Insulin-stimulated Blood Flow Research Articles

Endothelial-Specific Reduction in Arf6 Impairs Insulin-Stimulated Vasodilation and Skeletal Muscle Blood Flow Resulting in Systemic Insulin Resistance in Mice.

Much of what we know about insulin resistance is based on studies from metabolically active tissues such as the liver, adipose tissue, and skeletal muscle. Emerging evidence suggests that the vascular endothelium plays a crucial role in systemic insulin resistance; however, the underlying mechanisms remain incompletely understood. Arf6 (ADP ribosylation factor 6) is a small GTPase that plays a critical role in endothelial cell function. Here, we tested the hypothesis that the deletion of endothelial Arf6 will result in systemic insulin resistance. We used mouse models of constitutive endothelial cell-specific Arf6 deletion (Arf6f/- Tie2Cre+) and tamoxifen-inducible Arf6 knockout (Arf6f/f Cdh5CreER+). Endothelium-dependent vasodilation was assessed using pressure myography. Metabolic function was assessed using a battery of metabolic assessments including glucose and insulin tolerance tests and hyperinsulinemic-euglycemic clamps. We used a fluorescence microsphere-based technique to measure tissue blood flow. Skeletal muscle capillary density was assessed using intravital microscopy. Endothelial Arf6 deletion impaired insulin-stimulated vasodilation in white adipose tissue and skeletal muscle feed arteries. The impairment in vasodilation was primarily due to attenuated insulin-stimulated nitric oxide bioavailability but independent of altered acetylcholine-mediated or sodium nitroprusside-mediated vasodilation. Endothelial cell-specific deletion of Arf6 also resulted in systematic insulin resistance in normal chow-fed mice and glucose intolerance in high-fat diet-fed obese mice. The underlying mechanisms of glucose intolerance were reductions in insulin-stimulated blood flow and glucose uptake in the skeletal muscle and were independent of changes in capillary density or vascular permeability. Results from this study support the conclusion that endothelial Arf6 signaling is essential for maintaining insulin sensitivity. Reduced expression of endothelial Arf6 impairs insulin-mediated vasodilation and results in systemic insulin resistance. These results have therapeutic implications for diseases that are associated with endothelial cell dysfunction and insulin resistance such as diabetes.

Sympathovagal Balance And Insulin-stimulated Blood Flow Relates To Blood Pressure Dipping In Metabolic Syndrome

Blunted dips in nocturnal systolic blood pressure (SBP) are independently related to cardiovascular disease (CVD). While alterations in blood flow and sympathovagal balance have been suggested to explain dysregulated nocturnal SBP, the role of blood flow and/or sympathovagal balance is unknown in patients with metabolic syndrome (MetS). PURPOSE: We hypothesized that insulin-stimulated blood flow and heart rate recovery (HRR) as well as exaggerated SBP exercise response relates to nocturnal SBP dipping. METHODS: Twenty-eight adults with MetS (ATP III criteria) (53.2 ± 6.5y; 35.8 ± 4.5 kg/m2) were categorized as “dippers” (≥10% change in SBP; 4 M/6F) or “non-dippers” (<10%; 5 M/13F). Twenty-four hour ambulatory blood pressure monitoring was recorded to assess SBP dipping. Participants were fed isocaloric mixed meals for the 24-hr blood pressure monitoring period. A graded, incremental exercise test (VO2peak) was conducted to assess SBP and HR slope response during exercise to estimate sympathetic activity. Rate pressure product (RPP) was also calculated. HRR (peak-post-exercise) at 1 and 2 min post exercise was then used to determine parasympathetic activity. Ultrasound was performed before and after a 2-hr euglycemic hyperinsulinemic clamp (40 mU/m2/min, 90 mg/dl) to assess blood flow. Metabolic panels (e.g. estimated glomerular filtration rate (eGFR), circulating sodium and potassium) were also tested. Three-day food logs were collected in a subgroup (n = 16) to estimate habitual diet. RESULTS: There were no statistical differences in ATP III criteria, age, or eGFR between dippers and non-dippers. Additionally, there were no differences between groups in blood or dietary sodium and potassium. Although HRR and SBP slope during exercise did not differ between dippers and non-dippers, HR slope trended (7.0 ± 1.2 vs. 8.2 ± 1.7, P = 0.09). Interestingly, SBP dipping negatively correlated with HRR 1 minute after exercise (r = -0.60, P = 0.004) as well as insulin stimulated blood flow (r = 0.73, P = 0.04). Also, RPP correlated with nocturnal SBP (r = 0.49, P = 0.02). CONCLUSIONS: HRR, insulin-stimulated blood flow and RPP appear to play a role in nocturnal blood pressure dipping. Together, this work suggests that sympathovagal balance is important for blood flow related CVD risk. Funding: NIH RO1-HL130296.

The Role of Endothelin‐1 Receptor Subtypes in Limiting Insulin‐Stimulated Blood Flow in Middle‐Aged Adults

Beyond its metabolic actions, insulin stimulates peripheral vasodilation, which serves to increase glucose delivery to skeletal muscle. Insulin activates two distinct signaling pathways leading to the production of nitric oxide versus endothelin‐1 (ET‐1). Several lines of research suggest that insulin‐stimulated ET‐1 production may limit insulin‐stimulated nitric oxide‐mediated vasodilation. Recently, it was demonstrated that basal ET‐1‐mediated vasoconstricton is elevated in middle‐aged adults. However, no studies have examined the role of ET‐1 during hyperinsulinemia in middle‐aged individuals. Therefore, we tested the hypothesis that ET‐1 receptor antagonism would improve insulin‐stimulated blood flow (BF) in a cohort of middle‐aged adults. In 14 subjects (age: 54±2 years, BMI: 26±1 kg/m2) with a range of insulin resistance (glucose infusion rate range: 2.0 to 7.2 mg/kg/min), two femoral arterial catheters were placed. One catheter in the left leg for measurement of arterial blood pressure (BP) and one in the right leg for determination of arterial glucose concentrations and intra‐arterial administration (25nmol/min) of BQ‐123 (ETA antagonist) and BQ‐788 (ETB antagonist). In addition, a femoral venous catheter (right leg) was placed for determination of venous glucose concentrations, and one antecubital venous catheter was placed for infusion of insulin and glucose. Femoral artery BF (right leg: Doppler Ultrasound), BP, and glucose uptake (arterial‐venous glucose concentration x BF) were measured during a hyperinsulinemic‐euglycemic clamp alone (insulin: 40 mU/m2/min), followed by hyperinsulinemia with intra‐arterial BQ‐123 administered for one hour (Insulin+BQ‐123). BQ‐788 was then co‐infused with BQ‐123 for an additional hour (insulin+BQ‐123+BQ‐788; n=10). The infusion of BQ‐123 during hyperinsulinemia augmented leg BF (Insulin: 399±42 mL/min; Insulin+BQ‐123: 500±47 mL/min, P&lt;0.001) and vascular conductance (Insulin: 4.3±0.5 mL/min/mmHg; Insulin+BQ‐123: 5.8±0.7 mL/min/mmHg, P&lt;0.001), and increased leg glucose uptake (Insulin: 76±13 mg/min; Insulin+BQ‐123: 103±15 mg/min, P=0.02). Subsequently, co‐infusion of BQ‐788 provided no further increase in leg BF (Insulin+BQ‐123: 485±60 mL/min; Insulin+BQ‐123+BQ‐788: 511±70 mL/min, P=0.42), vascular conductance, or glucose uptake. Sub‐analysis of subjects with type 2 diabetes (T2D, n=7) versus non‐T2D (n=7) suggested no difference in responsiveness to either drug intervention between groups (both P&gt;0.05). These data suggest ET‐1 signaling through ETA receptors plays an important role in limiting insulin‐stimulated BF in middle‐age, whereas ETB receptor signaling has little effect. These findings provide novel insight into the role of ET‐1 during hyperinsulinemia, favoring selective targeting of the ETA receptor for improving insulin‐stimulated BF and glucose uptake with aging.Support or Funding InformationSupported by UTA College of Nursing and Health Innovation

Obesity, type 2 diabetes, and impaired insulin-stimulated blood flow: role of skeletal muscle NO synthase and endothelin-1.

Although impairments in endothelial signaling are hypothesized to reduce insulin-stimulated blood flow in type 2 diabetes (T2D), human studies examining these links are limited. We provide the first measures of nitric oxide synthase and endothelin-1 expression from skeletal muscle tissue containing native microvessels in individuals with and without T2D before and during insulin stimulation. Higher basal skeletal muscle expression of endothelin-1 and reduced endothelial nitric oxide phosphorylation (peNOS)/eNOS may contribute to reduced insulin-stimulated blood flow in obese T2D patients.

Acute inactivity impairs glycemic control but not blood flow to glucose ingestion.

Insulin-stimulated increases in skeletal muscle blood flow play a role in glucose disposal. Indeed, 7 d of aerobic exercise in patients with Type 2 diabetes increased blood flow responses to an oral glucose tolerance test (OGTT) and improved insulin sensitivity. More recent work suggests that reduced daily physical activity impairs glycemic control (GC) in healthy individuals. Herein, we sought to determine whether an acute reduction in daily activity (from >10,000 to <5000 steps per day) for 5 d (RA5) in healthy individuals reduced insulin-stimulated blood flow and GC in parallel and if a 1-d return to activity (RTA1) improved these outcomes. OGTT were performed as a stimulus to increase insulin in 14 healthy, recreationally active men (24 ± 1.1 yr) at baseline, RA5, and RTA1. Measures of insulin sensitivity (Matsuda index) and femoral and brachial artery blood flow were made during the OGTT. Free-living measures of GC including peak postprandial glucose (peak PPG) were also made via continuous glucose monitoring. Femoral and brachial artery blood flow increased during the OGTT but neither was significantly impacted by changes in physical activity (P > 0.05). However, insulin sensitivity was decreased by RA5 (11.3 ± 1.5 to 8.0 ± 1.0, P < 0.05). Likewise, free-living GC measures of peak PPG (113 ± 3 to 123 ± 5 mg·dL(-1), P < 0.05) was significantly increased at RA5. Interestingly, insulin sensitivity and GC as assessed by peak PPG were not restored after RTA1 (P > 0.05). Thus, acute reductions in physical activity impaired GC and insulin sensitivity; however, blood flow responses to an OGTT were not affected. Further, a 1-d return to activity was not sufficient to normalize GC after 5 d of reduced daily physical activity.

Decreased blood flow but unaltered insulin sensitivity of glucose uptake in skeletal muscle of chronic smokers

Chronic cigarette smoking is associated with dysfunction of the vascular endothelium. Smokers have also been shown to be insulin-resistant, at least in some studies. Since insulin-induced vasodilation is dependent on endothelial cell nitric oxide (NO) synthesis, we tested the hypothesis that decreased skeletal muscle blood flow causes insulin resistance in smokers. We studied 37 young normotensive normolipidemic nondiabetic men, of which 14 were smokers and 23 lifelong nonsmokers. The groups were similar with respect to age, body mass index (BMI), and maximal oxygen uptake (VO2max). Basal and insulin-stimulated femoral muscle blood flow was measured using [(15)O]H2O and insulin-stimulated muscle glucose uptake using [18F]fluoro-2-deoxy-D-glucose ([18F]FDG) and positron emission tomography (PET). Whole-body glucose uptake was measured using the hyperinsulinemic (insulin infusion 5 mU/kg x min)-euglycemic clamp technique. In the basal state, muscle blood flow was 51% lower in smokers (17 +/- 3 mL/kg muscle x min) versus nonsmokers (35 +/- 17 mL/kg x min, P < .0001). Insulin increased muscle blood flow comparably in both groups; the mean rate of insulin-stimulated blood flow was 30 +/- 10 and 55 +/- 38 mL/kg x min (P = .049), respectively. Whole-body and skeletal muscle glucose uptake were similar in both groups during insulin infusion. We conclude that muscle blood flow is lower in chronic smokers compared with nonsmokers under both fasting and hyperinsulinemic conditions. The insulin-induced increase in muscle blood flow and insulin-stimulated glucose uptake appear normal, suggesting that the vasodilatory and metabolic effects of insulin are intact in smokers and the reduced muscle blood flow per se does not cause insulin resistance in these subjects.

Insulin-induced vasodilatation: physiology or pharmacology?

Ever since the pioneering studies performed by Andres, Rabinowitz and Zierler 40 years ago [1‐4], investigators interested in insulin action considered insulin resistance in obesity [3], hypertension [5], insulin-dependent (IDDM) [6] and non-insulin-dependent diabetes mellitus (NIDDM) [7] to be primarily a problem of glucose utilization in peripheral tissues, which was independent of glucose delivery. However, a series of studies published since 1990, notably by Alain Baron and his associates, demonstrated that under certain conditions insulin increased leg blood flow [8], and that defects in this action of insulin were likely to contribute significantly to insulin resistance of glucose uptake caused by obesity [8], hypertension [9], IDDM [10] and NIDDM [11]. These latter findings contradicted the vast majority of earlier data and created a controversy regarding the role, or lack thereof, of defects in insulin stimulated blood flow in the pathogenesis of insulin resistance of glucose uptake. It is the purpose of the ensuing review to try to discuss the reasons for the discrepant findings. We will first review a series of studies performed in our own laboratory, which were designed to resolve the “flow controversy”. These studies examined whether the blood flow response to insulin is influenced by: i) the method used to quantitate blood flow; ii) the anatomical location of the flow measurement; iii) dose and duration of the insulin exposure; iv) individual factors such as limb muscularity, physical fitness and endothelial function, defined as the ability to vasodilate in response to nitric oxide (NO) synthesis-dependent vasodilatators. We will try to establish whether this physiological knowledge helps to explain the discrepancies observed in the studies performed in various insulin resistant conditions. We will then review the studies directly testing the hypothesis that an increase in blood flow enhances glucose uptake, and discuss the possibility that various vasoactive agents may change total flow similarly but have distinct effects on flow distribution. Finally, we will review studies relating insulin’s vascular effects to endothelial function, which have opened up an entirely new perspective for understanding how insulin resistance might predispose to vascular disease.

Insulin resistance in essential hypertension is characterized by impaired insulin stimulation of blood flow in skeletal muscle.

To determine whether insulin-stimulated blood flow in patients with mild essential hypertension is altered. Eleven untreated mildly hypertensive patients [aged 35 +/- 2 years, body mass index 25.1 +/- 0.4 kg/m2, mean arterial pressure 110 +/- 2 mmHg (means +/- SEM) and 10 matched normotensive subjects (mean arterial pressure 94 +/- 3 mmHg). Blood flow was quantitated directly in skeletal muscle both basally and during supraphysiologic hyperinsulinemia (serum insulin approximately = 450 mU/l) using radiowater ([15O]H2O) and positron emission tomography. Whole-body and femoral muscle glucose uptakes were determined using the euglycemic insulin clamp technique, [18F]-2-fluoro-2-deoxy-D-glucose and positron emission tomography. Rates of whole-body and femoral muscle glucose uptake were significantly lower in the hypertensive than in the normotensive group. Insulin increased muscle blood flow by 91% in the normotensive group, but only by 33% in the hypertensive group. The ability of insulin to stimulate blood flow in patients with mild essential hypertension is impaired.

Physical fitness and endothelial function (nitric oxide synthesis) are independent determinants of insulin-stimulated blood flow in normal subjects.

Insulin induces vasodilation via stimulation of nitric oxide (NO) synthesis. This action of insulin exhibits considerable interindividual variation. We determined whether the response of blood flow to endothelium-dependent vasoactive agents correlates with that to insulin or whether other factors, such as physical fitness, limb muscularity, or vasodilatory capacity, better explain variations in insulin-stimulated blood flow. Direct measurements of the forearm blood flow response to three 2-h sequential doses of insulin (1, 2, and 5 mU/ kg.min), endothelium-dependent (acetylcholine and NG-monomethyl-L-arginine) and endothelium-independent (sodium nitroprusside) vasoactive agents, and ischemia (reactive hyperemic forearm blood flow) were performed in 22 normal subjects (age, 24 +/- 1 yr; body mass index, 22.2 +/- 0.6 kg/m2; maximal aerobic power, 40 +/- 2 mL/kg.min). The highest insulin dose increased blood flow by 111 +/- 17%. The fraction of basal blood flow inhibited by NG-monomethyl-L-arginine (NO synthesis-dependent flow) varied from 6-47%. Maximal aerobic power (r = 0.52; P < 0.02), the percentage of forearm muscle (r = 0.50; P < 0.02), and the NO synthesis-dependent flow (r = 0.42; P < 0.05), but not reactive hyperemic, acetylcholine-stimulated, or sodium nitroprusside-stimulated flow, were significantly correlated with insulin-stimulated (5 mU/kg.min) blood flow. In multiple linear regression analysis, 52% of the variation (multiple R = 0.72; P < 0.001) in insulin-stimulated blood flow was explained by NO synthesis-dependent flow (P < 0.005) and the percentage of forearm muscle (P < 0.005). We conclude that endothelial function (NO synthesis-dependent basal blood flow) and forearm muscularity are independent determinants of insulin-stimulated blood flow.

Physical fitness and endothelial function (nitric oxide synthesis) are independent determinants of insulin-stimulated blood flow in normal subjects

Physical fitness and endothelial function (nitric oxide synthesis) are independent determinants of insulin-stimulated blood flow in normal subjects

Physical fitness, muscle morphology, and insulin-stimulated limb blood flow in normal subjects.

The response of limb blood flow to insulin is highly variable even in normal subjects. We examined whether physical fitness or differences in muscle morphology contribute to this variation. Maximal aerobic power, muscle fiber composition and capillarization, and the response of forearm glucose extraction and blood flow to a sequential hyperinsulinemic euglycemic clamp (serum insulin 374 +/- 10, 816 +/- 23, and 2,768 +/- 78 pmol/l) were determined in 16 normal males (age 25 +/- 1 yr, body mass index 24 +/- 1 kg/m2). Maximal aerobic power correlated positively with the proportion of type I fibers (r = 0.67, P < 0.01) and negatively with the proportion of type IIb fibers (r = -0.73, P < 0.01). Fiber composition but not blood flow correlated significantly with forearm and whole body glucose uptake. All doses of insulin significantly increased forearm blood flow, maximally by 123 +/- 21%. The ratio of capillaries per fiber was significantly correlated with basal and insulin-stimulated blood flow (0.58- 0.76, P < 0.05-0.01). Mean arterial blood pressure and the insulin-induced increase in blood flow were inversely correlated (r = -0.59, P < 0.05). We conclude that variation in glucose extraction is significantly determined by muscle fiber composition, whereas variation in insulin-stimulated blood flow is closely associated with muscle capillarization.

Insulin-mediated increase in calf blood flow is not related to blood pressure levels in patients with type 2 diabetes mellitus.

An association between insulin resistance and blood pressure levels has been found both in non-diabetic subjects and in type 2 diabetic patients. In non-diabetic subjects, infusion of insulin increases skeletal muscle blood flow, and it has been suggested that an attenuated flow-response to insulin may cause insulin resistance and may also serve as a mechanism relating insulin resistance to elevated blood pressure. We wanted to explore whether these findings could be extended to patients with type 2 diabetes. The effect of insulin infusion during a hyperinsulinaemic euglycaemic glucose clamp investigation on calf blood flow and its relationship to average diastolic blood pressure (DBPAVG), insulin sensitivity (GFRIlbm), body fat and total exercise capacity (TEC) was examined in 11 patients with type 2 diabetes. We found that insulin infusion increased median calf blood flow from 2.3 to 2.7 ml/min.100 g (p = 0.01). The change in blood flow (delta flow) did not correlate to the subjects' average blood pressure levels or their insulin sensitivity. Significant correlations between delta flow and leg fat %, fasting and clamp-insulin levels and total exercise capacity were observed. In conclusion, we found that insulin infusion increased calf blood flow significantly in patients with type 2 diabetes. However, our hypothesis that a negative correlation exists between blood pressure levels and insulin stimulated blood flow, was invalidated. Leg fat was identified as an important predictor of delta flow, GDRILBM and TEC in these patients.