- Email: [email protected]
Sodium balance and antidiuresis in thiazide-treated rats with diabetes insipidus
European Journal of Pharmacology, 89 (1983) 283-286
283
Elsevier Biomedical Press
Short communication S O D I U M BALANCE AND ANTIDIURESIS IN T H I A Z I D E - T R E A T E D RATS W I T H DIABETES INSIPIDUS STEPHEN J. WALTER * and DAVID G. SHIRLEY
Department of Physiology, Charing Cross Hospital Medical School, London W6 8RF, England Received 3 January 1983, accepted 7 March 1983
S.J. WALTER and D.G. SHIRLEY, Sodium balance and antidiuresis in thiazide-treated rats with diabetes insipidus, European J. Pharmacol. 89 (1983) 283-286. A sodium deficit was induced in Brattleboro rats by lowering the dietary sodium content. Urine volume was only slightly reduced. Addition of hydrochlorothiazide to the food caused a small, transient sodium deficit but led to a marked, sustained antidiuresis. The acute administration of a large dose of hydrochlorothiazide produced a larger, sustained sodium deficit but no lasting antidiuresis. These results indicate that sodium depletion cannot account for the antidiuresis of hydrochlorothiazide treatment in diabetes insipidus. Diabetes insipidus
Antidiuresis
Hydrochlorothiazide
1. Introduction The administration of thiazide diuretics to patients or animals with diabetes insipidus results in a paradoxical antidiuresis (Harvard, 1965). It is generally considered that the reduction in urine volume is a consequence of a sodium depletion resulting from the natriuretic action of the drugs (Martinez-Maldonado et al., 1973; Skadhauge, 1973). Although experiments using rats with diabetes insipidus have provided support for this hypothesis for the period immediately after thiazide administration (Shirley et al., 1978), the sustained antidiuresis which accompanies chronic thiazide treatment is more difficult to explain. We have found that urinary sodium excretion is increased only on the first day of treatment and that the resulting sodium depletion remains statistically significant for only 48 h (Walter et al., 1982). It remains possible to argue, however, that the large scatter of data which occurs in such balance studies could mask a small sodium deficit, and that
* To whom all correspondence should be addressed. 0014-2999/83/0000 0000/$03.00 © 1983 Elsevier Biomedical Press
Sodium balance
this sodium depletion might be an essential feature of the long-term antidiuretic action of thiazides. The present experiments were designed to investigate this possibility.
2. Materials and methods All experiments were performed on male Brattleboro rats with hereditary hypothalamic diabetes insipidus, which weighed 200-250 g at the start of the period of investigation. Each rat was placed in a metabolism cage (Metabowl, Jencons Ltd., Leighton Buzzard, U.K.) and allowed free access to food and deionised water. The basic diet (Ewos Ltd., Watford, U.K.) contained sodium and potassium at a concentration of 4 mmol and 3 mmol per kg dry weight respectively. This dry food was made into a paste with a solution containing KC1 and sometimes NaC1, so that the final paste had a potassium content of 180 m m o l / k g dry weight and a sodium content of either 4 or 122 m m o l / k g dry weight. During the first 7 days in the metabolism cages, all rats received food with a sodium content of 122
284 m m o l / k g dry weight. The first 4 of these days were regarded as an equilibration period and the next 3 days as a control period. For the final 12 days of the experiment, the sodium content of the diet was reduced to 4 m m o l / k g dry weight. After 6 days on the low-sodium diet the rats were divided into 2 groups. One group (acute HCTZ) remained on the same diet for the final period of 6 days, but were given, by gavage, 25 mg (80 /~mol) of hydrochlorothiazide (Merck, Sharp and Dohme) dissolved in 5 ml of 50 mmol/1 KOH on each of the first 2 of these days. On the same 2 days, rats in the second group (chronic HCTZ) received 5 ml of 50 mmol/1 KOH containing no drug. Hydrochlorothiazide was administered to the chronic HCTZ group on all 6 days, mixed with the food at a concentration of 500 mg (1.7 mmol) per kg dry weight. The mean daily intake of the drug in this group was approximately 6 mg (20 ymol). Throughout the period in metabolism cages, measurements of body weight, food intake and urine volume were made daily and the Na and K contents of the urine and faeces were determined. At the end of the experiment (6 days after the start of drug treatment) the extracellular fluid volume (inulin space) of 6 rats from each group was determined (Walter et al., 1982). A final blood sample was obtained from all rats for plasma analysis. Measurements of urine, plasma, food and faecal Na + and K + concentrations, plasma osmolality, plasma protein concentration and packed cell volume were performed essentially as described previously (Walter et al., 1982). A minor modification was that in order to measure accurately very low Na + concentrations an appropriately modified calibration curve was established for the flame photometer.
3. Results
3.1. Effect of low Na diet (fig. 1) On the first day on which they were given food containing a low sodium content, all rats excreted more sodium than they ate and therefore went into a negative sodium balance, the mean deficit being
Na content of food (mmollkg)
I--
l
4
1
(a) Urine volume (ml/day)
''Jll
(b) UrinaryNa excretion (~mol/day) 25OO
5O0
o
(c) Cumulative Na balance relative to day 0 (~umol) 0
-1 O 1
2 3 4 5 6 Days Fig. 1. (a) Urine volume, (b) urinary sodium excretion and (c) cumulative sodium balance in Brattleboro rats maintained on a diet with sodium content of 122 or 4 mmol per kg dry weight. Values are given as means and standard errors. The number of rats was 18.
approximately 350 /~mol/rat. Urine volume was reduced by approximately 20 ml (P < 0.001, paired t-test) compared with the previous day. On subsequent days, sodium excretion in both urine and faeces was markedly reduced. Nevertheless, after 6 days on the low sodium diet there remained a sodium deficit of approximately 200 ~mol per rat. By this time, urine volume was only slightly (though significantly) lower than in the control period. Although the potassium content of the diet was not reduced, and the rats continued in positive potassium balance, the introduction of the low Na
285 diet resulted in a significant reduction in faecal K excretion (normal Na diet: 63 + 5 /~mol K / d a y ; low Na diet: 28 + 2 ktmol K / d a y ; P < 0.001). This fall in faecal K output was maintained throughout the period of reduced sodium intake.
the food for 6 further days (chronic HCTZ); the other half were given hydrochlorothiazide by gavage on each of the first two of these days (acute HCTZ), but received no drug thereafter.
3.2.1. Urine volume (fig. 2) 3.2. Effect of hydrochlorothiazide administration Six days after the introduction of the low Na diet, half the rats received hydrochlorothiazide in
(a) Urine volume (mllday) 20O
Both methods of hydrochlorothiazide administration resulted in a prompt antidiuresis. The reduction in urine volume was maintained provided that hydrochlorothiazide treatment was continued (chronic HCTZ). In contrast, after the second and final intragastric dose of hydrochlorothiazide (acute HCTZ), urine volume quickly returned to control values.
3.2.2. Electrolyte balance (fig. 2)
I (b) Urinary Na excretion (/Jmollday) 300 200 10O
(c) Cumulative Na balance relative to day O (/Jmol) O
-400
-1
0
1
2 3 4 5 6 Days Fig. 2. (a) Urine volume, (b) urinary sodium excretion and (c) cumulative sodium balance in Brattleboro rats maintained on a diet with a sodium content of 4 mmol per kg dry weight. On days 1-6, one group of rats (chronic HCTZ) received hydrochlorothiazide (20 #mol/day) in the food (closed columns). On days 1 and 2, the other group (acute HCTZ) received hydrochlorothiazide (80 /~mol) intragastrically (stippled columns), but thereafter received no drug (open columns). Values are given as means and standard errors. The number of rats in each group was 9.
On the first day of thiazide treatment there was a significant rise in urinary sodium excretion in each group of rats, the increase being almost identical in the two groups. However, on the second day of treatment sodium excretion was significantly greater (P < 0.001) in rats given the drug by gavage (acute HCTZ). In consequence the sodium deficit became significantly greater in the acute HCTZ group than in rats receiving the drug within the food (chronic HCTZ), and remained so during the final 4 days in the metabolism cages. Although hydrochlorothiazide treatment appeared to cause some increase in urinary K excretion, there was no statistically significant potassium deficit in either group of animals.
3.2.3. Body weight, extracellular fluid volume, packed cell volume and plasma analyses Body weight increased only slowly throughout the period on the low Na diet. At no time during the study were the body weights of the two groups of animals significantly different from one another. Extracellular fluid volume (ECFV) was measured in 6 rats from each group at the end of their period in metabolism cages. The ECFV of rats receiving chronic hydrochlorothiazide treatment (48.2 + 1.9 ml; 19.7 + 0.4 ml/100g body weight) was significantly lower than that of the acute HCTZ group (53.8 + 1.6 ml; 21.9 + 0.5 ml/100g body weight, P < 0.02). This reduced ECFV was associated with significant increases in packed cell volume (51.9 + 1.2 vs. 49.1 ___0.3, P < 0.05) and
286 plasma protein c o n c e n t r a t i o n (70.4_+ 1.7 vs. 64.0 + 1.5 g / l , P < 0.02). There were no significant differences between the two groups of rats with regard to plasma osmolality or N a + or K + concentrations.
4. Discussion As previously observed in Brattleboro rats on a n o r m a l sodium diet (Walter et al., 1982), chronic hydrochlorothiazide treatment resulted in an antidiuresis which persisted for as long as the drug was administered. W h e n hydrochlorothiazide was given as a very large dose on two consecutive days (acute H C T Z ) , an antidiuresis was also observed, b u t as soon as the drug was w i t h d r a w n the urine flow rate r e t u r n e d to normal. This occurred despite the fact that from the second day of treatm e n t the acute H C T Z group was in a greater N a deficit than the chronic H C T Z rats. It should be emphasized that these results were o b t a i n e d in a situation where the daily sodium intake and o u t p u t were very small. Errors of meas u r e m e n t were c o n s e q u e n t l y also small, a n d the b a l a n c e data correspondingly reliable. In view of this it seems clear that the antidiuresis resulting from chronic hydrochlorothiazide a d m i n i s t r a t i o n c a n n o t be explained simply as a consequence of a s o d i u m depletion. This conclusion is strengthened by the findings d u r i n g the period preceding hydrochlorothiazide a d m i n i s t r a t i o n . Lowering the sodium c o n t e n t of the diet resulted in a significant N a deficit (as great as that f o u n d even in the early stages of hydrochlorothiazide treatment), a n d yet the reduction in urine flow was trivial c o m p a r e d with that caused by thiazide. In a previous investigation we had already questioned the role of sodium depletion in mediating the antidiuresis which occurs during chronic thiazide treatment (Walter et al., 1982). The present study has shown that the sodium depletion a n d the antidiuresis can be completely dissociated. The cause of the antidiuresis, therefore, still needs to be explained. The reduction in urine volume seen in Brattleboro rats receiving chronic hydrochlorothiazide t r e a t m e n t whilst on a n o r m a l Na
diet is associated with a fall in ECFV (Walter el al., 1982) which might be responsible for the changes in renal function which bring about the antidiuresis (Shirley et al., 1982). In the present experiments (using a low Na diet) the rats with a sustained antidiuresis (chronic H C T Z ) had a lower E C F V than those of the acute H C T Z group, whose urine volume had returned to pre-drug levels. Rather surprisingly, this reduced E C F V was not associated with a detectable difference in body weight between the two groups, suggesting that hydrochlorothiazide might somehow cause a redistribution of body water from the extracellular to the intracellular fluid c o m p a r t m e n t . This hypothesis, which receives some support from the observed increases in packed cell volume and plasma protein concentration, is consistent with previous evidence of a thiazide-induced redistribution of body sodium (Walter et al., 1982).
Acknowledgements We thank the University of London Central Research Fund for financial support, Dr. K. Thomsen for helpful discussion and Mr. J. Skinner for technical assistance.
References Harvard, C.W.H., 1965, Thiazide-induced antidiuresis in diabetes insipidus, Proc. Roy. Soc. Med. 58, 1005. Martinez-Maldonado, M., G. Eknoyan and W.N. Suki. 1973. Diuretics in nonedematous states, Arch. Intern. Med. 131, 797. Shirley, D.G., S.J. Walter and J.F. Laycock, 1978, The role of sodium depletion in hydrochlorothiazide-inducedantidiuresis in Brattleboro rats with diabetes insipidus, Clin. Sci. Mol. Med. 54, 209. Shirley, D.G., S.J. Walter and J.F. Laycock, 1982, The antidiuretic effect of chronic hydrochlorothiazide treatment in rats with diabetes insipidus: renal mechanisms, Clin. Sci. 63, 533. Skadhauge, E., 1973,Diureticsand diabetes insipidus, in: Modern Diuretic Therapy in the Treatment of Cardiovascular and Renal Disease, eds. A.F. Lant and G.M. Wilson (Exerpta Medica: Amsterdam) p. 306. Walter, S.J., J. Skinner, J.F. Laycock and D.G. Shirley, 1982, The antidiuretic effect of chronic hydrochlorothiazide treatment in rats with diabetes insipidus: water and electrolyte balance, Clin. Sci. 63, 525.
283
Elsevier Biomedical Press
Short communication S O D I U M BALANCE AND ANTIDIURESIS IN T H I A Z I D E - T R E A T E D RATS W I T H DIABETES INSIPIDUS STEPHEN J. WALTER * and DAVID G. SHIRLEY
Department of Physiology, Charing Cross Hospital Medical School, London W6 8RF, England Received 3 January 1983, accepted 7 March 1983
S.J. WALTER and D.G. SHIRLEY, Sodium balance and antidiuresis in thiazide-treated rats with diabetes insipidus, European J. Pharmacol. 89 (1983) 283-286. A sodium deficit was induced in Brattleboro rats by lowering the dietary sodium content. Urine volume was only slightly reduced. Addition of hydrochlorothiazide to the food caused a small, transient sodium deficit but led to a marked, sustained antidiuresis. The acute administration of a large dose of hydrochlorothiazide produced a larger, sustained sodium deficit but no lasting antidiuresis. These results indicate that sodium depletion cannot account for the antidiuresis of hydrochlorothiazide treatment in diabetes insipidus. Diabetes insipidus
Antidiuresis
Hydrochlorothiazide
1. Introduction The administration of thiazide diuretics to patients or animals with diabetes insipidus results in a paradoxical antidiuresis (Harvard, 1965). It is generally considered that the reduction in urine volume is a consequence of a sodium depletion resulting from the natriuretic action of the drugs (Martinez-Maldonado et al., 1973; Skadhauge, 1973). Although experiments using rats with diabetes insipidus have provided support for this hypothesis for the period immediately after thiazide administration (Shirley et al., 1978), the sustained antidiuresis which accompanies chronic thiazide treatment is more difficult to explain. We have found that urinary sodium excretion is increased only on the first day of treatment and that the resulting sodium depletion remains statistically significant for only 48 h (Walter et al., 1982). It remains possible to argue, however, that the large scatter of data which occurs in such balance studies could mask a small sodium deficit, and that
* To whom all correspondence should be addressed. 0014-2999/83/0000 0000/$03.00 © 1983 Elsevier Biomedical Press
Sodium balance
this sodium depletion might be an essential feature of the long-term antidiuretic action of thiazides. The present experiments were designed to investigate this possibility.
2. Materials and methods All experiments were performed on male Brattleboro rats with hereditary hypothalamic diabetes insipidus, which weighed 200-250 g at the start of the period of investigation. Each rat was placed in a metabolism cage (Metabowl, Jencons Ltd., Leighton Buzzard, U.K.) and allowed free access to food and deionised water. The basic diet (Ewos Ltd., Watford, U.K.) contained sodium and potassium at a concentration of 4 mmol and 3 mmol per kg dry weight respectively. This dry food was made into a paste with a solution containing KC1 and sometimes NaC1, so that the final paste had a potassium content of 180 m m o l / k g dry weight and a sodium content of either 4 or 122 m m o l / k g dry weight. During the first 7 days in the metabolism cages, all rats received food with a sodium content of 122
284 m m o l / k g dry weight. The first 4 of these days were regarded as an equilibration period and the next 3 days as a control period. For the final 12 days of the experiment, the sodium content of the diet was reduced to 4 m m o l / k g dry weight. After 6 days on the low-sodium diet the rats were divided into 2 groups. One group (acute HCTZ) remained on the same diet for the final period of 6 days, but were given, by gavage, 25 mg (80 /~mol) of hydrochlorothiazide (Merck, Sharp and Dohme) dissolved in 5 ml of 50 mmol/1 KOH on each of the first 2 of these days. On the same 2 days, rats in the second group (chronic HCTZ) received 5 ml of 50 mmol/1 KOH containing no drug. Hydrochlorothiazide was administered to the chronic HCTZ group on all 6 days, mixed with the food at a concentration of 500 mg (1.7 mmol) per kg dry weight. The mean daily intake of the drug in this group was approximately 6 mg (20 ymol). Throughout the period in metabolism cages, measurements of body weight, food intake and urine volume were made daily and the Na and K contents of the urine and faeces were determined. At the end of the experiment (6 days after the start of drug treatment) the extracellular fluid volume (inulin space) of 6 rats from each group was determined (Walter et al., 1982). A final blood sample was obtained from all rats for plasma analysis. Measurements of urine, plasma, food and faecal Na + and K + concentrations, plasma osmolality, plasma protein concentration and packed cell volume were performed essentially as described previously (Walter et al., 1982). A minor modification was that in order to measure accurately very low Na + concentrations an appropriately modified calibration curve was established for the flame photometer.
3. Results
3.1. Effect of low Na diet (fig. 1) On the first day on which they were given food containing a low sodium content, all rats excreted more sodium than they ate and therefore went into a negative sodium balance, the mean deficit being
Na content of food (mmollkg)
I--
l
4
1
(a) Urine volume (ml/day)
''Jll
(b) UrinaryNa excretion (~mol/day) 25OO
5O0
o
(c) Cumulative Na balance relative to day 0 (~umol) 0
-1 O 1
2 3 4 5 6 Days Fig. 1. (a) Urine volume, (b) urinary sodium excretion and (c) cumulative sodium balance in Brattleboro rats maintained on a diet with sodium content of 122 or 4 mmol per kg dry weight. Values are given as means and standard errors. The number of rats was 18.
approximately 350 /~mol/rat. Urine volume was reduced by approximately 20 ml (P < 0.001, paired t-test) compared with the previous day. On subsequent days, sodium excretion in both urine and faeces was markedly reduced. Nevertheless, after 6 days on the low sodium diet there remained a sodium deficit of approximately 200 ~mol per rat. By this time, urine volume was only slightly (though significantly) lower than in the control period. Although the potassium content of the diet was not reduced, and the rats continued in positive potassium balance, the introduction of the low Na
285 diet resulted in a significant reduction in faecal K excretion (normal Na diet: 63 + 5 /~mol K / d a y ; low Na diet: 28 + 2 ktmol K / d a y ; P < 0.001). This fall in faecal K output was maintained throughout the period of reduced sodium intake.
the food for 6 further days (chronic HCTZ); the other half were given hydrochlorothiazide by gavage on each of the first two of these days (acute HCTZ), but received no drug thereafter.
3.2.1. Urine volume (fig. 2) 3.2. Effect of hydrochlorothiazide administration Six days after the introduction of the low Na diet, half the rats received hydrochlorothiazide in
(a) Urine volume (mllday) 20O
Both methods of hydrochlorothiazide administration resulted in a prompt antidiuresis. The reduction in urine volume was maintained provided that hydrochlorothiazide treatment was continued (chronic HCTZ). In contrast, after the second and final intragastric dose of hydrochlorothiazide (acute HCTZ), urine volume quickly returned to control values.
3.2.2. Electrolyte balance (fig. 2)
I (b) Urinary Na excretion (/Jmollday) 300 200 10O
(c) Cumulative Na balance relative to day O (/Jmol) O
-400
-1
0
1
2 3 4 5 6 Days Fig. 2. (a) Urine volume, (b) urinary sodium excretion and (c) cumulative sodium balance in Brattleboro rats maintained on a diet with a sodium content of 4 mmol per kg dry weight. On days 1-6, one group of rats (chronic HCTZ) received hydrochlorothiazide (20 #mol/day) in the food (closed columns). On days 1 and 2, the other group (acute HCTZ) received hydrochlorothiazide (80 /~mol) intragastrically (stippled columns), but thereafter received no drug (open columns). Values are given as means and standard errors. The number of rats in each group was 9.
On the first day of thiazide treatment there was a significant rise in urinary sodium excretion in each group of rats, the increase being almost identical in the two groups. However, on the second day of treatment sodium excretion was significantly greater (P < 0.001) in rats given the drug by gavage (acute HCTZ). In consequence the sodium deficit became significantly greater in the acute HCTZ group than in rats receiving the drug within the food (chronic HCTZ), and remained so during the final 4 days in the metabolism cages. Although hydrochlorothiazide treatment appeared to cause some increase in urinary K excretion, there was no statistically significant potassium deficit in either group of animals.
3.2.3. Body weight, extracellular fluid volume, packed cell volume and plasma analyses Body weight increased only slowly throughout the period on the low Na diet. At no time during the study were the body weights of the two groups of animals significantly different from one another. Extracellular fluid volume (ECFV) was measured in 6 rats from each group at the end of their period in metabolism cages. The ECFV of rats receiving chronic hydrochlorothiazide treatment (48.2 + 1.9 ml; 19.7 + 0.4 ml/100g body weight) was significantly lower than that of the acute HCTZ group (53.8 + 1.6 ml; 21.9 + 0.5 ml/100g body weight, P < 0.02). This reduced ECFV was associated with significant increases in packed cell volume (51.9 + 1.2 vs. 49.1 ___0.3, P < 0.05) and
286 plasma protein c o n c e n t r a t i o n (70.4_+ 1.7 vs. 64.0 + 1.5 g / l , P < 0.02). There were no significant differences between the two groups of rats with regard to plasma osmolality or N a + or K + concentrations.
4. Discussion As previously observed in Brattleboro rats on a n o r m a l sodium diet (Walter et al., 1982), chronic hydrochlorothiazide treatment resulted in an antidiuresis which persisted for as long as the drug was administered. W h e n hydrochlorothiazide was given as a very large dose on two consecutive days (acute H C T Z ) , an antidiuresis was also observed, b u t as soon as the drug was w i t h d r a w n the urine flow rate r e t u r n e d to normal. This occurred despite the fact that from the second day of treatm e n t the acute H C T Z group was in a greater N a deficit than the chronic H C T Z rats. It should be emphasized that these results were o b t a i n e d in a situation where the daily sodium intake and o u t p u t were very small. Errors of meas u r e m e n t were c o n s e q u e n t l y also small, a n d the b a l a n c e data correspondingly reliable. In view of this it seems clear that the antidiuresis resulting from chronic hydrochlorothiazide a d m i n i s t r a t i o n c a n n o t be explained simply as a consequence of a s o d i u m depletion. This conclusion is strengthened by the findings d u r i n g the period preceding hydrochlorothiazide a d m i n i s t r a t i o n . Lowering the sodium c o n t e n t of the diet resulted in a significant N a deficit (as great as that f o u n d even in the early stages of hydrochlorothiazide treatment), a n d yet the reduction in urine flow was trivial c o m p a r e d with that caused by thiazide. In a previous investigation we had already questioned the role of sodium depletion in mediating the antidiuresis which occurs during chronic thiazide treatment (Walter et al., 1982). The present study has shown that the sodium depletion a n d the antidiuresis can be completely dissociated. The cause of the antidiuresis, therefore, still needs to be explained. The reduction in urine volume seen in Brattleboro rats receiving chronic hydrochlorothiazide t r e a t m e n t whilst on a n o r m a l Na
diet is associated with a fall in ECFV (Walter el al., 1982) which might be responsible for the changes in renal function which bring about the antidiuresis (Shirley et al., 1982). In the present experiments (using a low Na diet) the rats with a sustained antidiuresis (chronic H C T Z ) had a lower E C F V than those of the acute H C T Z group, whose urine volume had returned to pre-drug levels. Rather surprisingly, this reduced E C F V was not associated with a detectable difference in body weight between the two groups, suggesting that hydrochlorothiazide might somehow cause a redistribution of body water from the extracellular to the intracellular fluid c o m p a r t m e n t . This hypothesis, which receives some support from the observed increases in packed cell volume and plasma protein concentration, is consistent with previous evidence of a thiazide-induced redistribution of body sodium (Walter et al., 1982).
Acknowledgements We thank the University of London Central Research Fund for financial support, Dr. K. Thomsen for helpful discussion and Mr. J. Skinner for technical assistance.
References Harvard, C.W.H., 1965, Thiazide-induced antidiuresis in diabetes insipidus, Proc. Roy. Soc. Med. 58, 1005. Martinez-Maldonado, M., G. Eknoyan and W.N. Suki. 1973. Diuretics in nonedematous states, Arch. Intern. Med. 131, 797. Shirley, D.G., S.J. Walter and J.F. Laycock, 1978, The role of sodium depletion in hydrochlorothiazide-inducedantidiuresis in Brattleboro rats with diabetes insipidus, Clin. Sci. Mol. Med. 54, 209. Shirley, D.G., S.J. Walter and J.F. Laycock, 1982, The antidiuretic effect of chronic hydrochlorothiazide treatment in rats with diabetes insipidus: renal mechanisms, Clin. Sci. 63, 533. Skadhauge, E., 1973,Diureticsand diabetes insipidus, in: Modern Diuretic Therapy in the Treatment of Cardiovascular and Renal Disease, eds. A.F. Lant and G.M. Wilson (Exerpta Medica: Amsterdam) p. 306. Walter, S.J., J. Skinner, J.F. Laycock and D.G. Shirley, 1982, The antidiuretic effect of chronic hydrochlorothiazide treatment in rats with diabetes insipidus: water and electrolyte balance, Clin. Sci. 63, 525.