Antisense knockdown of neuronal nitric oxide synthase antagonizes nitrous oxide-induced behavior

doi:10.1016/S0006-8993(03)02289-3

Brain Research

Volume 968, Issue 1, 4 April 2003, Pages 167-170

https://doi.org/10.1016/S0006-8993(03)02289-3 Get rights and content

Abstract

The behavioral effects of nitrous oxide (N₂O) were antagonized by non-specific inhibitors of nitric oxide synthase (NOS). To identify the isoform of NOS involved in this response, mice were pretreated with an antisense oligodeoxynucleotide (AS-ODN) against neuronal NOS, and then tested in a light/dark exploration paradigm. The AS-ODN but not the mismatch ODN significantly antagonized N₂O induced behavior and also reduced NOS activity in the cerebellum and hippocampus. These results implicate neuronal NOS in the N₂O response.

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Acknowledgements

This work was support by NIH Grant DA-10343.

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Cited by (14)

Nitrous oxide as a putative novel dual-mechanism treatment for bipolar depression: Proof-of-concept study design and methodology
2020, Contemporary Clinical Trials Communications
Citation Excerpt :
To measure the integrity of the blind, clinical raters and participants will complete questionnaires guessing which treatment the participant received following the 60-min post-treatment interview. B12 and nitric oxide (NO) levels will be examined as predictors of response owing to known associations with the mechanism of action of N2O [45–49]. B12 will be examined in screening clinical blood work.
Depressive symptoms predominate in the course of bipolar disorder (BD) and there is an urgent need to evaluate novel application of repurposed compounds that act on pre-specified treatment targets. Several lines of reasoning suggest that nitrous oxide (N₂O) is an ideal medication to study as a potential treatment and as a strategy to identify the underlying pathophysiology of bipolar depression. N₂O is a potent cerebral vasodilator and there is compelling evidence of reduced frontal cerebral blood flow (CBF; i.e. hypoperfusion) in depression. Therefore, N₂O may increase CBF and thereby improve symptoms of depression. The goal of this randomized, double-blind trial is to study the effect of a single administration of N₂O versus the active comparator midazolam on mood and CBF in adults with treatment-resistant bipolar depression.
Participants with BD-I/-II currently experiencing a major depressive episode will be randomized to one of two conditions (n = 20/group): 1) inhaled N₂O plus intravenous saline, or 2) inhaled room air plus intravenous midazolam. Montgomery-Asberg Depression Rating Scale scores will serve as the primary endpoint. CBF will be measured via arterial spin labelling magnetic resonance imaging.
N₂O is a potential novel treatment for bipolar depression, as it causes cerebral vasodilation. This proof-of-concept study will provide valuable information regarding the acute impact of N₂O on mood and on CBF. If N₂O proves to be efficacious in future larger-scale trials, its ubiquity, safety, low cost, and ease of use suggest that it has great potential to become a game-changing acute treatment for bipolar depression.
Regulatory effects of anesthetics on nitric oxide
2016, Life Sciences
Citation Excerpt :
The antinociceptive effect of N2O is dependent on NO. Several studies show that NO may play a critical role in N2O-induced antinociception in the CNS [133–138]. The action involves sequential activation of nNOS [135,139–141]. Similarly, N2O induces anxiolytic-like behavior, which is mediated by NO/nNOS enzyme activity [142,143].
Nitric oxide (NO) is a free radical gas in the biological system, which is produced by nitric oxide synthase (NOS) family. NO acts as a biological mediator and plays important roles in different systems in humans. The NO/NOS system exerts a broad spectrum of signaling functions involved in vasodilation, inflammation, oxidative stress, cardioprotection and neuroprotection. It has been demonstrated that intravenous and volatile anesthetics (such as propofol, ketamine, midazolam, isoflurane, sevoflurane, and desflurane, etc.) modulate NO production through multiple mechanisms that may influence physiological and pathophysiological processes. This review focuses on the effects of different anesthetics on NO/NOS regulation in different disease conditions. Possible cellular mechanisms and intermediate role of NO/NOS in anesthetic-mediated organ protection are also discussed. It would be interesting to clarify the impact of anesthetics on NO/NOS regulation. This review gives an overview of the effects of different anesthetics on NO/NOS regulation and function in different physiologic and pathophysiologic states.
An anxiolytic-like effect of hyperbaric oxygen in the mouse light/dark exploration test
2012, Life Sciences
Citation Excerpt :
The anxiolytic-like effects of chlordiazepoxide in the mouse elevated plus-maze were antagonized by systemic pretreatment with the nitric oxide synthase-inhibitor L-NG-nitro arginine (L-NOARG); this inhibitory effect of L-NOARG was stereospecifically and completely reversed by intracerebroventricular administration of l-arginine but not d-arginine (Quock and Nguyen, 1992). The anxiolytic-like effects of nitrous oxide (N2O) were similarly antagonized by pretreatment with a variety of NOS-inhibitors (Caton et al., 1994; Li and Quock, 2001; Li et al., 2003b) or an antisense oligodeoxynucleotide (AS-ODN) against neuronal nitric oxide synthase (nNOS) (Li et al., 2003a). Conversely, the intracerebroventricular administration of a NO-donor into the brain was found to induce anxiolytic-like behavioral effects in the mouse light/dark exploration test (Li and Quock, 2002).
We studied whether hyperbaric oxygen (HBO₂) treatment, which is known to increase production of nitric oxide (NO) in the brain, might also produce an NO-dependent anxiolytic-like behavioral response.
Male NIH Swiss mice (20–25 g) were subjected to a 60-min HBO₂ treatment at different absolute atmospheres, and anxiety was assessed using the light/dark exploration test at different time intervals following the cessation of HBO₂ treatment. To ascertain the underlying mechanism of action, other groups of mice were pretreated with the NO synthase inhibitor N^G-monomethyl-l-arginine acetate, the NO scavenger 2-(4-carboxyphenyl)-4,5-dihydro-4,4,5,5-tetramethyl-1H-imidazolyl-1-oxy-3-oxide (carboxy-PTIO), the soluble guanylyl cyclase-inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) or the benzodiazepine antagonist flumazenil to determine their influence on the HBO₂-induced anxiolytic-like effect.
A 60-min HBO₂ treatment at 3.0 absolute atmospheres increased the time spent by mice in the light compartment that lasted up to 90 min following the end of HBO₂ treatment. This anxiolytic effect of HBO₂ was significantly reduced by pretreatment with L-NMMA, carboxy-PTIO, ODQ and flumazenil.
Based on these findings, we conclude that a 60-min HBO₂ treatment is capable of inducing an anxiolytic effect that possibly involves NO, cyclic GMP and the benzodiazepine binding site.
Involvement of a NO-cyclic GMP-PKG signaling pathway in nitrous oxide-induced antinociception in mice
2011, European Journal of Pharmacology
Citation Excerpt :
It is interesting that a NO–cyclic GMP–PKG pathway is also involved in another pharmacological effect of N2O, the relief of anxiety. The anxiolytic effect of N2O in the mouse light/dark exploration test was significantly reduced by NOS-inhibitors and nNOS-directed antisense oligodeoxynucleotides (Li et al., 2003a,b), ODQ (Li et al., 2004b) and Rp-8-pCPT-cGMPS (Li et al., 2005). The findings of this study support the hypothesis that a NO–cyclic GMP–PKG pathway is involved in N2O-induced antinociception.
The antinociceptive effect of nitrous oxide (N₂O) is dependent on nitric oxide (NO); however, the next step in the pathway activated by NO is undetermined. The present study was conducted to test the hypothesis that a N₂O action involves sequential activation of NO synthase, soluble guanylyl cyclase and protein kinase G to induce an antinociceptive effect in mice. The antinociceptive responsiveness of male NIH Swiss mice to N₂O was assessed using the acetic acid abdominal constriction test. Different groups of mice were pretreated with either saline, the NO scavenger 2-(4-carboxyphenyl)-4,5-dihydro-4,4,5,5-tetramethyl-1H-imidazolyl-1-oxy-3-oxide (carboxy-PTIO), the guanylyl cyclase-inhibitor 1H-[1,2,4]-oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ), the protein kinase G-inhibitor Rp-isomer of 8-(4-chlorophenylthio)-guanosine-3′,5′-cyclic monophosphorothioate (Rp-8-pCPT-cGMPS) or the selective phosphodiesterase V-inhibitor 1,2-dihydro-2-[(2-methyl-4-pyridinyl)methyl]-1-oxo-8-(2-pyrimidinylmethoxy)-4-(3,4,5-trimethoxyphenyl)-2,7-naphthyridine-3-carboxylic acid methyl ester hydrochloride (T 0156). Vehicle (saline)-pretreated mice responded to N₂O in a concentration-dependent manner. This antinociceptive effect was antagonized by systemic pretreatment with carboxy-PTIO and ODQ and central pretreatment with Rp-8-pCPT-cGMPS. In each case, the dose–response curve for N₂O was progressively shifted to the right by increasing the dose of each pretreatment drug. On the other hand, N₂O-induced antinociception was enhanced by systemic pretreatment with T 0156; the dose–response curve for N₂O was shifted to the left. The ATP-sensitive potassium channel blocker glibenclamide was without influence on the antinociceptive effect of N₂O. These results support the hypothesis that N₂O-induced antinociception in mice is mediated by a NO–cyclic GMP–PKG pathway.
Antagonism of the antinociceptive effect of nitrous oxide by inhibition of enzyme activity or expression of neuronal nitric oxide synthase in the mouse brain and spinal cord
2010, European Journal of Pharmacology
Previous studies have implicated nitric oxide (NO) in the antinociceptive response to the anesthetic gas nitrous oxide (N₂O). The present study was conducted to confirm this NO involvement using pharmacological and gene knockdown and knockout strategies to inhibit the supraspinal and spinal production of NO. Antinociceptive responsiveness to 70% N₂O was assessed using the acetic acid (0.6%) abdominal constriction test in NIH Swiss mice following intracerebroventricular (i.c.v.) or intrathecal (i.t.) pretreatment with the NOS-inhibitor l-N^G-nitro arginine methyl ester (L-NAME) or an antisense oligodeoxynucleotide (AS-ODN) directed against neuronal NOS (nNOS). Experiments were also conducted in mice homozygous for a defective nNOS gene (nNOS^−/−). Mice that were pretreated i.c.v. or i.t. with L-NAME (1.0 µg) both exhibited 80–90% reduction in the magnitude of the N₂O-induced antinociceptive response. Mice that were pretreated i.c.v. or i.t. with nNOS AS-ODN (3 × 25 µg) exhibited a 60–80% antagonism of the antinociceptive response. Compared to wild-type mice, nNOS knockout mice showed a 60% reduction in N₂O-induced antinociception. These findings consistently demonstrate that transient or developmental suppression of nNOS expression significantly reduces antinociceptive responsiveness to N₂O. NO of both supraspinal and spinal origin, therefore, plays an important role in the antinociceptive response to N₂O.
Antisense oligos to neuronal nitric oxide synthase aggravate motoneuron death induced by spinal root avulsion in adult rat
2006, Experimental Neurology
The present study used nitric oxide synthase (nNOS) antisense oligos (nNOS AS-ODN) to assess the role of nNOS in motoneuron death induced by spinal root avulsion. A right seventh cervical (C7) spinal root avulsion was performed on adult male Sprague–Dawley rats. Two weeks later, FITC-labeled random oligos (FITC-R-ODN), nNOS AS-ODN, R-ODN or TE buffer was applied to the lesioned side of the C7 spinal segment and refreshed every 3 days. FITC-R-ODN was first detected inside the injured motoneurons at 10 h, accumulated to a maximum by 24 h and faded out from 72 h. Following avulsion, nNOS AS-ODN decreased the number of nNOS-positive motoneurons in the lesioned segment compared either with buffer (P < 0.001 at 15 days, 3 and 4 weeks post-injury) or with R-ODN control (P = 0.002 at 15 days, P < 0.001 at 3 and 4 weeks post-injury). Interestingly, nNOS AS-ODN also decreased the number of surviving motoneurons compared either with buffer (P = 0.005 at 15 days, P < 0.001 at 3 or 4 weeks) or with R-ODN control (P < 0.001 at 3 or 4 weeks). Meanwhile, there were no significant differences between R-ODN and buffer control either in the number of nNOS-positive motoneurons (P = 0.245 at 15 days, P = 0.089 at 3 weeks and P = 0.162 at 4 weeks) or in the number of surviving motoneurons (P = 0.426 at 15 days, P = 0.321 at 3 weeks or P = 0.344 at 4 weeks). These findings indicate that nNOS AS-ODN, applied from 2 weeks after avulsion, aggravates the motoneuron death due to root avulsion by specifically down-regulating nNOS gene expression and that the expression of nNOS in adult spinal motoneurons in response to root avulsion may play a beneficial role in the survival of injured neurons.

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Short communicationAntisense knockdown of neuronal nitric oxide synthase antagonizes nitrous oxide-induced behavior

Abstract

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Acknowledgements

Pharmacol. Biochem. Behav.

Physiol. Behav.

Neurosci. Biobehav. Rev.

Cell

Pharmacol. Biochem. Behav.

Short communication
Antisense knockdown of neuronal nitric oxide synthase antagonizes nitrous oxide-induced behavior