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Effects of atrial natriuretic peptide on adrenergically induced norepinephrine release and vasoconstriction in the dog kidney
Eu~~~Q~ J5w-d of Phur~ucff~o~, 199 11991) 255-258 0 I991 Elsevier Science Publishers I3.V. ~14-2~9/9f/SO3.50 ~D~~~~ 0014299~1004608
EJP 20847
Short communication
Received 19 April 1991, accepted I May 1991
The effects of atria1 natriuretic peptide (ANPI on the neural control of renal blood flow were examined in anesthetized dogs, Intrarenal arterial infusion of ANP (a-hANP, 10 and 50 ng/kg per min) suppressed the decreas.. in renal blood flow but not the increase in renal venous plasma norepinephrine concentration induced by renal nerve stimulation 0 and 2 Hz, for 1 min) ?.NP also attenuated the blood flow response to intrarenal arterial injection of methoxamine U-20 pg). These results sugge< hat ANP acts at a postsynaptic site to suppress adrenergically induced vasoconstriction in the dog kidney.
ANP (atria1 natriuretic pol~eptide);
Kidney; Norepinephrine
1. Introduction
Atria1 natriuretic peptide (ANP), a potent natriuretic and vasodilatory humoral factor (Go&z, 19881, has been reported to inhibit vasoconstriction and neurotransmitter release induced by adrenergic stimuli. ANP suppresses pressor responses to a-adrenoceptor agonists in pithed rats (Haass et al., 1985; ZukowskaGrojec et al., 1986). ANP also attenuates the elevation of plasma norepinephrine (NE) levels induced by spinal cord stimulation in pithed rats ~Zukowska-Grojec et al., 1986) and 13H]NE overflow evoked by periarterial nerve stimulation in the isolated rat mesenteric vascuIature (Nakamura and Inagami, 1986). However, we found that in the dog kidney in vivo ANP failed to affect NE release during renal nerve stimulation at a Iow frequency that caused little change in renal hemodynamics (Hisa et al., 1989). A recent report has also shown that ANP does not exert an inhibitory effect on NE reiease, whereas it attenuate? the elevation of perfusion pressure induced by periarterial nerve stimulation in the isofated rat kidney (Schwartz and Eikenburg, 1991). In the present study, we examined further the possible inhibitory actions of ANP on the neural control of vascular tone in the dog kidney. The effects of ANP on NE release, the decrease in renal blood ffow induced by graded renal nerve stimulation, and the blood flow
Correspondence to: H. Hisa, Department of Pharmacology, Pharmaceutical Institute, Tohoku University, Aobayama, Sendai 980, Japan.
release: Vasoconstriction
response to an cu,-adrenoceptor agonist, methoxamine, were assessed in anesthetized dogs.
2. Materials and methods 2.1. Animal preparation
Mongrel dogs of either sex, weighing 9-19 kg, were anesthetized with sodium pentobarbitai (30 mg/kg i-v.), then intubated and ventilated a~i~cially with room air. Decamethonium bromide (0.25 mg,/kg i.v.) was given to prevent spontaneous active respiratory movement. Anesthesia was maintained with a continuous i.v. infusion of sodium pentobarbital at a rate of 5 mg/kg per h throughout the experiments. The right brachial artery was cannulated to measure systemic blood pressure with a pressure transducer (Nihon Kohden Co., Tokyo, Japan, TP-200T). The left kidney was exposed by a retroperitoneal ffank incision. AI? visible renal nerves were dissected away from the renal vessels and cut after ligation. An electromagnetic flow probe (2.5-3.5 mm in diameter, Nihon Kohden) was attached at the renal artery to measure renal b!ood flow with a square-wave flowmeter (Nihon Kohden, MF-27). A curved 25-gauge needle connected to a polyethylene tube was inserted into the renal artery for A??P infusion. In methoxamine experiments, a second 25gauge needle was placed in the renal artery for drug injection. In renal nerve stimulation experiments, platinum eiectrodes were placed on the nerve bundles and a catheter was inserted into the renal vein via the go-
nadal vein to collect renal venous blood samples. Heart rate was rno~~to~d with an ele~tr~ardiograph. Mean arterial pressure, heart rate and renal blood flow were recorded with a polygraph system CNihon Kohden). After ~mpletiox~ of surgery, 60-90 min were allowed for SFabilizatio~. The animals were divided into three groups.
Renal nerve stimulation (duration, 1 ms; supramaximal v&age, 10-20 VI for 1 min at 1 and 2 Hz (Group It n = 7) or an intrarenal arterial bolus injection of methoxamine (Nippon Shinyaku, Kyoto, Japan) at 510 and 20 kg (Group 2, n = 7) was applied at 7- to lo-min intervals. Renal venous blood for the determination of plasma NE concentrations was withdrawn before and during the last 10 s of each renal nerve stimulation. Five minutes after the control responses had been obtained, ANP (&ANP, Osaka Protein Research Foundation, Osaka, Japan) was infused into the renal artery, using a motor-driven infusion pump (Harvard Apparatus Co., Inc., Millis, MA, 97.51, at increasing rates of 10 and 50 ng/kg per min. Ten minutes later, renaI nerve stimulation and bled sapling or methoxamine injection was repeated during ANP infusion at each dose. In Group 3 (n = 61, prazosin (0.2 mg/kg; Sigma Co., St. Louis, MO, U.S.A.) was injected iv. and, 15-20 min later, the effect of ANP (50 ng/kg per min) on the NE release response to 2-Hz renal nerve stimulation was examined as in Group 1.
3. Results In renal nerve stimulation experiments (Group 0, basal hemodynamic values (obtained before the start of l-Hz renal nerve sti~nulation) in the control period were mean arterial pressure, 110 + 10 mm Hg; heart rate, 117 t 11 bpm; renal blood flow, 138 f 18 ml/min. Basal values for the renal venous plasma NE concentration and the renal NE efflux were 0.10 + 0.02 ng/ml and 8.1 it 2.8 ng/min, respectively. Intrarenal arterial infusion of ANP reduced basal renal blood flow and mean arterial pressure slightly (renal blood flow, 129 +_ 16 and 128 & 16 ml/min; mean arterial pressure, 107 f 10 and 103 19 mm Hg; during ANP infusion at 10 and 50 ng/kg per min, respectively). Heart rate, plasma NE concentration and NE efflux remained unchanged. Renal nerve stimulation reduced renal blood flow and increased renal venous plasma NE ~ncentrations and renal efflux of NE in a frequency-dependent manner (Group 1, fig. 1A). The decrease in renal blood flow induced by ~-HZ renal nerve stimulation was attenuated during ANP infusion at 10 ng/kg per min. ANP infusion at 50 ng/kg per min si~ifi~ant~y suppressed the response of renal blood flow to l- and ~-HZ renal nerve stimulation. Even at 50 ng/kg per min, ANP failed to affect the nerve stimulation-in-
2.3. Meusureinems Blood samples were transferred to chilled tubes containing disodium EDTA (2 mg/ml of blood) and then centrifuged to obtain plasma samples. Catecholamines were extracted from plasma by the alumina absorption method, and plasma NE concentrations were determined by high-performance liquid chromato~aphy with an amperometric detector CBioanalytical Systems, LC-304!, as described previously (Hayashi et al., 19871. The renal efflux of NE was calculated by multipl~ng the renal venous plasma NE concentration tjl the renal plasma flow.
0
WP
60
lug
40
$t!
20 0
2.4. Data analysis r?91values are expressed as means + SE. Effects of ANP on tne blood flow and NE release responses were analyzed by an analysis of variance for multifactor repeated measures. Dunnett’s test was used for multiple comparisons between the control value and the value obtained during ANP infusion at each dose. Differences at a P value less than 0.05 were considered to be statisticafly significant.
Fig. 1. Effects of atrial natriuretic peptide on renal va~onsfrjct~on and norepinephrine release induced by renal nerve stimulation (A, Group 1, n = 7) and on renal vasoconstriction induced by intrarenal arterial injection of methoxamiae (B, Group 2, n = 7). Values are means+S.E. RNS, renal nerve stimulation; MTX, methoxamine; d, changes from basal values; RBF, renal blood flow; NEC, renal venous plasma norep~nephrine concentration; NEE, renal norepinephrine efflux; ANP. atria1 natriuretic peptide (ru-hANP). ANP was infused into the renal artery at increasing rates of 10 and 50 ngfkg per min. * P < 0.05 compared with corresponding control (dose zero) values.
257 TABLE 1 Effects of atria1 natriuretic peptide on norepinephrine release induced by renal nerve stimulation in the presence of prazosin (Group 3). Values are means +S.E. n = 6. A, changes from basal values in response to renal nerve stimulation (2 Hz); RBF, renal blood flow; NEC, renal venous plasma norepinephrine con~ntration; NEE, renal norepinephrine efflux; ANP, atrial natriuretic peptide Icu-hANP). Prazosin (0.2 mg/kg) was given iv. 15-20 min before the start of the experiments. ANP was infused into the renal artery at 50 ng/kg per min.
Control ANP
ARBF (ml/mitt)
ANEC
Ins/ml)
ANEE tng/min)
-i2*2 -10*2
1.32+025 1.09 f 0.25
45.0+3.4 43.5 + 3.6
duced increase in plasma NE concentration. The increase in NE efflux induced by Z-Hz renal nerve stimulation tended to be potentiated during ANP infusion, but the change was not statistically significant. Intrarenal bolus injection of methoxamine decreased renal blood flow, which was also suppressed dose dependently by ANP infusion (Group 2, fig. 1B). The reproducibility of the blood flow rtsponses to the consecutive application of nerve stimulation and methoxamine injection has been confirmed in a previous study from our laboratory (Chiba et al., 1990). In pr~osin-pretreated animals (Group 31, Z-Hz renal nerve stimulation caused a very slight decrease in renal blood flow (table 1). ANP infusion did not cause a further suppression of the blood flow response to nerve stimulation. The nerve stimulation-induced increase in plasma NE concentration or NE efflux was unaffected during ANP infusion.
4. Discussion ANP has been suggested to suppress adrenergicaliy induced vasoconstriction and neural NE reiease in whole animals and in isolated blood vessels (Haass et al., 1985; Zukowska-Grojec et al., 1986; Nakamaru and Inagami, 1986). In the in vivo dog kidney, however, we have previously shown that intrarenal infusion of ANP at 10 ng/kg per min fails to affect NE release during renal nerve stimulation at 1 Hz for 10 min (Hisa et al., 1989). To examine further the possible inhibitory effects of ANP on the renal sympathetic nervous system, we used a higher dose of ANP (50 ng/kg per min) and a higher frequency of renal nerve stimulation (2 Hz) in the present study. Nerve stimulation was pe~ormed for a short period (1 min), because the decreased renal blood flow recovers during continuous nerve stimulation, probably because of the production of vasodilatory prostaglandins (Hayashi et al., 19871, which would
complicate the interpretation of the vascular effect of ANP. lntrarenai infusion of ANP at 10 and 50 ng/kg per min dose dependently attenuated the decre;se in renal blood flow induced by renal nerve stimulation. This result, whjch confirms our recent obse~atjo~ (Hisa et al., 1990), suggests that ANP interferes with the neural control of vascular tone in the dog kidney. In contrast, the nerve stimulation-induced increase in renal venous plasma NE concentration was unaffected during ANP infusion. ANP tended to potentiate the increase in renal NE efflux induced by 2-k renal nerve stjmulation. This may be due to the substantially attenuated blood flow response, since ANP did not affect the NE efflux response when renal vasoconstriction was almost abolished by pretreatment with prazosin. A study carried out in our laboratory has demonstrated that preparations of the dog kidney in vivo are sensitive to the presynaptic inhibitory effects of humorai factors such as adenosine (Yoneda et al., 1990). Thus. ANP does not seem to exert a modulatory action on neural NE release in the dog kidney. ANP may act on postsynaptic sites to inhibit the neurally induced renal vasoconstriction recently shown in the pump-perfused rat kidney (Schwartz and Eikenburg, 1991). It is generally accepted that neurally released NE causes vasoconstriction via activation of tu,-adrenoceptors. In our study, ANP attenuated the renal blood flow response to intrarenal injection of methoxamine, which preferentially activates tr,-adrenoceptors in the canine renal vasculature (Chiba et al., 1990). This observation suggests that ANP inhibits the postsynaptic Eu,-adrenoceptor mechanism, thereby regulating the neural control of renal vascular tone. However, studies performed with pithed rats have demonstrated that ANP does not inhibit cur-adrenoceptor-mediated systemic vasoconstriction (Haass et al., 1985; Zukowska-Grojec et al., 1986). Although ANP slightly suppressed the pressor response io sympathetic stimulation in yohimbine-treated rats, this was related to a blunted NE release response by a presynaptic action of ANP (Zukowska-Grojec et al., 1986). We observed, however, that even in the presence of yohimbine ANP attenuated the nerve stimulation-induced renal blood flow response without suppressing the NE release response (data are not shown). These discrepancies may be ascribable to differences in the systemic vs. the regional (renal) sympathetic nervous system. Although the present study focuses on the neural control of vascular tone, it should be noted that ANP also inhibits angiotensin II-induced renal vasoconstriction in dogs (Hisa et al,, 1990) and in rats (Schwartz and Eikenburg, 1991). ANP therefore does not seem to be specific for ar-adrenoceptor mechanisms. ANP could act on common pathway(s) mediating vasoconstriction in response to neural and humoral stimuli in the kid-
a~ismby \vhicl~ ANP inhibits renal vasortey. ~~s~r~ct~~~ ~a~aot be ~ctermined from the present study. This issue requires further elucidation. iBn~~c~~s~~~~,the present study demonstrates that ~~~~ea~~s~y a~rn~~~stered ANP can suppress neurally ~~~~ced ~~s~o~strict~o~ without affecting neurotransmitter release in the dog kidney in vivo.
This work was supported in part by a Grant-in-Aid for Scientific Research Japan) No. 02771695 and by the Sapporo Bioscience Foundation (Japan). Part of this work was presented at the 11th International Congress of Pharmacology. Amsterdam, 1990.
rences Chiha. K.. Y. Hayashi. H. Hisa. M. Suzuki-Kusaba and S. Satoh, W&l. Effects of a novel ru,-adrenoceptor antagonist. SGB-1534, on adrenergically induced renal vasoconstriction in dogs. Eur. J. Phxmxol. i76. 263. Goetz, K.L.. l9SS. Physiology and pathology of atrial peptides. Am. J. Phpiol. 2%. El.
Haass, M., I.J. Kopin, D.S. Goldstein and Z. Zukowska~Grojec, 1985, Differential inhibition of alpha adrenoceptor-mediated pressor responses by rat atrial natriuretic peptide in the pithed rat, J. Pharmacol. Exp. Ther. 235, 122. Hayashi, Y.. H. Hisa and S. Satoh. 1987, Role of prostaglandin in norepinephrine and renin release in canine kidney, Am. J. Physiol. 253. F929. Hisa, H., Y. Tomura and S. Satoh 1989, Atrial natriuretic factor suppresses neural stimulation of renin release in dogs, Am. J. Physiol. 257, E332. Hisa, H., Y. Tomun, A. Takahara, K. Chiba and S. Satoh, 1990, Atria1 natriuretic peptide suppresses renal vasoconstriction induced by angiotensin II, norepinephrine and renal nerve stimulation in dogs, Eur. J. Pharmacol. 183, 1063 (Abstract). Nakamaru, M. and T. Inagami, 1986, Atria1 natriuretic factor inhibits norepinephrine release evoked by sympathetic nerve stimulation in isolated perfused rat mesenteric arteries, Eur. J. Pharmacol. 123, 459. Schwartz, D.D. and D.C. Eikenburg, 1991, Effects of synthetic ANF (rANF(3-28)) on sympathetic neurotransmission in the isolated perfused rat kidney, Life Sci. 48, 781. Yoneda, H., H. Hisa and S. Satoh, 1990, Effects of adenosine on adrenergically induced renal vasoconsriction in dogs, Eur. J. Pharmacol. 176, 109. Zukowska-Grojec. Z., M. Haass, I.J. Kopin and N. Zamir, 1986, Interaction of atrial natriuretic peptide with the sympathetic and endocrine systems in the pithed rat, J. Pharmacol. Exp. Ther. 239, 480.
EJP 20847
Short communication
Received 19 April 1991, accepted I May 1991
The effects of atria1 natriuretic peptide (ANPI on the neural control of renal blood flow were examined in anesthetized dogs, Intrarenal arterial infusion of ANP (a-hANP, 10 and 50 ng/kg per min) suppressed the decreas.. in renal blood flow but not the increase in renal venous plasma norepinephrine concentration induced by renal nerve stimulation 0 and 2 Hz, for 1 min) ?.NP also attenuated the blood flow response to intrarenal arterial injection of methoxamine U-20 pg). These results sugge< hat ANP acts at a postsynaptic site to suppress adrenergically induced vasoconstriction in the dog kidney.
ANP (atria1 natriuretic pol~eptide);
Kidney; Norepinephrine
1. Introduction
Atria1 natriuretic peptide (ANP), a potent natriuretic and vasodilatory humoral factor (Go&z, 19881, has been reported to inhibit vasoconstriction and neurotransmitter release induced by adrenergic stimuli. ANP suppresses pressor responses to a-adrenoceptor agonists in pithed rats (Haass et al., 1985; ZukowskaGrojec et al., 1986). ANP also attenuates the elevation of plasma norepinephrine (NE) levels induced by spinal cord stimulation in pithed rats ~Zukowska-Grojec et al., 1986) and 13H]NE overflow evoked by periarterial nerve stimulation in the isolated rat mesenteric vascuIature (Nakamura and Inagami, 1986). However, we found that in the dog kidney in vivo ANP failed to affect NE release during renal nerve stimulation at a Iow frequency that caused little change in renal hemodynamics (Hisa et al., 1989). A recent report has also shown that ANP does not exert an inhibitory effect on NE reiease, whereas it attenuate? the elevation of perfusion pressure induced by periarterial nerve stimulation in the isofated rat kidney (Schwartz and Eikenburg, 1991). In the present study, we examined further the possible inhibitory actions of ANP on the neural control of vascular tone in the dog kidney. The effects of ANP on NE release, the decrease in renal blood ffow induced by graded renal nerve stimulation, and the blood flow
Correspondence to: H. Hisa, Department of Pharmacology, Pharmaceutical Institute, Tohoku University, Aobayama, Sendai 980, Japan.
release: Vasoconstriction
response to an cu,-adrenoceptor agonist, methoxamine, were assessed in anesthetized dogs.
2. Materials and methods 2.1. Animal preparation
Mongrel dogs of either sex, weighing 9-19 kg, were anesthetized with sodium pentobarbitai (30 mg/kg i-v.), then intubated and ventilated a~i~cially with room air. Decamethonium bromide (0.25 mg,/kg i.v.) was given to prevent spontaneous active respiratory movement. Anesthesia was maintained with a continuous i.v. infusion of sodium pentobarbital at a rate of 5 mg/kg per h throughout the experiments. The right brachial artery was cannulated to measure systemic blood pressure with a pressure transducer (Nihon Kohden Co., Tokyo, Japan, TP-200T). The left kidney was exposed by a retroperitoneal ffank incision. AI? visible renal nerves were dissected away from the renal vessels and cut after ligation. An electromagnetic flow probe (2.5-3.5 mm in diameter, Nihon Kohden) was attached at the renal artery to measure renal b!ood flow with a square-wave flowmeter (Nihon Kohden, MF-27). A curved 25-gauge needle connected to a polyethylene tube was inserted into the renal artery for A??P infusion. In methoxamine experiments, a second 25gauge needle was placed in the renal artery for drug injection. In renal nerve stimulation experiments, platinum eiectrodes were placed on the nerve bundles and a catheter was inserted into the renal vein via the go-
nadal vein to collect renal venous blood samples. Heart rate was rno~~to~d with an ele~tr~ardiograph. Mean arterial pressure, heart rate and renal blood flow were recorded with a polygraph system CNihon Kohden). After ~mpletiox~ of surgery, 60-90 min were allowed for SFabilizatio~. The animals were divided into three groups.
Renal nerve stimulation (duration, 1 ms; supramaximal v&age, 10-20 VI for 1 min at 1 and 2 Hz (Group It n = 7) or an intrarenal arterial bolus injection of methoxamine (Nippon Shinyaku, Kyoto, Japan) at 510 and 20 kg (Group 2, n = 7) was applied at 7- to lo-min intervals. Renal venous blood for the determination of plasma NE concentrations was withdrawn before and during the last 10 s of each renal nerve stimulation. Five minutes after the control responses had been obtained, ANP (&ANP, Osaka Protein Research Foundation, Osaka, Japan) was infused into the renal artery, using a motor-driven infusion pump (Harvard Apparatus Co., Inc., Millis, MA, 97.51, at increasing rates of 10 and 50 ng/kg per min. Ten minutes later, renaI nerve stimulation and bled sapling or methoxamine injection was repeated during ANP infusion at each dose. In Group 3 (n = 61, prazosin (0.2 mg/kg; Sigma Co., St. Louis, MO, U.S.A.) was injected iv. and, 15-20 min later, the effect of ANP (50 ng/kg per min) on the NE release response to 2-Hz renal nerve stimulation was examined as in Group 1.
3. Results In renal nerve stimulation experiments (Group 0, basal hemodynamic values (obtained before the start of l-Hz renal nerve sti~nulation) in the control period were mean arterial pressure, 110 + 10 mm Hg; heart rate, 117 t 11 bpm; renal blood flow, 138 f 18 ml/min. Basal values for the renal venous plasma NE concentration and the renal NE efflux were 0.10 + 0.02 ng/ml and 8.1 it 2.8 ng/min, respectively. Intrarenal arterial infusion of ANP reduced basal renal blood flow and mean arterial pressure slightly (renal blood flow, 129 +_ 16 and 128 & 16 ml/min; mean arterial pressure, 107 f 10 and 103 19 mm Hg; during ANP infusion at 10 and 50 ng/kg per min, respectively). Heart rate, plasma NE concentration and NE efflux remained unchanged. Renal nerve stimulation reduced renal blood flow and increased renal venous plasma NE ~ncentrations and renal efflux of NE in a frequency-dependent manner (Group 1, fig. 1A). The decrease in renal blood flow induced by ~-HZ renal nerve stimulation was attenuated during ANP infusion at 10 ng/kg per min. ANP infusion at 50 ng/kg per min si~ifi~ant~y suppressed the response of renal blood flow to l- and ~-HZ renal nerve stimulation. Even at 50 ng/kg per min, ANP failed to affect the nerve stimulation-in-
2.3. Meusureinems Blood samples were transferred to chilled tubes containing disodium EDTA (2 mg/ml of blood) and then centrifuged to obtain plasma samples. Catecholamines were extracted from plasma by the alumina absorption method, and plasma NE concentrations were determined by high-performance liquid chromato~aphy with an amperometric detector CBioanalytical Systems, LC-304!, as described previously (Hayashi et al., 19871. The renal efflux of NE was calculated by multipl~ng the renal venous plasma NE concentration tjl the renal plasma flow.
0
WP
60
lug
40
$t!
20 0
2.4. Data analysis r?91values are expressed as means + SE. Effects of ANP on tne blood flow and NE release responses were analyzed by an analysis of variance for multifactor repeated measures. Dunnett’s test was used for multiple comparisons between the control value and the value obtained during ANP infusion at each dose. Differences at a P value less than 0.05 were considered to be statisticafly significant.
Fig. 1. Effects of atrial natriuretic peptide on renal va~onsfrjct~on and norepinephrine release induced by renal nerve stimulation (A, Group 1, n = 7) and on renal vasoconstriction induced by intrarenal arterial injection of methoxamiae (B, Group 2, n = 7). Values are means+S.E. RNS, renal nerve stimulation; MTX, methoxamine; d, changes from basal values; RBF, renal blood flow; NEC, renal venous plasma norep~nephrine concentration; NEE, renal norepinephrine efflux; ANP. atria1 natriuretic peptide (ru-hANP). ANP was infused into the renal artery at increasing rates of 10 and 50 ngfkg per min. * P < 0.05 compared with corresponding control (dose zero) values.
257 TABLE 1 Effects of atria1 natriuretic peptide on norepinephrine release induced by renal nerve stimulation in the presence of prazosin (Group 3). Values are means +S.E. n = 6. A, changes from basal values in response to renal nerve stimulation (2 Hz); RBF, renal blood flow; NEC, renal venous plasma norepinephrine con~ntration; NEE, renal norepinephrine efflux; ANP, atrial natriuretic peptide Icu-hANP). Prazosin (0.2 mg/kg) was given iv. 15-20 min before the start of the experiments. ANP was infused into the renal artery at 50 ng/kg per min.
Control ANP
ARBF (ml/mitt)
ANEC
Ins/ml)
ANEE tng/min)
-i2*2 -10*2
1.32+025 1.09 f 0.25
45.0+3.4 43.5 + 3.6
duced increase in plasma NE concentration. The increase in NE efflux induced by Z-Hz renal nerve stimulation tended to be potentiated during ANP infusion, but the change was not statistically significant. Intrarenal bolus injection of methoxamine decreased renal blood flow, which was also suppressed dose dependently by ANP infusion (Group 2, fig. 1B). The reproducibility of the blood flow rtsponses to the consecutive application of nerve stimulation and methoxamine injection has been confirmed in a previous study from our laboratory (Chiba et al., 1990). In pr~osin-pretreated animals (Group 31, Z-Hz renal nerve stimulation caused a very slight decrease in renal blood flow (table 1). ANP infusion did not cause a further suppression of the blood flow response to nerve stimulation. The nerve stimulation-induced increase in plasma NE concentration or NE efflux was unaffected during ANP infusion.
4. Discussion ANP has been suggested to suppress adrenergicaliy induced vasoconstriction and neural NE reiease in whole animals and in isolated blood vessels (Haass et al., 1985; Zukowska-Grojec et al., 1986; Nakamaru and Inagami, 1986). In the in vivo dog kidney, however, we have previously shown that intrarenal infusion of ANP at 10 ng/kg per min fails to affect NE release during renal nerve stimulation at 1 Hz for 10 min (Hisa et al., 1989). To examine further the possible inhibitory effects of ANP on the renal sympathetic nervous system, we used a higher dose of ANP (50 ng/kg per min) and a higher frequency of renal nerve stimulation (2 Hz) in the present study. Nerve stimulation was pe~ormed for a short period (1 min), because the decreased renal blood flow recovers during continuous nerve stimulation, probably because of the production of vasodilatory prostaglandins (Hayashi et al., 19871, which would
complicate the interpretation of the vascular effect of ANP. lntrarenai infusion of ANP at 10 and 50 ng/kg per min dose dependently attenuated the decre;se in renal blood flow induced by renal nerve stimulation. This result, whjch confirms our recent obse~atjo~ (Hisa et al., 1990), suggests that ANP interferes with the neural control of vascular tone in the dog kidney. In contrast, the nerve stimulation-induced increase in renal venous plasma NE concentration was unaffected during ANP infusion. ANP tended to potentiate the increase in renal NE efflux induced by 2-k renal nerve stjmulation. This may be due to the substantially attenuated blood flow response, since ANP did not affect the NE efflux response when renal vasoconstriction was almost abolished by pretreatment with prazosin. A study carried out in our laboratory has demonstrated that preparations of the dog kidney in vivo are sensitive to the presynaptic inhibitory effects of humorai factors such as adenosine (Yoneda et al., 1990). Thus. ANP does not seem to exert a modulatory action on neural NE release in the dog kidney. ANP may act on postsynaptic sites to inhibit the neurally induced renal vasoconstriction recently shown in the pump-perfused rat kidney (Schwartz and Eikenburg, 1991). It is generally accepted that neurally released NE causes vasoconstriction via activation of tu,-adrenoceptors. In our study, ANP attenuated the renal blood flow response to intrarenal injection of methoxamine, which preferentially activates tr,-adrenoceptors in the canine renal vasculature (Chiba et al., 1990). This observation suggests that ANP inhibits the postsynaptic Eu,-adrenoceptor mechanism, thereby regulating the neural control of renal vascular tone. However, studies performed with pithed rats have demonstrated that ANP does not inhibit cur-adrenoceptor-mediated systemic vasoconstriction (Haass et al., 1985; Zukowska-Grojec et al., 1986). Although ANP slightly suppressed the pressor response io sympathetic stimulation in yohimbine-treated rats, this was related to a blunted NE release response by a presynaptic action of ANP (Zukowska-Grojec et al., 1986). We observed, however, that even in the presence of yohimbine ANP attenuated the nerve stimulation-induced renal blood flow response without suppressing the NE release response (data are not shown). These discrepancies may be ascribable to differences in the systemic vs. the regional (renal) sympathetic nervous system. Although the present study focuses on the neural control of vascular tone, it should be noted that ANP also inhibits angiotensin II-induced renal vasoconstriction in dogs (Hisa et al,, 1990) and in rats (Schwartz and Eikenburg, 1991). ANP therefore does not seem to be specific for ar-adrenoceptor mechanisms. ANP could act on common pathway(s) mediating vasoconstriction in response to neural and humoral stimuli in the kid-
a~ismby \vhicl~ ANP inhibits renal vasortey. ~~s~r~ct~~~ ~a~aot be ~ctermined from the present study. This issue requires further elucidation. iBn~~c~~s~~~~,the present study demonstrates that ~~~~ea~~s~y a~rn~~~stered ANP can suppress neurally ~~~~ced ~~s~o~strict~o~ without affecting neurotransmitter release in the dog kidney in vivo.
This work was supported in part by a Grant-in-Aid for Scientific Research Japan) No. 02771695 and by the Sapporo Bioscience Foundation (Japan). Part of this work was presented at the 11th International Congress of Pharmacology. Amsterdam, 1990.
rences Chiha. K.. Y. Hayashi. H. Hisa. M. Suzuki-Kusaba and S. Satoh, W&l. Effects of a novel ru,-adrenoceptor antagonist. SGB-1534, on adrenergically induced renal vasoconstriction in dogs. Eur. J. Phxmxol. i76. 263. Goetz, K.L.. l9SS. Physiology and pathology of atrial peptides. Am. J. Phpiol. 2%. El.
Haass, M., I.J. Kopin, D.S. Goldstein and Z. Zukowska~Grojec, 1985, Differential inhibition of alpha adrenoceptor-mediated pressor responses by rat atrial natriuretic peptide in the pithed rat, J. Pharmacol. Exp. Ther. 235, 122. Hayashi, Y.. H. Hisa and S. Satoh. 1987, Role of prostaglandin in norepinephrine and renin release in canine kidney, Am. J. Physiol. 253. F929. Hisa, H., Y. Tomura and S. Satoh 1989, Atrial natriuretic factor suppresses neural stimulation of renin release in dogs, Am. J. Physiol. 257, E332. Hisa, H., Y. Tomun, A. Takahara, K. Chiba and S. Satoh, 1990, Atria1 natriuretic peptide suppresses renal vasoconstriction induced by angiotensin II, norepinephrine and renal nerve stimulation in dogs, Eur. J. Pharmacol. 183, 1063 (Abstract). Nakamaru, M. and T. Inagami, 1986, Atria1 natriuretic factor inhibits norepinephrine release evoked by sympathetic nerve stimulation in isolated perfused rat mesenteric arteries, Eur. J. Pharmacol. 123, 459. Schwartz, D.D. and D.C. Eikenburg, 1991, Effects of synthetic ANF (rANF(3-28)) on sympathetic neurotransmission in the isolated perfused rat kidney, Life Sci. 48, 781. Yoneda, H., H. Hisa and S. Satoh, 1990, Effects of adenosine on adrenergically induced renal vasoconsriction in dogs, Eur. J. Pharmacol. 176, 109. Zukowska-Grojec. Z., M. Haass, I.J. Kopin and N. Zamir, 1986, Interaction of atrial natriuretic peptide with the sympathetic and endocrine systems in the pithed rat, J. Pharmacol. Exp. Ther. 239, 480.