Characterization of the effects of two new arginine/citrulline analogues on constitutive and inducible nitric oxide synthases in rat aorta.
1. New potent inhibitors of nitric oxide synthase (NOS) may be useful in the treatment of septic shock, a disorder characterized by a vascular hyporeactivity to catecholamines caused by an overproduction of nitric oxide (NO-). We examined the effects of L-thiocitrulline (L-TC) and S-methyl-L-thiocitrulline (L-SMTC), novel NOS inhibitors, on the constitutive and inducible NOS in rat aorta and compared those effects with inhibition due to NG-methyl-L-arginine (L-NMA). 2. Phenylephrine evoked similar concentration-contraction curves in the control rings and in the rings treated with these different NOS inhibitors (10 microM), whereas 100 microM of L-NMA, L-TC or L-SMTC increased significantly, and to a similar extent, contractions evoked by phenylephrine in aortic rings with endothelium without significantly affecting the maximal responses. 3. Relaxations evoked by acetylcholine, adenosine triphosphate, or calcium ionophore were significantly inhibited in a dose-dependent manner by L-NMA, L-SMTC, or L-TC (10-100 microM). The potencies of these inhibitors in reducing the relaxations of these vasodilators were not significantly different. 4. In endotoxin-treated preparations with endothelium, the three L-arginine analogues (10 microM) significantly potentiated contractile responses to phenylephrine (pEC50: 6.73 +/- 0.12 and 7.3 +/- 0.12, 7.34 +/- 0.13, or 7.22 +/- 0.14; in the absence and the presence of L-NMA, L-TC, or L-SMTC respectively) and increased maximal contractions from 1.53 +/- 0.15 g to 1.95 +/- 0.13 g, 2.08 +/- 0.12 g, and 2.03 +/- 0.13 g with L-NMA, L-TC, and L-SMTC respectively. A higher concentration of these NOS inhibitors (100 microM)further increased contractions evoked by this alpha1-agonist without further enhancing the maximal contractions; however, contractions evoked by 10 nM phenylephrine were significantly greater in the presence of L-SMTC or L-TC than in the presence of L-NMA (100 microM) (L-NMA: 0.4 +/- 0.11 g; L-TC:0.78 +/- 0.14 g and L-SMTC: 0.82+/-0.17 g). The effects of these inhibitors on NO- synthesis induced by endotoxin were significantly reversed by addition of L-arginine (1 mM) but not by L-citrulline (1 mM). InLPS-treated rings with endothelium, all three NOS inhibitors (100 microM) shifted the concentration contraction curves evoked by phenylephrine significantly to the left (pEC5o: 7.19 +/- 0.03 and 7.79 +/- 0.08,8.01 +/- 0.07, or 8.02 +_ 0.07, in the absence and the presence of L-NMA, L-TC, or L-SMTC, respectively)and increased significantly maximal contractions from 2.05 +/- 0.05 g to 2.38 +/- 0.14 g, 2.5 +/- 0.12 g, and 2.4 +_ 0.21 g with L-NMA, L-TC, and L-SMTC, respectively. L-TC and L-SMTC were significantly more potent than L-NMA in potentiating contractions evoked by 10 nM and 30 nM phenylephrine.5. L-TC and L-SMTC produced dose-dependent increases in tone in LPS-treated aortic rings with and without endothelium. In LPS-treated rings with endothelium, L-NMA induced contractions but in preparations without endothelium low concentrations of L-NMA induced small contractions while high concentrations of this inhibitor evoked relaxations. In both preparations L-TC and L-SMTC were significantly more potent than L-NMA in increasing vascular tone.6. These results suggest that L-SMTC, L-TC and L-NMA were equipotent on basal and agonist stimulated NO- synthesis produced by the constitutive isoform of NOS, whereas the two new L-arginine analogues were more potent than L-NMA in inhibiting the production of NO- induced by endotoxin in rat aorta.
Joly
G A
GA
Department of Genitourinary Oncology, University of Texas M.D. Anderson Cancer Center, Houston 77030, USA.
Narayanan
K
K
Griffith
O W
OW
Kilbourn
R G
RG
D000109:Acetylcholine / Q000494:pharmacology; D000244:Adenosine Diphosphate / Q000494:pharmacology; D000704:Analysis of Variance; D000818:Animals; D001013:Aorta, Thoracic / Q000187:drug effects* / Q000201:enzymology; D001120:Arginine / Q000031:analogs & derivatives / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002956:Citrulline / Q000031:analogs & derivatives / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004790:Enzyme Induction / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007476:Ionophores / Q000494:pharmacology; D008297:Male; D009119:Muscle Contraction / Q000187:drug effects; D009126:Muscle Relaxation / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000187:drug effects*; D009569:Nitric Oxide / Q000037:antagonists & inhibitors*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010656:Phenylephrine / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D013890:Thiourea / Q000031:analogs & derivatives / Q000494:pharmacology; D014662:Vasoconstrictor Agents / Q000494:pharmacology; D019323:omega-N-Methylarginine
D004791:Enzyme Inhibitors; D007476:Ionophores; D014662:Vasoconstrictor Agents; C086715:thiocitrulline; D010656:Phenylephrine; D019323:omega-N-Methylarginine; D002956:Citrulline; D009569:Nitric Oxide; D000001:Calcimycin; D000244:Adenosine Diphosphate; D001120:Arginine; D019001:Nitric Oxide Synthase; D013890:Thiourea; C086717:S-methylthiocitrulline; D000109:Acetylcholine