Prostaglandin I2 does not contribute to the hypotensive effect of the superoxide dismutase mimetic Tempo in rats with aortic coarctation-induced hypertension.

This study was designed to investigate the contribution of prostaglandins to the vasodepressor effect of the superoxide dismutase mimetic Tempo in rats made hypertensive by ligation of the abdominal aorta at a point between the left and right renal arteries. Rings of thoracic aorta taken from rats with aortic coarctation released more 6-keto-PGF1alpha (a non-enzymatic product of PGI2 degradation) in the presence than in the absence of Tempo (1 mmol/L; 35.3 +/- 10.1 versus 13.6 +/- 2.6 pg/mg tissue). However, Tempo administered intravenously (2 mg/kg bolus injection plus infusion at 3 mg/kg/h) to rats with aortic coarctation did not increase significantly the concentration of 6-keto-PGF1alpha in vena cava blood. Treatment with Tempo did not affect the arterial pressure of un-operated normotensive rats but promptly decreased the arterial pressure of rats with aortic coarctation-induced hypertension (from 178 +/- 2 to 125 +/- 6 mmHg). The vasodepressor effect of Tempo in hypertensive animals was not affected by pretreatment with indomethacin to inhibit prostaglandin synthesis. These data argue against the hypothesis that PGI2 contributes to the acute hypotensive effect of Tempo in rats with aortic coarctation.

Modulation by 20-HETE of phenylephrine-induced mesenteric artery contraction in spontaneously hypertensive and Wistar-Kyoto rats.

Small mesenteric arteries of spontaneously hypertensive (SHR) and Wistar-Kyoto rats (WKY) were compared for the production of 20-HETE and the effects of 20-HETE and N-methylsulfonyl-12,12-dibromododec-11-enamide (DDMS, 30 micromol/L), a 20-HETE synthesis inhibitor, on contractile responsiveness to phenylephrine (0.1 to 50.0 micromol/L). 20-HETE production was higher in vessels of SHR compared with WKY (1.34+/-0.16 versus 0.27+/-0.09 pmol/mg tissue, P<0.05). Phenylephrine elicited concentration-dependent vascular contraction; the R(max) was similar in vessels of SHR and WKY, but the former were more sensitive as denoted by the lower EC(50) (1.10+/-0.14 versus 1.89+/-0.33 micromol/L, P<0.05). DDMS caused a rightward shift in the concentration-response curve to phenylephrine, increasing (P<0.05) the EC(50) by 258% and 134% in vessels of SHR and WKY, respectively. In contrast, in DDMS-treated vessels, 20-HETE (0.01 to 10.0 micromol/L) caused a leftward shift in the phenylephrine concentration-response curve, decreasing (P<0.05) the EC(50) without affecting the R(max). Importantly, the minimal concentration of 20-HETE that decreased the EC(50) of phenylephrine was much smaller in vessels of SHR that of WKY (0.01 versus 1.0 micromol/L). We conclude that 20-HETE increases the sensitivity of mesenteric arterial vessels to phenylephrine, vessels of SHR are more sensitive to this action of the eicosanoid than vessels of WKY, and vessels of SHR produce more 20-HETE than do vessels of WKY. Hence, 20-HETE of vascular origin may be a determinant of the increased reactivity to constrictor agonists in the vasculature of SHR.

Human heme oxygenase-1 gene transfer lowers blood pressure and promotes growth in spontaneously hypertensive rats.

Heme oxygenase (HO) catalyzes the conversion of heme to biliverdin, with release of free iron and carbon monoxide. Both heme and carbon monoxide have been implicated in the regulation of vascular tone. A retroviral vector containing human HO-1 cDNA (LSN-HHO-1) was constructed and subjected to purification and concentration of the viral particles to achieve 5x10(9) to 1x10(10) colony-forming units per milliliter. The ability of concentrated infectious viral particles to express human HO-1 (HHO-1) in vivo was tested. A single intracardiac injection of the concentrated infectious viral particles (expressing HHO-1) to 5-day-old spontaneously hypertensive rats resulted in functional expression of the HHO-1 gene and attenuation of the development of hypertension. Rats expressing HHO-1 showed a significant decrease in urinary excretion of a vasoconstrictor arachidonic acid metabolite and a reduction in myogenic responses to increased intraluminal pressure in isolated arterioles. Unexpectedly, HHO-1 chimeric rats showed a simultaneous significant proportionate increase in somatic growth. Thus, delivery of HHO-1 gene by retroviral vector attenuates the development of hypertension and promotes body growth in spontaneously hypertensive rats.

Vasoregulatory function of the heme-heme oxygenase-carbon monoxide system.

Arterial vessels express one or more heme oxygenase (HO) isoenzymes that catalyze the metabolism of heme to carbon monoxide (CO) and biliverdin. Carbon monoxide promotes vasorelaxation through mechanisms that, depending on the vessels, involve activation of soluble guanylate cyclase, stimulation of calcium-activated potassium channels, or diminished synthesis of constrictor mediators, such as, endothelin and 20-HETE. Inhibitors of HO elicit vasoconstriction in vivo and in isolated pressurized arterioles. Inhibitors of HO also enhance myogenic vasoconstriction, as well as the constriction induced by phenylephrine in several vessels. The blood pressure of awake rats is increased by acute treatment with HO inhibitors, a response that is accompanied by attenuation of baroreflex activity. All in all, it would appear that a product of HO activity manufactured by arterial vessels, presumably CO, promotes vasodilation and decreases the reactivity of vascular smooth muscle to myogenic stimuli and constrictor agonists. In doing so, CO of vascular origin may contribute to the implementation of antihypertensive mechanisms. Carbon monoxide produced in central nervous system structures, for example, the nucleus tractus solitarii, also appears to support a blood pressure-lowering mechanism linked to inhibitory modulation of baroreceptor reflex activity.

Carbon monoxide produced by isolated arterioles attenuates pressure-induced vasoconstriction.

Studies were conducted on isolated rat gracilis muscle arterioles to examine the role of vascular heme oxygenase (HO)-derived carbon monoxide (CO) on myogenic constrictor responses to stepwise increments in intraluminal pressure. The arterioles express HO-2 but not HO-1 and manufacture CO. Both HO-2 protein expression and CO production are reduced in arterioles maintained for 18 h before experimentation in media containing HO-2 antisense oligodeoxynucleotides (AS-ODN). Pressurization of arterioles mounted on a myograph over the pressure range of 40--100 mmHg elicits reduction of internal diameter. At pressures >40 mmHg, the internal diameter of vessels treated with either HO-2 AS-ODN, the HO inhibitor chromium mesoporphyrin (CrMP), or the K(+) channel blocker tetraethylammonium (TEA) are smaller than the corresponding control values. The inclusion of exogenous CO, but not of biliverdin, in the superfusion buffer attenuates pressure-induced vasoconstriction in CrMP-treated vessels. However, exogenous CO does not attenuate pressure-induced vasoconstriction in vessels treated with both CrMP and TEA. Collectively, these data suggest that CO of vascular origin attenuates pressure-induced arteriolar constriction via a mechanism involving a TEA-sensitive K(+) channel.

Carbon monoxide of vascular origin attenuates the sensitivity of renal arterial vessels to vasoconstrictors.

Rat renal interlobar arteries express heme oxygenase 2 (HO-2) and manufacture carbon monoxide (CO), which is released into the headspace gas. CO release falls to 30% and 54% of control, respectively, after inhibition of HO activity with chromium mesoporphyrin (CrMP) or of HO-2 expression with antisense oligodeoxynucleotides (HO-2 AS-ODN). Patch-clamp studies revealed that CrMP decreases the open probability of a tetraethylammonium-sensitive (TEA-sensitive) 105 pS K channel in interlobar artery smooth muscle cells, and that this effect of CrMP is reversed by CO. Assessment of phenylephrine-induced tension development revealed reduction of the EC(50) in vessels treated with HO-2 AS-ODN, CrMP, or TEA. Exogenous CO greatly minimized the sensitizing effect on agonist-induced contractions of agents that decrease vascular CO production, but not the sensitizing effect of K channel blockade with TEA. Collectively, these data suggest that vascular CO serves as an inhibitory modulator of vascular reactivity to vasoconstrictors via a mechanism that involves a TEA-sensitive K channel.

Contributions of prostacyclin and nitric oxide to carbon monoxide-induced cerebrovascular dilation in piglets.

Carbon monoxide (CO) is an endogenous dilator in the newborn cerebral microcirculation. Other dilators include prostanoids and nitric oxide (NO), and interactions among the systems are likely. Experiments on anesthetized piglets with cranial windows address the hypothesis that CO-induced dilation of pial arterioles involves interaction with the prostanoid and NO systems. Topical application of CO or the heme oxygenase substrate heme-L-lysinate (HLL) produced dilation. Indomethacin, N(omega)-nitro-L-arginine (L-NNA), and either iberiotoxin or tetraethylammonium chloride (TEA) were used to inhibit prostanoids, NO, and Ca(2+)-activated K(+) (K(Ca)) channels, respectively. Indomethacin, L-NNA, iberiotoxin, or TEA blocked cerebral vasodilation to CO and HLL. Vasodilations to both CO and HLL were returned to indomethacin-treated piglets by topical application of iloprost. Vasodilations to both CO and HLL were returned to L-NNA-treated piglets by sodium nitroprusside but not iloprost. In iberiotoxin- or TEA-treated piglets, dilations to CO and HLL could not be restored by either iloprost or sodium nitroprusside. The dilator actions of CO involve prostacyclin and NO as permissive enablers. The permissive actions of prostacyclin and NO may alter the K(Ca) channel response to CO because neither iloprost nor sodium nitroprusside could restore dilation to CO when these channels were blocked.

Expression of myogenic constrictor tone in the aorta of hypertensive rats.

We investigated the effect of intraluminal pressure or stretch on the development of tone in the descending thoracic aorta from rats with aortic coarctation-induced hypertension of 7-14 days duration. Increments of pressure >100 mmHg decreased the diameter of thoracic aortas from hypertensive but not from normotensive rats. The pressure-induced constriction was not demonstrable in vessels superfused with calcium-free buffer. Stretched rings of aorta from hypertensive rats exhibited a calcium-dependent constrictor tone accompanied by elevated calcium influx that varied in relation to the degree of stretch. Blockers of L-type calcium channels and inhibitors of protein kinase C reduced both basal tone and calcium influx in aortic rings of hypertensive rats. Hence, the thoracic aorta of hypertensive rats expresses a pressure- and stretch-activated constrictor mechanism that relies on increased calcium influx through L-type calcium channels via a protein kinase C-regulated pathway. The expression of such a constrictor mechanism is suggestive of acquired myogenic behavior.

CYP4A1 antisense oligonucleotide reduces mesenteric vascular reactivity and blood pressure in SHR.

The cytochrome P-450 4A (CYP4A)-derived arachidonic acid metabolite 20-hydroxyeicosatetraenoic acid (20-HETE) affects renal tubular and vascular functions and has been implicated in the control of arterial pressure. We examined the effect of antisense oligonucleotide (ODN) to CYP4A1, the low K(m) arachidonic acid omega-hydroxylating isoform, on vascular 20-HETE synthesis, vascular reactivity, and blood pressure in the spontaneously hypertensive rat (SHR). Administration of CYP4A1 antisense ODN decreased mean arterial blood pressure from 137 +/- 3 to 121 +/- 4 mmHg (P < 0.05) after 5 days of treatment, whereas treatment with scrambled antisense ODN had no effect. Treatment with CYP4A1 antisense ODN reduced the level of CYP4A-immunoreactive proteins along with 20-HETE synthesis in mesenteric arterial vessels. Mesenteric arteries from rats treated with antisense ODN exhibited decreased sensitivity to the constrictor action of phenylephrine (EC(50) 0.69 +/- 0.17 vs. 1.77 +/- 0.40 microM). Likewise, mesenteric arterioles from antisense ODN-treated rats revealed attenuation of myogenic constrictor responses to increases of transmural pressure. The decreased vascular reactivity and myogenic responses were reversible with the addition of 20-HETE. These data suggest that CYP4A1-derived 20-HETE facilitates myogenic constrictor responses in the mesenteric microcirculation and contributes to pressor mechanisms in SHR.

Vasopressin and PGE(2) regulate activity of apical 70 pS K(+) channel in thick ascending limb of rat kidney.

Vasopressin and prostaglandin E(2) (PGE(2)) are involved in regulating NaCl reabsorption in the thick ascending limb (TAL) of the rat kidney. In the present study, we used the patch-clamp technique to study the effects of vasopressin and PGE(2) on the apical 70 pS K(+) channel in the rat TAL. Addition of vasopressin increased the channel activity, defined as NP(o), from 1.11 to 1.52 (200 pM) and 1.80 (500 pM), respectively. The effect of vasopressin can be mimicked by either forskolin (1-5 microM) or 8-bromo-cAMP/dibutyryl-cAMP (8-Br-cAMP/DBcAMP) (200-500 microM). Moreover, the effects of cAMP and vasopressin were not additive and application of 10 microM H-89 abolished the effect of vasopressin. This suggests that the effect of vasopressin is mediated by a cAMP-dependent pathway. Applying 10 nM PGE(2) alone had no significant effect on the channel activity. However, PGE(2) (10 nM) abolished the stimulatory effect of vasopressin. The PGE(2)-induced inhibition of the vasopressin effect was the result of decreasing cAMP production because addition of 200 microM 8-Br-cAMP/DBcAMP reversed the PGE(2)-induced inhibition. In addition to antagonizing the vasopressin effect, high concentrations of PGE(2) reduced channel activity in the absence of vasopressin by 33% (500 nM) and 51% (1 microM), respectively. The inhibitory effect of high concentrations of PGE(2) was not the result of decreasing cAMP production because adding the membrane-permeant cAMP analog failed to restore the channel activity. In contrast, inhibiting protein kinase C (PKC) with calphostin C (100 nM) abolished the effect of 1 microM PGE(2). We conclude that PGE(2) inhibits apical K(+) channels by two mechanisms: 1) low concentrations of PGE(2) attenuate the vasopressin-induced stimulation mainly by reducing cAMP generation, and 2) high concentrations of PGE(2) inhibit the channel activity by a PKC-dependent pathway.

Lipoxygenase-dependent mechanisms in hypertension.

This study was designed to examine the contribution of lipoxygenase products to mechanisms of vascular contraction and elevated blood pressure in rats with aortic coarctation-induced hypertension. In cytosolic fractions of aortae taken from hypertensive rats, 12-lipoxygenase protein was increased as compared to normotensive controls. Aortic rings from hypertensive, but not from normotensive rats, exhibited a basal tone which was reduced 74+/-12 and 71+/-22%, respectively, by the lipoxygenase inhibitors cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate (CDC, 10(-5) mol/L) and 5,8,11-eicosatriynoic acid (ETI, 10(-5) mol/L). CDC (8 mg/kg s.c.) did not affect the blood pressure of normotensive rats but decreased that of hypertensive rats from 182+/-6 to 151+/-10 mm Hg. The blood pressure lowering effect of CDC was blunted in hypertensive rats pretreated with indomethacin or antibodies against 5,6-dihydro-prostaglandin I2. These data suggest contribution of lipoxygenase-derived products to mechanisms underlying aortic smooth muscle basal tone and elevated blood pressure in rats with aortic coarctation-induced hypertension. The vasodepressor effect of CDC depends on a mechanism involving vasodilatory prostaglandins.

Amlodipine enhances NO production induced by an ACE inhibitor through a kinin-mediated mechanism in canine coronary microvessels.

Our previous study found that angiotensin-converting enzyme (ACE) inhibitors and amlodipine induce NO release from coronary microvessels through a kinin-dependent mechanism. The goal of this study was to determine whether amlodipine could potentiate NO formation during ACE inhibition. Coronary microvessels were isolated from 16 mongrel dogs. Nitrite, the hydration product of NO, from coronary microvessels was quantified by using the Griess reaction. Bradykinin and kallikrein all significantly increased nitrite release from coronary microvessels in a concentration-dependent manner. The ACE inhibitor, ramiprilat, potentiated these effects. Amlodipine also markedly potentiated nitrite production by ramiprilat. For instance, amlodipine (10(-10) M) enhanced nitrite release induced by ramiprilat (10(-7) M) from 122 +/- 9 to 168 +/- 14 pmol/mg (p < 0.05 vs. ramiprilat). Nitrite release potentiated by ramiprilat and amlodipine was entirely blocked by N(omega)-nitro-L-arginine methyl ester (L-NAME, an inhibitor of NO synthase), HOE 140 (Icatibant, a specific B2-kinin receptor antagonist), and dichloroisocoumarin (DCIC, a serine protease inhibitor that blocks local kinin formation). These results clearly show that there is a synergistic effect on NO formation when amlodipine is combined with ACE inhibition. Our data suggest that kinin-mediated coronary NO production may contribute importantly to the beneficial therapeutic action of ACE inhibitors, especially in combination with amlodipine in the treatment of heart disease.

Amlodipine promotes kinin-mediated nitric oxide production in coronary microvessels of failing human hearts.

Recently, we found that amlodipine can release nitric oxide (NO) from canine coronary microvessels, which raises the question of whether amlodipine can also promote coronary NO production in failing human hearts. The goal of this study was to define the effect of amlodipine on NO production in failing human hearts and to determine the role of kinins in the control of NO production induced by amlodipine. Six explanted human hearts with end-stage heart failure were obtained immediately at transplant surgery. Coronary microvessels were isolated as previously described, and nitrite, the stable metabolite of NO in aqueous solution, was measured using the Griess Reaction. Amlodipine (10(-10) to 10(-5) mol/L) significantly increased nitrite production in coronary microvessels in a dose-dependent manner. The increase in nitrite in response to the highest dose of amlodipine (79%) was similar in magnitude to either that of the angiotensin-converting enzyme inhibitor ramiprilat (74%) or the neutral endopeptidase inhibitors phosphoramidon (61%) and thiorphan (72%). Interestingly, the increase in nitrite production induced by amlodipine was entirely abolished by N(omega)-nitro-L-arginine methyl ester and also HOE-140 (a bradykinin-2 antagonist) and dichloroisocoumarin (a serine protease inhibitor that blocks kallikrein activity). These results indicate that amlodipine can promote coronary NO production in failing human hearts and that this effect is dependent on a kinin-mediated mechanism.

Carbon monoxide and cerebral microvascular tone in newborn pigs.

The present study addresses the hypothesis that CO produced from endogenous heme oxygenase (HO) can dilate newborn cerebral arterioles. HO-2 protein was highly expressed in large and small blood vessels, as well as parenchyma, of newborn pig cerebrum. Topically applied CO dose-dependently dilated piglet pial arterioles in vivo over the range 10(-11)-10(-9) M (maximal response). CO-induced cerebrovascular dilation was abolished by treatment with the Ca2+-activated K+ channel inhibitors tetraethylammonium chloride and iberiotoxin. The HO substrate heme-L-lysinate also produced tetraethylammonium-inhibitable, dose-dependent dilation from 5 x 10(-8) to 5 x 10(-7) M (maximal). The HO inhibitor chromium mesoporphyrin blocked dilation of pial arterioles in response to heme-L-lysinate. In addition to inhibiting dilation to heme-L-lysinate, chromium mesoporphyrin also blocked pial arteriolar dilations in response to hypoxia but did not alter responses to hypercapnia or isoproterenol. We conclude that CO dilates pial arterioles via activation of Ca2+-activated K+ channels and that endogenous HO-2 potentially can produce sufficient CO to produce the dilation.

Contribution of endogenous carbon monoxide to regulation of diameter in resistance vessels.

Endogenous carbon monoxide was proposed to subserve vasodepressor functions. If so, inhibition of heme oxygenase may be expected to promote vascular contraction. This hypothesis was examined in large and small arteries and in isolated first-order gracilis muscle arterioles of rat. The heme oxygenase inhibitors chromium mesoporphyrin (CrMP) and cobalt protoporphyrin (0.175-102 micromol/l) decreased the diameter of pressurized (80 mmHg) gracilis muscle arterioles, whereas magnesium protoporphyrin, a weak heme oxygenase inhibitor, did not. CrMP also elicited development of isometric tension in the muscular branch of the femoral artery but not in the aorta or femoral artery. Arteriolar constrictor responses to CrMP varied in relation to the intravascular pressure, were blunted in preparations exposed to exogenous carbon monoxide (100 micromol/l), and were unaffected by an endothelin receptor antagonist. Importantly, CrMP amplified the constrictor response to increases of pressure in gracilis arterioles. Accordingly, the constrictor effect of heme oxygenase inhibitors is attributable to magnification of myogenic tone due to withdrawal of a vasodilatory mechanism mediated by endogenous carbon monoxide. The study suggests that the vascular carbon monoxide system plays a role in the regulation of basal tone in resistance vessels.

Carbon monoxide stimulates the apical 70-pS K+ channel of the rat thick ascending limb.

We have investigated the expression of heme oxygenase (HO) in the rat kidney and the effects of HO-dependent heme metabolites on the apical 70-pS K+ channel in the thick ascending limb (TAL). Reverse transcriptase-PCR (RT-PCR) and Western blot analyses indicate expression of the constitutive HO form, HO-2, in the rat cortex and outer medulla. Patch-clamping showed that application of 10 microM chromium mesoporphyrin (CrMP), an inhibitor of HO, reversibly reduced the activity of the apical 70-pS K+ channel, defined by NPo, to 26% of the control value. In contrast, addition of 10 microM magnesium protoporphyrin had no significant effect on channel activity. HO involvement in regulation of the apical 70-pS K+ channel of the TAL, was further indicated by the addition of 10 microM heme-L-lysinate, which significantly stimulated the channel activity in cell-attached patches by 98%. The stimulatory effect of heme on channel activity was also observed in inside-out patches in the presence of 0.5-1 mM reduced nicotinamide adenine dinucleotide phosphate. This was completely abolished by 10 microM CrMP, suggesting that a HO-dependent metabolite of heme mediated the effect. This was further supported by exposure of the cytosolic membrane of inside-out patches to a carbon monoxide-bubbled bath solution, which increased channel activity. Moreover, carbon monoxide completely abolished the effect of 10 microM CrMP on the channel activity. In contrast, 10 microM biliverdin, another HO-dependent metabolite of heme, had no effect. We conclude that carbon monoxide produced from heme via an HO-dependent metabolic pathway stimulates the apical 70-pS K+ channel in the rat TAL.

Contribution of cytochrome P-450 4A1 and 4A2 to vascular 20-hydroxyeicosatetraenoic acid synthesis in rat kidneys.

20-Hydroxyeicosatetraenoic acids (20-HETE), a biologically active cytochrome P-450 (CYP) metabolite of arachidonic acid in the rat kidney, can be catalyzed by CYP4A isoforms including CYP4A1, CYP4A2, and CYP4A3. To determine the contribution of CYP4A isoforms to renal 20-HETE synthesis, specific antisense oligonucleotides (ODNs) were developed, and their specificity was examined in vitro in Sf9 cells expressing CYP4A isoforms and in vivo in Sprague-Dawley rats. Administration of CYP4A2 antisense ODNs (167 nmol. kg body wt-1. day-1 iv for 5 days) decreased vascular 20-HETE synthesis by 48% with no effect on tubular synthesis, whereas administration of CYP4A1 antisense ODNs inhibited vascular and tubular 20-HETE synthesis by 52 and 40%, respectively. RT-PCR of microdissected renal microvessel RNA indicated the presence of CYP4A1, CYP4A2, and CYP4A3 mRNAs, and a CYP4A1-immunoreactive protein was detected by Western analysis of microvessel homogenates. Blood pressure measurements revealed a reduction of 17 +/- 6 and 16 +/- 4 mmHg in groups receiving CYP4A1 and CYP4A2 antisense ODNs, respectively. These studies implicate CYP4A1 as a major 20-HETE synthesizing activity in the rat kidney and further document the feasibility of using antisense ODNs to specifically inhibit 20-HETE synthesis and thereby investigate its role in the regulation of renal function and blood pressure.

Kinin-mediated coronary nitric oxide production contributes to the therapeutic action of angiotensin-converting enzyme and neutral endopeptidase inhibitors and amlodipine in the treatment in heart failure.

Increasing evidence suggests that angiotensin-converting enzyme (ACE) inhibitors can increase vascular nitric oxide (NO) production. Recent studies have found that combined inhibition of ACE and neutral endopeptidase (NEP) may have a greater beneficial effect in the treatment of heart failure than inhibition of ACE alone. Amlodipine, a calcium channel antagonist, has also been reported to have a favorable effect in the treatment of patients with cardiac dysfunction. The purpose of this study was to determine whether and the extent to which all of these agents used in the treatment of heart failure stimulate vascular NO production. Heart failure was induced by rapid ventricular pacing in conscious dogs. Coronary microvessels were isolated from normal and failing dog hearts. Nitrite, the stable metabolite of NO, was measured by the Griess reaction. ACE and NEP inhibitors and amlodipine significantly increased nitrite production from coronary microvessels in both normal and failing dog hearts. However, nitrite release was reduced after heart failure. For instance, the highest concentration of enalaprilat, thiorphan, and amlodipine increased nitrite release from 85 +/- 4 to 156 +/- 9, 82 +/- 7 to 139 +/- 8, and 74 +/- 4 to 134 +/-10 pmol/mg (all *p <.01 versus control), respectively, in normal dog hearts. Nitrite release in response to the highest concentration of these two inhibitors and amlodipine was reduced by 41% and 31% and 32% (all #p <.01 versus normal), respectively, in microvessels after heart failure. The increase in nitrite induced by either ACE or NEP inhibitors or amlodipine was entirely abolished by Nw-nitro-L-arginine methyl ester, HOE 140 (a B2-kinin receptor antagonist), and dichloroisocoumarin (a serine protease inhibitor) in both groups. Our results indicate that: 1) there is an impaired endothelial NO production after pacing-induced heart failure; 2) both ACE and NEP are largely responsible for the metabolism of kinins and modulate canine coronary NO production in normal and failing heart; and 3) amlodipine releases NO even after heart failure and this may be partly responsible for the favorable effect of amlodipine in the treatment of heart failure. Thus, the restoration of reduced coronary vascular NO production may contribute to the beneficial effects of these agents in the treatment of heart failure.

Involvement of nitric oxide and potassium channels in the reduction of basal tone produced by blockade of thromboxane A2/prostaglandin H2 receptors in aortic rings of hypertensive rats.

This study was designed to investigate involvement of potassium channels in the action of nitric oxide facilitating reduction of basal tone by thromboxane A2/prostaglandin H2 receptor blockade with ifetroban in rings of thoracic aorta taken from rats with aortic coarctation-induced hypertension. Ifetroban-induced reduction of basal tone in aortic rings without drug pretreatment was attenuated (P<0.05) in rings pretreated with the nitric oxide synthesis inhibitor N(omega-nitro-L-arginine methyl ester (L-NAME; 3 x 10(-4) mol/L; 0.55+/-0.09 g versus 0.23+/-0.07 g). The vasorelaxing effect of ifetroban also was decreased (P<0.05) in preparations pretreated with a potassium channel blocker, either tetraethylammonium (TEA; 10(-2) mol/L) or 4-aminopyridine (4-AP; 3 x 10(-3) mol/L). Ifetroban-induced reduction of basal tone was not attenuated in preparations pretreated first with L-NAME and then with sodium nitroprusside (SNP; 6+/-1 nmol/L) to compensate for the loss of endogenous nitric oxide. However, the facilitatory effect of SNP on ifetroban-induced relaxation of aortic rings pretreated with L-NAME alone was not demonstrable in rings pretreated with L-NAME plus TEA or 4-AP. These observations suggest that a mechanism involving nitric oxide and potassium channels facilitates the reduction in basal tone produced by ifetroban in aortic rings of rats with aortic coarctation-induced hypertension.

Role of nitric oxide in the control of renal oxygen consumption and the regulation of chemical work in the kidney.

Inhibition of NO synthesis has recently been shown to increase oxygen extraction in vivo, and NO has been proposed to play a significant role in the regulation of oxygen consumption by both skeletal and cardiac muscle in vivo and in vitro. It was our aim to determine whether NO also has such a role in the kidney, a tissue with a relatively low basal oxygen extraction. In chronically instrumented conscious dogs, administration of an inhibitor of NO synthase, nitro-L-arginine (NLA, 30 mg/kg i.v.), caused a maintained increase in mean arterial pressure and renal vascular resistance and a decrease in heart rate (all P<0.05). At 60 minutes, urine flow rate and glomerular flow rate decreased by 44+/-12% and 45+/-7%, respectively; moreover, the amount of sodium reabsorbed fell from 16+/-1.7 to 8.5+/-1.1 mmol/min (all P<0.05). At this time, oxygen uptake and extraction increased markedly by 115+/-37% and 102+/-34%, respectively (P<0.05). Oxygen consumption also significantly increased from 4.5+/-0.6 to 7.1+/-0.9 mL O2/min. Most important, the ratio of oxygen consumption to sodium reabsorbed increased dramatically from 0.33+/-0.07 to 0.75+/-0.11 mL O2/mmol Na+ (P<0.05), suggesting a reduction in renal efficiency for transporting sodium. In vitro, both a NO-donating agent and the NO synthase-stimulating agonist bradykinin significantly decreased both cortical and medullary renal oxygen consumption. In conclusion, NO plays a role in maintaining a balance between oxygen consumption and sodium reabsorption, the major ATP-consuming process in the kidney, in conscious dogs, and NO can inhibit mitochondrial oxygen consumption in canine renal slices in vitro.