Acta Physiologica Sinica

Dec. 2001, 53 (6), 451¡«455

Differential responses of regional vascular beds to local injection of agmatine in rats

LI Qing, FAN Zhenª²Zhong, WANG Yiª²He, HE Ruiª²Rong*

(Department of Physiology, Institute of Basic Medicine, Hebei Medical University, Shijiazhuang 050017)

Abstract: In 66 anaesthetized rats, the effects of local injection of agmatine on femoral, renal, and mesenteric vascular beds were investigated respectively by constant flow perfusion method. The results are as follows. (1) Agmatine (0.1, 0.5, 1 mg/kg) increased the perfusion pressure (PP) of femoral vascular bed in a doseª²dependent manner. The effect of agmatine (1 mg/kg) was completely blocked by pretreatment with idazoxan (0.5 mg/kg), an antagonist for imidazoline receptors (IR) and ¦Á2ª²adrenergic receptors (¦Á2ª²AR), and yohimbine (1 mg/kg), a selective ¦Á2ª²AR antagonist. (2) Agmatine also increased the PP of renal vascular bed in a doseª²dependent manner, and agmatine at high dose (1 mg/kg) caused a biphasic increase of PP in renal vascular bed. Idazoxan blocked these effects completely, while yohimbine led the agmatineª²induced effect to a decrease in the PP of renal vascular bed. (3) Agmatine decreased the PP of mesenteric vascular bed in a doseª²dependent manner, an effect which was completely blocked by idazoxan, but unaffected by yohimbine. From the results obtained, it is concluded that agmatine differentially affects the vascular tone in the femoral, renal, and mesenteric vascular beds.

Key words: agmatine; perfusion pressure; vascular bed; imidazoline receptor; ¦Á2ª²adrenoceptor receptor

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ѧ¿Æ·ÖÀàºÅ: Q463

Agmatine, a polycationic amine synthesized through Lª²arginine decarboxylation which is catalyzed by the enzyme arginine decarboxylase, has been identified as an endogenous clonidineª²displacing substance (CDS) in the mam~malian brain£Û1£Ý and may serve as a neuro~trans~mitter/neuromodulator in the central nervous system£Û2£Ý. Our previous studies demonstrated that intravenous administration of agmatine resulted in decreases in heart rate, blood pressure, cardiac output, myocardial contractility and total peripheral resistance in the anaesthetised rats£Û3£Ý, and in Dahl salt sensitive hypertensive and Dah1 saltª²resistant normotensive rats£Û4£Ý. But it is uncertain if the agmatineª²induced decrease of the total peripheral resistance is centrally or peripherally mediated. Recently, we have shown that agmatine relaxed isolated aortic artery precontracted by phenylephrine or CaCl2£Û5£Ý. However, the direct effects of agmatine on femoral, renal, and mesenteric vascular beds have not yet been elucidated. The present study was undertaken to define the differential responses of femoral, renal, and mesenteric vascular beds to local injection of agmatine. ¡¡¡¡

1MATERIALS AND METHODS

1.1 AnimalsSpragueª²Dawly rats, (male, 320¡À20 g, n=66, Grade ¢ò, Certificate No.04036) obtained from the Experimental Animal Center of Hebei Province.

1.2 Surgical procedureRats were anaesthetized with urethane (1.0 g/kg) intraperitoneally. Supplemental doses of anesthetics were given as necessary. The trachea was cannulated for ventilation and polyethylene cannula containing heparinized (20 U/ml) saline was introduced into the femoral artery for monitoring blood pressure with a pressure transducer (MPUª²0.5, Nihon Kohden), and the heart rate (HR) was monitored by a heart rate counter (ATª²600G, Nihon Kohden) triggered by differential signals of arterial pressure pulse. Body temperature was maintained at 37¡«38¡æ throughout the experiment.

1.2.1 Perfusion of femoral vascular bedThe constant flow perfusion method was adopted to measure the femoral vascular tone. A catheter was inserted into the common carotid artery, from which blood was perfused with a peristaltic pump (1210 A, Harvard) into the femoral artery by constant flow.

1.2.2 Perfusion of renal vascular bedThe left renal artery was exposed via a retroperitoneal approach. A catheter was inserted into the common carotid artery, from which blood was perfused with a peristaltic pump into the renal artery by constant flow.

1.2.3 Perfusion of mesenteric vascular bedThe left mesenteric artery was exposed via a retroperitoneal approach. A catheter was inserted into the common carotid artery, from which blood was perfused with a peristaltic pump into the mesenteric artery by constant flow.

1.3 Experimental protocolsAfter operation, the perfusion pressure was adjusted to be similar to BP, and the rats were allowed to stabilize for more than 30 min. A stable recording was obtained for more than 5 min, the drug (a fixed volume of 20 ¦Ìl over a period of 10 s at a constant speed) was injected into the perfusion circuit directly. Before application of drug, the vehicle (normal saline) was used as control. The changes in BP, HR, and the perfusion pressure were recorded on a polygraph (RMª²6000, Nihon Kohden). The experimental protocols were: (1) different doses of agmatine (0.1, 0.5, 1 mg/kg) were injected into the femoral (n=10), renal (n=10), and mesenteric (n=10) arteries, respectively. BP, HR, and perfusion pressure was examined. (2) Idazoxan (0.5 mg/kg) was injected into the perfusion circuit followed by injection of agmatine (1 mg/kg). BP, HR, and perfusion pressure was recorded in femoral (n=6), renal (n=6), and mesenteric (n=6) vascular beds, respectively. (3) Yohimbine (1 mg/kg) was injected into the perfusion circuit followed by injection of agmatine (1 mg/kg). BP, HR, and perfusion pressure was recorded in femoral (n=6), renal (n=6), and mesenteric (n=6) vascular beds, respectively.

1.4 DrugsAgmatine, idazoxan, and yohimbine (Sigma Chemical Co, St Louis, MO) were dissolved in saline. The drugs were freshly prepared before use.

1.5 StatisticsData were expressed as mean¡ÀSE and analyzed by ANOVA. Dunnett¡äs t test are used as appropriate. P<0.05 was considered significant.

2RESULTS

2.1 Effects of agmatine on femoral vascular bed

¡¡Intrafemoral arterial injection of agmatine at 0.1, 0.5, 1 mg/kg increased the perfusion pressure of femoral vascular bed in a doseª²dependent manner (Fig.1A, Table 1A). The effects of agmatine began at 20¡À5 s, reached the maximal response within 74¡À8 s, and lasted for 5¡À2 min. Either idazoxan (0.5 mg/kg) or yohimbine (1 mg/kg) blocked completely the vasoconstrictor response induced by agmatine (1 mg/kg) (Fig.2).

2.2 Effects of agmatine on the renal vascular bed

¡¡Intrarenal arterial injection of agmatine at 0.1, 0.5, 1 mg/kg increased the perfusion pressure of renal vascular bed in a doseª²dependent manner (Fig. 1B£¬ Table 1B). The effects of agmatine began at 22¡À4 s, reached the max~imal response within 61¡À8 s, and lasted for 4¡À2 min.

Table¡¡£±.Changes in HR, MAP, and PP after injecting agmatine (0.1, 0.5, 1 mg/kg) into the femoral, renal, and mesenteric vascular beds

A. Intrafemoral arterial injection of agmatine; B. Intrarenal arterial injection of agmatine; C. Intramesenteric arterial injection of agmatine. ~*P<0.05;~**P<0.01 vs pretreatment; ~¢LP<0.05, ~¢L¢LP<0.01 vs control.

Fig.1.Effects of agmatine (0.1, 0.5, 1 mg/kg) on HR, BP, and the perfusion pressure in femoral, renal, and mesenteric vascular beds. HR, heart rate; MAP, mean arterial pressure; PP, perfusion pressure. A. Intrafemoral arterial injection of agmatine; B. Intrarenal arterial injection of agmatine; C. Intramesenteric arterial injection of agmatine.

Fig.2.Effects of idazoxan (0.5 mg/kg), and yohimbine (1 mg/kg) on the agmatine (1 mg / kg)ª²induced changes in HR, MAP, and PP of femoral (n=6), renal (n=6), and mesenteric(n=6) vascular beds. ~*P<0.05; ~**P<0.01 vs pretreatment. ¡õ, intrafemoral arterial injection of agmatine; ý«, intrarenal arterial injection of agmatine; ¡ö, intramesenteric arterial injection of agmatine.

Specifically, following injection of a high dose of agmatine (1 mg/kg), the renal perfusion pressure was initially increased significantly (P<0.01), returned to the baseline within 8¡À3 min, and subsequently showed a delayed increase at 10¡À3 min (Fig.3). Idazoxan (0.5 mg/kg) blocked completely both the early and delayed vasoconstrictor responses induced by agmatine (1 mg/kg), while pretreatment with yohimbine resulted in a change in the response to agmatine from vasoconstriction to vasodilatation (Fig.2).

2.3 Effect of agmatine on mesenteric vascular bed

¡¡Intramesenteric arterial injection of agmatine at 0.1, 0.5 and 1 mg/kg decreased the perfusion pressure of mesenteric vascular bed in a doseª²dependent manner (Fig.1C, Table.1C). The effects of agmatine began at

Fig.3.Biphasic increase in renal PP following intrarenal arterial injection of agmatine at high dose (1 mg/kg). Note that the second phase increase in renal PP occurred 12 min later following agmatine injection.

25¡À4 s, reached the maximal response within 78¡À8 s, and lasted for 6¡À2 min. The agmatineª²induced¡¡vasodilatation was abolished completely by idazoxan and unaffected by yohimbine (Fig.2).

3DISCUSSION

Agmatine is present in the vascular bed, where it is synthesized by endothelial cells and stored in endothelial and smooth muscle cells£Û6£Ý. Most vessels contain ¦Á2ª²adrenergic receptors (¦Á2ª²AR) and/or imidazoline receptors (IR). Dyke et al. proved that vascular ¦Á2ª²ARs are expressed in vascular smooth muscle, endothelium and perivascular sympathetic nerves£Û7£Ý. Stimulation of vascular ¦Á2ª²AR produces a direct vasoconstriction. Moreover, Vascular smooth muscle and endothelium also express IR£Û8£Ý, and the activation of IR may induce vasodilatation in some vascular beds£Û9£Ý. In the present study, we demonstrated that agmatine could doseª²dependently increase the perfusion pressures in both perfused femoral and renal vascular beds, and decrease the perfusion pressure in mesenteric vascular bed. These effects of agmatine could be abolished completely by idazoxan, an antagonist for IR and ¦Á2ª²AR£Û10£Ý, suggesting that ¦Á2ª²AR and/or IR¡¡were involved. To characterize the receptor(s) responsible for agmatineª²induced vasoconstriction, we used yohimbine, a selective ¦Á2ª²AR antagonist, in our experiments. In perfused femoral vascular bed, the vasoconstrictor response to agmatine was also blocked completely by yohimbine. From such a result and the findings of Cheung£Û11£Ý, it is suggested that the femoral vascular bed contains only ¦Á2ª²AR or a greater proportion of ¦Á2ª²AR than IR. For the renal vascular bed, pretreatment with ¦Á2ª²AR blocker yohimbine resulted in a change in the response to agmatine from vasoconstriction to vasodilatation. From such results, it is conceivable that the action of ¦Á2ª²AR predominates over that of IR in the renal vascular bed, and so long as the action of agmatine on ¦Á2ª²AR was blocked by yohimbine, its action on IR would be unmasked with a resultant vasodilatation. In contrast, idazoxan completely blocked the actions of agmatine on renal vessels. Interestingly, agmatine at high dose (1 mg/kg) induced a delayed vasoconstriction in perfused renal vascular bed, while such a delayed vasoconstrictor response was not observed in the femoral vascular bed. The mechanism responsible for the occurrence of delayed response remained to be clarified. In perfused mesenteric vascular bed, agmatine decreased the perfusion pressure, indicating a vasodilatation. To examine the receptor(s) responsible for the vasodilatation induced by agmatine in mesenteric vascular bed, yohimbine and idazoxan were used. The results showed that yohimbine had no effect on the vasodilatation induced by agmatine, while idazoxan completely abolished the vasodilative response to agmatine. The findings indicated that the vasodilatation induced by agmatine resulted from the activation of IR, and implied that the mesenteric vascular bed contained only IR, or mainly IR.

In summary, the present study provides evidence that agmatine may affect differentially the vascular tone in the femoral, renal, and mesenteric vascular beds.

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Received 2001ª²04ª²02Accepted 2001ª²05ª²12

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Email: syho@Hebmu.edu.cn