Kazuhide Ayajiki

Cerebral Blood Flow Regulation by Nitric Oxide: Recent Advances

Nitric oxide (NO) is undoubtedly quite an important intercellular messenger in cerebral and peripheral hemodynamics. This molecule, formed by constitutive isomers of NO synthase, endothelial nitric-oxide synthase, and neuronal nitric-oxide synthase, plays pivotal roles in the regulation of cerebral blood flow and cell viability and in the protection of nerve cells or fibers against pathogenic factors associated with cerebral ischemia, trauma, and hemorrhage. Cerebral blood flow is increased and cerebral vascular resistance is decreased by NO derived from endothelial cells, autonomic nitrergic nerves, or brain neurons under resting and stimulated conditions. Somatosensory stimulation also evokes cerebral vasodilatation mediated by neurogenic NO. Oxygen and carbon dioxide alter cerebral blood flow and vascular tone mainly via constitutively formed NO. Endothelial dysfunction impairs cerebral hemodynamics by reducing the bioavailability of NO and increasing the production of reactive oxygen species (ROS). The NO-ROS interaction is an important issue in discussing blood flow and cell viability in the brain. Recent studies on brain circulation provide quite useful information concerning the physiological roles of NO produced by constitutive isoforms of nitric-oxide synthase and how NO may promote cerebral pathogenesis under certain conditions, including cerebral ischemia/stroke, cerebral vasospasm after subarachnoid hemorrhage, and brain injury. This information would contribute to better understanding of cerebral hemodynamic regulation and its dysfunction and to development of novel therapeutic measures to treat diseases of the central nervous system.

Evidence for a Causal Role of the Renin-Angiotensin System in Vascular Dysfunction Associated With Insulin Resistance

Abstract—Excess production of superoxide anion in response to angiotensin II plays a central role in the transduction of signal molecules and the regulation of vascular tone. We examined the ability of insulin resistance to stimulate superoxide anion production and investigated the identity of the oxidases responsible for its production. Rats were fed diets containing 60% fructose (fructose-fed rats) or 60% starch (control rats) for 8 weeks. In aortic homogenates from fructose-fed rats, the superoxide anion generated in response to NAD(P)H was more than 2-fold higher than that of control rats. Pretreatment of the aorta from fructose-fed rats with inhibitors of NADPH oxidase significantly reduced superoxide anion production. In the isolated aorta, contraction induced by angiotensin II was more potent in fructose-fed rats compared with control rats. Losartan normalized blood pressure, NAD(P)H oxidase activity, endothelial function, and angiotensin II-induced vasoconstriction in fructose-fed rats. To elucidate the molecular mechanisms of the enhanced constrictor response to angiotensin II, expressions of angiotensin II receptor and subunits of NADPH oxidase were examined with the use of angiotensin II type 1a receptor knockout (AT1a KO) mice. Expression of AT1a receptor mRNA was enhanced in fructose-fed mice, whereas expression of either AT1b or AT2 was unaltered. In addition, protein expression of each subunit of NADPH oxidase was increased in fructose-fed mice, whereas the expression was significantly decreased in fructose-fed AT1a KO mice. The novel observation of insulin resistance-induced upregulation of AT1 receptor expression could explain the association of insulin resistance with endothelial dysfunction and hypertension.

Cerebral vasodilatation induced by stimulation of the pterygopalatine ganglion and greater petrosal nerve in anesthetized monkeys

Although brain cell viability depends largely on cerebral circulation, mechanisms of blood flow control, such as autoregulation, or of the pathogenesis of functionally impaired blood supply to brain regions, such as in cerebral vasospasm after subarachnoid hemorrhage, have not been clearly defined. Our recent studies support the hypothesis that nitric oxide, released from nitrergic nerves, plays a crucial role as a neurotransmitter in vasodilating cerebral arteries from primate and subprimate mammals. In the present study, we demonstrated, by using arterial angiography, that electrical stimulation of the pterygopalatine ganglion produced vasodilatation of ipsilateral cerebral arteries of anesthetized Japanese monkeys. The response was abolished by intravenous injections of N(G)-nitro-L-arginine, a nitric oxide synthase inhibitor. Denervation of the ganglion elicited cerebral vasoconstriction, indicating that vasodilator nerves from the vasomotor center were tonically active. Stimulation of the greater petrosal nerve, upstream of the pterygopalatine ganglion, also elicited cerebral vasodilatation, which was abolished by treatment with the nitric oxide synthase inhibitor and with hexamethonium, indicating that the nerve is in connection via synapses with the nitrergic nerve innervating cerebral arteries. Endogenous nitric oxide released from the nerve may contribute to the maintenance of blood flow in major cerebral arteries necessary to supply blood to the different brain regions. Without this influence, cerebral arteries might be constricted to the extent that blood flow is impeded. This is the first direct evidence indicating an important role of nitric oxide liberated by pre- and postganglionic nerve stimulation in the control of cerebral arterial tone in primates.

Impairment by damage of the pterygopalatine ganglion of nitroxidergic vasodilator nerve function in canine cerebral and retinal arteries.

Histochemical study revealed that transcutaneous injection of ethanol into the vicinity of the pterygopalatine ganglion greatly decreased the positive staining for NADPH diaphorase activity after 1 week in the ipsilateral ganglion of a dog and abolished the staining of perivascular nerves in the middle and posterior cerebral arteries. Transmural electrical stimulation or nicotine produced a relaxation in middle and posterior cerebral arteries isolated from the side with the nontreated ganglion (control side), whereas the relaxation was abolished or reversed to a contraction in the arteries from the side with the ethanol-treated ganglion. Nitric oxide-induced relaxations did not differ in the arteries from both sides. The response to nerve stimulation of the control arteries was suppressed by treatment with NG-nitro-L-arginine (L-NA), an inhibitor of nitric oxide synthase, and the inhibition was reversed by L-arginine. Nicotine produced a contraction followed by a relaxation in central retinal arterial strips obtained from the control side; the relaxation was abolished and the contraction was potentiated in the arteries from the treated side. The nicotine-induced relaxation was abolished by L-NA, and the contraction was suppressed by phentolamine. On the other hand, the nicotine-induced relaxation in superficial temporal arteries, susceptible to L-NA, was not attenuated by treatment with ethanol. The findings obtained so far support our hypothesis that nitric oxide released from the vasodilator nerve acts as a transmitter to produce arterial smooth muscle relaxation and suggest that the nerve fibers to the cerebral and retinal arteries arise from the pterygopalatine ganglion.

Preganglionic and Postganglionic Neurons Responsible for Cerebral Vasodilation Mediated by Nitric Oxide in Anesthetized Dogs

The authors performed investigations to functionally determine the route of efferent innervation in vivo responsible for cerebral vasodilation mediated by nitric oxide (NO). In anesthetized beagles, electrical stimulation of the pterygopalatine ganglion vasodilated ipsilateral cerebral arteries such as the middle cerebral and posterior communicating arteries. Intravenous injections of NG-nitro-L-arginine (L-NA) markedly inhibited the response to nerve stimulation, and the effect was reversed by L-arginine. Stimulation of the proximal portion of the greater superficial petrosal nerve, upstream of the pterygopalatine ganglion, also produced cerebral vasodilation, which was abolished by L-NA and restored by L-arginine. Treatment with hexamethonium abolished the response to stimulation of the petrosal nerve but did not affect the response to pterygopalatine ganglion stimulation. Destruction of the pterygopalatine ganglion by cauterization constricted the cerebral arteries. Postganglionic denervation abolished the vasodilation, lacrimation, and nasal secretion induced on the ipsilateral side by stimulation of the pterygopalatine ganglion and petrosal nerve. The vasodilator response was suppressed by L-NA but unaffected by atropine, whereas lacrimation and nasal secretion were abolished solely by atropine. It is concluded that postganglionic neurons from the pterygopalatine ganglion play crucial roles in cerebral vasodilation mediated by NO from the nerve, and preganglionic neurons, possibly from the superior salivatory nucleus through the greater superficial petrosal nerve, innervate the pterygopalatine ganglion. Tonic discharges from the vasomotor center participate significantly in the maintenance of cerebral vasodilation.

Mechanisms underlying arginine vasopressin-induced relaxation in monkey isolated coronary arteries.

OBJECTIVE The present study was undertaken to examine whether arginine vasopressin (AVP) relaxes primate coronary artery and to analyse the mechanisms of its action in reference to endothelial nitric oxide and AVP receptor subtype. METHODS Isometrical tension responses to AVP and desmopressin were recorded in isolated monkey coronary arteries. RESULTS AVP (10(-9) to 10(-7) mol/l) induced a concentration-related relaxation; endothelium-denudation abolished the response. Treatment with N(G)-nitro-L-arginine, but not the D-enantiomer, abolished the endothelium-dependent relaxation, which was restored by L-arginine. Treatment with SR49059 and [Pmp1,Tyr(Me)2]-Arg8-vasopressin, selective inhibitors of V1 receptor subtype, attenuated the relaxant response to AVP, whereas the relaxation induced by sodium nitroprusside was not affected by SR49059. Desmopressin, a V2 receptor agonist, up to 10(-8) mol/l did not elicit relaxation. CONCLUSIONS It is concluded that AVP-induced monkey coronary arterial relaxation is mediated via nitric oxide synthesized from L-arginine in association with stimulation of V1 receptor subtypes in the endothelium.

Inhibition of nitroxidergic nerve function by neurogenic acetylcholine in monkey cerebral arteries

1. Modification by endogenous or exogenous acetylcholine and vasoactive intestinal polypeptide (VIP) of vasodilatation mediated by nitric oxide (NO) released from nitroxidergic nerves was studied in isolated monkey cerebral arteries. In arterial strips denuded of endothelium, transmural electrical stimulation (2‐20 Hz) produced relaxations that were abolished by tetrodotoxin. 2. The relaxation response was attenuated by acetylcholine, and the attenuation was reversed by atropine. Attenuation was also observed with AF‐DX 116, an antagonist of the muscarinic acetylcholine receptor subtype, M2. NO‐induced relaxation was not affected by acetylcholine. Neurogenic relaxation was also inhibited by physostigmine and potentiated by atropine. 3. VIP in concentrations that elicited slight relaxation did not alter the response to nerve stimulation. In the strips showing tachyphylaxis to VIP, the neurogenic response was not inhibited. 4. Histochemical studies of whole‐mount preparations revealed nerve fibres with NO synthase and VIP immunoreactivity, and also acetylcholinesterase, suggesting the presence of perivascular nitroxidergic, VIPergic and cholinergic innervation. 5. It is concluded that the actions of nitroxidergic nerve fibres on the monkey cerebral artery are inhibited by nerve‐released acetylcholine acting on prejunctional muscarinic receptors, possibly of the M2 subtype. Despite the presence of VIP immunoreactive nerve fibres and the ability of exogenous VIP to relax the artery, there is no evidence supporting either a prejunctional modulation of nitroxidergic nerve function by VIP or a role for VIP as a vasodilatory neurotransmitter.

Nitric oxide mediates, and acetylcholine modulates, neurally induced relaxation of bovine cerebral arteries

Helical strips of bovine basilar arteries denuded of the endothelium responded to transmural electrical stimulation with frequency-dependent relaxations that were abolished or markedly attenuated by treatment with tetrodotoxin, oxyhemoglobin and Methylene Blue. Relaxations induced by vasoactive intestinal polypeptide and calcitonin gene-related peptide were not affected by oxyhemoglobin and Methylene Blue. The neurally induced relaxation was not attenuated in the artery made unresponsive to these peptides by successive application. The relaxation caused by nerve stimulation was markedly inhibited by treatment with NG-nitro-L-arginine, a nitric oxide synthase inhibitor, which did not inhibit the relaxation caused by exogenously applied nitric oxide. The inhibition was reversed by L-arginine but not by the D-enantiomer. Exogenously applied acetylcholine did not alter the tone of endothelium-denuded arteries. Neurally induced relaxations were attenuated by treatment with acetylcholine and physostigmine and were significantly potentiated by atropine. It may be concluded that the relaxation induced by nerve stimulation is mediated by nitric oxide, but not by vasoactive intestinal polypeptide or calcitonin gene-related peptide, derived from vasodilator nerves innervating the bovine basilar artery, and the nerve function is inhibited prejunctionally via muscarinic receptor activation by acetylcholine released from cholinergic nerves but is not influenced by vasoactive intestinal polypeptide.

Interaction of Endothelial Nitric Oxide and Angiotensin in the Circulation

Discovery of the unexpected intercellular messenger and transmitter nitric oxide (NO) was the highlight of highly competitive investigations to identify the nature of endothelium-derived relaxing factor. This labile, gaseous molecule plays obligatory roles as one of the most promising physiological regulators in cardiovascular function. Its biological effects include vasodilatation, increased regional blood perfusion, lowering of systemic blood pressure, and antithrombosis and anti-atherosclerosis effects, which counteract the vascular actions of endogenous angiotensin (ANG) II. Interactions of these vasodilator and vasoconstrictor substances in the circulation have been a topic that has drawn the special interest of both cardiovascular researchers and clinicians. Therapeutic agents that inhibit the synthesis and action of ANG II are widely accepted to be essential in treating circulatory and metabolic dysfunctions, including hypertension and diabetes mellitus, and increased availability of NO is one of the most important pharmacological mechanisms underlying their beneficial actions. ANG II provokes vascular actions through various receptor subtypes (AT1, AT2, and AT4), which are differently involved in NO synthesis and actions. ANG II and its derivatives, ANG III, ANG IV, and ANG-(1-7), alter vascular contractility with different mechanisms of action in relation to NO. This review article summarizes information concerning advances in research on interactions between NO and ANG in reference to ANG receptor subtypes, radical oxygen species, particularly superoxide anions, ANG-converting enzyme inhibitors, and ANG receptor blockers in patients with cardiovascular disease, healthy individuals, and experimental animals. Interactions of ANG and endothelium-derived relaxing factor other than NO, such as prostaglandin I2 and endothelium-derived hyperpolarizing factor, are also described.

Conversion of angiotensin I to angiotensin II in dog isolated renal artery : role of two different angiotensin II-generating enzymes

The functional role of the endothelium in conversion of angiotensin (Ang) I to Ang II was studied in helical strips of dog renal arteries. In the arteries precontracted with PGF2α, Angs I and II caused a moderate relaxation, which was abolished by treatment with saralasin and reversed to a contraction by indomethacin. Removal of the endothelium attenuated the response to Ang I but did not abolish it. The Ang I-induced relaxation in the arteries without endothelium was not significantly attenuated by an Ang-converting enzyme inhibitor. SA446, but was markedly suppressed by chymostatin. On the other hand, in the arteries with endothelium. the relaxation was suppressed but not abolished by SA446, and the remaining relaxation was abolished by additional treatment with chymostatin. Relaxation induced by prostaglandin I2 was unaffected by these enzyme inhibitors. These results strongly suggest that the conversion of Ang I to Ang II is due mainly to the Ang-converting enzyme in the endothelium and to the chymostatin-sensitive Ang II-generating enzyme in subendothelial tissues.