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Effects of efonidipine hydrochloride (NZ-105), a calcium antagonist, on renal function in conscious spontaneously hypertensive rats
~ ) Pergamon
0306-3623(94)00192-8
Gen. Pharmac. Vol.26, No. 2, pp. 333-337,1995 Copyright© 1995ElsevierScienceLtd Prin~d in Great Britain.All fights reserved 0306-3623/95$9.50+ 0.00
Effects of Efonidipine Hydrochloride (NZ-105), a Calcium Antagonist, on Renal Function in Conscious Spontaneously Hypertensive Rats T. Y O T S U M O T O , Y. M A S U D A , * C. S H U D O , H. S U G I T A , T. Y A M A S H I T A a n d S. T A N A K A Shiraoka Research Station of Biological Science, Nissan Chemical Industries Ltd., 1470, Shiraoka, Minamisaitama, Saitama, 349-02, Japan [Tel: (81)-480-92-2513; Fax: (81)-480-92-2516] (Received 20 June 1994)
Abstract--1. We investigated the effects of short- and long-term administration of efonidipine hydrochloride (NZ-105), 1,4-dihydropyridine derivative, in conscious spontaneously hypertensive rats (SHR). 2. Oral administration of NZ-105 for 12 weeks caused diuretic and natriuretic effects, which were not attenuated during the experimental period. 3. In the short-term experiment for investigating the mechanism of the diuretic effect, intravenous injection of NZ-105 (0.03 mg/kg of body weight) significantly increased the urine volume (UV), renal plasma flow (RPF) and glomerular filtration rate (GFR). The increment rate of UV and RPF was 105.4+ 17.8% and 111.7_+72.8%, respectively, which were larger than the increment rate of GFR (38.5 _+ 14.0%). 4. The diuretic or natriuretic effect of NZ-105 was suggested to be due to both the inhibition of sodium reabsorption and, at least in part. the increase of GFR. Key Words: NZ-105, calcium antagonist, renal function
INTRODUCTION Efonidipine hydrochloride (NZ-105), is a 1,4-dihydropyridine derivative which exerts a long-lasting hypotensive effect with a very slow onset (Masuda et al., 1990). We previously demonstrated that NZ-105 possessed diuretic and natriuretic effects by 15-day successive oral administration in spontaneously hypertensive rats (SHRs) (Masuda et al., 1990). Moreover, we have already reported that intravenous injection of NZ-105 in anesthetized SHR causes a diuretic effect without a change of the glomerular filtration rate (GFR) (Yokoyama et al., 1992). On the other hand, in clinical situations, NZ-105 revealed diuretic effects accompanied with an increase of G F R (personal communication, K. Taniguchi et al.).
*To whom all correspondence should be addressed.
In the present study, we further studied the above mentioned findings to explore the long-term diuretic effect and to investigate its mechanism. Firstly, we investigated the diuretic effects by oral administration of NZ-105 for 12 consecutive weeks in SHRs. Secondary, in conscious SHRs, we investigated the effects of intravenous injection on the renal hemodynamics to study the diuretic mechanism. MATERIALS AND METHODS Chronic experiment procedure Twenty-eight-week-old male SHRs (Charles River, Japan) were used. The SHRs were divided into three groups so that the systolic blood pressure (SBP) was evenly distributed. SBP was measured by the tail-cuff method (UR-5000, UEDA). The animals were given orally NZ-105 (5 and 20mg/kg) or solvent (0.5% methyl cellulose) once a day for 12 weeks. Food and water were given ad libitum. Every 4 weeks, urine was
333
334
T. Yotsumoto et al.
collected. Briefly, the rats were orally administered the drugs and loaded saline at the volume of 25 ml/kg body weight immediately after being placed in metabolism cages individually. The urine was collected for 6 hr in the absence of food and water intake. On the last day of the administration period (at 12 weeks), the urine was collected up to 6 hr and from 6 to 24 hr after the administration. Urine volume (UV) was measured gravimetrically, and urinary sodium (UNa ÷) or potassium (UK ÷) was analyzed by means of a flame photometer (Hitachi 710).
Short-term experiment procedure SHR (200-270 g, Charles River, Japan) were anesthetized with sodium pentobarbital (50 mg/kg, i.p.). Polyethylene catheters were placed into the bladder for collecting urine, and into the vena cava via the left femoral vein for infusion of inulin and p-aminohippuric acid (PAH). Other polyethylene catheters were placed into the abdominal aorta via the left femoral artery for withdrawal of blood. Except for the bladder catheter, the free ends of the catheters were exteriorized in the neck from under the skin. The wounds were surgically sutured. Each animal was placed in a Ballman cage and the experiments were started after they had recovered from anesthesia, at least 2 hr after surgery. A priming dose of inulin (200mg/kg) and PAH (8mg/kg) was given, followed by a sustaining infusion of isotonic saline containing inulin (36 mg/ml) and PAH (3 mg/ml) at a rate of 3.3 ml/hr. After an equilibration period (170rain), urine samples were collected for 20 min as a control period. After the control period, the drug or vehicle (5% cremophore-saline) was injected. Then urine samples were collected during two 20 rain clearance periods. In this experiment, arterial blood samples were obtained at the midpoint of the control period (20min before the drug or vehicle administration) and at each 20-rain clearance period. UV was determined gravimetrically. Urine and plasma samples were analyzed for inulin and PAH. Inulin concentrations in samples were quantified photometrically with anthrone. PAH was quantified photometrically with N-l-naphthylethylenediamine. The G F R and renal plasma flow (RPF) were estimated from the renal clearance of inulin and PAH, respectively.
Drugs NZ-105 was synthesized by the Central Research Laboratories of Nissan Chemical Industries Ltd. For
oral administration, NZ-105 was suspended in 0.5% methyl cellulose solution (Shinetsu Chemical, SM400) and administered at a volume of 2ml/kg through a gastric tube. For intravenous injection, NZ-105 was dissolved in 5% cremophore-saline (1 mg/ml), diluted with saline for the injection at a dose of 0.03 mg/kg. The solution was injected in a volume of 0.4 ml/kg via an administration catheter.
Statistical analysis All results were expressed as the mean + SEM. In long-term experiment results, statistical analyses were performed using the Bartlett test of variance. Parametrical analyses were performed by one-way analysis of variance. If these analysis results were significant, comparisons among groups were analyzed by Tukey's multiple method. On the other hand, non-parametrical analysis was performed with the Kruskal-Wallis test. If significant, it was performed using Tukey's method. For acute experiment results, statistical analysis was performed using the F-test of variance, followed by the Student's t-test or Aspin-Welch's t-test.
RESULTS
Chronic experiment Figure 1 shows the effect of chronic administration of NZ-105 for UV, UNa + and UK + in SHRs. The UV was significantly increased by the administration of 20 mg/kg of NZ-105 daily for 12 weeks, and by that of 5 mg/kg for 12 weeks compared with the control. Similarly the UNa + was also significantly increased at the 12th week for the high close of NZ-105, and at the 4th and 12th weeks for the low dose of NZ-105. The UK + was not significantly changed, except for the 12th week value for the high dose of NZ-105. The diuretic and natriuretic effects of NZ-105 were not attenuated during the 12-week period of the repeated administration. Table l shows the UV and the amount of urinary electrolyte excretion (UNa +, U K +) for the 0-6 hr, 6-24 hr and 0-24hr period at the 12th week after chronic administration of NZ-105. NZ-105 caused a significant and dose-dependent increase in UV, and the amount of UNa + in the 0-6 hr period, compared with the control group. UK + was significantly increased only by the high dose of NZ-105. Each parameter determined for the 6-24 hr urine tended to decrease compared with the 0 - 6 h r urine of the NZ-105 treated rats, which were slightly lower than those of the control group.
Renal effects of efonidipine Acute experiment Figure 2 shows the effects of intravenous injection of NZ-105 on UV, R P F and G F R in the conscious SHR. The U V was significantly increased in the first and second clearance periods after administration of the drug, compared with the control group. The R P F and G F R were significantly increased in the first clearance period, but not the second period. The incremental rate of UV, R P F and G F R from - 2 0 - 0 min (control period) to the 0-20 min (the first clearance period) was 105.4 + 17.8%, 111.7 + 72.8% and 38.5 + 14.0%, respectively.
335
Table I. The effect of the vehicle or efonidipine hydrochloride (NZ-105) for UV, the amount of UNa+V and U K + V in the 0 ~ , 6-24 and 0-24 hr period in SHR after administration for 12 consecutive weeks Drug (mg/kg)
Time (hr)
UV (ml/kg)
UNa÷V (,u Eq/kg)
UK÷V (/~Eq/kg)
Control
0~ Total
6.4 _+ 1.5 12.7_+2.0 19.1 _+ 3.1
839.1 _+ 204.5 1702.8 -+ 157.2 2542.0 _+ 306.6
857.9 _+ 176.9 1614.0_+139.5 2471.9 _+ 266.4
NZ-105 15]
~6 6-24 Total
17.5 _+ 1.3"* 9.8_+0.7 27.3_+ 1.5
2220.0 _+ 237.0** 1052.3 + 104.1 1310.0_+ 120.9 1313.2_+ 116.6 3530.1 _+ 233.6* 2365.4_+ 160.4
NZ-105 [201
0-6 6-24 Total
3 0 . 4 + 1.8"* 3738.4+232.1"* 1751.5_+99.7"* 12.4_+1.2 1197.2 + 116.1 1480.3_+89.4 42.8_+ 1,5"* 4935.6_+ 173.4"* 3231.7_+ 121.1"
0.5%MC 6-24
All values are the mean _+ SEM from 11 to 13 animals. *P < 0.05, **P < 0.01, compared with the values of the control group.
DISCUSSION
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Fig. 1. Urine volume, urinary Na ÷ and K* excretion for a 6-hr period on the day before and 4, 8 and 12 weeks after repeated administration of efonidipine hydrochloride (NZ105) for up to 12 weeks. NZ-105 was orally administered at 5 mg/kg ([], n = I 1), 20 ( , , n = 13), or vehicle (K], n = 13) in conscious spontaneously hypertensive rats. *P < 0.05, **P < 0.01; significantly different from each control value.
It is well known that chronic treatment with vasodilators, such as hydralazine and minoxidil, cause renal sodium and water retention (Ledingham, 1981). To prevent edema, the simultaneous use of diuretics is needed in a clinical situation. On the other hand, it has been shown that several 1,4-dihydropyridine class calcium antagonists have not only an anti-hypertensive effect by their direct vasodilating action, but also diuretic and natriuretic effects (Johns, 1985, 1988; Abe et al., 1983; M o r i m o t o et al., 1989; Oizumi et al., 1989), which are beneficial characteristics for an anti-hypertensive drug. We have already shown that NZ-105 caused antihypertensive and diuretic effects, and that those effects of NZ-105, but not nicardipine, were attenuated for a 15-day period in conscious SHR. In the present study, NZ-105 caused a diuretic and natriuretic effect which is consistent with the previous result, which was not attenuated for 12 weeks of repeated administration (Masuda et al., 1990). Nordlander et al. (1985) reported that the acute administration of felodipine caused a diuretic and natriuretic effect in SHRs, while chronic administration for 6 months maintained a diuretic effect, but not a natriuretic effect. They speculated that the reason may be a readjustment for the natriuretic effect induced by chronic administration of felodipine. Our present findings indicate that the readjustment for the natriuretic effect of NZ-105 may be weaker than that of felodipine. On the other hand, NZ-105 caused a marked diuretic effect during the 0 - 6 hr period, and this effect tended to decrease at the 6-24 hr period compared with the control group (Table 1). This may be due to a compensatory mechanism for the diuretic effect caused during the early period (0-6 hr).
336
T. Yotsumoto et al.
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20-40
Time (min) Fig. 2. Effects of intravenous administration of efonidipine hydrochloride (NZ-105) on urine volume (UV), renal plasma flow (RPF) and glomerular filtration rate (GFR) in conscious spontaneously hypertensive rats. NZ-105 was administered at 0.03 mg/kg (m, n = 5), and vehicle (I:], n = 6). *P < 0.05; significantlydifferent from each control value.
The mechanism for diuretic and natriuretic effects of 1,4-dihydropyridine class calcium antagonists has been reported to be an increase of G F R (Abe et al., 1983) and/or inhibition of water and sodium reabsorption at the proximal tubules (H~iberle et al., 1987), distal tubules and/or collecting ducts (Dibona and Sawin, 1984). In recent years, there have been many reports showing that 1,4-dihydropyridine class calcium antagonists increased G F R in humans (Baba et al., 1987; Yokoyama and Kaburagi, 1983), but not
in rats, which are usually examined in an anesthetized situation (Johns, 1985; Morimoto et al., 1989; Dibona and Sawin, 1984). In our previous experiment, the G F R was not altered by intravenous injection of NZ-105 in anesthetized SHRs (Yokoyama et al., 1992), while the G F R was increased in the clinical situation. To examine whether this discrepancy is due to anesthesia or differences in animal strains, we investigated the effects of NZ-105 for GFR, UV and RPF by using conscious SHRs. Consequently, as shown in Fig. 2, intravenous injection of NZ-105 (0.03 mg/kg) caused an increase in UV accompanied with a transient increment of GFR. However, the percent change of UV (105.4 ± 17.8%) was obviously much larger than that of G F R (38.5 + 14.0%). Although the exact cause of the difference between the case of conscious SHRs and anesthetized SHRs is unknown, it may be due to the lack of reflex via autonomic nervous system induced by anesthesia. The decrease of blood pressure followed by the decrease of intraglomerulus pressure raises the GFR, because of the tubuloglomerular feedback (TGF) mechanism in a conscious situation. However, in an anesthetized situation, G F R probably did not change due to the lack of T G F mechanism. We cannot exclude other possibilities such as NZ-105 having a differential effect on the afferent and efferent arteries, as speculated by Johns (1985). Our previous report using analysis of lithium clearance showed that intravenous injection of NZ105 inhibited the water and Na ÷ reabsorption mainly at the loop of Henle and the nephron segments beyond the proximal tubules. Therefore, the diuretic and natriuretic effects of NZ-105 were suggested to be caused mainly by the inhibition of reabsorption at the tubules rather than an increment of GFR. CONCLUSION These results show that the chronic treatment of NZ-105 causes diuretic and natriuretic effects and that these effects were not attenuated for the 12-week period, and that the mechanism of the action was mainly the inhibition of reabsorption at the nephron segments and partly the increase of GFR. Thus NZ-105 possessing these characteristics may be a beneficial drug in clinical situations. SUMMARY Efonidipine hydrochloride (NZ-105) is a 1,4-dihydropyridine derivative which has diuretic and natriuretic effects. In the present study, the effects of shortand long-term administration of NZ-105 were investigated in conscious spontaneously hypertensive
Renal effects of efonidipine rats (SHRs). Oral administration o f NZ-105 for 12 weeks caused diuretic and natriuretic effects, which were not attenuated during the experimental period. In the short-term experiment for investigating the mechanism of the diuretic effect, the intravenous injection of NZ-105 (0.03 mg/kg of body weight) significantly increased the urine volume (UV), renal plasma flow (RPF) and glomerular filtration rate (GFR). The increment rates of U V and R P F were 105.4_ 17.8% and I l l . 7 + 72.8%, respectively, which were larger than the increment rate of G F R (38.5_+ 14.0%). The diuretic or natriuretic effect of NZ-105 was suggested to have been caused, not only by the inhibition of sodium reabsorption but also, at least in part, the increase of G F R . In conclusion, chronic treatment of NZ-105 caused diuretic and natriuretic effects, and these effects were not attenuated for the 12-week period. The mechanism of action was mainly the inhibition of reabsorption at the nephron segments and partly the increase of G F R . NZ- 105 possessing these characteristics may be a beneficial drug in clinical situations. REFERENCES Abe Y., Komori T., Miura K., Takeda T., Imanishi M., Okahara T. and Yamamoto K. (1983) Effects of the calcium antagonist nicardipine on renal function and renin release in dogs. J. Cardiovasc. Pharmacol. 5, 254-259. Baba T., Ishizaki T., Murabayashi S., Aoyagi K., Tamasawa N. and Takabe K. (1987) Multiple oral doses of nicardipine, a calcium-entry blocker: effects on renal function, plasma renin activity, and aldosterone concen-
OP
26/2--H
337
tration in mild-to-moderate essential hypertension. C/in. Pharmacol. Ther. 42, 232-239. Dibona G. F. and Sawin L. L. (1984) Renal tubular site of action of felodipine. J. Pharmacol. exp. Ther. 228, 420-424. H/iberle D. A., Kawata T. and Davis J. M. (1987) The site of action of nitrendipine in the rat kidney. J. Cardiovasc. Pharmacol. 9 (Suppl. I), $17-$23. Johns E. J. (1985) The influence of diltiazem and nifedipine on renal function in the rat. Br. J. Pharmacol. 84, 707-713. Johns E. J. (1988) A study of the renal actions of amlodipine in the normotensive and spontaneously hypertensive rat. Br. J. Pharmacol. 94, 311-318. Ledingham J. G. G. (1981) Implications of antihypertensive therapy on sodium balance and sodium and water retension. Br. J. c/in. Pharmacol. 12, 15S-21S. Masuda Y., Takeguchi M., Arakawa C., Sakai T., Hibi M., Tanaka S., Shigenobu K. and Kasuya Y. (1990) Antihypertensive and diuretic effects of NZ-105, a novel dihydropyridine derivative. Arch. int. Pharmacodyn. 304, 247-264. Morimoto S., Ohyama T., Hisaki K. and Matsumura Y. (1989) Effects of CV-4093, a new dihydropyridine calcium channel blocker, on renal hemodynamics and function in stroke-prone spontaneously hypertensive rats (SHRSP). Jpn. J. Pharmacol. 51, 257-265. Nordlander M., Dibona G. F., Ljung B., Yao T. and Thoren P. (1985) Renal and cardiovascular effects of acute and chronic administration of felodipine to SHR. Eur. J. Pharmacol. 113, 25-36. Oizumi K., Nishino H., Miyamoto M., Fukushige J., Fukami M. and Koike H. (1989) Beneficial renal effects of CS-905, a novel dihydropyridine calcium blocker, in SHR. Jpn. J. Pharmacol. 51, 501-508. Yokoyama S. and Kaburagi T. (1983) Clinical effects of intravenous nifedipine on renal function. J. Cardiovasc. Pharmacol. 5, 67-71. Yokoyama T., Masuda Y., Sakai T., Tanaka S. and Tomita K. (1992) Effects of NZ-105, a new calcium antagonist, on renal function in anesthetized spontaneously hypertensive rats. J. Cardiovasc. Pharmacol. 19, 851-856.
0306-3623(94)00192-8
Gen. Pharmac. Vol.26, No. 2, pp. 333-337,1995 Copyright© 1995ElsevierScienceLtd Prin~d in Great Britain.All fights reserved 0306-3623/95$9.50+ 0.00
Effects of Efonidipine Hydrochloride (NZ-105), a Calcium Antagonist, on Renal Function in Conscious Spontaneously Hypertensive Rats T. Y O T S U M O T O , Y. M A S U D A , * C. S H U D O , H. S U G I T A , T. Y A M A S H I T A a n d S. T A N A K A Shiraoka Research Station of Biological Science, Nissan Chemical Industries Ltd., 1470, Shiraoka, Minamisaitama, Saitama, 349-02, Japan [Tel: (81)-480-92-2513; Fax: (81)-480-92-2516] (Received 20 June 1994)
Abstract--1. We investigated the effects of short- and long-term administration of efonidipine hydrochloride (NZ-105), 1,4-dihydropyridine derivative, in conscious spontaneously hypertensive rats (SHR). 2. Oral administration of NZ-105 for 12 weeks caused diuretic and natriuretic effects, which were not attenuated during the experimental period. 3. In the short-term experiment for investigating the mechanism of the diuretic effect, intravenous injection of NZ-105 (0.03 mg/kg of body weight) significantly increased the urine volume (UV), renal plasma flow (RPF) and glomerular filtration rate (GFR). The increment rate of UV and RPF was 105.4+ 17.8% and 111.7_+72.8%, respectively, which were larger than the increment rate of GFR (38.5 _+ 14.0%). 4. The diuretic or natriuretic effect of NZ-105 was suggested to be due to both the inhibition of sodium reabsorption and, at least in part. the increase of GFR. Key Words: NZ-105, calcium antagonist, renal function
INTRODUCTION Efonidipine hydrochloride (NZ-105), is a 1,4-dihydropyridine derivative which exerts a long-lasting hypotensive effect with a very slow onset (Masuda et al., 1990). We previously demonstrated that NZ-105 possessed diuretic and natriuretic effects by 15-day successive oral administration in spontaneously hypertensive rats (SHRs) (Masuda et al., 1990). Moreover, we have already reported that intravenous injection of NZ-105 in anesthetized SHR causes a diuretic effect without a change of the glomerular filtration rate (GFR) (Yokoyama et al., 1992). On the other hand, in clinical situations, NZ-105 revealed diuretic effects accompanied with an increase of G F R (personal communication, K. Taniguchi et al.).
*To whom all correspondence should be addressed.
In the present study, we further studied the above mentioned findings to explore the long-term diuretic effect and to investigate its mechanism. Firstly, we investigated the diuretic effects by oral administration of NZ-105 for 12 consecutive weeks in SHRs. Secondary, in conscious SHRs, we investigated the effects of intravenous injection on the renal hemodynamics to study the diuretic mechanism. MATERIALS AND METHODS Chronic experiment procedure Twenty-eight-week-old male SHRs (Charles River, Japan) were used. The SHRs were divided into three groups so that the systolic blood pressure (SBP) was evenly distributed. SBP was measured by the tail-cuff method (UR-5000, UEDA). The animals were given orally NZ-105 (5 and 20mg/kg) or solvent (0.5% methyl cellulose) once a day for 12 weeks. Food and water were given ad libitum. Every 4 weeks, urine was
333
334
T. Yotsumoto et al.
collected. Briefly, the rats were orally administered the drugs and loaded saline at the volume of 25 ml/kg body weight immediately after being placed in metabolism cages individually. The urine was collected for 6 hr in the absence of food and water intake. On the last day of the administration period (at 12 weeks), the urine was collected up to 6 hr and from 6 to 24 hr after the administration. Urine volume (UV) was measured gravimetrically, and urinary sodium (UNa ÷) or potassium (UK ÷) was analyzed by means of a flame photometer (Hitachi 710).
Short-term experiment procedure SHR (200-270 g, Charles River, Japan) were anesthetized with sodium pentobarbital (50 mg/kg, i.p.). Polyethylene catheters were placed into the bladder for collecting urine, and into the vena cava via the left femoral vein for infusion of inulin and p-aminohippuric acid (PAH). Other polyethylene catheters were placed into the abdominal aorta via the left femoral artery for withdrawal of blood. Except for the bladder catheter, the free ends of the catheters were exteriorized in the neck from under the skin. The wounds were surgically sutured. Each animal was placed in a Ballman cage and the experiments were started after they had recovered from anesthesia, at least 2 hr after surgery. A priming dose of inulin (200mg/kg) and PAH (8mg/kg) was given, followed by a sustaining infusion of isotonic saline containing inulin (36 mg/ml) and PAH (3 mg/ml) at a rate of 3.3 ml/hr. After an equilibration period (170rain), urine samples were collected for 20 min as a control period. After the control period, the drug or vehicle (5% cremophore-saline) was injected. Then urine samples were collected during two 20 rain clearance periods. In this experiment, arterial blood samples were obtained at the midpoint of the control period (20min before the drug or vehicle administration) and at each 20-rain clearance period. UV was determined gravimetrically. Urine and plasma samples were analyzed for inulin and PAH. Inulin concentrations in samples were quantified photometrically with anthrone. PAH was quantified photometrically with N-l-naphthylethylenediamine. The G F R and renal plasma flow (RPF) were estimated from the renal clearance of inulin and PAH, respectively.
Drugs NZ-105 was synthesized by the Central Research Laboratories of Nissan Chemical Industries Ltd. For
oral administration, NZ-105 was suspended in 0.5% methyl cellulose solution (Shinetsu Chemical, SM400) and administered at a volume of 2ml/kg through a gastric tube. For intravenous injection, NZ-105 was dissolved in 5% cremophore-saline (1 mg/ml), diluted with saline for the injection at a dose of 0.03 mg/kg. The solution was injected in a volume of 0.4 ml/kg via an administration catheter.
Statistical analysis All results were expressed as the mean + SEM. In long-term experiment results, statistical analyses were performed using the Bartlett test of variance. Parametrical analyses were performed by one-way analysis of variance. If these analysis results were significant, comparisons among groups were analyzed by Tukey's multiple method. On the other hand, non-parametrical analysis was performed with the Kruskal-Wallis test. If significant, it was performed using Tukey's method. For acute experiment results, statistical analysis was performed using the F-test of variance, followed by the Student's t-test or Aspin-Welch's t-test.
RESULTS
Chronic experiment Figure 1 shows the effect of chronic administration of NZ-105 for UV, UNa + and UK + in SHRs. The UV was significantly increased by the administration of 20 mg/kg of NZ-105 daily for 12 weeks, and by that of 5 mg/kg for 12 weeks compared with the control. Similarly the UNa + was also significantly increased at the 12th week for the high close of NZ-105, and at the 4th and 12th weeks for the low dose of NZ-105. The UK + was not significantly changed, except for the 12th week value for the high dose of NZ-105. The diuretic and natriuretic effects of NZ-105 were not attenuated during the 12-week period of the repeated administration. Table l shows the UV and the amount of urinary electrolyte excretion (UNa +, U K +) for the 0-6 hr, 6-24 hr and 0-24hr period at the 12th week after chronic administration of NZ-105. NZ-105 caused a significant and dose-dependent increase in UV, and the amount of UNa + in the 0-6 hr period, compared with the control group. UK + was significantly increased only by the high dose of NZ-105. Each parameter determined for the 6-24 hr urine tended to decrease compared with the 0 - 6 h r urine of the NZ-105 treated rats, which were slightly lower than those of the control group.
Renal effects of efonidipine Acute experiment Figure 2 shows the effects of intravenous injection of NZ-105 on UV, R P F and G F R in the conscious SHR. The U V was significantly increased in the first and second clearance periods after administration of the drug, compared with the control group. The R P F and G F R were significantly increased in the first clearance period, but not the second period. The incremental rate of UV, R P F and G F R from - 2 0 - 0 min (control period) to the 0-20 min (the first clearance period) was 105.4 + 17.8%, 111.7 + 72.8% and 38.5 + 14.0%, respectively.
335
Table I. The effect of the vehicle or efonidipine hydrochloride (NZ-105) for UV, the amount of UNa+V and U K + V in the 0 ~ , 6-24 and 0-24 hr period in SHR after administration for 12 consecutive weeks Drug (mg/kg)
Time (hr)
UV (ml/kg)
UNa÷V (,u Eq/kg)
UK÷V (/~Eq/kg)
Control
0~ Total
6.4 _+ 1.5 12.7_+2.0 19.1 _+ 3.1
839.1 _+ 204.5 1702.8 -+ 157.2 2542.0 _+ 306.6
857.9 _+ 176.9 1614.0_+139.5 2471.9 _+ 266.4
NZ-105 15]
~6 6-24 Total
17.5 _+ 1.3"* 9.8_+0.7 27.3_+ 1.5
2220.0 _+ 237.0** 1052.3 + 104.1 1310.0_+ 120.9 1313.2_+ 116.6 3530.1 _+ 233.6* 2365.4_+ 160.4
NZ-105 [201
0-6 6-24 Total
3 0 . 4 + 1.8"* 3738.4+232.1"* 1751.5_+99.7"* 12.4_+1.2 1197.2 + 116.1 1480.3_+89.4 42.8_+ 1,5"* 4935.6_+ 173.4"* 3231.7_+ 121.1"
0.5%MC 6-24
All values are the mean _+ SEM from 11 to 13 animals. *P < 0.05, **P < 0.01, compared with the values of the control group.
DISCUSSION
2°1
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or
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8
12
weeks
Fig. 1. Urine volume, urinary Na ÷ and K* excretion for a 6-hr period on the day before and 4, 8 and 12 weeks after repeated administration of efonidipine hydrochloride (NZ105) for up to 12 weeks. NZ-105 was orally administered at 5 mg/kg ([], n = I 1), 20 ( , , n = 13), or vehicle (K], n = 13) in conscious spontaneously hypertensive rats. *P < 0.05, **P < 0.01; significantly different from each control value.
It is well known that chronic treatment with vasodilators, such as hydralazine and minoxidil, cause renal sodium and water retention (Ledingham, 1981). To prevent edema, the simultaneous use of diuretics is needed in a clinical situation. On the other hand, it has been shown that several 1,4-dihydropyridine class calcium antagonists have not only an anti-hypertensive effect by their direct vasodilating action, but also diuretic and natriuretic effects (Johns, 1985, 1988; Abe et al., 1983; M o r i m o t o et al., 1989; Oizumi et al., 1989), which are beneficial characteristics for an anti-hypertensive drug. We have already shown that NZ-105 caused antihypertensive and diuretic effects, and that those effects of NZ-105, but not nicardipine, were attenuated for a 15-day period in conscious SHR. In the present study, NZ-105 caused a diuretic and natriuretic effect which is consistent with the previous result, which was not attenuated for 12 weeks of repeated administration (Masuda et al., 1990). Nordlander et al. (1985) reported that the acute administration of felodipine caused a diuretic and natriuretic effect in SHRs, while chronic administration for 6 months maintained a diuretic effect, but not a natriuretic effect. They speculated that the reason may be a readjustment for the natriuretic effect induced by chronic administration of felodipine. Our present findings indicate that the readjustment for the natriuretic effect of NZ-105 may be weaker than that of felodipine. On the other hand, NZ-105 caused a marked diuretic effect during the 0 - 6 hr period, and this effect tended to decrease at the 6-24 hr period compared with the control group (Table 1). This may be due to a compensatory mechanism for the diuretic effect caused during the early period (0-6 hr).
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2..~0-
•~. 150 ~,~ I0050-
60-
"~ 40-
20-
0 14
°mm
m
-20-0
0-20
20-40
Time (min) Fig. 2. Effects of intravenous administration of efonidipine hydrochloride (NZ-105) on urine volume (UV), renal plasma flow (RPF) and glomerular filtration rate (GFR) in conscious spontaneously hypertensive rats. NZ-105 was administered at 0.03 mg/kg (m, n = 5), and vehicle (I:], n = 6). *P < 0.05; significantlydifferent from each control value.
The mechanism for diuretic and natriuretic effects of 1,4-dihydropyridine class calcium antagonists has been reported to be an increase of G F R (Abe et al., 1983) and/or inhibition of water and sodium reabsorption at the proximal tubules (H~iberle et al., 1987), distal tubules and/or collecting ducts (Dibona and Sawin, 1984). In recent years, there have been many reports showing that 1,4-dihydropyridine class calcium antagonists increased G F R in humans (Baba et al., 1987; Yokoyama and Kaburagi, 1983), but not
in rats, which are usually examined in an anesthetized situation (Johns, 1985; Morimoto et al., 1989; Dibona and Sawin, 1984). In our previous experiment, the G F R was not altered by intravenous injection of NZ-105 in anesthetized SHRs (Yokoyama et al., 1992), while the G F R was increased in the clinical situation. To examine whether this discrepancy is due to anesthesia or differences in animal strains, we investigated the effects of NZ-105 for GFR, UV and RPF by using conscious SHRs. Consequently, as shown in Fig. 2, intravenous injection of NZ-105 (0.03 mg/kg) caused an increase in UV accompanied with a transient increment of GFR. However, the percent change of UV (105.4 ± 17.8%) was obviously much larger than that of G F R (38.5 + 14.0%). Although the exact cause of the difference between the case of conscious SHRs and anesthetized SHRs is unknown, it may be due to the lack of reflex via autonomic nervous system induced by anesthesia. The decrease of blood pressure followed by the decrease of intraglomerulus pressure raises the GFR, because of the tubuloglomerular feedback (TGF) mechanism in a conscious situation. However, in an anesthetized situation, G F R probably did not change due to the lack of T G F mechanism. We cannot exclude other possibilities such as NZ-105 having a differential effect on the afferent and efferent arteries, as speculated by Johns (1985). Our previous report using analysis of lithium clearance showed that intravenous injection of NZ105 inhibited the water and Na ÷ reabsorption mainly at the loop of Henle and the nephron segments beyond the proximal tubules. Therefore, the diuretic and natriuretic effects of NZ-105 were suggested to be caused mainly by the inhibition of reabsorption at the tubules rather than an increment of GFR. CONCLUSION These results show that the chronic treatment of NZ-105 causes diuretic and natriuretic effects and that these effects were not attenuated for the 12-week period, and that the mechanism of the action was mainly the inhibition of reabsorption at the nephron segments and partly the increase of GFR. Thus NZ-105 possessing these characteristics may be a beneficial drug in clinical situations. SUMMARY Efonidipine hydrochloride (NZ-105) is a 1,4-dihydropyridine derivative which has diuretic and natriuretic effects. In the present study, the effects of shortand long-term administration of NZ-105 were investigated in conscious spontaneously hypertensive
Renal effects of efonidipine rats (SHRs). Oral administration o f NZ-105 for 12 weeks caused diuretic and natriuretic effects, which were not attenuated during the experimental period. In the short-term experiment for investigating the mechanism of the diuretic effect, the intravenous injection of NZ-105 (0.03 mg/kg of body weight) significantly increased the urine volume (UV), renal plasma flow (RPF) and glomerular filtration rate (GFR). The increment rates of U V and R P F were 105.4_ 17.8% and I l l . 7 + 72.8%, respectively, which were larger than the increment rate of G F R (38.5_+ 14.0%). The diuretic or natriuretic effect of NZ-105 was suggested to have been caused, not only by the inhibition of sodium reabsorption but also, at least in part, the increase of G F R . In conclusion, chronic treatment of NZ-105 caused diuretic and natriuretic effects, and these effects were not attenuated for the 12-week period. The mechanism of action was mainly the inhibition of reabsorption at the nephron segments and partly the increase of G F R . NZ- 105 possessing these characteristics may be a beneficial drug in clinical situations. REFERENCES Abe Y., Komori T., Miura K., Takeda T., Imanishi M., Okahara T. and Yamamoto K. (1983) Effects of the calcium antagonist nicardipine on renal function and renin release in dogs. J. Cardiovasc. Pharmacol. 5, 254-259. Baba T., Ishizaki T., Murabayashi S., Aoyagi K., Tamasawa N. and Takabe K. (1987) Multiple oral doses of nicardipine, a calcium-entry blocker: effects on renal function, plasma renin activity, and aldosterone concen-
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