Dilation Of Porcine Retinal Arterioles Research Articles

Dilation of porcine retinal arterioles to nobiletin, a polymethoxyflavonoid: Roles of nitric oxide and voltage-dependent potassium channel

We examined the effects of nobiletin, a polymethoxyflavonoid, on the retinal microvascular diameter to determine if they depend on the endothelium and/or smooth muscle to reveal the signaling mechanisms involved in this vasomotor activity. Porcine retinal arterioles were isolated, cannulated, and pressurized without flow in vitro. Video microscopic techniques recorded diametric responses to nobiletin. The retinal arterioles dilated in a nobiletin concentration-dependent (100 pM-10 μM) manner and decreased by 50% after endothelial removal. The nitric oxide (NO) synthase inhibitor, Nω-nitro-L-arginine methyl ester (L-NAME), reduced nobiletin-induced vasodilation comparable to denudation. Blockade of soluble guanylyl cyclase by 1H-[1,2,4] oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ) produced a similar inhibitory effect as that by L-NAME. Nobiletin-induced vasodilation was also inhibited by the nonselective potassium channel inhibitor, tetraethylammonium (TEA), and the voltage-gated K (Kv) inhibitor, 4-aminopyridine. Co-administration of L-NAME and TEA almost eliminated nobiletin-induced vasodilation. Nobiletin elicits both endothelium-dependent and -independent dilation of retinal arterioles mediated by NO release and Kv channel activation, respectively.

Histamine-Induced Dilation of Isolated Porcine Retinal Arterioles: Role of Endothelium-Derived Hyperpolarizing Factor.

Although endothelium-dependent nitric oxide (NO)-mediated dilation of retinal arterioles has been well described, the role of endothelium-derived hyperpolarizing factor (EDHF) in the retinal arteriolar response remains unclear. In the current study, we examined the contribution of EDHF to the retinal arteriolar dilation to the inflammatory agent histamine and investigated the signaling mechanisms underlying this vasomotor activity. Porcine retinal arterioles were isolated, cannulated, and pressurized without flow for functional study by using video microscopic techniques. The immunohistochemical staining was performed to determine histamine receptor subtypes. Histamine (0.1-30 μM) produced concentration-dependent dilation of retinal arterioles in a manner sensitive to H1- and H2-receptor antagonists chlorpheniramine and famotidine, respectively. Histamine-induced vasodilation was almost abolished after endothelial removal. In the intact vessels, vasodilation to histamine was partially inhibited by the inhibitors of cyclooxygenase (indomethacin), NO synthase (NG-nitro-L-arginine methyl ester, L-NAME), or Ca2+ -activated K+ (KCa) channels (apamin plus charybdotoxin). Combination of the above inhibitors abolished histamine-induced vasodilation. Residual vasodilation in the presence of indomethacin and L-NAME was further reduced by the cytochrome P450 enzyme inhibitor sulfaphenazole but not by the gap junction inhibitor carbenoxolone or the hydrogen peroxide scavenger catalase. Immunohistochemical signals for H1- and H2-receptor expression were found only in the endothelium. The endothelium plays an essential role in the dilation of porcine retinal arterioles to histamine via H1- and H2-receptor activation. The EDHF derived from cytochrome P450 contributed in part to this vasodilation via KCa channel activation, in addition to the endothelial release of NO and prostanoids.

Homocysteine Inhibition of Endothelium-Dependent Nitric Oxide-Mediated Dilation of Porcine Retinal Arterioles via Enhanced Superoxide Production

Elevated plasma concentration of homocysteine, a sulfur-containing amino acid, is an emerging risk factor for cardiovascular diseases. Recent epidemiologic studies have confirmed that elevated homocysteine levels are associated with ocular vascular diseases; however, the direct effect of homocysteine on ocular microvascular reactivity remains unknown. We investigated whether homocysteine affects endothelium-dependent nitric oxide (NO)-mediated dilation of retinal arterioles and whether oxidative stress and distinct protein kinase signaling pathways are involved in the homocysteine-mediated effect. Porcine retinal arterioles were isolated, cannulated, and pressurized without flow in vitro. Diameter changes were recorded using videomicroscopy techniques. Intraluminal treatment with homocysteine (1 mM, 180 minutes) significantly attenuated arteriolar dilation in response to the endothelium-dependent NO-mediated agonists bradykinin and A23187 but not in response to the endothelium-independent NO donor sodium nitroprusside. In the presence of the superoxide scavenger 4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPOL), the nicotinamide adenine dinucleotide phosphate-oxidase (NAD(P)H oxidase inhibitor apocynin, p38 kinase inhibitor SB203580, and peroxisome proliferator-activated receptor-γ (PPAR-γ) agonist pioglitazone, the detrimental effect of homocysteine on bradykinin-induced dilation was prevented; however, neither the xanthine oxidase inhibitor allopurinol, the JNK inhibitor SP600125, or pioglitazone with PPAR-γ inhibitor GW9662 had that effect. Homocysteine inhibits endothelium-dependent NO-mediated dilation in the retinal arterioles by producing superoxide from NAD(P)H oxidase, which appears to be linked with p38 kinase. By impairing endothelium-dependent NO-mediated vasoreactivity, homocysteine potentially facilitates development of retinal vascular diseases. In addition, pioglitazone can prevent homocysteine-induced endothelial dysfunction possibly by activating PPAR-γ.

Dilation of Porcine Retinal Arterioles to Cilostazol: Roles of eNOS Phosphorylation via cAMP/Protein Kinase A and AMP-Activated Protein Kinase and Potassium Channels

Cilostazol, a selective inhibitor of phosphodiesterase 3, has antiplatelet aggregation and peripheral vasodilation effects. We examined the effects of cilostazol on the retinal microvascular diameter to determine its dependence on the endothelium and/or smooth muscle to reveal the signaling mechanisms involved in this vasomotor activity. Porcine retinal arterioles were isolated, cannulated, and pressurized without flow in vitro. Video microscopic techniques recorded the diametric responses to cilostazol. The retinal arterioles dilated in response to cilostazol in a dose-dependent (100 pM-10 μM) manner; the dilation decreased by 60% after endothelial removal. The nitric oxide (NO) synthase inhibitor, NG-nitro-L-arginine methyl ester (L-NAME), inhibited cilostazol-induced vasodilation comparable to denudation. Inhibition of soluble guanylyl cyclase and blockade of protein kinase A (PKA) were comparable to L-NAME. Compound C, an AMP-activated protein kinase (AMPK) inhibitor, partially inhibited cilostazol-induced vasodilation, which exhibited a weaker inhibitory effect on cilostazol-induced vasodilation than blockade of PKA. The large-conductance Ca²⁺-activated K channel (BK(Ca) channel) blocker, iberiotoxin, also inhibited cilostazol-induced vasodilation. The residual vasodilation decreased further with co-administration of L-NAME and iberiotoxin. Cilostazol elicits endothelium-dependent and -independent dilation of the retinal arterioles mediated by NO release and BK(Ca) channel activation, respectively. Endothelial nitric oxide synthase (eNOS) phosphorylation via the cAMP/PKA and AMPK pathways and consequent activation of the soluble guanylyl cyclase/cyclic guanosine monophosphate pathway might play an important role in cilostazol-induced vasodilation of the retinal arterioles.