Skeletal muscle microvascular recruitment by physiological hyperinsulinemia precedes increases in total blood flow.

Supraphysiological doses of insulin enhance total limb blood flow and recruit capillaries in skeletal muscle. Whether these processes change in response to physiological hyperinsulinemia is uncertain. To examine this, we infused either saline (n = 6) or insulin (euglycemic clamp, 3.0 mU x min(-1) x kg(-1), n = 9) into anesthetized rats for 120 min. Femoral artery flow was monitored continuously using a Doppler flow probe, and muscle microvascular recruitment was assessed by metabolism of infused 1-methylxanthine (1-MX) and by contrast-enhanced ultrasound (CEU). Insulin infusion raised plasma insulin concentrations by approximately 10-fold. Compared with saline, physiological hyperinsulinemia increased femoral artery flow (1.02 +/- 0.10 vs. 0.68 +/- 0.09 ml/min; P < 0.05), microvascular recruitment (measured by 1-MX metabolism [6.6 +/- 0.5 vs. 4.5 +/- 0.48 nmol/min; P < 0.05] as well as by CEU [167.0 +/- 39.8 vs. 28.2 +/- 13.8%; P < 0.01]), and microvascular flow velocity (beta, 0.14 +/- 0.02 vs. 0.09 +/- 0.02 s(-1)). Subsequently, we studied the time dependency of insulin's vascular action in a second group (n = 5) of animals. Using CEU, microvascular volume was measured at 0, 30, and 90 min of insulin infusion. Insulin augmented microvascular perfusion within 30 min (52.8 +/- 14.8%), and this persisted at 90 min (64.6 +/- 9.9%). Microvascular recruitment occurred without changes to femoral artery flow or beta. We conclude that insulin increases tissue perfusion by recruiting microvascular beds, and at physiological concentrations this precedes increases in total muscle blood flow by 60-90 min.     

Glucagon-like peptide 1 recruits microvasculature and increases glucose use in muscle via a nitric oxide-dependent mechanism

Glucagon-like peptide 1 (GLP-1) increases tissue glucose uptake and causes vasodilation independent of insulin. We examined the effect of GLP-1 on muscle microvasculature and glucose uptake. After confirming that GLP-1 potently stimulates nitric oxide (NO) synthase (NOS) phosphorylation in endothelial cells, overnight-fasted adult male rats received continuous GLP-1 infusion (30 pmol/kg/min) for 2 h plus or minus NOS inhibition. Muscle microvascular blood volume (MBV), microvascular blood flow velocity (MFV), and microvascular blood flow (MBF) were determined. Additional rats received GLP-1 or saline for 30 min and muscle insulin clearance/uptake was determined. GLP-1 infusion acutely increased muscle MBV (P < 0.04) within 30 min without altering MFV or femoral blood flow. This effect persisted throughout the 120-min infusion period, leading to a greater than twofold increase in muscle MBF (P < 0.02). These changes were paralleled with increases in plasma NO levels, muscle interstitial oxygen saturation, hind leg glucose extraction, and muscle insulin clearance/uptake. NOS inhibition blocked GLP-1-mediated increases in muscle MBV, glucose disposal, NO production, and muscle insulin clearance/uptake. In conclusion, GLP-1 acutely recruits microvasculature and increases basal glucose uptake in muscle via a NO-dependent mechanism. Thus, GLP-1 may afford potential to improve muscle insulin action by expanding microvascular endothelial surface area.     

Insulin sensitivity of muscle capillary recruitment in vivo

We have reported that insulin exerts two vascular actions in muscle; it both increases blood flow and recruits capillaries. In parallel hyperinsulinemic-euglycemic clamp studies, we compared the insulin dose response of muscle microvascular recruitment and femoral blood flow as well as hindleg glucose uptake in fed, hooded Wistar and fasted Sprague-Dawley rats. Using insulin doses between 0 and 30 mU(-1). min(-1). kg(-1), we measured microvascular recruitment at 2 h by 1-methylxanthine (1-MX) metabolism or contrast-enhanced ultrasound (CEU), and muscle glucose uptake was measured by either arteriovenous differences or using 2-deoxyglucose. We also examined the time course for reversal of microvascular recruitment following cessation of a 3 mU. min(-1). kg(-1) insulin infusion. In both groups, whether measured by 1-MX metabolism or CEU, microvascular recruitment was fully activated by physiologic hyperinsulinemia and occurred at lower insulin concentrations than those that stimulated glucose uptake or hindleg total blood flow. The latter processes were insulin dose dependent throughout the entire dose range studied. Upon stopping the insulin infusion, increases in microvascular volume persisted for 15-30 min after insulin concentrations returned to basal levels. We conclude that the precapillary arterioles that regulate microvascular recruitment are more insulin sensitive than resistance arterioles that regulate total flow.     

Acute vasoconstriction-induced insulin resistance in rat muscle in vivo

Insulin-mediated changes in blood flow are associated with altered blood flow distribution and increased capillary recruitment in skeletal muscle. Studies in perfused rat hindlimb have shown that muscle metabolism can be regulated by vasoactive agents that control blood flow distribution within the hindlimb. In the present study, the effects of a vasoconstrictive agent that has no direct effect on skeletal muscle metabolism but that alters perfusion distribution in rat hindlimb was investigated in vivo to determine its effects on insulin-mediated vascular action and glucose uptake. We measured the effects of alpha-methylserotonin (alpha-met5HT) on mean arterial blood pressure, heart rate, femoral blood flow, hindlimb vascular resistance, and glucose uptake in control and euglycemic insulin-clamped (10 mU x min(-1) x kg(-1)) anesthetized rats. Blood flow distribution within the hindlimb muscles was assessed by measuring the metabolism of 1-methylxanthine (1-MX), an exogenously added substrate for capillary xanthine oxidase. Alpha-met5HT (20 microg x min(-1) x kg(-1)) infusion alone increased mean arterial blood pressure by 25% and increased hindlimb vascular resistance but caused no change in femoral blood flow. These changes were associated with decreased hindlimb 1-MX metabolism indicating less capillary flow. Insulin infusion caused decreased hindlimb vascular resistance that was associated with increased femoral blood flow and 1-MX metabolism. Treatment with alpha-met5HT infusion commenced before insulin infusion prevented the increase in femoral blood flow and inhibited the stimulation of 1-MX metabolism. Alpha-met5HT infusion had no effect on hindlimb glucose uptake but markedly inhibited the insulin stimulation of glucose uptake (P < 0.05) and was associated with decreased glucose infusion rates to maintain euglycemia (P < 0.05). A significant correlation (P < 0.05) was observed between 1-MX metabolism and hindlimb glucose uptake but not between femoral blood flow and glucose uptake. The results indicate that in vivo, certain types of vasoconstriction in muscle such as elicited by 5HT2 agonists, which prevent normal insulin recruitment of capillary flow, cause impaired muscle glucose uptake. Moreover, if vasoconstriction of this kind results from stress-induced increase in sympathetic outflow, then this may provide a clue as to the link between hypertension and insulin resistance that is often observed in humans.     

Obesity blunts insulin-mediated microvascular recruitment in human forearm muscle

We have previously shown that skeletal muscle capillaries are rapidly recruited by physiological doses of insulin in both humans and animals. This facilitates glucose and insulin delivery to muscle, thus augmenting glucose uptake. In obese rats, both insulin-mediated microvascular recruitment and glucose uptake are diminished; however, this action of insulin has not been studied in obese humans. Here we used contrast ultrasound to measure microvascular blood volume (MBV) (an index of microvascular recruitment) in the forearm flexor muscles of lean and obese adults before and after a 120-min euglycemic-hyperinsulinemic (1 mU . min(-1) . kg(-1)) clamp. We also measured brachial artery flow, fasting lipid profile, and anthropomorphic variables. Fasting plasma glucose (5.4 +/- 0.1 vs. 5.1 +/- 0.1 mmol/l, P = 0.05), insulin (79 +/- 11 vs. 38 +/- 6 pmol/l, P = 0.003), and percent body fat (44 +/- 2 vs. 25 +/- 2%, P = 0.001) were higher in the obese than the lean adults. After 2 h of insulin infusion, whole-body glucose infusion rate was significantly lower in the obese versus lean group (19.3 +/- 3.2 and 37.4 +/- 2.6 mumol . min(-1) . kg(-1) respectively, P < 0.001). Compared with baseline, insulin increased MBV in the lean (18.7 +/- 3.3 to 25.0 +/- 4.1, P = 0.019) but not in the obese group (20.4 +/- 3.6 to 18.8 +/- 3.8, NS). Insulin increased brachial artery diameter and flow in the lean but not in the obese group. We observed a significant, negative correlation between DeltaMBV and BMI (R = -0.482, P = 0.027) in response to insulin. In conclusion, obesity eliminated the insulin-stimulated muscle microvascular recruitment and increased brachial artery blood flow seen in lean individuals.     

Lipid infusion impairs physiologic insulin-mediated capillary recruitment and muscle glucose uptake in vivo

Infusion of triglycerides and heparin causes insulin resistance in muscle. Because the vascular actions of insulin, particularly capillary recruitment, may contribute to the increase in glucose uptake by skeletal muscle, we investigated the effects of Intralipid/heparin infusion on the hemodynamic actions of insulin during clamp conditions. Saline or 10% Intralipid/heparin (33 U/ml) was infused into anesthetized rats at 20 microl/min for 6 h. At 4 h into the saline infusion, a 2-h hyperinsulinemic (3 mU. min(-1).kg(-1))-euglycemic clamp was conducted (Ins group). At 4 h into the lipid infusion, a 2-h saline control (Lip group) or 2-h hyperinsulinemic-euglycemic clamp (Lip + Ins group) was conducted. Arterial blood pressure, heart rate, femoral blood flow (FBF), hindleg vascular resistance, glucose infusion rate (GIR), hindleg glucose uptake (HGU), and muscle 2-deoxyglucose uptake (R'g) were measured. Capillary recruitment, as measured by metabolism of infused 1-methylxanthine (1-MX), was also assessed. When compared with either Lip or Lip + Ins, Ins had no effect on arterial blood pressure, heart rate, FBF, or vascular resistance but increased GIR, HGU, and R'g of soleus, plantaris, extensor digitorum longus, and gastrocnemius red muscles and hindlimb 1-MX metabolism. GIR, HGU, and R'g of soleus, plantaris, gastrocnemius red, and the combined muscles and 1-MX metabolism were less in Lip + Ins than in Ins rats. HGU correlated closely with hindleg capillary recruitment (r = 0.86, P < 0.001) but not total hindleg blood flow. In conclusion, acute elevation of plasma free fatty acids blocks insulin-mediated glucose uptake and capillary recruitment.     

Glucagon-Like Peptide 1 Recruits Muscle Microvasculature and Improves Insulin’s Metabolic Action in the Presence of Insulin Resistance

Glucagon-like peptide 1 (GLP-1) acutely recruits muscle microvasculature, increases muscle delivery of insulin, and enhances muscle use of glucose, independent of its effect on insulin secretion. To examine whether GLP-1 modulates muscle microvascular and metabolic insulin responses in the setting of insulin resistance, we assessed muscle microvascular blood volume (MBV), flow velocity, and blood flow in control insulin-sensitive rats and rats made insulin-resistant acutely (systemic lipid infusion) or chronically (high-fat diet [HFD]) before and after a euglycemic-hyperinsulinemic clamp (3 mU/kg/min) with or without superimposed systemic GLP-1 infusion. Insulin significantly recruited muscle microvasculature and addition of GLP-1 further expanded muscle MBV and increased insulin-mediated glucose disposal. GLP-1 infusion potently recruited muscle microvasculature in the presence of either acute or chronic insulin resistance by increasing muscle MBV. This was associated with an increased muscle delivery of insulin and muscle interstitial oxygen saturation. Muscle insulin sensitivity was completely restored in the presence of systemic lipid infusion and significantly improved in rats fed an HFD. We conclude that GLP-1 infusion potently expands muscle microvascular surface area and improves insulin’s metabolic action in the insulin-resistant states. This may contribute to improved glycemic control seen in diabetic patients receiving incretin-based therapy.     

Delayed transcapillary delivery of insulin to muscle interstitial fluid after oral glucose load in obese subjects

Obese subjects exhibit a delay in insulin action and delivery of insulin to muscle interstitial fluid during glucose/insulin infusion. The aim of the present study was to follow the distribution of insulin to skeletal muscle after an oral glucose load in obese subjects. We conducted an oral glucose tolerance test (OGTT) in 10 lean and 10 obese subjects (BMI 23 +/- 0.6 vs. 33 +/- 1.2 kg/m(2); P < 0.001). Insulin measurements in muscle interstitial fluid were combined with forearm arteriovenous catheterization and blood flow measurements. In the obese group, interstitial insulin was significantly (35-55%) lower than plasma insulin (P < 0.05) during the 1st h after the OGTT, whereas in lean subjects, no significant difference was found between interstitial and plasma insulin levels during the same time period. The permeability surface area product for glucose, representing capillary recruitment, increased in the lean group (P < 0.05) but not in the obese group (NS). Obese subjects had a significantly higher plasma insulin level at 90-120 min after oral glucose (398 +/- 57 vs. 224 +/- 37 pmol/l in control subjects; P < 0.05). The significant gradient between plasma insulin and muscle interstitial insulin during the first hour after OGTT suggests a slow delivery of insulin in obese subjects. The hindered transcapillary transport of insulin may be attributable to a defect in insulin-mediated capillary recruitment.     

Delayed transcapillary transport of insulin to muscle interstitial fluid in obese subjects

Insulin-resistant subjects have a slow onset of insulin action, and the underlying mechanism has not been determined. To evaluate whether a delayed transcapillary transport is part of the peripheral insulin resistance, we followed the kinetics of infused insulin and inulin in plasma and muscle interstitial fluid in obese insulin-resistant patients and control subjects. A total of 10 lean and 10 obese men (BMI 24 +/- 0.8 vs. 32 +/- 0.8 kg/m(2), P < 0.001) was evaluated during a hyperinsulinemic-euglycemic clamp (insulin infusion rate 120 mU. m(-2). min(-1)) combined with an inulin infusion. Measurements of insulin and inulin in plasma were taken by means of arterial-venous catheterization of the forearm and microdialysis in brachioradialis muscle combined with forearm blood flow measurements with vein occlusion pletysmography. The obese subjects had a significantly lower steady-state glucose infusion rate and, moreover, demonstrated a delayed appearance of insulin (time to achieve half-maximal concentration [T(1/2)] 72 +/- 6 vs. 46 +/- 6 min in control subjects, P < 0.05) as well as inulin (T(1/2) 83 +/- 3 vs. 53 +/- 7 min, P < 0.01) in the interstitial fluid. Also, the obese subjects had a delayed onset of insulin action (T(1/2) 70 +/- 9 vs. 45 +/- 5 min in control subjects, P < 0.05), and their forearm blood flow rate was significantly lower. These results demonstrate a delayed transcapillary transport of insulin and inulin from plasma to the muscle interstitial fluid and a delayed onset of insulin action in insulin-resistant obese subjects.     

Microvascular recruitment is an early insulin effect that regulates skeletal muscle glucose uptake in vivo

Insulin increases glucose disposal into muscle. In addition, in vivo insulin elicits distinct nitric oxide synthase-dependent vascular responses to increase total skeletal muscle blood flow and to recruit muscle capillaries (by relaxing resistance and terminal arterioles, respectively). In the current study, we compared the temporal sequence of vascular and metabolic responses to a 30-min physiological infusion of insulin (3 mU. min(-1). kg(-1), euglycemic clamp) or saline in rat skeletal muscle in vivo. We used contrast-enhanced ultrasound to continuously quantify microvascular volume. Insulin recruited microvasculature within 5-10 min (P < 0.05), and this preceded both activation of insulin-signaling pathways and increases in glucose disposal in muscle, as well as changes in total leg blood flow. Moreover, l-NAME (N(omega)-nitro-l-arginine-methyl ester), a specific inhibitor of nitric oxide synthase, blocked this early microvascular recruitment (P < 0.05) and at least partially inhibited early increases in muscle glucose uptake (P < 0.05). We conclude that insulin rapidly recruits skeletal muscle capillaries in vivo by a nitric oxide-dependent action, and the increase in capillary recruitment may contribute to the subsequent glucose uptake.     

Contraction stimulates nitric oxide independent microvascular recruitment and increases muscle insulin uptake

We examined whether contraction-induced muscle microvascular recruitment would expand the surface area for insulin and nutrient exchange and thereby contribute to insulin-mediated glucose disposal. We measured in vivo rat hindlimb microvascular blood volume (MBV) using contrast ultrasound and femoral blood flow (FBF) using Doppler ultrasound in response to a stimulation frequency range. Ten minutes of 0.1-Hz isometric contraction more than doubled MBV (P < 0.05; n = 6) without affecting FBF (n = 7), whereas frequencies >0.5 Hz increased both. Specific inhibition of nitric oxide (NO) synthase with N(omega)-l-nitro-arginine-methyl ester (n = 5) significantly elevated mean arterial pressure by approximately 30 mmHg but had no effect on basal FBF or MBV. We next examined whether selectively elevating MBV without increasing FBF (0.1-Hz contractions) increased muscle uptake of albumin-bound Evans blue dye (EBD). Stimulation at 0.1 Hz (10 min) elicited more than twofold increases in EBD content (micrograms EBD per gram dry tissue) in stimulated versus contralateral muscle (n = 8; 52.2 +/- 3.8 vs. 20 +/- 2.5, respectively; P < 0.001). We then measured muscle uptake of EBD and (125)I-labeled insulin (dpm per gram dry tissue) with 0.1-Hz stimulation (n = 6). Uptake of EBD (19.1 +/- 3.8 vs. 9.9 +/- 1; P < 0.05) and (125)I-insulin (5,300 +/- 800 vs. 4,244 +/- 903; P < 0.05) was greater in stimulated muscle versus control. Low-frequency contraction increases muscle MBV by a NO-independent pathway and facilitates muscle uptake of albumin and insulin in the absence of blood flow increases. This microvascular response may, in part, explain enhanced insulin action in exercising skeletal muscle.     

Exercise training improves insulin-mediated capillary recruitment in association with glucose uptake in rat hindlimb.

Exercise training is considered to be beneficial in the treatment and prevention of insulin insensitivity, and much of the effect occurs in muscle. We have recently shown that capillary recruitment by insulin in vivo is associated with and may facilitate insulin action to increase muscle glucose uptake. In the present study, we examined the effect of 14 days of voluntary exercise training on euglycemic-hyperinsulinemic clamped (10 mU. min(-1). kg(-1) for 2 h), anesthetized rats. Whole-body glucose infusion rate (GIR), hindleg glucose uptake, femoral blood flow (FBF), vascular resistance, and capillary recruitment, as measured by metabolism of infused 1-methylxanthine (1-MX), were assessed. In sedentary animals, insulin caused a significant (P < 0.05) increase in FBF (1.6-fold) and capillary recruitment (1.7-fold) but a significant decrease in vascular resistance. In addition, hindleg glucose uptake was increased (4.3-fold). Exercise training increased insulin-mediated GIR (24%), hindleg glucose uptake (93%), and capillary recruitment (62%) relative to sedentary animals. Neither capillary density nor total xanthine-oxidase activity in skeletal muscle were increased as a result of the training regimen used. We concluded that exercise training improves insulin-mediated increases in capillary recruitment in combination with augmented muscle glucose uptake. Increased insulin-mediated glucose uptake may in part result from the improved hemodynamic control attributable to exercise training.     

Interaction between insulin sensitivity and muscle perfusion on glucose uptake in human skeletal muscle: evidence for capillary recruitment

Insulin and glucose delivery (muscle perfusion) can modulate insulin-mediated glucose uptake. This study was undertaken to determine 1) to what extent insulin sensitivity modulates the effect of perfusion on glucose uptake and 2) whether this effect is achieved via capillary recruitment. We measured glucose disposal rates (GDRs) and leg muscle glucose uptake (LGU) in subjects exhibiting a wide range of insulin sensitivity, after 4 h of steady-state (SS) euglycemic hyperinsulinemia (>6,000 pmol/l) and subsequently after raising the rate of leg blood flow (LBF) 2-fold with a superimposed intrafemoral artery infusion of methacholine chloride (Mch), an endothelium-dependent vasodilator. LBF was determined by thermodilution: LGU = arteriovenous glucose difference (AVGdelta) x LBF. As a result of the 114+/-12% increase in LBF induced by Mch, the AVGdelta decreased 32+/-4%, and overall rates of LGU increased 40+/-5% (P < 0.05). We found a positive relationship between the Mch-modulated increase in LGU and insulin sensitivity (GDR) (r = 0.60, P < 0.02), suggesting that the most insulin-sensitive subjects had the greatest enhancement of LGU in response to augmentation of muscle perfusion. In separate groups of subjects, we also examined the relationship between muscle perfusion rate and glucose extraction (AVGdelta). Perfusion was either pharmacologically enhanced with Mch or reduced by intra-arterial infusion of the nitric oxide inhibitor N(G)-monomethyl-L-arginine during SS euglycemic hyperinsulinemia. Over the range of LBF, changes in AVGdelta were smaller than expected based on the noncapillary recruitment model of Renkin. Together, the data indicate that 1) muscle perfusion becomes more rate limiting to glucose uptake as insulin sensitivity increases and 2) insulin-mediated increments in muscle perfusion are accompanied by capillary recruitment. Thus, insulin-stimulated glucose uptake displays both permeability- and perfusion-limited glucose exchange properties.     

Effect of acute ketosis on the endothelial function of type 1 diabetic patients: the role of nitric oxide.

In type 1 diabetic patients, acute loss of metabolic control is associated with increased blood flow, which is believed to favor the development of long-term complications. The mechanisms for inappropriate vasodilation are partially understood, but a role of endothelium-derived nitric oxide (NO) production can be postulated. We assessed, in type 1 diabetic patients, the effect of the acute loss of metabolic control and its restoration on forearm endothelial function in 13 type 1 diabetic patients who were studied under conditions of mild ketosis on two different occasions. In study 1, after basal determination, a rapid amelioration of the metabolic picture was obtained by insulin infusion. In study 2, seven type 1 diabetic patients underwent the same experimental procedure, except that fasting plasma glucose was maintained constant throughout. Basal plasma venous concentrations of nitrites/nitrates (NO2- + NO3-) were determined both before and after intravenous insulin infusion. Endothelium-dependent and -independent vasodilation of the brachial artery was assessed by an intra-arterial infusion of N(G)-monomethyl-L-arginine (L-NMMA) and sodium nitroprusside (SNP), respectively. The same parameters were determined in 13 control subjects at baseline conditions and during a hyperinsulinemic-euglycemic glucose clamp. Baseline forearm blood flow (4.89 +/- 0.86 vs. 3.65 +/- 0.59 ml x (100 ml tissue)(-1) x min(-1)) and NO2- + NO3- concentration (30 +/- 8 vs. 24 +/- 3 micromol/l) were higher in type 1 diabetic patients than in control subjects (P < 0.05). Insulin infusion was associated with lower forearm blood flow and plasma (NO2- + NO3-) concentration (P < 0.05), irrespective of the prevailing glucose levels, as compared with patients under ketotic conditions. The responses to L-NMMA were significantly lower in type 1 diabetic patients during euglycemia and hyperglycemic hyperinsulinemia (-11 +/- 5 and -10 +/- 4%, respectively, of the ratio of the infused arm to the control arm) than in control subjects at baseline (-18 +/- 6%, P < 0.05) and during hyperinsulinemia (-32 +/- 11%, P < 0.01). We conclude that the acute loss of metabolic control is associated with a functional disturbance of the endothelial function characterized by hyperemia and increased NO release during ketosis and blunted NO-mediated vasodilatory response during restoration of metabolic control by intravenous insulin. This functional alteration is unlikely to be explained by hyperglycemia itself.     

Marked resistance of the ability of insulin to decrease arterial stiffness characterizes human obesity

We tested the hypothesis that insulin has effects on large artery stiffness in addition to its slow vasodilatory effect on resistance vessels in skeletal muscle, and whether such an effect might be altered in obesity. Eight nonobese (aged 25 +/- 1 years, BMI 22.7 +/- 0.4 kg/m2) and eight obese (aged 27 +/- 2 years, BMI 30.6 +/- 0.9 kg/m2) men were studied under normoglycemic-hyperinsulinemic (sequential 2-h insulin infusions of 1 [step 1] and 2 [step 2] mU x kg(-1) x min(-1)) conditions, and another seven men participated in a saline control study. Central aortic pressure waves were synthesized from those recorded in the periphery using applanation tonometry and a validated reverse transfer function every 30 min. This allowed determination of augmentation (the pressure difference between early and late systolic pressure peaks) and the augmentation index (augmentation divided by pulse pressure), a measure of arterial stiffness. Whole-body glucose uptake was reduced by 48 (step 1) and 41% (step 2) (P < 0.01) in the obese subjects versus the nonobese subjects. Basal forearm blood flow averaged 2.5 +/- 0.2 and 2.6 +/- 0.2 ml x dl(-1) x min(-1) in the obese and nonobese subjects, respectively (NS). Insulin induced a significant increase in forearm blood flow after 2.5 h (3.6 +/- 0.4 ml x dl(-1) x min(-1), P < 0.05 vs. basal) in the nonobese subjects and after 4 h in the obese subjects (3.2 +/- 0.2, P < 0.05). In contrast to these slow changes in peripheral blood flow, augmentation and the augmentation index decreased significantly in the nonobese subjects after 1 h (-3.0 +/- 1.6 mmHg and -10.0 +/- 5.4%, respectively, P < 0.001 vs. basal), but remained unchanged until 3 h in the obese subjects. Percent fat (r = 0.86, P < 0.0001) and whole-body glucose uptake (r = -0.72, P < 0.01) correlated with the change in the augmentation index by insulin. These data demonstrate temporal dissociation in insulin's vascular actions. Insulin's effect to decrease arterial stiffness in nonobese subjects (a decrease in wave reflection) is observed under physiological conditions and precedes a slow vasodilatory effect in the periphery. In the obese subjects, insulin's normal effect to decrease central wave reflection is severely blunted. The degree of impairment in this novel vascular action of insulin is closely correlated with the degree of obesity and insulin action on glucose uptake.     

Insulin regulation of skeletal muscle PDK4 mRNA expression is impaired in acute insulin-resistant states

We previously showed that insulin has a profound effect to suppress pyruvate dehydrogenase kinase (PDK) 4 expression in rat skeletal muscle. In the present study, we examined whether insulin's effect on PDK4 expression is impaired in acute insulin-resistant states and, if so, whether this change is accompanied by decreased insulin's effects to stimulate Akt and forkhead box class O (FOXO) 1 phosphorylation. To induce insulin resistance, conscious overnight-fasted rats received a constant infusion of Intralipid or lactate for 5 h, while a control group received saline infusion. Following the initial infusions, each group received saline or insulin infusion (n = 6 or 7 each) for an additional 5 h, while saline, Intralipid, or lactate infusion was continued. Plasma glucose was clamped at basal levels during the insulin infusion. Compared with the control group, Intralipid and lactate infusions decreased glucose infusion rates required to clamp plasma glucose by approximately 60% (P < 0.01), confirming the induction of insulin resistance. Insulin's ability to suppress PDK4 mRNA level was impaired in skeletal muscle with Intralipid and lactate infusions, resulting in two- to threefold higher PDK4 mRNA levels with insulin (P < 0.05). Insulin stimulation of Akt and FOXO1 phosphorylation was also significantly decreased with Intralipid and lactate infusions. These data suggest that insulin's effect to suppress PDK4 gene expression in skeletal muscle is impaired in insulin-resistant states, and this may be due to impaired insulin signaling for stimulation of Akt and FOXO1 phosphorylation. Impaired insulin's effect to suppress PDK4 expression may explain the association between PDK4 overexpression and insulin resistance in skeletal muscle.     

Angiotensin II receptors modulate muscle microvascular and metabolic responses to insulin in vivo

OBJECTIVE

Angiotensin (ANG) II interacts with insulin-signaling pathways to regulate insulin sensitivity. The type 1 (AT₁R) and type 2 (AT₂R) receptors reciprocally regulate basal perfusion of muscle microvasculature. Unopposed AT₂R activity increases muscle microvascular blood volume (MBV) and glucose extraction, whereas unopposed AT₁R activity decreases both. The current study examined whether ANG II receptors modulate muscle insulin delivery and sensitivity.

RESEARCH DESIGN AND METHODS

Overnight-fasted rats were studied. In protocol 1, rats received a 2-h infusion of saline, insulin (3 mU/kg/min), insulin plus PD123319 (AT₂R blocker), or insulin plus losartan (AT₁R blocker, intravenously). Muscle MBV, microvascular flow velocity, and microvascular blood flow (MBF) were determined. In protocol 2, rats received ¹²⁵I-insulin with or without PD123319, and muscle insulin uptake was determined.

RESULTS

Insulin significantly increased muscle MBV and MBF. AT₂R blockade abolished insulin-mediated increases in muscle MBV and MBF and decreased insulin-stimulated glucose disposal by ~30%. In contrast, losartan plus insulin increased muscle MBV by two- to threefold without further increasing insulin-stimulated glucose disposal. Plasma nitric oxide increased by >50% with insulin and insulin plus losartan but not with insulin plus PD123319. PD123319 markedly decreased muscle insulin uptake and insulin-stimulated Akt phosphorylation.

CONCLUSIONS

We conclude that both AT₁Rs and AT₂Rs regulate insulin’s microvascular and metabolic action in muscle. Although AT₁R activity restrains muscle metabolic responses to insulin via decreased microvascular recruitment and insulin delivery, AT₂R activity is required for normal microvascular responses to insulin. Thus, pharmacologic manipulation aimed at increasing the AT₂R-to-AT₁R activity ratio may afford the potential to improve muscle insulin sensitivity and glucose metabolism.Skeletal muscle microvascular perfusion distribution is determined by precapillary terminal arteriolar tone. Dilating these arterioles increases microvascular perfusion and expands the capillary exchange surface area, whereas constriction leads to the opposite (1,2). Microvascular insulin resistance and dysfunction are closely related with metabolic insulin resistance in diabetes (2–4). Insulin-mediated microvascular recruitment precedes insulin-stimulated glucose uptake in skeletal muscle (5), and blockade of insulin’s microvascular action with N^ω-nitro-l-arginine methyl ester (l-NAME) decreases steady-state insulin-stimulated glucose disposal by ~40% (5,6).To act on muscle, insulin must first traverse the microvasculature perfusing the muscle and then be transported through the vascular endothelium into muscle interstitium. Recent evidence suggests that altered muscle microvascular perfusion profoundly affects insulin delivery and action in muscle (2). Many physiological factors regulate muscle microvascular perfusion in vivo, including insulin, mixed meals, and muscle contraction (7–12). Increased muscle microvascular recruitment induced by muscle contraction is associated with increased muscle insulin uptake (11).The renin-angiotensin system (RAS) plays a central role in maintaining hemodynamic stability (13,14), and angiotensin (ANG) II can interact with the insulin-signaling pathways to regulate insulin sensitivity. In cultured cells, ANG II acts via the ANG II type 1 receptor (AT₁R) to impair insulin actions (15–17). On the other hand, acutely raising ANG II systemically improves insulin-stimulated muscle glucose utilization in humans (18–20) and increases muscle microvascular recruitment independent of blood pressure changes in rodents (21). Both the AT₁R and ANG II type 2 receptor (AT₂R) are present on endothelial cells, vascular smooth-muscle cells, and other vessel-associated cells throughout skeletal muscle microcirculation (13,22,23). ANG II stimulates cell proliferation and vasoconstriction via the AT₁Rs and promotes vasodilation through the AT₂R (24,25). We recently have reported that both the AT₁Rs and the AT₂Rs significantly regulate basal microvascular tone and glucose use by muscle (21). Although basal AT₂R activity increases muscle microvascular blood volume (MBV) (an index of microvascular surface area and perfusion) and glucose extraction, basal AT₁R activity decreases both (21).In the current study, we assessed whether ANG II receptors modulate muscle insulin delivery and sensitivity in vivo. Our results indicate that both AT₁Rs and AT₂Rs regulate insulin’s microvascular and metabolic action in muscle. Although AT₁R activity restrains muscle metabolic responses to insulin via decreased microvascular recruitment and insulin delivery, AT₂R activity is required for normal microvascular responses to insulin.     

Interrelationships among insulin's antilipolytic and glucoregulatory effects and plasma triglycerides in nondiabetic and diabetic patients with endogenous hypertriglyceridemia

We tested the hypothesis that the previously observed association among hypertriglyceridemia, hyperinsulinemia, and insulin resistance could be explained by a defect in insulin's antilipolytic effect. Insulin action was measured in 10 nondiabetic and 8 diabetic patients with hypertriglyceridemia (fasting plasma triglyceride 800 +/- 154 and 1105 +/- 445 mg/dl, respectively, P NS; fasting plasma glucose 99 +/- 3 and 161 +/- 12 mg/dl, respectively, P less than .001) and in 8 weight-matched normolipemic nondiabetic individuals (fasting plasma triglyceride and glucose 110 +/- 21 and 91 +/- 3 mg/dl). The slope of the decay in plasma free fatty acid (FFA) during insulin infusion was used as an index of insulin's antilipolytic effect. Insulin stimulation of glucose uptake in vivo during intravenous hyperinsulinemic clamp and in vitro in adipocytes were measures of insulin's glucoregulatory action. Both glucoregulatory and antilipolytic effects were similarly reduced in both hypertriglyceridemic groups compared with normal subjects. The plasma triglyceride concentration correlated positively with the slope of FFA suppression by insulin (r = .81, P less than .0001) and the fasting FFA concentration (r = .65, P less than .0001). In multiple linear regression analysis, insulin's antilipolytic effect and the fasting FFA concentration explained 83% of the variation in the plasma triglyceride concentration. These associations were independent of insulin's glucoregulatory effect and the fasting plasma insulin concentration. The data indicate that patients with endogenous hypertriglyceridemia are resistant to both the antilipolytic and glucoregulatory actions of insulin and that increased flux of FFA as a result of the latter, rather than hyperinsulinemia, is responsible for elevation of very-low-density lipoprotein production.(ABSTRACT TRUNCATED AT 250 WORDS)     

Insulin-mediated hemodynamic changes are impaired in muscle of Zucker obese rats

Insulin-mediated hemodynamic effects in muscle were assessed in relation to insulin resistance in obese and lean Zucker rats. Whole-body glucose infusion rate (GIR), femoral blood flow (FBF), hindleg glucose extraction (HGE), hindleg glucose uptake (HGU), 2-deoxyglucose (DG) uptake into muscles of the lower leg (R(g)), and metabolism of infused 1-methylxanthine (1-MX) to measure capillary recruitment were determined for isogylcemic (4.8 +/- 0.2 mmol/l, lean; 11.7 +/- 0.6 mmol/l, obese) insulin-clamped (20 mU. min(-1). kg(-1) x 2 h) and saline-infused control anesthetized age-matched (20 weeks) lean and obese animals. Obese rats (445 +/- 5 g) were less responsive to insulin than lean animals (322 +/- 4 g) for GIR (7.7 +/- 1.4 vs. 22.2 +/- 1.1 mg. min(-1). kg(-1), respectively), and when compared with saline-infused controls there was no increase due to insulin by obese rats in FBF, HGE, HGU, and R(g) of soleus, plantaris, red gastrocnemius, white gastrocnemius, extensor digitorum longus (EDL), or tibialis muscles. In contrast, lean animals showed marked increases due to insulin in FBF (5.3-fold), HGE (5-fold), HGU (8-fold), and R(g) ( approximately 5.6-fold). Basal (saline) hindleg 1-MX metabolism was 1.5-fold higher in lean than in obese Zucker rats, and insulin increased in only that of the lean. Hindleg 1-MX metabolism in the obese decreased slightly in response to insulin, thus postinsulin lean was 2.6-fold that of the postinsulin obese. We conclude that muscle insulin resistance of obese Zucker rats is accompanied by impaired hemodynamic responses to insulin, including capillary recruitment and FBF.     

Systemic and local adrenergic regulation of muscle glucose utilization during hypoglycemia in healthy subjects

Adrenergic responses are crucial for hypoglycemic recovery. Epinephrine increases glucose production, lipolysis, and peripheral insulin resistance as well as blood flow and glucose delivery. Sympathetic activation causes vasoconstriction and reduces glucose delivery. To determine the effects of alpha- and beta-adrenergic activity on muscle glucose uptake during hypoglycemia, we studied forearm blood flow (FBF) (plethysmography), arteriovenous glucose difference (AV-diff), and forearm glucose uptake (FGU) during insulin infusion with 60 min of euglycemia followed by 60 min of hypoglycemia. Twelve healthy subjects (27 plus minus 5 years of age) were randomized to intravenous propranolol (IV PROP, 80 microg/min), intravenous phentolamine (IV PHEN, 500 microg/min), intra-arterial propranolol (IA PROP, 25 microg/min), intra-arterial phentolamine (IA PHEN, 12 microg/min per 100 ml forearm tissue), and saline (SAL). FBF increased during hypoglycemia with SAL (P < 0.001) but not with IA or IV PROP. FGU (P = 0.015) and AV-diff (P = 0.099) fell during hypoglycemia with IA PROP but not with IV PROP. FBF increased during hypoglycemia with IA and IV PHEN (P < 0.005). AV-diff fell during hypoglycemia with IA and IV PHEN (P < 0.01), but FGU was unchanged. Blood pressure fell (P < 0.001), and adrenergic and neuroglycopenic symptoms increased with IV PHEN (P < 0.01). Thus, systemic but not local propranolol prevents a decrease in forearm glucose extraction during hypoglycemia, suggesting that epinephrine increases peripheral muscular insulin resistance through systemic effects. alpha-Adrenergic activation inhibits vasodilation and helps maintain brain glucose delivery.