Activities of 7-Nitroindazole and 1-(2-(Trifluoromethylphenyl)-imidazole Independent of Neuronal Nitric-Oxide Synthase Inhibition

Abstract

7-Nitroindazole (NI) is a widely used inhibitor of neuronal nitricoxide synthase (nNOS) used to study the role of the neuronal NO pathway in the nervous system. 7-NI prevents convulsions, including 2-amino-4-methylphosphinobutyric acid (glufosinate)-induced convulsions, in experimental models. Herein, we examined nNOS involvement in glufosinate-induced convulsions and the specificity of 7-NI for nNOS. Another nNOS inhibitor, 1-[2-(trifluoromethyl)phenyl]imidazole (TRIM), inhibited NOS activity in vivo, and it prevented glufosinate-induced convulsions. In contrast, an endothelial NOS inhibitor, N 5 -(1-iminoethyl)-Lornithine, inhibited NOS activity in vivo, but it did not prevent the convulsions. These results suggest the involvement of nNOS in glufosinate-induced convulsions. However, a nonspecific NOS inhibitor, N -nitro-L-arginine methyl ester, inhibited NOS activity in vivo, but it failed to prevent glufosinate-induced convulsions. 6-NI and indazole, which did not inhibit NOS activity in vivo, suppressed glufosinate-induced convulsions. Moreover, glufosinate elicited convulsions in nNOS-deficient mice. These results suggest the anticonvulsant effects of 7-NI and TRIM on glufosinate-induced convulsions do not involve nNOS inhibition, instead possibly being related to an undefined property of nitrogen-containing chemical structures.

Regulation of Neuronal Glutathione Synthesis

“…In a study conducted by our group, pretreatment with 7-nitroindazole (7-NI), an nNOS inhibitor, also prevented both GSH depletion and nitrotyrosine formation induced by MPTP. Although we recently found 7-NI to be non-specific to nNOS (131), a previous report demonstrated attenuation of MPTP neurotoxicity in nNOS-deficient mice (132). These results suggest a major role of ONOO − derived from nNOS in MPTP-induced GSH depletion.…”

Neuronal Nitric Oxide Synthase Inhibition Prevents Cerebral Palsy following Hypoxia-Ischemia in Fetal Rabbits: Comparison between JI-8 and 7-Nitroindazole

“…However, the margin for inhibition between nNOS and eNOS is at best 4-fold. It has been speculated that 7-NI may act through an unknown mechanism other than nNOS inhibition [Matsumura et al, 2008].…”

On the selectivity of neuronal NOS inhibitors

“…S-Methyl- l -thiocitrulline (Furfine et al ., 1994) is also used, albeit less frequently, to selectively inhibit nNOS in vitro and in vivo , yet studies show that the compound is <10-fold selective for isolated nNOS over eNOS (Furfine et al ., 1994; Boer et al ., 2000; both using human NOS isozymes). The use of other compounds with moderate or good selectivity for isolated nNOS, such as S , S ′-1,4-phenylene-bis(1,2-ethanediyl)bis-isothiourea (100-fold selectivity over eNOS; tested using isolated human NOS by Boer et al ., 2000), 1[2-(trifluoromethyl)phenyl]-1 H -imidazole (TRIM; 38-fold selectivity over eNOS determined with homogenates of mouse cerebellum and bovine aortic endothelial cells; Handy et al ., 1995) and AR-R17477 (28-fold over eNOS using human isozymes, Brzozowski et al ., 2011; 100-fold for rat nNOS over human eNOS in partially purified preparations, Reif et al ., 2000), is also hampered by problems such as a lack of specificity (isothioureas, see Babu and Griffith, 1998a; TRIM, Matsumura et al ., 2008) and/or a lack of availability (AR-R17477).…”