Angiotensin Ⅱ Receptors Modulate Muscle Microvascular Responses To Insulin

Part1Angiotensin Ⅱ Receptors Regulate Basal Skeletal Muscle Microvascular RecruitmentBackground The major function of the muscle microcirculation is to regulate tissue perfusion to ensure adequate delivery of nutrients, oxygen, and hormones despite changing needs based on activity and feeding patterns. To do this, the endothelium provides an exchange surface area between the plasma compartment and tissue interstitium that can change based upon tissue needs. Microvascular perfusion is determined by relaxing precapillary terminal arterioles, which expands exchange surface area (capillary recruitment). A number of physiological and pathological factors can regulate this process.The renin-angiotensin system (RAS) plays pivotal roles in maintaining vascular health and hemodynamic stability. Ang Ⅱ exerts its vascular actions via2G protein-coupled receptors, the type1receptor (AT1R) and type2receptor (AT2R). In both resistance microvessels and large capacitance vessels, activation of AT1R promotes vasoconstriction and smooth muscle proliferation, whereas AT2R stimulation activates an vasodilator cascade composed of bradykinin, NO, and cGMP, leading to vasodilation and opposing the vasoconstrictor actions of Ang Ⅱ via the AT1R.Both AT1Rs and AT2Rs are present throughout the skeletal muscle microvasculature, where substrate exchanges occur. Whether they modulate basal muscle microvascular perfusion and substrate metabolism is not known.Objective To examine whether angiotensin Ⅱ type1and type2receptors regulate basal skeletal muscle microvascular volume, glucose use and insulin uptake in vivo.Methods In protocol1, Male SD (n=20) were anesthetized and studied under1of the following4groups after an overnight fasted:(1) group1received IV injection of losartan (AT1R blocker,0.3mg/kg);(2) group2received systemic infusion of PD123319(AT2R blocker,50ug/kg per minute);(3) group3received losartan injection and PD123319infusion;(4) group4received losartan injection30minutes after beginning systemic infusion of NO synthase inhibitor (L-NAME50ug/kg per minute). Muscle microvascular blood volume (MBV, an indicator of microvascular perfusion and surface exchange area), was measured using contrast-enhanced ultrasound (CEU). Throughout the study, mean arterial blood pressure (MAP) was monitored via a sensor connected to the carotid arterial catheter.In protocol2, Male SD (n=10) were anesthetized and studied under1of the following2groups after an overnight fasted:(1) group1received IV injection of losartan (0.3mg/kg);(2) group2received systemic infusion of PD123319(50ug/kg per minute). Femoral artery blood flow (FBF) was measured using and ultrasound flow probe.In protocol3, Male SD (n=20) were anesthetized and studied under1of the following4groups after an overnight fasted:(1) group1received IV injection of losartan (0.3mg/kg);(2) group2received systemic infusion of PD123319(50ug/kg per minute);(3) group3received losartan injection and PD123319infusion;(4) group4received losartan injection30minutes after beginning systemic infusion of L-NAME (50ug/kg per minute). Femoral venous blood glucose and arterial blood glucose concentrations were determined and the hindleg glucose uptakes were calculated.In protocol4, Male SD (n=15) were anesthetized and studied under1of the following3groups after an overnight fasted:(1) group1received saline as control;(2) group1received IV injection of losartan (0.3mg/kg);(3) group3received losartan injection30minutes after beginning systemic infusion of L-NAME (50ug/kg per minute). Each rat received a bolus IV injection of125I-insulin5min prior to the end of study, and muscle125I-insulin uptake was determined.Results (1) Administration of the AT1R blocker losartan increased muscle MBV by～3-fold (P<0.001). This was associated with a significant increase in hindleg glucose extraction and muscle125I-insulin uptake (P<0.001). There was no change in either FBF or MAP after losartan injection. By contrast, infusing AT2R antagonist PD123319significantly decreased muscle MBV by～80%(P<0.001). This was associated with a significant decrease in hindleg glucose extraction (P<0.01). There was no change in either FBF or MAP after PD123319infusion.(2)AT2R antagonism blocked the losartan-induced increase in muscle MBV and glucose uptake (P>0.05, compared with basal).(3) Inhibition of NO synthase blocked the losartan-induced increase in muscle MBV, glucose uptake and125I-insulin uptake (P>0.05, compared with basal).Conclusions (1)Angiotensin II acts on both AT1R and AT2R to regulate basal muscle microvascular perfusion. Basal AT1R tone restricts muscle MBV, glucose extraction and insulin uptake, whereas basal AT2R activity increases muscle MBV, extraction and insulin uptake.(2) AT1R blockade with losartan increases muscle microvasvular recruitment, glucose extraction and insulin uptake via AT2R and nitric oxide-dependent mechanism. Part2Angiotensin II Receptors Modulate Basal Skeletal Muscle Metabolic and Microvascular Responses to InsulinBackground Skeletal muscle is particularly important in the metabolic actions of insulin, since it is responsible for almost90%of insulin-mediated whole-body glucose disposal in humans. Evidence suggests that insulin delivery to skeletal muscle interstitium is the rate-limiting step in insulin stimulated muscle glucose uptake. Insulin promotes its own delivery to muscle interstitium by relaxing pre-capillary arterioles to increase the microvascular exchange surface perfused within skeletal muscle (microvascular recruitment). Insulin resistance causes endothelial dysfunction, impairs insulin-mediated increases in microvascular recruitment and its own delivery to muscle interstitium. Microvascular insulin resistance and dysfunction are closely related with metabolic insulin resistance. Blockade of insulin's microvascular action with L-NAME decreases steady state insulin-stimulated glucose disposal by-40%.The renin-angiotensin system (RAS) plays central roles in main-taining hemodynamic stability and angiotensin II (ANG II) can interact with the insulin signaling pathways to regulate insulin sensitivity. Both the AT1R and AT2R are present throughout skeletal muscle microcirculation. We have previously demonstrated that basal AT1R and AT2R regulate muscle MBV, glucose use and insulin delivery. The changes of muscle MBV, glucose use and insulin delivery is tightly associated with microvascular and metabolic actions of insulin. Whether AT1R and AT2R regulate insulin's microvascular and metabolic action in skeletal muscle is not known.Objective To examine the selective effect of AT1R and AT2R activities on skeletal muscle metabolic insulin sensitivity and microvascular insulin sensitivity in vivo.Methods In protocol1, male SD rats (n=15) were anesthetized and studied under1of the following3groups after an overnight fasted:(1) Group1rats received an intravenous infusion of regular insulin (3mU/kg/min) for120min;(2)Group2received systemic infusion of insulin and a bolus i.v. injection of losartan (AT1R blocker,0.3mg/kg);(3) Group3received systemic infusion of insulin and PD123319(AT2R blocker,50μg/kg/min) for120min. Arterial blood glucose was determined every10min and30%dextrose was infused at a variable rate to maintain blood glucose within10%of basal. Skeletal muscle microvascular blood volume (MBV), microvascular flow velocity (MFV), and microvascular blood flow (MBF) were determined using contrast-enhancedultrasound (CEU). Throughout the study, mean arterial blood pressure (MAP) was monitored via a sensor connected to the carotid arterial catheter. In protocol2, Male SD rats (n=15) were anesthetized and studied under1of the following3groups after an overnight fasted:(1) Group1rats received an intravenous infusion of regular insulin (3mU/kg/min) for120min;(2)Group2received systemic infusion of insulin and a bolus i.v. injection of losartan (0.3mg/kg);(3) Group3received systemic infusion of insulin and PD123319(50μg/kg/min) for120min. Arterial blood glucose was determined every10min and30%dextrose was infused at a variable rate to maintain blood glucose within10%of basal. Femoral artery blood flow (FBF) was measured using a flow probe.Results (1) Insulin significantly increased muscle MBV by1.5～2-fold(p<0.05). The steady-state glucose infusion rate (GIR) reached about～8mg/kg.min.(2) losartan+insulin increased muscle MBV by>3-fold (p<0.05) without further increasing GIR.(3)AT2R blockade with PD123319abolished insulin mediated increases in muscle MBV and decreased insulin-stimulated glucose disposal by～30%(p<0.05,com-pared with insulin control group).Conclusions Both AT1Rs and AT2Rs regulate insulin's micro-vascular and metabolic action in muscle. While AT1R activity restrains muscle metabolic responses to insulin via decreased microvascular rec-ruitment, AT2R activity is required for normal microvascular responses to insulin. Chapter3Losartan recruits muscle microvasculature and prevents lipid-induced micarovascular and metabolic insulin resistanceBackground Patients with diabetes have increased plasma concentrations of free fatty acids which cause insulin resistance in multiple insulin responsive tissues, including the microvasculature. Microvascular insulin resistance contributes significantly to metabolic insulin resistance seen in diabetes. Blockade of angiotensin Ⅱ type1receptor (AT1R) with losartan recruits muscle microvasculature in the basal condition and may improve insulin sensitivity. Whether losartan recruits muscle microvasculature and rescues insulin's metabolic action in the presence of high FFA concentrations is not known.Objective To examine whether losartan recruits muscle micro-vasculature and rescues insulin's metabolic action in the presence of high free fatty acids.Methods In protocol1, After an overnight fast, male Sprague-Dawley rats received a systemic infusion of either saline or intralipid+heparin (3.3%and30U/ml) for3hrs, with a3mU/min/kg euglycemic insulin clamp superimposed for the last2hrs. A third group received the same infusion of intralipid+heparin and insulin, with a losartan injection (0.3mg/kg, i.v. bolus)5min before starting the insulin infusion. A fourth group received the same infusion protocols as the third group, with a50ug/kg/min L-NAME infusion superimposed. Muscle microvascular blood volume (MBV) and microvascular flow velocity (MFV) were measured using contrast-enhanced ultrasound before and at30,60and120min of insulin infusion. Muscle microvascular blood flow (MBF) was calculated as the product of MBV and MFV. Steady state whole body glucose disposal rates were calculated. Throughout the study, mean arterial blood pressure (MAP) was monitored via a sensor connected to the carotid arterial catheter.In protocol2, Male SD (n=15) were anesthetized and studied under1of the following3groups after an overnight fasted:(1) group1received saline as control;(2) group2received a systemic infusion of intralipid+heparin (3.3%and30U/ml);(3) group3received received a systemic infusion of intralipid+heparin and losartan injection (0.3mg/kg, i.v. bolus). Each rat received a bolus IV injection of125I-insulin5min prior to the end of study, and muscle125I-insulin uptake was determined.Results (1)Insulin infusion doubled muscle MBV at30min and this effect lasted for the entire120min (p<0.05). Lipid infusion abolished the insulin-mediated increase in muscle MBV, and lowered insulin-stimulated whole body glucose disposal (p<0.001). Losartan injection in the presence of lipid and insulin infusion resulted in2-3-fold increase in MBV (p<0.05), and rescued insulin's actions to stimulate whole body glucose disposal (p=0.001).(2)Lipid infusion did not inhibit muscle123I-insulin uptake compared with saline control, losartan did not further change muscle125I-insulin uptake in the presence of lipid infusion.Conclusions Losartan potently recruits muscle microvasculature and attenuates metabolic insulin resistance induced by lipid infusion. This suggests that angiotensin II type1receptor blockade may improve muscle insulin sensitivity via microvascular recruitment.