Hyperglycemia and the angiotensin II AT 1 receptor The hyperglycemia induced by angiotensin II in rats is mediated by AT 1 receptors

We have shown that the renin-angiotensin system (RAS) is involved in glucose homeostasis during acute hemorrhage. Since almost all of the physiological actions described for angiotensin II were mediated by AT1 receptors, the present experiments were designed to determine the participation of AT1 receptors in the hyperglycemic action of angiotensin II in freely moving rats. The animals were divided into two experimental groups: 1) animals submitted to intravenous administration of angiotensin II (0.96 nmol/100 g body weight) which caused a rapid increase in plasma glucose reaching the highest values at 5 min after the injection (33% of the initial values, P<0.01), and 2) animals submitted to intravenous administration of DuP-753 (losartan), a nonpeptide antagonist of angiotensin II with AT 1-receptor type specificity (1.63 μmol/100 g body weight as a bolus, iv, plus a 30-min infusion of 0.018 μmol 100 g body weight -1 min-1 before the injection of angiotensin II), which completely blocked the hyperglycemic response to angiotensin II (P<0.01). This inhibitory effect on glycemia was already demonstrable 5 min (8.9 ± 0.28 mM, angiotensin II, N = 9 vs 6.4 ± 0.22 mM, losartan plus angiotensin II, N = 11) after angiotensin II injection and persisted throughout the 30-min experiment. Controls were treated with the same volume of saline solution (0.15 M NaCl). These data demonstrate that the angiotensin II receptors involved in the direct and indirect hyperglycemic actions of angiotensin II are mainly of the AT1-type. Correspondence

In addition to affecting fluid volume, electrolytes and hemodynamic states, the renin-angiotensin system (RAS) is also involved in the regulation of metabolic and endocrine function, especially blood glucose homeostasis (1)(2)(3).Several in vitro hepatocyte studies have shown that angiotensin II stimulates glycogen phosphorylase activity (4)(5)(6) and gluconeogenesis (7)(8)(9)(10).Recently, we have shown that RAS involvement in blood glucose regulation is of physiological significance, with angiotensin II producing a dose-dependent hyperglycemic response (1).In contrast, intravenous infusion of an angiotensin II peptide-analog antagonist, [1sar,8-thr]-angiotensin II (sarthran), had an inhibitory effect on hemorrhage hyperglycemia (1,2).These were the first data demonstrating that the RAS elicits a physiological glycemic response to angiotensin, in addition to activating the sympathetic nervous system and adrenomedullary secretion (2,3).Therefore, the next step was to determine which type of angiotensin II receptor is involved in this well-confirmed hyperglycemic action of angiotensin II.Since most of the physiological functions described for angiotensin II are mediated by the AT 1 -receptor type (11), the present study was designed to investigate the effect of DuP-753 (losartan), a non-peptide AT 1 -selective antagonist (11,12), on the hyperglycemic response to intravenous injection of angiotensin II.Adult male Wistar rats (12-14 weeks) had free access to Purina rat chow and tap water and were housed under controlled temperature with 14 h of light (5:00-19:00 h) per day.At the age of 11 weeks, the rats were placed in individual cages and handled frequently.One week later, they were anesthetized with ether and a silastic catheter was inserted through the jugular vein into the right atrium for blood sampling.This catheter was filled with saline solution and rinsed every other day with heparinized saline solution (25 IU/ml).All animals were allowed to recover for one week before being utilized in the experiments.
On the day of the experiment, the rats had their venous catheter connected to a peristaltic pump 1 h prior to intravenous infusion of angiotensin II (Sigma, St. Louis, MO).After 30 min, DuP-753 (Du Pont Merck Pharmaceutical Company, Wilmington, DE) was administered intravenously over a 30-min period (1.63 µmol/100 g body weight as a bolus plus a continuous infusion of 0.018 µmol 100 g body weight -1 min -1 ).Controls submitted to 30-min saline infusion before angiotensin II injection (Sal/Ang II group) were treated with the same volume (0.2 ml as a bolus plus an infusion of 0.007 ml 100 g body weight -1 min -1 ) of saline solution (0.15 M NaCl).At time zero, losartan or saline infusion was stopped and angiotensin II (0.96 nmol/100 g body weight) or saline (0.15 M NaCl, 0.2 ml/ 100 g body weight) was injected over a period of 2 min.Blood samples (0.4 ml) were col- lected at -30 min (immediately before losartan or saline pretreatment) and 0, 5, 10, 15 and 30 min after the injection of angiotensin II or saline.The volume was replaced with saline solution after each sample.The experiments were done between 12:00 and 17:00 h.Blood was centrifuged at 4 o C and plasma was stored at -20 o C until the time for the glucose assay, carried out in duplicate by the oxidase method (GodAna, Labtest, BR, Lagoa Santa, MG).The data are reported as means ± SEM.The integrated area under the glucose curve was calculated by the trapezoidal rule.Differences between groups were determined by analysis of variance followed by the Newman-Keuls test.Glycemia after angiotensin II injection was compared to basal values by the paired Student t-test.A probability of P<0.05 was considered to be significant.
As illustrated in Figure 1A, following the injection of 0.96 nmol/100 g body weight of angiotensin II (Sal/Ang II group, 9 rats) there was an immediate increase in plasma glucose levels, reaching the highest value at 5 min after injection (8.9 ± 0.28 mM, at 5 min vs 6.7 ± 0.23 mM, basal value), when the increase was about 33% of the initial values (P<0.01).At 10 min the values were still high (16.6%,P<0.01), and at 30 min postinjection plasma glucose levels were returning to normal.The increase of plasma glucose following angiotensin II injection was completely blocked (P<0.01) by infusion of the angiotensin II antagonist losartan (DuP-753/Ang II group, 11 rats).This effect of losartan infusion persisted throughout the 30-min experimental period.Plasma glucose of saline-pretreated (Sal/Sal group) and losartan-pretreated (DuP-753/Sal group) rats did not change during control tests without angiotensin II injection (Figure 1A,B).
The present data show that the hyperglycemia induced by angiotensin II is com-pletely blocked by DuP-753, a non-peptide antagonist of angiotensin II with AT 1 -receptor type specificity (11,12).In fact, this receptor type seems to mediate the actions of angiotensin II in the liver that contains only angiotensin II receptors which can be blocked by DuP-753 (11,13,14).It has been shown recently that angiotensin II increases hepatic glucose production by a receptor-mediated mechanism that is not related to the pressor response to the hormone (15).Angiotensin II induces transduction signs (phosphoinositide turnover and calcium mobilization) and activates glycogen phosphorylase and adenylate cyclase through AT 1 receptors in hepatocytes (13,14).In addition, losartan has been shown to block the increased production of glucose by angiotensin II infused during a single-pass perfusion of rat liver (16).However, we have recently demonstrated that the hyperglycemic response to angiotensin II is also dependent on sympathetic adrenomedullary system activation.Therefore, a hyperglycemic response to angiotensin II attributed to this indirect action of the peptide could occur despite the blockade of the hepatocyte AT 1 receptors by losartan administration.It is important to stress that the stimulatory actions of angiotensin II on adrenal catecholamine release are also inhibited by losartan, despite the predominance of the AT 2 receptor type in the adrenal medulla (12,17,18).Therefore, the results of these studies are in agreement with our present data and previous studies (1-3), and indicate that the hyperglycemic effect of angiotensin II produced by its stimulatory actions on the sympathetic adrenomedullary system and on hepatic glucose output is losartan sensitive.
In summary, the present results show that the angiotensin II receptors involved in the direct and indirect hyperglycemic actions of the hormone are both mainly of the AT 1 -type.