An oral glucose tolerance test does not affect cerebral blood flow: role of NOS.

Animal data indicate that insulin triggers a robust nitric oxide synthase (NOS)-mediated dilation in cerebral arteries similar to the peripheral tissue vasodilation observed in healthy adults. Insulin's role in regulating cerebral blood flow (CBF) in humans remains unclear but may be important for understanding the links between insulin resistance, diminished CBF, and poor brain health outcomes. We tested the hypothesis that an oral glucose challenge (oral glucose tolerance test, OGTT), which increases systemic insulin and glucose, would acutely increase CBF in healthy adults due to NOS-mediated vasodilation, and that changes in CBF would be greater in anterior regions where NOS expression or activity may be greater. In a randomized, single-blind approach, 18 young healthy adults (24 ± 5 yr) underwent magnetic resonance imaging (MRI) with a placebo before and after an OGTT (75 g glucose), and 11 of these adults also completed an NG-monomethyl-l-arginine (l-NMMA) visit. Four-dimensional (4-D) flow MRI quantified macrovascular CBF and arterial spin labeling (ASL) quantified microvascular perfusion. Subjects completed baseline imaging with a placebo (or l-NMMA), then consumed an OGTT followed by MRI scans and blood sampling every 10-15 min for 90 min. Contrary to our hypothesis, total CBF (P = 0.17) and global perfusion (P > 0.05) did not change at any time point up to 60 min after the OGTT, and no regional changes were detected. l-NMMA did not mediate any effect of OGTT on CBF. These data suggest that insulin-glucose challenge does not acutely alter CBF in healthy adults.

Recent insights into mechanisms of hypoxia-induced vasodilatation in the human brain.

The cerebral vasculature manages oxygen delivery by adjusting arterial blood in-flow in the face of reductions in oxygen availability. Hypoxic cerebral vasodilatation, and the associated hypoxic cerebral blood flow reactivity, involve many vascular, erythrocytic and cerebral tissue mechanisms that mediate elevations in cerebral blood flow via micro- and macrovascular dilatation. This contemporary review focuses on in vivo human work - with reference to seminal preclinical work where necessary - on hypoxic cerebrovascular reactivity, particularly where recent advancements have been made. We provide updates with the following information: in humans, hypoxic cerebral vasodilatation is partially mediated via a - likely non-obligatory - combination of: (1) nitric oxide synthases, (2) deoxygenation-coupled S-nitrosothiols, (3) potassium channel-related vascular smooth muscle hyperpolarization, and (4) prostaglandin mechanisms with some contribution from an interrelationship with reactive oxygen species. And finally, we discuss the fact that, due to the engagement of deoxyhaemoglobin-related mechanisms, reductions in O2 content via haemoglobin per se seem to account for ∼50% of that seen with hypoxic cerebral vasodilatation during hypoxaemia. We further highlight the issue that methodological impediments challenge the complete elucidation of hypoxic cerebral reactivity mechanisms in vivo in healthy humans. Future research is needed to confirm recent advancements and to reconcile human and animal findings. Further investigations are also required to extend these findings to address questions of sex-, heredity-, age-, and disease-related differences. The final step is to then ultimately translate understanding of these mechanisms into actionable, targetable pathways for the prevention and treatment of cerebral vascular dysfunction and cerebral hypoxic brain injury.

Reduced basal macrovascular and microvascular cerebral blood flow in young adults with metabolic syndrome: potential mechanisms.

Ninety-million Americans suffer metabolic syndrome (MetSyn), increasing the risk of diabetes and poor brain outcomes, including neuropathology linked to lower cerebral blood flow (CBF), predominantly in anterior regions. We tested the hypothesis that total and regional CBF is lower in MetSyn more so in the anterior brain and explored three potential mechanisms. Thirty-four controls (25 ± 5 yr) and 19 MetSyn (30 ± 9 yr), with no history of cardiovascular disease/medications, underwent four-dimensional flow magnetic resonance imaging (MRI) to quantify macrovascular CBF, whereas arterial spin labeling quantified brain perfusion in a subset (n = 38/53). Contributions of cyclooxygenase (COX; n = 14), nitric oxide synthase (NOS, n = 17), or endothelin receptor A signaling (n = 13) were tested with indomethacin, NG-monomethyl-L-arginine (L-NMMA), and Ambrisentan, respectively. Total CBF was 20 ± 16% lower in MetSyn (725 ± 116 vs. 582 ± 119 mL/min, P < 0.001). Anterior and posterior brain regions were 17 ± 18% and 30 ± 24% lower in MetSyn; reductions were not different between regions (P = 0.112). Global perfusion was 16 ± 14% lower in MetSyn (44 ± 7 vs. 36 ± 5 mL/100 g/min, P = 0.002) and regionally in frontal, occipital, parietal, and temporal lobes (range 15-22%). The decrease in CBF with L-NMMA (P = 0.004) was not different between groups (P = 0.244, n = 14, 3), and Ambrisentan had no effect on either group (P = 0.165, n = 9, 4). Interestingly, indomethacin reduced CBF more in Controls in the anterior brain (P = 0.041), but CBF decrease in posterior was not different between groups (P = 0.151, n = 8, 6). These data indicate that adults with MetSyn exhibit substantially reduced brain perfusion without regional differences. Moreover, this reduction is not due to loss of NOS or gain of ET-1 signaling but rather a loss of COX vasodilation.NEW & NOTEWORTHY We tested the impact of insulin resistance (IR) on resting cerebral blood flow (CBF) in adults with metabolic syndrome (MetSyn). Using MRI and research pharmaceuticals to study the role of NOS, ET-1, or COX signaling, we found that adults with MetSyn exhibit substantially lower CBF that is not explained by changes in NOS or ET-1 signaling. Interestingly, adults with MetSyn show a loss of COX-mediated vasodilation in the anterior but not posterior circulation.

Preserved ß-adrenergic-mediated vasodilation in skeletal muscle of young adults with obesity despite shifts in cyclooxygenase and nitric oxide synthase.

Central adiposity is associated with greater sympathetic support of blood pressure. ß-adrenergic receptors (ß-AR) buffer sympathetically mediated vasoconstriction and ß-AR-mediated vasodilation is attenuated in preclinical models of obesity. With this information, we hypothesized ß-AR vasodilation would be lower in obese compared with normal weight adults. Because ß-AR vasodilation in normal weight adults is limited by cyclooxygenase (COX) restraint of nitric oxide synthase (NOS), we further explored the contributions of COX and NOS to ß-AR vasodilation in this cohort. Forearm blood flow (FBF, Doppler ultrasound) and mean arterial blood pressure (MAP, brachial arterial catheter) were measured and forearm vascular conductance (FVC) was calculated (FVC = FBF/MAP). The rise in FVC from baseline (ΔFVC) was quantified during graded brachial artery infusion of isoproterenol (Iso, 1-12 ng/100 g/min) in normal weight (n = 36) and adults with obesity (n = 22) (18-40 yr old). In a subset of participants, Iso-mediated vasodilation was examined before and during inhibition of NOS [NG-monomethyl-l-arginine (l-NMMA)], COX (ketorolac), and NOS + COX (l-NMMA + ketorolac). Iso-mediated increases in FVC did not differ between groups (P = 0.57). l-NMMA attenuated Iso-mediated ΔFVC in normal weight (P = 0.03) but not adults with obesity (P = 0.27). In normal weight adults, ketorolac increased Iso-mediated ΔFVC (P < 0.01) and this response was lost with concurrent l-NMMA (P = 0.67). In contrast, neither ketorolac (P = 0.81) nor ketorolac + l-NMMA (P = 0.40) altered Iso-mediated ΔFVC in adults with obesity. Despite shifts in COX and NOS, ß-AR vasodilation is preserved in young adults with obesity. These data highlight the presence of a compensatory shift in microvascular control mechanisms in younger humans with obesity.NEW & NOTEWORTHY We examined ß-adrenergic receptor-mediated vasodilation in skeletal muscle of humans with obesity and normal weight. Results show that despite shifts in the contribution of cyclooxygenase and nitric oxide synthase, ß-adrenergic-mediated vasodilation is relatively preserved in young, otherwise healthy adults with obesity. These data highlight the presence of subclinical changes in microvascular control mechanisms early in the obesity process and suggest duration of obesity and/or the addition of primary aging may be necessary for overt dysfunction.

Nitric oxide synthase inhibition in healthy adults reduces regional and total cerebral macrovascular blood flow and microvascular perfusion.

The importance of nitric oxide (NO) in regulating cerebral blood flow (CBF) remains unresolved, due in part to methodological approaches, which lack a comprehensive assessment of both global and regional effects. Importantly, NO synthase (NOS) expression and activity appear greater in some anterior brain regions, suggesting region-specific NOS influence on CBF. We hypothesized that NO contributes to basal CBF in healthy adults, in a regionally distinct pattern that predominates in the anterior circulation. Fourteen healthy adults (7 females; 24 ± 5 years) underwent two magnetic resonance imaging (MRI) study visits with saline (placebo) or the NOS inhibitor, L-NMMA, administered in a randomized, single-blind approach. 4D flow MRI quantified total and regional macrovascular CBF, whereas arterial spin labelling (ASL) MRI quantified total and regional microvascular perfusion. L-NMMA (or volume-matched saline) was infused intravenously for 5 min prior to imaging. L-NMMA reduced CBF (L-NMMA: 722 ± 100 vs. placebo: 771 ± 121 ml/min, P = 0.01) with similar relative reductions (5-7%) in anterior and posterior cerebral circulations, due in part to the reduced cross-sectional area of 9 of 11 large cerebral arteries. Global microvascular perfusion (ASL) was reduced by L-NMMA (L-NMMA: 42 ± 7 vs. placebo: 47 ± 8 ml/100g/min, P = 0.02), with 7-11% reductions in both hemispheres of the frontal, parietal and temporal lobes, and in the left occipital lobe. We conclude that NO contributes to macrovascular and microvascular regulation including larger artery resting diameter. Contrary to our hypothesis, the influence of NO on cerebral perfusion appears regionally uniform in healthy young adults. KEY POINTS: Cerebral blood flow (CBF) is vital for brain health, but the signals that are key to regulating CBF remain unclear. Nitric oxide (NO) is produced in the brain, but its importance in regulating CBF remains controversial since prior studies have not studied all regions of the brain simultaneously. Using modern MRI approaches, a drug that inhibits the enzymes that make NO (L-NMMA) reduced CBF by up to 11% in different brain regions. NO helps maintain proper CBF in healthy adults. These data will help us understand whether the reductions in CBF that occur during ageing or cardiovascular disease are related to shifts in NO signalling.

Differential contribution of cyclooxygenase to basal cerebral blood flow and hypoxic cerebral vasodilation.

Cyclooxygenase (COX) is proposed to regulate cerebral blood flow (CBF); however, accurate regional contributions of COX are relatively unknown at baseline and particularly during hypoxia. We hypothesized that COX contributes to both basal and hypoxic cerebral vasodilation, but COX-mediated vasodilation is greater in the posterior versus anterior cerebral circulation. CBF was measured in 9 healthy adults (28 ± 4 yr) during normoxia and isocapnic hypoxia (fraction of inspired oxygen = 0.11), with COX inhibition (oral indomethacin, 100mg) or placebo. Four-dimensional flow magnetic resonance imaging measured cross-sectional area (CSA) and blood velocity to quantify CBF in 11 cerebral arteries. Cerebrovascular conductance (CVC) was calculated (CVC = CBF × 100/mean arterial blood pressure) and hypoxic reactivity was expressed as absolute and relative change in CVC [ΔCVC/Δ pulse oximetry oxygen saturation (SpO2)]. At normoxic baseline, indomethacin reduced CVC by 44 ± 5% (P < 0.001) and artery CSA (P < 0.001), which was similar across arteries. Hypoxia (SpO2 80%-83%) increased CVC (P < 0.01), reflected as a similar relative increase in reactivity (% ΔCVC/-ΔSpO2) across arteries (P < 0.05), in part because of increases in CSA (P < 0.05). Indomethacin did not alter ΔCVC or ΔCVC/ΔSpO2 to hypoxia. These findings indicate that 1) COX contributes, in a largely uniform fashion, to cerebrovascular tone during normoxia and 2) COX is not obligatory for hypoxic vasodilation in any regions supplied by large extracranial or intracranial arteries.

Reactive oxygen species and cyclooxygenase products explain the majority of hypoxic cerebral vasodilation in healthy humans.

AIM: The role of reactive oxygen species (ROS) in human cerebral blood flow (CBF) during hypoxia is largely unknown. Additionally, it is unknown whether ROS interact with cyclooxygenase-derived signals during hypoxia to increase CBF. We hypothesized ROS inhibition would reduce hypoxic CBF, and combined inhibition of cyclooxygenase (COX) and ROS would decrease hypoxic CBF more than ROS suppression alone. METHODS: We measured middle cerebral artery velocity with transcranial Doppler ultrasound in 12 healthy adults during normoxia and 2 isocapnic hypoxia trials. Intravenous ascorbic acid infusion during the first hypoxia trial suppressed ROS. Oral indomethacin inhibited COX between hypoxia trials. The second bout of hypoxia tested the combined effects of ROS and COX inhibition. Middle cerebral artery velocity was normalized for blood pressure as cerebrovascular conductance index. RESULTS: Hypoxia increased cerebrovascular conductance index in both trials (P < 0.05). Ascorbic acid infusion did not alter cerebrovascular conductance index during hypoxia. Combined ascorbic acid and indomethacin significantly reduced hypoxia-mediated increases in cerebrovascular conductance index from 17 ± 2 to 4 ± 1 cm s-1 100 mm Hg-1 (P < 0.05). CONCLUSION: ROS are not obligatory for hypoxic cerebral vasodilation. Current data indicate ROS and COX together may account for the majority of the increase in CBF through the middle cerebral artery during hypoxia. These data are the first to demonstrate compensatory hypoxic vasodilatory signalling in human cerebral circulation.

Phosphodiesterase-5 inhibition preserves exercise-onset vasodilator kinetics when NOS activity is reduced.

Nitric oxide (NO)-mediated vasodilation contributes to the rapid rise in muscle blood flow at exercise onset. This occurs via increased cyclic guanosine monophosphate (cGMP), which is catabolized by phosphodiesterase-5 (PDE-5). Whether PDE-5 limits exercise vasodilation onset kinetics is unknown. We hypothesized the time course of exercise vasodilation would be 1) accelerated during PDE-5 inhibition (sildenafil citrate, SDF) and 2) decelerated during NO synthase inhibition ( NG-monomethyl-l-arginine, l-NMMA), and 3) the effect of SDF on vasodilation onset kinetics would be attenuated with concurrent l-NMMA. Data from 29 healthy adults were analyzed. Individuals completed 5 min of moderate-intensity forearm exercise under control conditions and during 1) oral SDF ( n = 8), 2) intra-arterial l-NMMA ( n = 15), or 3) combined SDF + l-NMMA ( n = 6). Forearm blood flow (FBF; Doppler ultrasound of the brachial artery) and mean brachial artery blood pressure (MAP) were measured continuously. Forearm vascular conductance (FVC, FBF ÷ MAP) was curve-fit with a monoexponential model, and vasodilation onset kinetics were assessed by mean response time (MRT, time to achieve 63% of steady state). SDF had no effect on MRT ( P = 0.90). NOS inhibition increased MRT ( P = 0.01). MRT during SDF+l-NMMA was not different from control exercise ( P = 0.76). PDE-5 inhibition alone has no effect on rapid-onset vasodilation. Whereas NOS inhibition decelerates vasodilator kinetics, when combined with SDF, vasodilator kinetics do not differ from control. These data suggest NO-independent activation of cGMP occurs at exercise onset; thus PDE-5 inhibition may improve vasodilation in pathologies where NO bioavailability is impaired. NEW & NOTEWORTHY We show that when NO bioavailability is reduced, PDE-5 inhibition can restore vasodilation onset kinetics of exercise-mediated vasodilation via NO-independent cGMP pathways. These data suggest PDE-5 inhibition may improve exercise vasodilation onset kinetics in pathologies where NO bioavailability is impaired.

Regional hypoxic cerebral vasodilation facilitated by diameter changes primarily in anterior versus posterior circulation.

The inability to quantify cerebral blood flow and changes in macrocirculation cross-sectional area in all brain regions impedes robust insight into hypoxic cerebral blood flow control. We applied four-dimensional flow magnetic resonance imaging to quantify cerebral blood flow (ml • min-1) and cross-sectional area (mm2) simultaneously in 11 arteries. In healthy adults, blood pressure, O2 Saturation (SpO2), and end-tidal CO2 were measured at baseline and steady-state hypoxia (FiO2 = 0.11). We investigated left and right: internal carotid, vertebral, middle, anterior, posterior cerebral arteries, and basilar artery. Hypoxia (SpO2 = 80±2%) increased total cerebral blood flow from 621±38 to 742±50 ml • min-1 ( p < 0.05). Hypoxia increased cerebral blood flow, except in the right posterior cerebral arteries. Hypoxia increased cross-sectional area in the anterior arteries (left and right internal carotid arteries, left and right middle, p < 0.05; left and right anterior p = 0.08) but only the right vertebral artery of the posterior circulation. Nonetheless, relative cerebral blood flow distribution and vascular reactivity (Δ%cerebral blood flow • ΔSpO2-1) were not different between arteries. Collectively, moderate hypoxia: (1) increased cerebral blood flow, but relative distribution remains similar to normoxia, (2) evokes similar vascular reactivity between 11 arteries, and (3) increased cross-sectional area primarily in the anterior arteries. This study provides the first wide-ranging, quantitative, functional and structural data regarding intracranial arteries during hypoxia in humans, highlighting cerebral blood flow regulation of microcirculation and macrocirculation differs between anterior and posterior circulation.

Potentiation of the NO-cGMP pathway and blood flow responses during dynamic exercise in healthy humans.

PURPOSE: Previous work has shown nitric oxide (NO) contributes to ~15% of the hyperemic response to dynamic exercise in healthy humans. This NO-mediated vasodilation occurs, in part, via increases in intracellular cyclic guanosine monophosphate (cGMP), which is catabolized by phosphodiesterase. We sought to examine the effect of phosphodiesterase-5 (PDE-5) inhibition on forearm blood flow (FBF) responses to dynamic handgrip exercise in healthy humans and the role of NO. We hypothesized exercise hyperemia would be augmented by sildenafil citrate (SDF, PDE-5 inhibitor). We further hypothesized any effect of SDF on exercise hyperemia would be abolished with intra-arterial infusion of the NO synthase (NOS) inhibitor L-NG-monomethyl arginine (L-NMMA). METHODS: FBF (Doppler ultrasound) was assessed at rest and during 5 min of dynamic forearm handgrip exercise at 15% of maximal voluntary contraction under control (saline) conditions and during 3 experimental protocols: (1) oral SDF (n = 10), (2) intra-arterial L-NMMA (n = 20), (3) SDF and L-NMMA (n = 10). FBF responses to intra-arterial sodium nitroprusside (NTP, NO donor) were also assessed. RESULTS: FBF increased with exercise (p < 0.01). Intra-arterial infusion of L-NMMA resulted in a reduction in exercise hyperemia (17 ± 1 to 15 ± 1 mL/dL/min, p < 0.01). Although the hyperemic response to NTP was augmented by SDF (area under the curve: 41 ± 7 vs 61 ± 11 AU, p < 0.01), there was no effect of SDF on exercise hyperemia (p = 0.33). CONCLUSIONS: Despite improving NTP-mediated vasodilation, oral SDF failed to augment exercise hyperemia in young, healthy adults. These observations reflect a minor contribution of NO and the cGMP pathway during exercise hyperemia in healthy young humans.

Insulin resistance is associated with lower arterial blood flow and reduced cortical perfusion in cognitively asymptomatic middle-aged adults.

Insulin resistance (IR) is associated with poor cerebrovascular health and increased risk for dementia. Little is known about the unique effect of IR on both micro- and macrovascular flow particularly in midlife when interventions against dementia may be most effective. We examined the effect of IR as indexed by the Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) on cerebral blood flow in macro- and microvessels utilizing magnetic resonance imaging (MRI) among cognitively asymptomatic middle-aged individuals. We hypothesized that higher HOMA-IR would be associated with reduced flow in macrovessels and lower cortical perfusion. One hundred and twenty cognitively asymptomatic middle-aged adults (57 ± 5 yrs) underwent fasting blood draw, phase contrast-vastly undersampled isotropic projection reconstruction (PC VIPR) MRI, and arterial spin labeling (ASL) perfusion. Higher HOMA-IR was associated with lower arterial blood flow, particularly within the internal carotid arteries (ICAs), and lower cerebral perfusion in several brain regions including frontal and temporal lobe regions. Higher blood flow in bilateral ICAs predicted greater cortical perfusion in individuals with lower HOMA-IR, a relationship not observed among those with higher HOMA-IR. Findings provide novel evidence for an uncoupling of macrovascular blood flow and microvascular perfusion among individuals with higher IR in midlife.

Greater Beta-Adrenergic Receptor Mediated Vasodilation in Women Using Oral Contraceptives.

BACKGROUND: ß-adrenergic receptors play an important role in mitigating the pressor effects of sympathetic nervous system activity in young women. Based on recent data showing oral contraceptive use in women abolishes the relationship between muscle sympathetic nervous system activity and blood pressure, we hypothesized forearm blood flow responses to a ß-adrenergic receptor agonist would be greater in young women currently using oral contraceptives (OC+, n = 13) when compared to those not using oral contraceptives (OC-, n = 10). METHODS: Women (18-35 years) were studied during the early follicular phase of the menstrual cycle (days 1-5) or placebo phase of oral contraceptive use. Forearm blood flow (FBF, Doppler ultrasound) and mean arterial blood pressure (MAP, brachial arterial catheter) were measured at baseline and during graded brachial artery infusion of the ß-adrenergic receptor agonist, Isoproterenol (ISO), as well as Acetylcholine (ACH, endothelium-dependent vasodilation) and Nitroprusside (NTP, endothelium-independent vasodilation). Forearm vascular conductance was calculated (FVC = FBF/MAP, ml/min/100 mmHg) and the rise in FVC from baseline during infusion quantified vasodilation (ΔFVC = FVCinfusion - FVCbaseline). RESULTS: ISO increased FVC in both groups (p < 0.01) and ISO-mediated ΔFVC was greater in OC+ compared to OC- (Main effect of group, p = 0.02). Expressing data as FVC and FBF resulted in similar conclusions. FVC responses to both ACH and NTP were also greater in OC+ compared to OC-. CONCLUSIONS: These data are the first to demonstrate greater ß-adrenergic receptor-mediated vasodilation in the forearm of women currently using oral contraceptives (placebo phase) when compared to those not using oral contraceptives (early follicular phase), and suggest oral contraceptive use influences neurovascular control.

Cerebral blood flow regulation in women across menstrual phase: differential contribution of cyclooxygenase to basal, hypoxic, and hypercapnic vascular tone.

In healthy young women, basal cerebral blood flow (CBF) and cerebrovascular reactivity may change across the menstrual cycle, but mechanisms remain untested. When compared with the early follicular phase of the menstrual cycle, we hypothesized women in late follicular phase would exhibit: 1) greater basal CBF, 2) greater hypercapnic increases in CBF, 3) greater hypoxic increases in CBF, and 4) increased cyclooxygenase (COX) signaling. We measured middle cerebral artery velocity (MCAv, transcranial Doppler ultrasound) in 11 healthy women (23 ± 1 yr) during rest, hypoxia, and hypercapnia. Subjects completed four visits: two during the early follicular (∼day 3) and two during the late follicular (∼day 14) phases of the menstrual cycle, with and without COX inhibition (oral indomethacin). Isocapnic hypoxia elicited an SPO2 = 90% and SPO2 = 80% for 5 min each. Separately, hypercapnia increased end-tidal CO2 10 mmHg above baseline. Cerebral vascular conductance index (CVCi = MCAv/MABP·100, where MABP is mean arterial blood pressure) was calculated and a positive change reflected vasodilation (ΔCVCi). Basal CVCi was greater in the late follicular phase (P < 0.001). Indomethacin decreased basal CVCi (∼37%) and abolished the phase difference (P < 0.001). Hypoxic ΔCVCi was similar between phases and unaffected by indomethacin. Hypercapnic ΔCVCi was similar between phases, and indomethacin decreased hypercapnic ΔCVCi (∼68%; P < 0.001) similarly between phases. In summary, while neither hypercapnic nor hypoxic vasodilation is altered by menstrual phase, increased basal CBF in the late follicular phase is fully explained by a greater contribution of COX. These data provide new mechanistic insight into anterior CBF regulation across menstrual phases and contribute to our understanding of CBF regulation in women.

ß-Adrenergic-mediated vasodilation in young men and women: cyclooxygenase restrains nitric oxide synthase.

We tested the hypothesis that women exhibit greater vasodilator responses to ß-adrenoceptor stimulation compared with men. We further hypothesized women exhibit a greater contribution of nitric oxide synthase and cyclooxygenase to ß-adrenergic-mediated vasodilation compared with men. Forearm blood flow (Doppler ultrasound) was measured in young men (n = 29, 26 ± 1 yr) and women (n = 33, 25 ± 1 yr) during intra-arterial infusion of isoproterenol (ß-adrenergic agonist). In subset of subjects, isoproterenol responses were examined before and after local inhibition of nitric oxide synthase [N(G)-monomethyl-l-arginine (l-NMMA); 6 male/10 female] and/or cyclooxygenase (ketorolac; 5 male/5 female). Vascular conductance (blood flow ÷ mean arterial pressure) was calculated to assess vasodilation. Vascular conductance increased with isoproterenol infusion (P < 0.01), and this effect was not different between men and women (P = 0.41). l-NMMA infusion had no effect on isoproterenol-mediated dilation in men (P > 0.99) or women (P = 0.21). In contrast, ketorolac infusion markedly increased isoproterenol-mediated responses in both men (P < 0.01) and women (P = 0.04) and this rise was lost with subsequent l-NMMA infusion (men, P < 0.01; women, P < 0.05). ß-Adrenergic vasodilation is not different between men and women and sex differences in the independent contribution of nitric oxide synthase and cyclooxygenase to ß-mediated vasodilation are not present. However, these data are the first to demonstrate ß-adrenoceptor activation of cyclooxygenase suppresses nitric oxide synthase signaling in human forearm microcirculation and may have important implications for neurovascular control in both health and disease.

Quantitative cerebrovascular 4D flow MRI at rest and during hypercapnia challenge.

UNLABELLED: Non-invasive measurement of cerebral blood flow (CBF) in humans is fraught with technologic, anatomic, and accessibility issues, which has hindered multi-vessel hemodynamic analysis of the cranial vasculature. Recent developments in cardiovascular MRI have allowed for the measurement of cine velocity vector fields over large imaging volumes in a single acquisition with 4D flow MRI. The purpose of this study was to develop an imaging protocol to simultaneously measure pulsatile flow in the circle of Willis as well as the carotid and vertebrate arteries at rest and during increased CO2 (hypercapnia). METHODS: 8 healthy adults (3 women, 26±0.4years) completed this study. Heart rate (pulse oximetry), arterial oxygen saturation (pulse oximetry), blood pressure (MAP, sphygmomanometry), and end-tidal CO2 (capnograph) were measured at rest (baseline) and during hypercapnia. Hypercapnia was induced via breathing a mixed gas of 3% CO2 and 21% O2 (balance N2) in the MR magnet. CBF and vessel cross-sectional area were quantified in 11 arteries using a 4D flow MRI scan, lasting 5-6min with a radially undersampled acquisition and an isotropic spatial resolution of 0.7mm. RESULTS: Baseline total CBF was 665±54ml • min(-1). Hypercapnia increased total CBF 9±3% to 721±61ml • min(-1). Hypercapnic increases in CBF ranged from 7 to 36% by artery, with the largest increases in the left anterior cerebral artery. Increases in artery cross-sectional area were observed in basilar and vertebral arteries. CONCLUSION: 4D flow MRI methods are sensitive enough to detect non-uniform changes in CBF and cross-sectional area to a mild yet clinically relevant CO2 stimulus. 4D flow MRI is a non-invasive reliable tool providing high spatio-temporal resolution in clinically feasible scan times without contrast agent. This approach can be used to interrogate regional cerebrovascular control in health and disease.

Cerebrovascular regulation in men and women: stimulus-specific role of cyclooxygenase.

Greater cerebral artery vasodilation mediated by cyclooxygenase (COX) in female animals is unexplored in humans. We hypothesized that young, healthy women would exhibit greater basal cerebral blood flow (CBF) and greater vasodilation during hypoxia or hypercapnia compared to men, mediated by a larger contribution of COX. We measured middle cerebral artery velocity (MCAv, transcranial Doppler ultrasound) in 42 adults (24 women, 18 men; 24 ± 1 years) during two visits, in a double-blind, placebo-controlled design (COX inhibition, 100 mg oral indomethacin, Indo). Women were studied early in the follicular phase of the menstrual cycle (days 1-5). Two levels of isocapnic hypoxia (SPO2 = 90% and 80%) were induced for 5-min each. Separately, hypercapnia was induced by increasing end-tidal carbon dioxide (PETCO 2) 10 mmHg above baseline. A positive change in MCAv (ΔMCAv) reflected vasodilation. Basal MCAv was greater in women compared to men (P < 0.01) across all conditions. Indo decreased baseline MCAv (P < 0.01) similarly between sexes. Hypoxia increased MCAv (P < 0.01), but ΔMCAv was not different between sexes. Indo did not alter hypoxic vasodilation in either sex. Hypercapnia increased MCAv (P < 0.01), but ΔMCAv was not different between sexes. Indo elicited a large decrease in hypercapnic vasodilation (P < 0.01) that was similar between sexes. During the early follicular phase, women exhibit greater basal CBF than men, but similar vasodilatory responses to hypoxia and hypercapnia. Moreover, COX is not obligatory for hypoxic vasodilation, but plays a vital and similar role in the regulation of basal CBF (~30%) and hypercapnic response (~55%) between sexes.

Exercise vasodilation is greater in women: contributions of nitric oxide synthase and cyclooxygenase.

PURPOSE: We hypothesized exercise vasodilation would be greater in women due to nitric oxide synthase (NOS) and cyclooxygenase (COX) signaling. METHODS: 45 healthy adults (23 women, W, 22 men, M, 26 ± 1 years) completed two 10-min trials of dynamic forearm exercise at 15 % intensity. Forearm blood flow (FBF; Doppler ultrasound), arterial pressure (brachial catheter), and forearm lean mass were measured to calculate relative forearm vascular conductance (FVCrel) = FBF 100 mmHg(-1) 100 g(-1) lean mass. Local intra-arterial infusion of L-NMMA or ketorolac acutely inhibited NOS and COX, respectively. In Trial 1, the first 5 min served as control exercise (CON), followed by 5 min of L-NMMA or ketorolac over the last 5 min of exercise. In Trial 2, the remaining drug was infused during 5-10 min, to achieve combined NOS-COX inhibition (double blockade, DB). RESULTS: Are mean ± SE. Women exhibited 29 % greater vasodilation in CON (ΔFVCrel, 19 ± 1 vs. 15 ± 1, p = 0.01). L-NMMA reduced ΔFVCrel (p < 0.001) (W: Δ -2.3 ± 1.3 vs. M: Δ -3.7 ± 0.8, p = 0.25); whereas, ketorolac modestly increased ΔFVCrel (p = 0.04) similarly between sexes (W: Δ 1.6 ± 1.1 vs. M: Δ 2.0 ± 1.6, p = 0.78). DB was also found to be similar between the sexes (p = 0.85). CONCLUSION: These data clearly indicate women produce a greater exercise vasodilator response. Furthermore, contrary to experiments in animal models, these data are the first to demonstrate vascular control by NOS and COX is similar between sexes.

Preserved Microvascular Endothelial Function in Young, Obese Adults with Functional Loss of Nitric Oxide Signaling.

Data indicate endothelium-dependent dilation (EDD) may be preserved in the skeletal muscle microcirculation of young, obese adults. Preserved EDD might be mediated by compensatory mechanisms, impeding insight into preclinical vascular dysfunction. We aimed to determine the functional roles of nitric oxide synthase (NOS) and cyclooxygenase (COX) toward EDD in younger obese adults. We first hypothesized EDD would be preserved in young, obese adults. Further, we hypothesized a reduced contribution of NOS in young, obese adults would be replaced by increased COX signaling. Microvascular EDD was assessed with Doppler ultrasound and brachial artery infusion of acetylcholine (ACh) in younger (27 ± 1 year) obese (n = 29) and lean (n = 46) humans. Individual and combined contributions of NOS and COX were examined with intra-arterial infusions of l-NMMA and ketorolac, respectively. Vasodilation was quantified as an increase in forearm vascular conductance (ΔFVC). Arterial endothelial cell biopsies were analyzed for protein expression of endothelial nitric oxide synthase (eNOS). ΔFVC to ACh was similar between groups. After l-NMMA, ΔFVC to ACh was greater in obese adults (p < 0.05). There were no group differences in ΔFVC to ACh with ketorolac. With combined NOS-COX inhibition, ΔFVC was greater in obese adults at the intermediate dose of ACh. Surprisingly, arterial endothelial cell eNOS and phosphorylated eNOS were similar between groups. Younger obese adults exhibit preserved EDD and eNOS expression despite functional dissociation of NOS-mediated vasodilation and similar COX signaling. Compensatory NOS- and COX-independent vasodilatory mechanisms conceal reduced NOS contributions in otherwise healthy obese adults early in life, which may contribute to vascular dysfunction.

Exercise-mediated vasodilation in human obesity and metabolic syndrome: effect of acute ascorbic acid infusion.

We tested the hypothesis that infusion of ascorbic acid (AA), a potent antioxidant, would alter vasodilator responses to exercise in human obesity and metabolic syndrome (MetSyn). Forearm blood flow (FBF, Doppler ultrasound) was measured in lean, obese, and MetSyn adults (n = 39, 32 ± 2 yr). A brachial artery catheter was inserted for blood pressure monitoring and local infusion of AA. FBF was measured during dynamic handgrip exercise (15% maximal effort) with and without AA infusion. To account for group differences in blood pressure and forearm size, and to assess vasodilation, forearm vascular conductance (FVC = FBF/mean arterial blood pressure/lean forearm mass) was calculated. We examined the time to achieve steady-state FVC (mean response time, MRT) and the rise in FVC from rest to steady-state exercise (Δ, exercise - rest) before and during acute AA infusion. The MRT (P = 0.26) and steady-state vasodilator responses to exercise (ΔFVC, P = 0.31) were not different between groups. Intra-arterial infusion of AA resulted in a significant increase in plasma total antioxidant capacity (174 ± 37%). AA infusion did not alter MRT or steady-state FVC in any group (P = 0.90 and P = 0.85, respectively). Interestingly, higher levels of C-reactive protein predicted longer MRT (r = 0.52, P < 0.01) and a greater reduction in MRT with AA infusion (r = -0.43, P = 0.02). We concluded that AA infusion during moderate-intensity, rhythmic forearm exercise does not alter the time course or magnitude of exercise-mediated vasodilation in groups of young lean, obese, or MetSyn adults. However, systemic inflammation may limit the MRT to exercise, which can be improved with AA.