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A High-Sodium Diet in Streptozotocin-Induced Diabetic Rats Impairs Endothelium-Derived Hyperpolarizing Factor-Mediated Vasodilation
Journal of Pharmacological Sciences
J Pharmacol Sci 104, 402 – 405 (2007)
©2007 The Japanese Pharmacological Society
Short Communication
A High-Sodium Diet in Streptozotocin-Induced Diabetic Rats Impairs Endothelium-Derived Hyperpolarizing Factor-Mediated Vasodilation Yasutoshi Hirabara1, Mina Araki1, Mio Fukuda1, Shiori Katafuchi1, Kenji Honda1, Ryo Saito1, and Yukio Takano1,* 1
Department of Physiology and Pharmacology, Faculty of Pharmaceutical Sciences, Fukuoka University, Fukuoka 814-0180, Japan
Received March 1, 2007; Accepted June 21, 2007
Abstract. We examined endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation of mesenteric arteries in high-sodium loaded streptozotocin (STZ)-induced diabetic rats. The study shows that acetylcholine (ACh)-induced, EDHF-mediated relaxation is relatively maintained in STZ-induced diabetic rats, but after a high-sodium diet was given, the function was significantly impaired in STZ-induced diabetic rats. Keywords: diabetes, mesenteric artery, endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation choline (ACh) is attenuated in animal models of diabetes (5, 6), whereas others have reported that the relaxation to ACh is normal (7, 8). Recently, the EDHF-mediated relaxation of mesenteric arteries in type 1 diabetic rats has been reported (9, 10). Moreover, Cheng et al. (11, 12) reported that the blood pressure of the diabetic Goto-Kakizaki rat (GK rat) worsens after being given a high-sodium diet. However, the detailed mechanism for this is not fully understood. Therefore, we examined EDHF-mediated relaxation of mesenteric arteries in high-sodium loaded STZ-induced diabetic rats. Male Wistar rats (180 – 200 g) were purchased from Kyudo (Kumamoto). The animals were maintained at 24 ± 2°C with a 12-h light-dark cycle (lights on at 07:00 h) and were given free access to commercial food and tap water. All procedures regarding animal care and use were carried out according to the guidelines of the Experimental Animal Care and Use Committee of Fukuoka University. Diabetic rats were obtained by a single injection of STZ (60 mg / kg) dissolved in 10 mM citrate buffer through the tail vein. As a control, age-matched rats were injected with the same volume of citrate buffer only. One week after the injection of STZ, diabetes was observed by measuring the glucose concentration with a GLUTEST SENSOR (Sanwa Chemical Co., Nagoya) in a blood sample obtained from the tail vein. Rats with blood glucose levels above 300 mg / dL were used as the
The endothelium cells play an important role in controlling vascular tone with the endothelium-derived relaxing factors (EDRF) or the endothelium-derived hyperpolarizing factor (EDHF). Nitric oxide (NO), one of the major EDRF substances, contributes to endothelium-dependent relaxation in the artery. Although the molecular identity of EDHF has not been determined, EDHF causes hyperpolarization by opening the K+ channel, leading to relaxation in small arteries (1, 2). Therefore, the failure of the EDHF may result not only in increased local vascular resistance but also possible damage to the small vessel (3). The prevalence of both type 1 and type 2 diabetes have been increasing and these diseases cause much distress in patients. For instance, a patient with diabetic neuropathy suffers from a variety of aberrant pain. Indeed, we have already reported on the allodynia in streptozotocin (STZ)-induced diabetic mice (4). In addition, diabetes is associated with complications such as renal dysfunction and cardiovascular disorders, which are induced by vascular damage. Therefore, a number of studies have examined vascular damage and vascular functions using animal models of diabetes. Some studies have demonstrated that the endotheliumdependent relaxation to vasodilators such as acetyl*Corresponding author. [email protected] Published online in J-STAGE: August 4, 2007 doi: 10.1254/jphs.SC0070091
402
Vasodilation in a High-Sodium Diet
diabetic rats. Control and diabetic rats were given free access to food and drinking water for 12 weeks after the injection of buffer or STZ. Then, for another 8 weeks, a high sodium diet (1% NaCl water and 8% NaCl chow; CLEA Japan, Tokyo) was given to control and diabetic rats to make them hypertensive. Systolic blood pressure (SBP) and heart rate (HR) were measured weekly in conscious rats by a tail-cuff blood pressure analyzer (BP-98A; Softron, Tokyo). Determination of the sensitivity of baroreceptor reflex was performed as described in our previous study (13). Experiments on mesenteric artery responses were performed as described previously (14). The main branches of the superior mesenteric arteries were excised, and arteries corresponding to the first to third branches enumerated from the main branch were used. The arteries were placed in chilled Krebs-Henseleit solution. The arteries were then freed from adipose and connective tissues under the microscope and cut into rings (outer diameter: 0.8 – 1.2 mm and length: 2 mm). Each ring was mounted on an L-shaped wire attached to a force-displacement transducer (T7-8-240; NEC Medicalsystems, Tokyo) in an organ bath (UC-5; UFER, Kyoto) that contained 5 mL of Krebs-Henseleit solution. The arterial rings were adjusted to a resting tension of 1.0 g and allowed to stabilize for 2 h during which time the medium was replaced every 10 min. At the end of the experimental period, the heart and left kidney were excised and weighed. Data are expressed as means ± S.E.M. Statistical analyses were performed using Student’s unpaired t-test or one-way analysis of variance followed by Dunnett’s multiple comparisons test. A value of P<0.05 was taken to indicate a statistically significant difference. Body weights in both STZ-induced diabetic and high sodium-loaded STZ-induced diabetic rats were significantly decreased compared with the age-matched control group. Blood glucose levels in STZ-induced diabetic and high sodium-loaded STZ-induced diabetic rats were significantly higher than those of the aged matched control group (control: 110 ± 5 mg / dL, STZ: 548 ± 28 mg / dL, salt diet: 101 ± 4 mg / dL, STZ-salt diet: 330 ± 24 mg / dL). Figure 1 shows the daily changes in SBP and HR in each animal model. There were no significant differences in SBP between the control and STZinduced diabetic rats, but the SBP increased after a high sodium diet was given. To study the central cardiovascular regulation in STZ-induced diabetic rats, we examined the sensitivity of the baroreceptor reflex, but there was no difference in changes in sensitivity of the baroreceptor reflex between the control and STZinduced diabetic rats (Fig. 1C).
403
Fig. 1. Cardiovascular characteristics. A: Systolic blood pressure (SBP), B: Heart rate (HR), C: Sensitivity of baroreceptor reflex. Open circles indicate Wistar control rats; open squares, STZ-induced diabetic rats; solid circles, high sodium-loaded Wistar rats; solid squares, high sodium-loaded STZ-induced diabetic rats. Each data point represents the mean ± S.E.M. from 4 – 6 rats. *P<0.05, **P<0.01 vs Wistar control rats; #P<0.05, ##P<0.01 vs high sodiumloaded Wistar rats (unpaired test).
As shown in Fig. 2, ACh elicited relaxation of rat mesenteric arteries during phenylephrine-induced contraction. The phenylephrine-induced contraction of the mesenteric artery was the same between controls and diabetic rats. The maximum relaxation and the sensitivity to ACh in STZ-induced diabetic rats did not change compared with control rats. However, the sensi-
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tivity to ACh was significantly attenuated in the high sodium-loaded group compared with the non-sodiumloaded group (Fig. 2A). Endothelium-independent relaxations to SNP were similar in STZ-diabetic rats and the control, but the sensitivity to SNP was significantly attenuated in high sodium-loaded STZ-diabetic rats (Fig. 2B). Next, to investigate the EDHF-mediated relaxation evoked by ACh in rat mesenteric artery, the NO synthetase inhibitor L-NAME (10−4 M) and cyclooxygenase inhibitor indomethacin (10−5 M) were added to the medium (12, 14). The concentration-response curves induced by ACh are shown in Fig. 3B. Interestingly, ACh-induced EDHF-mediated relaxation was hardly observed in high sodium-loaded STZ-induced diabetic rats. On the other hand, the maximum contraction and sensitivity to phenylephrine did not change (Fig. 3A). The present study shows that the blood pressure and vascular functions are relatively maintained in STZinduced diabetic rats. The maximum relaxation and the
Fig. 2. Relaxation to ACh (A) or SNP (B) after eliciting precontraction with phenylephrine (0.5 × 10−6 – 0.5 × 10−5 M) in isolated endothelium-intact mesenteric arterial rings. The symbols are the same as in Fig. 1. A: Endothelium-dependent vascular relaxation to ACh was impaired in high sodium-loaded animal groups (*P<0.05, **P<0.01 vs Wistar control rats). B: Endothelium-independent vascular relaxation to SNP was slightly impaired in high sodiumloaded STZ-induced diabetic rats (*P<0.05 vs Wistar control rats). Each data point represents the mean ± S.E.M. from 5 – 8 experiments.
sensitivity to ACh in STZ-induced diabetic rats did not change compared with control rats. Moreover, there were no differences in changes in sensitivity of the baroreceptor reflex between the control and STZinduced diabetic rats (Fig. 1C), indicating that the central cardiovascular regulation is maintained in STZinduced diabetic rats. However, these functions were impaired in rats fed the high-sodium diet. In particular, ACh-induced, EDHF-mediated relaxation of the mesenteric artery is hardly observed in high sodiumloaded STZ-induced diabetic rats. This finding supports other results with the mesenteric artery from type 2 diabetic GK rats (11, 12). The present findings that the vascular responses to ACh in STZ-induced diabetic rats did not change agree with studies in type 1 diabetic model animals (7, 8). Others, however, reported that the endothelium-dependent relaxation to ACh is attenuated in STZ-induced diabetic rats (5, 6). Although we have no direct evidence that can clearly explain this discrepancy, the differences seem to have been caused by size or place of the mesenteric arteries that we used. The reason for this is
Fig. 3. Relaxation of mesenteric arterial rings to ACh after preincubation with the NOS inhibitor L-NAME and cyclooxygenase inhibitor indomethacine. The symbols are the same as in Fig. 1. A: Pre-contraction was elicited with phenylephrine (1.0 × 10−7 – 1.0 × 10−6 M) in isolated endothelium-intact mesenteric arterial rings. B: The high-sodium diet impaired ACh-induced EDHF-mediated relaxation (**P<0.01 vs Wistar control rats).
Vasodilation in a High-Sodium Diet
based on the findings that the attenuation of endothelium-dependent relaxation was not observed in larger blood vessels such as femoral and carotid arteries (15). The endothelium-independent vascular relaxation to SNP was unchanged in STZ-induced diabetic rats (Fig. 2B). This finding indicates that the sensitivity of the smooth muscle of mesenteric arteries to NO remained unchanged in STZ-induced diabetic rats. In our animal model, a high sodium diet induced cardiac and kidney hypertrophy (data not shown). Interestingly, ACh-induced, EDHF-mediated relaxation was not observed in high sodium-loaded STZ-induced diabetic rats. This result supports the clinical symptom. Cheng et al. reported that the angiotensin-converting enzyme inhibitor enalapril and vasopeptidase omapatrilat improved the endothelial-dependent vascular relaxation in GK rats (12), suggesting that the renin-angiotensin system is involved in the development of endothelial dysfunction in diabetes with high dietary sodium. In fact, angiotensin II has been shown to be generated in the artery wall in response to increased vascular wall tension. The other possibility seems to be due to damage of the vascular smooth muscle because the high-sodium diet induces the damage to the end-organ and inflammation associated with increased oxidative stress. Indeed, the present study shows that SNP-induced relaxation was attenuated in high sodium-loaded STZinduced diabetic rats compared with the non-sodium loaded group, but attenuation of relaxation induced by SNP was small compared with the acetylcholineinduced relaxation. In contrast, ACh-induced, EDHFmediated relaxation was hardly observed in high sodium-loaded STZ-induced diabetic rats. Moreover, phenylephrine-induced contraction was maintained in high sodium-loaded STZ-induced diabetic rats. Thus, we cannot explain the impairment of ACh-induced, EDHFmediated relaxation only by the damage of the smooth muscle. In addition to the damage of the smooth muscle, the function of EDHF, such as potassium channels, seems to be lost during the high sodium diet (3). The present paper demonstrated that the injury of ACh-induced, EDHF-mediated relaxation induced in diabetic rats was enhanced in high sodium-loaded STZinduced diabetic rats. This indicated that high sodium intake remarkably increases the risk of endothelium dysfunction. However, as the exact mechanism has not been established, further studies are needed to elucidate the substances underlying the impairment of AChinduced, EDHF-mediated relaxation.
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References 1 Chen G, Suzuki H, Weston AH. Acetylcholine releases endothelium-derived hyperpolarizing factor and EDRF from rat blood vessels. Br J Pharmacol. 1988;95:1165–1174. 2 Komori K, Vanhotte PM. Endothelium-derived hyperpolarizing factor. Blood Vessels. 1990;27:238–245. 3 Matsumoto T, Miyamori K, Kobayashi T, Kamata K. Specific impairment of endothelium-derived hyperpolarizing factor-type relaxation in mesenteric arteries from streptozotocin-indued diabetic mice. Vascul Pharmacol. 2006;44:450–460. 4 Koga K, Honda K, Ando S, Harasawa I , Kamiya H, Takano Y. Intrathecal clonidine inhibits mechanical allodynia via activation of the spinal muscarinic M1 receptor in streptozotocin-induced diabetic mice. Eur J Pharmacol. 2004;505:75–82. 5 Fukao M, Hattori Y, Kanno M, Sakuma I, Kitabatake A. Alteration in endothelium-dependent hyperpolarization and relaxation in mesenteric arteries from streptozotocin-induced diabetic rats. Br J Pharmacol. 1997;121:1383–1391. 6 Poston L, Taylor PD. Endothelium-mediated vascular function in insulin-dependent diabetes mellitus. Clin Sci. 1995;88:245– 255. 7 Harris KM, Macleod KM. Influence of the endothelium on contractile responses of arteries from diabetic rats. Eur J Pharmacol. 1988;153:55–64. 8 Mulhern M, Docherty JR. Effects of experimental diabetes on the responsiveness of rat aorta. Br J Pharmacol. 1989;97:1007– 1012. 9 Makino A, Ohuchi K, Kamata K. Mechanisms underlying the attenuation of endothelium-dependent vasodilatation in the mesenteric arterial bed of the streptozotocin-induced diabetic rat. Br J Pharmacol. 2000;130:549–556. 10 Wigg SJ, Tare M, Tonta MA, O’brien RC, Meredith IT, Parkington HC. Comparison of effects of diabetes mellitus on an EDHF-dependent and EDHF-independent artery. Am J Physiol Heart Circ Physiol. 2001;281:H232–H240. 11 Cheng ZJ, Vaskonen T, Tikkanen I, Nurminen K, Ruskoaho H, Vapaatalo H, et al. Endothelial dysfunction and salt-sensitive hypertension in spontaneously diabetic Goto-Kakizaki rats. Hypertension. 2001;37:433–439. 12 Cheng ZJ, Gronholm T, Louhelainen M, Finckenberg P, Merasto S, Tikkanen I, et al. Vascular and renal effects of vasopeptidase inhibition and angiotensin-converting enzyme blockade in spontaneously diabetic Goto-Kakizaki rats. J Hypertens. 2005; 23:1757–1770. 13 Nakayama Y, Takano Y, Eguchi K, Migita K, Saito R, Tujimoto G, et al. Modulation of the arterial baroreceptor reflex by the vasopressin receptor in the area postrema of the hypertensive rats. Neurosci Lett. 1997; 226:179–182. 14 Mizuta A, Takano Y, Honda K, Saito R, Matsumoto T, Kamiya H. Nitric oxide is a mediator of tachykinin NK3 receptorinduced relaxation in rat mesenteric artery. Br J Pharmacol. 1995;116:2919–2922. 15 Shi Y, Ku DD, Man RYK, Vanhotte PM. Augmented endothelium-derived hyperpolarizating factor-mediated relaxations attenuate endothelial dysfunction in femoral and mesentericm but not in carotid arteries from type 1 diabetic rats. J Pharmacol Exp Ther. 2006;318:276–281.
J Pharmacol Sci 104, 402 – 405 (2007)
©2007 The Japanese Pharmacological Society
Short Communication
A High-Sodium Diet in Streptozotocin-Induced Diabetic Rats Impairs Endothelium-Derived Hyperpolarizing Factor-Mediated Vasodilation Yasutoshi Hirabara1, Mina Araki1, Mio Fukuda1, Shiori Katafuchi1, Kenji Honda1, Ryo Saito1, and Yukio Takano1,* 1
Department of Physiology and Pharmacology, Faculty of Pharmaceutical Sciences, Fukuoka University, Fukuoka 814-0180, Japan
Received March 1, 2007; Accepted June 21, 2007
Abstract. We examined endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation of mesenteric arteries in high-sodium loaded streptozotocin (STZ)-induced diabetic rats. The study shows that acetylcholine (ACh)-induced, EDHF-mediated relaxation is relatively maintained in STZ-induced diabetic rats, but after a high-sodium diet was given, the function was significantly impaired in STZ-induced diabetic rats. Keywords: diabetes, mesenteric artery, endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation choline (ACh) is attenuated in animal models of diabetes (5, 6), whereas others have reported that the relaxation to ACh is normal (7, 8). Recently, the EDHF-mediated relaxation of mesenteric arteries in type 1 diabetic rats has been reported (9, 10). Moreover, Cheng et al. (11, 12) reported that the blood pressure of the diabetic Goto-Kakizaki rat (GK rat) worsens after being given a high-sodium diet. However, the detailed mechanism for this is not fully understood. Therefore, we examined EDHF-mediated relaxation of mesenteric arteries in high-sodium loaded STZ-induced diabetic rats. Male Wistar rats (180 – 200 g) were purchased from Kyudo (Kumamoto). The animals were maintained at 24 ± 2°C with a 12-h light-dark cycle (lights on at 07:00 h) and were given free access to commercial food and tap water. All procedures regarding animal care and use were carried out according to the guidelines of the Experimental Animal Care and Use Committee of Fukuoka University. Diabetic rats were obtained by a single injection of STZ (60 mg / kg) dissolved in 10 mM citrate buffer through the tail vein. As a control, age-matched rats were injected with the same volume of citrate buffer only. One week after the injection of STZ, diabetes was observed by measuring the glucose concentration with a GLUTEST SENSOR (Sanwa Chemical Co., Nagoya) in a blood sample obtained from the tail vein. Rats with blood glucose levels above 300 mg / dL were used as the
The endothelium cells play an important role in controlling vascular tone with the endothelium-derived relaxing factors (EDRF) or the endothelium-derived hyperpolarizing factor (EDHF). Nitric oxide (NO), one of the major EDRF substances, contributes to endothelium-dependent relaxation in the artery. Although the molecular identity of EDHF has not been determined, EDHF causes hyperpolarization by opening the K+ channel, leading to relaxation in small arteries (1, 2). Therefore, the failure of the EDHF may result not only in increased local vascular resistance but also possible damage to the small vessel (3). The prevalence of both type 1 and type 2 diabetes have been increasing and these diseases cause much distress in patients. For instance, a patient with diabetic neuropathy suffers from a variety of aberrant pain. Indeed, we have already reported on the allodynia in streptozotocin (STZ)-induced diabetic mice (4). In addition, diabetes is associated with complications such as renal dysfunction and cardiovascular disorders, which are induced by vascular damage. Therefore, a number of studies have examined vascular damage and vascular functions using animal models of diabetes. Some studies have demonstrated that the endotheliumdependent relaxation to vasodilators such as acetyl*Corresponding author. [email protected] Published online in J-STAGE: August 4, 2007 doi: 10.1254/jphs.SC0070091
402
Vasodilation in a High-Sodium Diet
diabetic rats. Control and diabetic rats were given free access to food and drinking water for 12 weeks after the injection of buffer or STZ. Then, for another 8 weeks, a high sodium diet (1% NaCl water and 8% NaCl chow; CLEA Japan, Tokyo) was given to control and diabetic rats to make them hypertensive. Systolic blood pressure (SBP) and heart rate (HR) were measured weekly in conscious rats by a tail-cuff blood pressure analyzer (BP-98A; Softron, Tokyo). Determination of the sensitivity of baroreceptor reflex was performed as described in our previous study (13). Experiments on mesenteric artery responses were performed as described previously (14). The main branches of the superior mesenteric arteries were excised, and arteries corresponding to the first to third branches enumerated from the main branch were used. The arteries were placed in chilled Krebs-Henseleit solution. The arteries were then freed from adipose and connective tissues under the microscope and cut into rings (outer diameter: 0.8 – 1.2 mm and length: 2 mm). Each ring was mounted on an L-shaped wire attached to a force-displacement transducer (T7-8-240; NEC Medicalsystems, Tokyo) in an organ bath (UC-5; UFER, Kyoto) that contained 5 mL of Krebs-Henseleit solution. The arterial rings were adjusted to a resting tension of 1.0 g and allowed to stabilize for 2 h during which time the medium was replaced every 10 min. At the end of the experimental period, the heart and left kidney were excised and weighed. Data are expressed as means ± S.E.M. Statistical analyses were performed using Student’s unpaired t-test or one-way analysis of variance followed by Dunnett’s multiple comparisons test. A value of P<0.05 was taken to indicate a statistically significant difference. Body weights in both STZ-induced diabetic and high sodium-loaded STZ-induced diabetic rats were significantly decreased compared with the age-matched control group. Blood glucose levels in STZ-induced diabetic and high sodium-loaded STZ-induced diabetic rats were significantly higher than those of the aged matched control group (control: 110 ± 5 mg / dL, STZ: 548 ± 28 mg / dL, salt diet: 101 ± 4 mg / dL, STZ-salt diet: 330 ± 24 mg / dL). Figure 1 shows the daily changes in SBP and HR in each animal model. There were no significant differences in SBP between the control and STZinduced diabetic rats, but the SBP increased after a high sodium diet was given. To study the central cardiovascular regulation in STZ-induced diabetic rats, we examined the sensitivity of the baroreceptor reflex, but there was no difference in changes in sensitivity of the baroreceptor reflex between the control and STZinduced diabetic rats (Fig. 1C).
403
Fig. 1. Cardiovascular characteristics. A: Systolic blood pressure (SBP), B: Heart rate (HR), C: Sensitivity of baroreceptor reflex. Open circles indicate Wistar control rats; open squares, STZ-induced diabetic rats; solid circles, high sodium-loaded Wistar rats; solid squares, high sodium-loaded STZ-induced diabetic rats. Each data point represents the mean ± S.E.M. from 4 – 6 rats. *P<0.05, **P<0.01 vs Wistar control rats; #P<0.05, ##P<0.01 vs high sodiumloaded Wistar rats (unpaired test).
As shown in Fig. 2, ACh elicited relaxation of rat mesenteric arteries during phenylephrine-induced contraction. The phenylephrine-induced contraction of the mesenteric artery was the same between controls and diabetic rats. The maximum relaxation and the sensitivity to ACh in STZ-induced diabetic rats did not change compared with control rats. However, the sensi-
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tivity to ACh was significantly attenuated in the high sodium-loaded group compared with the non-sodiumloaded group (Fig. 2A). Endothelium-independent relaxations to SNP were similar in STZ-diabetic rats and the control, but the sensitivity to SNP was significantly attenuated in high sodium-loaded STZ-diabetic rats (Fig. 2B). Next, to investigate the EDHF-mediated relaxation evoked by ACh in rat mesenteric artery, the NO synthetase inhibitor L-NAME (10−4 M) and cyclooxygenase inhibitor indomethacin (10−5 M) were added to the medium (12, 14). The concentration-response curves induced by ACh are shown in Fig. 3B. Interestingly, ACh-induced EDHF-mediated relaxation was hardly observed in high sodium-loaded STZ-induced diabetic rats. On the other hand, the maximum contraction and sensitivity to phenylephrine did not change (Fig. 3A). The present study shows that the blood pressure and vascular functions are relatively maintained in STZinduced diabetic rats. The maximum relaxation and the
Fig. 2. Relaxation to ACh (A) or SNP (B) after eliciting precontraction with phenylephrine (0.5 × 10−6 – 0.5 × 10−5 M) in isolated endothelium-intact mesenteric arterial rings. The symbols are the same as in Fig. 1. A: Endothelium-dependent vascular relaxation to ACh was impaired in high sodium-loaded animal groups (*P<0.05, **P<0.01 vs Wistar control rats). B: Endothelium-independent vascular relaxation to SNP was slightly impaired in high sodiumloaded STZ-induced diabetic rats (*P<0.05 vs Wistar control rats). Each data point represents the mean ± S.E.M. from 5 – 8 experiments.
sensitivity to ACh in STZ-induced diabetic rats did not change compared with control rats. Moreover, there were no differences in changes in sensitivity of the baroreceptor reflex between the control and STZinduced diabetic rats (Fig. 1C), indicating that the central cardiovascular regulation is maintained in STZinduced diabetic rats. However, these functions were impaired in rats fed the high-sodium diet. In particular, ACh-induced, EDHF-mediated relaxation of the mesenteric artery is hardly observed in high sodiumloaded STZ-induced diabetic rats. This finding supports other results with the mesenteric artery from type 2 diabetic GK rats (11, 12). The present findings that the vascular responses to ACh in STZ-induced diabetic rats did not change agree with studies in type 1 diabetic model animals (7, 8). Others, however, reported that the endothelium-dependent relaxation to ACh is attenuated in STZ-induced diabetic rats (5, 6). Although we have no direct evidence that can clearly explain this discrepancy, the differences seem to have been caused by size or place of the mesenteric arteries that we used. The reason for this is
Fig. 3. Relaxation of mesenteric arterial rings to ACh after preincubation with the NOS inhibitor L-NAME and cyclooxygenase inhibitor indomethacine. The symbols are the same as in Fig. 1. A: Pre-contraction was elicited with phenylephrine (1.0 × 10−7 – 1.0 × 10−6 M) in isolated endothelium-intact mesenteric arterial rings. B: The high-sodium diet impaired ACh-induced EDHF-mediated relaxation (**P<0.01 vs Wistar control rats).
Vasodilation in a High-Sodium Diet
based on the findings that the attenuation of endothelium-dependent relaxation was not observed in larger blood vessels such as femoral and carotid arteries (15). The endothelium-independent vascular relaxation to SNP was unchanged in STZ-induced diabetic rats (Fig. 2B). This finding indicates that the sensitivity of the smooth muscle of mesenteric arteries to NO remained unchanged in STZ-induced diabetic rats. In our animal model, a high sodium diet induced cardiac and kidney hypertrophy (data not shown). Interestingly, ACh-induced, EDHF-mediated relaxation was not observed in high sodium-loaded STZ-induced diabetic rats. This result supports the clinical symptom. Cheng et al. reported that the angiotensin-converting enzyme inhibitor enalapril and vasopeptidase omapatrilat improved the endothelial-dependent vascular relaxation in GK rats (12), suggesting that the renin-angiotensin system is involved in the development of endothelial dysfunction in diabetes with high dietary sodium. In fact, angiotensin II has been shown to be generated in the artery wall in response to increased vascular wall tension. The other possibility seems to be due to damage of the vascular smooth muscle because the high-sodium diet induces the damage to the end-organ and inflammation associated with increased oxidative stress. Indeed, the present study shows that SNP-induced relaxation was attenuated in high sodium-loaded STZinduced diabetic rats compared with the non-sodium loaded group, but attenuation of relaxation induced by SNP was small compared with the acetylcholineinduced relaxation. In contrast, ACh-induced, EDHFmediated relaxation was hardly observed in high sodium-loaded STZ-induced diabetic rats. Moreover, phenylephrine-induced contraction was maintained in high sodium-loaded STZ-induced diabetic rats. Thus, we cannot explain the impairment of ACh-induced, EDHFmediated relaxation only by the damage of the smooth muscle. In addition to the damage of the smooth muscle, the function of EDHF, such as potassium channels, seems to be lost during the high sodium diet (3). The present paper demonstrated that the injury of ACh-induced, EDHF-mediated relaxation induced in diabetic rats was enhanced in high sodium-loaded STZinduced diabetic rats. This indicated that high sodium intake remarkably increases the risk of endothelium dysfunction. However, as the exact mechanism has not been established, further studies are needed to elucidate the substances underlying the impairment of AChinduced, EDHF-mediated relaxation.
405
References 1 Chen G, Suzuki H, Weston AH. Acetylcholine releases endothelium-derived hyperpolarizing factor and EDRF from rat blood vessels. Br J Pharmacol. 1988;95:1165–1174. 2 Komori K, Vanhotte PM. Endothelium-derived hyperpolarizing factor. Blood Vessels. 1990;27:238–245. 3 Matsumoto T, Miyamori K, Kobayashi T, Kamata K. Specific impairment of endothelium-derived hyperpolarizing factor-type relaxation in mesenteric arteries from streptozotocin-indued diabetic mice. Vascul Pharmacol. 2006;44:450–460. 4 Koga K, Honda K, Ando S, Harasawa I , Kamiya H, Takano Y. Intrathecal clonidine inhibits mechanical allodynia via activation of the spinal muscarinic M1 receptor in streptozotocin-induced diabetic mice. Eur J Pharmacol. 2004;505:75–82. 5 Fukao M, Hattori Y, Kanno M, Sakuma I, Kitabatake A. Alteration in endothelium-dependent hyperpolarization and relaxation in mesenteric arteries from streptozotocin-induced diabetic rats. Br J Pharmacol. 1997;121:1383–1391. 6 Poston L, Taylor PD. Endothelium-mediated vascular function in insulin-dependent diabetes mellitus. Clin Sci. 1995;88:245– 255. 7 Harris KM, Macleod KM. Influence of the endothelium on contractile responses of arteries from diabetic rats. Eur J Pharmacol. 1988;153:55–64. 8 Mulhern M, Docherty JR. Effects of experimental diabetes on the responsiveness of rat aorta. Br J Pharmacol. 1989;97:1007– 1012. 9 Makino A, Ohuchi K, Kamata K. Mechanisms underlying the attenuation of endothelium-dependent vasodilatation in the mesenteric arterial bed of the streptozotocin-induced diabetic rat. Br J Pharmacol. 2000;130:549–556. 10 Wigg SJ, Tare M, Tonta MA, O’brien RC, Meredith IT, Parkington HC. Comparison of effects of diabetes mellitus on an EDHF-dependent and EDHF-independent artery. Am J Physiol Heart Circ Physiol. 2001;281:H232–H240. 11 Cheng ZJ, Vaskonen T, Tikkanen I, Nurminen K, Ruskoaho H, Vapaatalo H, et al. Endothelial dysfunction and salt-sensitive hypertension in spontaneously diabetic Goto-Kakizaki rats. Hypertension. 2001;37:433–439. 12 Cheng ZJ, Gronholm T, Louhelainen M, Finckenberg P, Merasto S, Tikkanen I, et al. Vascular and renal effects of vasopeptidase inhibition and angiotensin-converting enzyme blockade in spontaneously diabetic Goto-Kakizaki rats. J Hypertens. 2005; 23:1757–1770. 13 Nakayama Y, Takano Y, Eguchi K, Migita K, Saito R, Tujimoto G, et al. Modulation of the arterial baroreceptor reflex by the vasopressin receptor in the area postrema of the hypertensive rats. Neurosci Lett. 1997; 226:179–182. 14 Mizuta A, Takano Y, Honda K, Saito R, Matsumoto T, Kamiya H. Nitric oxide is a mediator of tachykinin NK3 receptorinduced relaxation in rat mesenteric artery. Br J Pharmacol. 1995;116:2919–2922. 15 Shi Y, Ku DD, Man RYK, Vanhotte PM. Augmented endothelium-derived hyperpolarizating factor-mediated relaxations attenuate endothelial dysfunction in femoral and mesentericm but not in carotid arteries from type 1 diabetic rats. J Pharmacol Exp Ther. 2006;318:276–281.