig. 1A, male rats (9 wk) weighing 260–320 g were sc implanted with an osmotic minipump containing DDAVP (n = 133) or saline (n = 10). Rats were housed individually in metabolic cages to collect urine, where they had free access to food (regular rat chow) and water. Blood (0.3 ml) was collected from the tail vein using a capillary tube to measure plasma sodium concentrations. On d 3, the rats, whose plasma sodium concentrations were in the range of 90–120 mEq/liter (n = 68), were divided into four groups of vehicle (1% HPMC) and tolvaptan 1-, 3-, and 10-mg/kg treatments by matching their individual plasma sodium concentrations. Tolvaptan or the vehicle was administrated by gavage from d 4–6. Gross behavioral changes of animals were observed and recorded to evaluate neurological symptoms. At the end of d 6, mortality was evaluated, and surviving animals were killed to measure brain sodium and water content.
FIG. 1. Experimental schedules of acute and chronic hyponatremia models. A, Acute model; B, chronic model.
Experimental schedule of chronic hyponatremia
As shown in Fig. 1B, at 8 wk of age, 21 rats weighing 250–270 g were divided into two groups and an osmotic minipump containing DDAVP (1 ng/h per body sc, n = 14) or saline (n = 7) was implanted. From d –3 to d 1, they received 70 ml/d of standard liquid food (Oriental Yeast Co., Ltd., Osaka, Japan). On all subsequent days, they received 40 ml/d of modified liquid food, which was prepared by exchanging water with a 14% dextrose solution (13). On d 4, the rats in the DDAVP-treated group were further divided into the vehicle (n = 7) and tolvaptan (n = 7) groups by matching their individual body weight and plasma sodium concentrations measured on d 3. Blood and urine were collected every day. Tolvaptan or the vehicle (1% HPMC) was administrated by gavage from d 4. Gross behavioral changes of animals were observed and recorded to evaluate neurological symptoms. On d 14, animals were killed, and the brain, heart, and kidney were isolated to measure organ weight and water content.
General procedures
Urinary sodium, potassium, and chloride and plasma sodium concentrations were measured using ion-specific electrodes (CX-3; Beckman Instruments, Fullerton, CA). Osmolality was measured by freezing-point depression (Model 3400; Fiske Associates, Needham Heights, MA). To measure water content, organs were dried for 48 h at 110 C on heating blocks. Brain sodium was extracted by 0.75 M nitric acid, and its concentration was analyzed by flame photometry (Hitachi 750, Tokyo, Japan).
Statistical analysis
Values were expressed as the means ± SEM. Differences were considered statistically significant at P < 0.05.
Acute hyponatremia
Differences between the DDAVP-infused groups were statistically tested using repeated measures ANOVA, followed by two-tailed Dunnett’s multiple comparison tests between the vehicle- and tolvaptan-treated groups. When comparing the saline-infused and vehicle-treated groups, two-tailed t tests were performed. A two-tailed Fisher’s exact test was used with Bonferroni’s correction to compare mortality between the groups.
Chronic hyponatremia
Differences between the DDAVP-infused vehicle and tolvaptan treatment groups were statistically analyzed using repeated measures ANOVA, followed by two-tailed t test.
Differences between the saline-infused and DDAVP-infused vehicle treatment group were statistically analyzed by a two-tailed t test.
Results
Effects of tolvaptan on acute severe hyponatremia
Mortality and plasma sodium.
DDAVP infusion and excess water loading markedly decreased plasma sodium levels in rats, and severe progressive hyponatremia developed (105.2 ± 1.0 mEq/liter, on d 3). Some of the animals consecutively decreased plasma sodium on subsequent days, then reached plasma sodium less than 90 mEq/liter and died. As shown in Table 1, of the 17 rats treated with the vehicle, seven rats died on d 5 and one died on d 6; total mortality was 47%. When tolvaptan (1, 3, and 10 mg/kg) was orally administrated on d 4, a dramatic improvement in mortality was seen. In the 1-mg/kg tolvaptan-treated group, two rats died on d 5 and four died on d 6 (total mortality = 38%), showing slightly prolonged survival. However, in the other tolvaptan-treated groups (3 and 10 mg/kg), plasma sodium levels gradually elevated after repeated tolvaptan administration (Fig. 2), and all rats survived d 6. In addition, statistically significant differences in mortality were seen between the vehicle- and tolvaptan-treated groups.
TABLE 1. Effects of tolvaptan on mortality in acute hyponatremia
FIG. 2. Time courses of plasma sodium concentrations in the acute model. Values are expressed as the mean ± SEM. Values are taken from the end of each experimental day. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed Dunnett’s multiple comparison test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. x, Saline-infused; , vehicle; , tolvaptan 1 mg/kg; , tolvaptan 3 mg/kg; and , tolvaptan 10 mg/kg.
Four hours after the first oral administration of tolvaptan (1, 3, and 10 mg/kg), mean plasma sodium concentrations increased by 0.5, 2.0, and 12.9 mEq/liter, respectively. For rats in the group given the highest dose of tolvaptan (10 mg/kg), the maximum change in individual plasma sodium concentration after 4 h was 21.6 mEq/liter, indicating that the concentration was rapidly corrected but remained within mild hyponatremic levels. Moreover, the maximum daily change was 24.6 mEq/liter and no abnormal animal behavior was observed, suggesting the correction rate was adequate. Furthermore, the plasma sodium concentration increased after repeated tolvaptan administrations (119.3 ± 2.3, 120.6 ± 2.7, 128.5 ± 1.4 mEq/liter on d 6), resulting in statistically significant differences (Fig. 2).
Urine parameters.
At 3 and 10 mg/kg, significant changes in urine volume and urine osmolality were seen 4 h after drug administration: Urine volume increased and osmolality decreased. The changes were also statistically significant at 24 h, except for urine volume in the 3-mg/kg tolvaptan group on d 6. Even at the lowest dose (1 mg/kg), diluted urine was excreted, and statistically significant differences were seen at some points (Fig. 3).
FIG. 3. Effects of tolvaptan on initial (0–4 h) and total daily urine volume and osmolality in the acute model. Values are expressed as the mean ± SEM. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed Dunnett’s multiple comparison test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. , Vehicle; , tolvaptan 1 mg/kg; , tolvaptan 3 mg/kg; and , tolvaptan 10 mg/kg.
Brain sodium and water content.
In the vehicle-treated group, significant changes in brain sodium and water content were observed after water retention: sodium content decreased and water content increased (Fig. 4). After 3 d of repeated tolvaptan dosing, the decreased brain sodium content increased even at the lower doses, and a statistically significant increment was observed in the highest dosed (10 mg/kg) group. However, despite significant changes in the vehicle-treated group, we observed no effects to brain water content.
FIG. 4. Effects of tolvaptan on brain sodium and water content in the acute model. Values are expressed as the mean ± SEM. Comparing the saline-infused and vehicle-treated groups for sodium content in the brain, a two-tailed t test was performed. Then, a two-tailed Dunnett’s multiple comparison test was performed to compare brain sodium contents. $$, P < 0.01 vs. saline-infused. *, P < 0.05 vs. vehicle.
Effects of tolvaptan on chronic hyponatremia
Plasma sodium control in chronic hyponatremia.
Figure 5 shows plasma sodium changes in chronic hyponatremia. Subcutaneous DDAVP infusion and liquid food consumption decreased plasma sodium concentrations by d 2 to about 110 mEq/liter, which was maintained thereafter, and severe, chronic hyponatremia was induced. Beginning on d 4, dose titrations of tolvaptan were administered. Plasma sodium concentrations gradually and significantly elevated with increasing tolvaptan doses (0.25, 0.5, 1, 2, 4, and 8 mg/kg). Mean elevated plasma sodium concentrations were as follows: 7.4 mEq/liter (0.25 mg/kg, d 4–5), 6.3 mEq/liter (0.5 mg/kg, d 5–6), 5.5 mEq/liter (1 mg/kg, d 6–7), 6.9 mEq/liter (2 mg/kg, d 7–8), and 4.4 mEq/liter (4 mg/kg, d 8–9). After d 9 in the tolvaptan-treated group, plasma sodium concentrations were within healthy levels. Therefore, elevated serum sodium levels were not observed when the tolvaptan dose was increased to 8 mg/kg. Finally, the plasma sodium concentration in the tolvaptan-treated group increased to 142.8 ± 0.4 mEq/liter (d 14). It contrasts with that of the vehicle-treated group, which was still low (110.6 ± 0.9 mEq/liter). Individual plasma sodium concentrations in this group had a maximum daily change of only 11.8 mEq/liter and no abnormal animal behavior (hyperactivity and hyperirritability, jumping, spasticity of the extremities, paralysis of the limbs, occasional seizure activity, and unresponsiveness), as previously described (20), was seen. These results suggest that the rate of correction was properly controlled.
FIG. 5. Effects of tolvaptan on plasma sodium levels in the chronic model. Values are expressed as the mean ± SEM. Seven animals comprise each group. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed t test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. x, Saline-infused; , vehicle; and , tolvaptan.
Aquaretic action of tolvaptan on chronic hyponatremia.
Oral administration of tolvaptan produced aquaresis. In the tolvaptan-treated group, the 0- to 4-h urine volume was 6–37 times greater, and urine osmolality was significantly lower than in the vehicle-treated group. Despite similar water intake for both groups, in the tolvaptan-treated group, the urine volume measured from 0–24 h was 1.1–1.5 times greater than that in the vehicle-treated group (except d 5 and 10), and significant differences were observed on d 6, 7, 8, and 13. At 0–24 h, urine osmolality was significantly lower in the tolvaptan-treated group than in the vehicle-treated group throughout the dosing period (Fig. 6). Figure 7 shows urine sodium excretion 0–24 h after oral administration of tolvaptan or the vehicle. In the tolvaptan-treated group, urine sodium excretion was 1.1–2.0 times less than in the vehicle-treated group (except on d 6), and the differences between the two groups were statistically significant on d 5, 7, 10, 11, and 13.
FIG. 6. Effects of tolvaptan on initial (0–4 h) and total daily urine volume and osmolality in the chronic model. Values are expressed as the mean ± SEM. Seven animals comprise each group. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed t test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. , Vehicle; and , tolvaptan.
FIG. 7. Effects of tolvaptan on urine sodium excretion in the chronic model. Values are expressed as the mean ± SEM. Seven animals comprise each group. The differences between the vehicle- and tolvaptan-treated groups were analyzed by repeated measures ANOVA, followed by a two-tailed t test at each time point. *, P < 0.05; **, P < 0.01 vs. vehicle. x, Saline-infused; , vehicle; and , tolvaptan.
Effects of tolvaptan on organ changes in chronic hyponatremia.
Organ weight and water content are shown in Fig. 8. Significant increases in wet weight of the heart and left kidney were observed in the vehicle-treated group, as compared with the saline-infused group. After dose titration of tolvaptan, the increment increases in wet weight of the heart and kidney were blunted, although the difference was not statistically significant in the heart. Brain, heart, and kidney water content also significantly increased in the vehicle-treated group as opposed to the saline-infused group. Tolvaptan completely reversed the vital water content increases of brain and heart seen in the vehicle-treated animals, with statistical significance. In the kidney, tolvaptan inhibited the water content increase by 43%, although statistical significance was not observed. No significant difference in body weight on d 14 was observed between the vehicle- (265.9 ± 4.1 g) and tolvaptan-treated (266.4 ± 2.6 g) groups. These data suggest that tolvaptan treatment inhibited organ water retention.
FIG. 8. Effects of tolvaptan on organ wet weight and water content in the chronic model. Values are expressed as the mean ± SEM. Seven animals comprise each group. At first, the differences between the saline-infused and vehicle groups were analyzed by a two-tailed t test, then a two-tailed t test was performed. $, P < 0.05; $$, P < 0.01 vs. saline-infused. **, P < 0.01 vs. vehicle.
Discussion
Brain adaptation to hypoosmolality is well known; when plasma sodium decreases, brain cells rapidly export electrolytes and organic osmolytes to prevent excess swelling (21). However, these adaptive mechanisms do not prevent the development of brain edema when serum sodium concentrations fall rapidly, so severe neurological symptoms or death may occur (22).
Therefore, acute severe hyponatremia needs to be rapidly corrected to less critical hyponatremia, then progressively corrected to normonatremic level. It has been reported that with rapid and controlled correction (estimated threshold of brain injury, <25 mEq/liter·d), neurological events and mortality may be avoided in acute severe hyponatremia (7). In chronic severe hyponatremia, however, the brain generally adapts itself to reduced concentrations of electrolytes and organic osmolytes. These adapted brain cells are unable to rapidly equilibrate themselves with extracellular osmolyte levels when hyponatremia is rapidly corrected (23). This situation leads to the contraction of cells, disturbance of the blood-brain barrier, and potentially irreversible neurological damage, which is manifested in extreme cases by paralysis, coma, or death.
In many cases of hyponatremia, plasma AVP levels are increased. When circulating AVP binds to V2 receptors existing in renal collecting ducts, renal free water reabsorption is stimulated via increasing intracellular cAMP and translocating aquaporin-2 from intracellular vesicles to the apical plasma (24). Tolvaptan is a specific AVP V2 receptor antagonist that blocks the binding of AVP at V2 receptors, resulting in aquaresis without depletion of electrolytes, and possibly improves diluted hyponatremia (19, 25).
We found that tolvaptan improved mortality in fluid-overloaded acute severe hyponatremia. In the vehicle-treated group, the rats failed to adapt themselves to hypoosmolality, which resulted in 47% mortality due to severe hyponatremia. In fact, a significant decrease in brain sodium and an increase of brain water content were observed during 6 d of the experiment. However, tolvaptan treatment reduced total mortality even at the lowest dose given, presumably because plasma sodium concentrations improved above some critical threshold. In the higher dose groups, death was not observed because plasma sodium concentrations rapidly elevated to mild hyponatremia levels. Furthermore, significant improvement in brain sodium content was observed although plasma sodium level was still low.
In the chronic hyponatremia study, the rate of increase of plasma sodium concentration was carefully controlled by using paced dose titration of tolvaptan. With this method, the severe state improved gradually to normonatremia without abnormal animal behavior suggesting neurological symptoms or death. In the chronic model, tolvaptan was effective at a lower dose (0.25 mg/kg) than in the acute model of hyponatremia (1 mg/kg). One of the reasons is the doses of DDAVP infused, 10 ng/h in the acute model and 1 ng/h in the chronic model.
After the normalization of plasma sodium, the dosing was extended for several days to confirm whether plasma sodium increased to hypernatremia or reverted back to hyponatremia. As a result, plasma sodium did not change any more and remained within the normal range during the extended dosing period. Similar results were seen in clinical studies (26, 27). In patients with hyponatremia secondary to congestive heart failure, plasma sodium concentrations increased to the normal range after administration of tolvaptan and remained within a normal range during treatment with tolvaptan. Interestingly, a different response of plasma sodium was observed in patients with normonatremia. After treatment, normonatremic patients had a transient increase in sodium levels, with the values returning to baseline within 3 wk of therapy. On the other hand, it has been reported that normalized plasma sodium reverted back to the state of hyponatremia, once the dosing ceased after normalization in similar animal models (16). These suggest that dose titration is a successful strategy for controlling the rate of increase of plasma sodium concentration, and that plasma sodium remains within a normal range while the dosing is extended.
It has been reported that tolvaptan produces aquaresis after single and multiple dose regimens (19) through elevated electrolyte-free water clearance in conscious rats (25). Through repeated tolvaptan administration, we were able to maintain aquaresis. Our results suggest that tolvaptan also increased free-water excretion, resulting in gradual increases in plasma sodium concentrations.
During tolvaptan treatment, 24-h urinary sodium excretion also decreased in both models, which might have partially contributed to the improvement of plasma sodium concentrations. Although the mechanisms are not clear, one possible explanation is the recovery from the situation of "solute depletion" by tolvaptan. It has been reported that the volume expansion state in syndrome of inappropriate antidiuretic hormone secretion is associated with decreased proximal tubular sodium reabsorption (28), so correction of volume expansion after tolvaptan treatment may contribute to the improvement of renal sodium reabsorption.
In the chronic model, the wet weight of the heart and kidney, and water content in the brain, heart, and kidney also significantly increased. In the vehicle-treated group, incremental increases in the heart and kidney weight almost entirely resulted from increased water content because the mean dry weight of these organs was the same as that of the saline-infused group (data not shown). After tolvaptan treatment, those changes significantly reduced organ water content, suggesting that tolvaptan improved water retention in these organs.
In conclusion, tolvaptan treatment showed aquaresis, and controlled the magnitude of plasma sodium elevation both in acute and chronic severe hyponatremia. Furthermore, tolvaptan improved mortality in acute severe hyponatremia and organ water retention in chronic severe hyponatremia. Thus, tolvaptan presents exciting therapeutic implications in the management of clinical hyponatremia.
Acknowledgments
We express sincere thanks to Dr. F. Czerwiec for his critical reading of this manuscript. We also thank Dr. J. Garfield and Mr. K. Komuro for their excellent assistance in the preparation of this manuscript.
References
Anderson RJ, Chung H, Kluge R, Schrier RW 1985 Hyponatremia: a prospective analysis of its epidemiology and the pathogenetic role of vasopressin. Ann Intern Med 102:164–168
Flear CTG, Gill GV, Burn J 1981 Hyponatremia: mechanisms and management. Lancet 2:26–81
Kleinfeld M, Casimir M, Borra S 1979 Hyponatremia as observed in a chronic disease facility. J Am Geriatrics Soc 27:156–161
Arieff AI 1986 Hyponatremia, convulsions, respiratory arrest, and permanent brain damage after elective surgery in healthy women. N Engl J Med 314:1529–1534
Ayus JC, Krothapalli RK, Armstrong DL, Norton HJ 1989 Symptomatic hyponatremia in rats: effect of treatment on mortality and brain lesions. Am J Physiol 257:F18–F22
Ayus JC, Arieff AI 1999 Chronic hyponatremic encephalopathy in postmenopausal women. Association of therapies with mobidity and mortality. JAMA 281:2299–2304
Ayus JC, Krothapalli RK, Arieff AI 1985 Changing concepts in treatment of severe symptomatic hyponatremia. Am J Med 78:897–901
Kleinschmitt-Demasters BK, Norenberg MD 1981 Rapid correction of hyponatremia causes demyelination: relation to central pontine myelinolysis. Science 211:1068–1070
Ayus JC, Krothapalli RK, Armstrong DL 1985 Rapid correction of severe hyponatremia in the rat: histopathological change in the brain. Am J Physiol 248:F711–F719
Verbalis JG, Martinez AJ, Drutarosky MD 1991 Neurological and neuropathological sequelae of correction of chronic hyponatremia. Kidney Int 39:1274–1282
Soupart A, Stenuit A, Perier O, Decaux G 1991 Limits of brain tolerance to daily increments in serum sodium in chronically hyponatremic rats treated with hypertonic saline or urea: advantages of urea. Clin Sci 80:77–84
Soupart A, Penninckx R, Stenuit A, Perier O, Decaux G 1992 Treatment of chronic hyponatremia in rats by intravenous saline: comparison of rate versus magnitude of correction. Kidney Int 41:1662–1667
Verbalis JG, Drutarosky MD 1988 Adaptation to chronic hypoosmolality in rats. Kidney Int 34:351–360
Saito T, Ishikawa S, Abe K, Kamoi K, Yamada K, Shimizu K, Saruta T, Yoshida S 1997 Acute aquaresis by the nonpeptide arginine vasopressin (AVP) antagonist OPC-31260 improves hyponatremia in patient with syndrome of inappropriate secretion of antidiuretic hormone (SIADH). J Clin Endocrinol Metab 82:1054–1057
Decaux G 2001 Long-term treatment of patient with inappropriate secretion of antidiuretic hormone by the vasopressin receptor antagonist conivaptan, urea, or furosemide. Am J Med 110:582–584
Fujisawa G, Ishikawa S, Tsuboi Y, Okada K, Saito T 1993 Therapeutic efficacy of non-peptide ADH antagonist OPC-31260 in SIADH rats. Kidney Int 44:19–23
Naito A, Hasegawa H, Kurasawa T, Ohtake Y, Matsukawa H, Ezure Y, Tsurita Y, Koike K, Shigenobu K 2000 The therapeutic efficacy of VP-343, a selective vasopressin V2 receptor antagonist, in the experimental SIADH rat model. Biol Pharm Bull 23:1323–1327
Naito A, Hasegawa H, Kurasawa T, Ohtake Y, Matsukawa H, Ezure Y, Koike K, Shigenobu K 2001 Histopathological study of kidney abnormalities in an experimental SIADH rat model and its application to the evaluation of the pharmacologic profile of VP-343, a selective vasopressin V2 receptor antagonist. Biol Pharm Bull 24:897–901
Yamamura Y, Nakamura S, Itoh S, Hirano T, Onogawa T, Yamashita T, Yamada Y, Tsujimae K, Aoyama M, Kotosai K, Ogawa H, Yamashita H, Kondo K, Tominaga M, Tsujimoto G, Mori T 1998 OPC-41061, a highly potent human vasopressin V2-receptor antagonist: pharmacological profile and aquaretic effect by single and multiple oral dosing in rats. J Pharmacol Exp Ther 287:860–867
Soupart A, Penninckx R, Crenier L, Stenuit A, Perier O, Decaux G 1994 Prevention of brain demyelination in rats after excessive correction of chronic hyponatremia by serum sodium lowering. Kidney Int 45:193–200
Verbalis JG, Gullans SR 1991 Hyponatremia causes large sustained reductions in brain content of multiple organic osmolytes in rats. Brain Res 567:274–282
Soupart A, Decaux G 1996 Therapeutic recommendations for management of severe hyponatremia: current concepts on pathogenesis and prevention of neurologic complications. Clin Nephrol 46:149–169
Verbalis JG, Gullans SR 1993 Rapid correction of hyponatremia produces differential effects on brain osmolyte reaccumulation in rats. Brain Res 606:19–27
Verbalis JG 2002 Vasopressin V2 receptor antagonists. J Mol Endocrinol 29:1–9
Hirano T, Yamamura Y, Nakamura S, Toshiyuki O, Mori T 2000 Effects of the V2-receptor antagonist OPC-41061 and the loop diuretic furosemide alone and in combination in rats. J Pharmacol Exp Ther 292:288–294
Gheorghiade M, Niazi I, Ouyang J, Czerwiec F, Kambayashi J, Zampino M, Orlandi C 2003 Vasopressin V2-receptor blockade with tolvaptan in patients with chronic heart failure: results from a double-blind, randomized trial. Circulation 107:2690–2696
Gheorghiade M, Gattis WA, O’Connor CM, Adams Jr KF, Elkayam U, Barbagelata A, Ghali JK, Benza RL, McGrew FA, Klapholz M, Ouyang J, Orlandi C 2004 Effects of tolvaptan, a vasopressin antagonist, in patients hospitalized with worsening heart failure: a randomized controlled trial. JAMA 291:1963–1971
Decaux G 2001 Difference in solute excretion during correction of hyponatremic patients with cirrhosis or syndrome of inappropriate secretion of antidiuretic hormone by oral vasopressin V2 receptor antagonist VPA-985. J Lab Clin Med 138:18–21(Toshiki Miyazaki, Yoshita)