Husain Shabeeh

Effects of Neuronal Nitric Oxide Synthase on Human Coronary Artery Diameter and Blood Flow In Vivo

Background— Nitric oxide (NO)-mediated local regulation of vascular tone is considered to involve endothelial NO synthase (eNOS). However, we recently reported that human forearm basal microvascular tone in vivo is tonically regulated by neuronal NO synthase (nNOS), in contrast to an acetylcholine-stimulated reduction in tone, which is eNOS dependent. Here, we investigated the in vivo effects of an nNOS-selective inhibitor, S-methyl-l-thiocitrulline (SMTC), on the human coronary circulation and on flow-mediated dilatation in the forearm. Methods and Results— In patients with angiographically normal coronary arteries, intracoronary infusion of SMTC (0.625 &mgr;mol/min) reduced basal coronary blood flow by 34.1±5.2% (n=10; P<0.01) and epicardial coronary diameter by 3.6±1.2% (P=0.02) but had no effect on increases in flow evoked by intracoronary substance P (20 pmol/min). The nonselective NOS inhibitor NG-monomethyl-l-arginine (25 &mgr;mol/min) also reduced basal coronary flow (by 22.3±5.3%; n=8; P<0.01) but, in contrast to SMTC, inhibited substance P-induced increases in flow (P<0.01). In healthy volunteers, local infusion of SMTC (0.2 &mgr;mol/min) reduced radial artery blood flow by 36.0±6.4% (n=10; P=0.03) but did not affect flow-mediated dilatation (P=0.55). In contrast, NG-monomethyl-l-arginine (2 &mgr;mol/min) infusion reduced radial blood flow to a similar degree (by 39.7±11.8%; P=0.02) but also inhibited flow-mediated dilatation by ≈80% (P<0.01). Conclusions— These data indicate that local nNOS-derived NO regulates basal blood flow in the human coronary vascular bed, whereas substance P-stimulated vasodilatation is eNOS mediated. Thus, nNOS and eNOS have distinct local roles in the physiological regulation of human coronary vascular tone in vivo.

Sympathetic activation increases NO release from eNOS but neither eNOS nor nNOS play an essential role in exercise hyperemia in the human forearm

Nitric oxide (NO) release from endothelial NO synthase (eNOS) and/or neuronal NO synthase (nNOS) could be modulated by sympathetic nerve activity and contribute to increased blood flow after exercise. We examined the effects of brachial-arterial infusion of the nNOS selective inhibitor S-methyl-l-thiocitrulline (SMTC) and the nonselective NOS inhibitor N(G)-monomethyl-l-arginine (l-NMMA) on forearm arm blood flow at rest, during sympathetic activation by lower body negative pressure, and during lower body negative pressure immediately after handgrip exercise. Reduction in forearm blood flow by lower body negative pressure during infusion of SMTC was not significantly different from that during vehicle (-28.5 ± 4.02 vs. -34.1 ± 2.96%, respectively; P = 0.32; n = 8). However, l-NMMA augmented the reduction in forearm blood flow by lower body negative pressure (-44.2 ± 3.53 vs. -23.4 ± 5.71%; n = 8; P < 0.01). When lower body negative pressure was continued after handgrip exercise, there was no significant effect of either l-NMMA or SMTC on forearm blood flow immediately after low-intensity exercise (P = 0.91 and P = 0.44 for l-NMMA vs. saline and SMTC vs. saline, respectively; each n = 10) or high-intensity exercise (P = 0.46 and P = 0.68 for l-NMMA vs. saline and SMTC vs. saline, respectively; each n = 10). These results suggest that sympathetic activation increases NO release from eNOS, attenuating vasoconstriction. Dysfunction of eNOS could augment vasoconstrictor and blood pressure responses to sympathetic activation. However, neither eNOS nor nNOS plays an essential role in postexercise hyperaemia, even in the presence of increased sympathetic activation.

Blood Pressure in Healthy Humans Is Regulated by Neuronal NO Synthase.

NO is physiologically generated by endothelial and neuronal NO synthase (nNOS) isoforms. Although nNOS was first identified in brain, it is expressed in other tissues, including perivascular nerves, cardiac and skeletal muscle. Increasing experimental evidence suggests that nNOS has important effects on cardiovascular function, but its composite effects on systemic hemodynamics in humans are unknown. We undertook the first human study to assess the physiological effects of systemic nNOS inhibition on basal hemodynamics. Seventeen healthy normotensive men aged 24±4 years received acute intravenous infusions of an nNOS-selective inhibitor, S-methyl-L-thiocitrulline, and placebo on separate occasions. An initial dose-escalation study showed that S-methyl-L-thiocitrulline (0.1–3.0 µmol/kg) induced dose-dependent changes in systemic hemodynamics. The highest dose of S-methyl-L-thiocitrulline (3.0 µmol/kg over 10 minutes) significantly increased systemic vascular resistance (+42±6%) and diastolic blood pressure (67±1 to 77±3 mm Hg) when compared with placebo (both P<0.01). There were significant decreases in heart rate (60±4 to 51±3 bpm; P<0.01) and left ventricular stroke volume (59±6 to 51±6 mL; P<0.01) but ejection fraction was unaltered. S-methyl-L-thiocitrulline had no effect on radial artery flow-mediated dilatation, an index of endothelial NOS activity. These results suggest that nNOS-derived NO has an important role in the physiological regulation of basal systemic vascular resistance and blood pressure in healthy humans.

Differential role of endothelial versus neuronal nitric oxide synthase in the regulation of coronary blood flow during pacing-induced increases in cardiac workload

Endothelial nitric oxide synthase (eNOS) was assumed to be the only source of nitric oxide (NO) involved in the regulation of human coronary blood flow (CBF). However, our recent first-in-human study using the neuronal NOS (nNOS)-selective inhibitor S-methyl-L-thiocitrulline (SMTC) showed that nNOS-derived NO also plays a role. In this study, we investigated the relative contribution of nNOS and eNOS to the CBF response to a pacing-induced increase in cardiac workload. Incremental right atrial pacing was undertaken in patients with angiographically normal coronary arteries during intracoronary infusion of saline vehicle and then either SMTC or N(G)-monomethyl-l-arginine (l-NMMA; which inhibits both eNOS and nNOS). Intracoronary SMTC (0.625 μmol/min) and l-NMMA (25 μmol/min) reduced basal CBF to a similar extent (-19.2 ± 3.2% and 25.0 ± 2.7%, respectively; n = 10 per group). Pacing-induced increases in CBF were significantly blunted by l-NMMA (maximum CBF: 83.5 ± 14.2 ml/min during saline vs. 61.6 ± 9.5 ml/min during l-NMMA; P < 0.01). By contrast, intracoronary SMTC had no effect on the maximum CBF during pacing (98.5 ± 12.9 ml/min during saline vs. 102.1 ± 16.6 ml/min during SMTC; P = not significant). l-NMMA also blunted the pacing-induced increase in coronary artery diameter (P < 0.001 vs. saline), whereas SMTC had no effect. Our results confirm a role of nNOS in the regulation of basal CBF in humans but show that coronary vasodilation in response to a pacing-induced increase in cardiac workload is exclusively mediated by eNOS-derived NO.

Impaired Neuronal Nitric Oxide Synthase–Mediated Vasodilator Responses to Mental Stress in Essential Hypertension

Neuronal NO synthase (nNOS) regulates blood flow in resistance vasculature at rest and during mental stress. To investigate whether nNOS signaling is dysfunctional in essential hypertension, forearm blood flow responses to mental stress were examined in 88 subjects: 48 with essential hypertension (42±14 years; blood pressure, 141±17/85±15 mm Hg; mean±SD) and 40 normotensive controls (38±14 years; 117±13/74±9 mm Hg). A subsample of 34 subjects (17 hypertensive) participated in a single blind 2-phase crossover study, in which placebo or sildenafil 50 mg PO was administered before an intrabrachial artery infusion of the selective nNOS inhibitor S-methyl-L-thiocitrulline (SMTC, 0.05, 0.1, and 0.2 &mgr;mol/min) at rest and during mental stress. In a further subsample (n=21) with an impaired blood flow response to mental stress, responses were measured in the presence and absence of the &agr;-adrenergic antagonist phentolamine. The blood flow response to mental stress was impaired in hypertensive compared with normotensive subjects (37±7% versus 70±8% increase over baseline; P<0.001). SMTC blunted responses to mental stress in normotensive but not in hypertensive subjects (reduction of 40±11% versus 3.0±14%, respectively, P=0.01, between groups). Sildenafil reduced the blood flow response to stress in normotensive subjects from 89±14% to 43±14% (P<0.03) but had no significant effect in hypertensive subjects. Phentolamine augmented impaired blood flow responses to mental stress from 39±8% to 67±13% (P<0.02). Essential hypertension is associated with impaired mental stress–induced nNOS-mediated vasodilator responses; this may relate to increased sympathetic outflow in hypertension. nNOS dysfunction may impair vascular homeostasis in essential hypertension and contribute to stress-induced cardiovascular events.

The human coronary vasodilatory response to acute mental stress is mediated by neuronal nitric oxide synthase.

Acute mental stress induces vasodilation of the coronary microvasculature. Here, we show that this response involves neuronal nitric oxide synthase in the human coronary circulation.

ROLE OF NEURONAL VS ENDOTHELIAL NITRIC OXIDE SYNTHASE IN THE CORONARY BLOOD FLOW RESPONSE TO PACING

Background Endothelial nitric oxide synthase (eNOS) has been assumed to be the major source of nitric oxide (NO) regulating human coronary blood flow (CBF). In recent first-in-human studies with a neuronal NOS (nNOS)-selective inhibitor, we provided evidence that nNOS-derived NO regulates basal coronary blood flow whereas eNOS mediates increases in flow in response to the endothelial agonist, substance P. The present study aimed to investigate the effects of nNOS vs eNOS inhibition on coronary blood flow response to increased heart rate. Methods We studied the effects of the nNOS-selective inhibitor, S-methyl-L-thiocitrulline (SMTC), and the non-selective NOS inhibitor, NG-monomethyl-L-arginine (L-NMMA) at doses previously shown to inhibit nNOS or both nNOS and eNOS, respectively. 18 patients already undergoing elective cardiac catheterisation for clinical reasons and found to have normal coronary arteries were included. An intracoronary Doppler flow wire was positioned in the coronary artery for measurement of blood flow velocity whereas coronary artery diameter was measured by quantitative angiography. An incremental pacing protocol that raised heart rate to a maximum of 150 bpm was undertaken in all patients via a temporary right atrial pacing wire. Pacing was performed in the presence of saline vehicle and then either L-NMMA or SMTC (one inhibitor per patient; n=9 each group). Results SMTC (0.625 μmol/min) and L-NMMA (25 μmol/min) both reduced basal CBF to a similar extent (−22.8%±1.24% vs −26.8%±2.16%; n=9 each; p=NS). During saline infusion, CBF increased with atrial pacing from 58.7±9.90 to 87.4±17.3 ml/min (n=9, p<0.05). During L-NMMA, the increase in CBF was significantly blunted compared to that during saline (ΔCBF 17.7±4.54 ml/min vs 28.7±8.08 ml/min during saline; n=9, p<0.05 by 2-way ANOVA). In patients receiving SMTC, however, the increase in CBF with pacing was similar to that during saline (ΔCBF 36.5±6.05 ml/min vs 25.2±6.65 ml/min during saline; n=9, p=NS by 2-way ANOVA). SMTC and L-NMMA both reduced basal coronary artery diameter to a similar extent (n=9). L-NMMA blunted the pacing-induced increase in coronary artery diameter (n=9, p<0.05 vs saline vehicle) whereas SMTC had no effect (n=9, p=NS). Conclusion These results suggest that increases in human coronary blood flow in response to incremental atrial pacing are mediated by eNOS-derived NO rather than nNOS-derived NO.

Diagnosis and management of stable angina in primary care

Stable angina is a common presentation in primary care. It is a symptom indicative of myocardial ischaemia rather than a disease itself. Two million people in the UK suffer from angina, and coronary heart disease accounts for around one in six deaths in the UK. Therefore, it is of pivotal importance that doctors should be informed about the symptoms of angina and their management.