Vallo Volke

Endogenous nitric oxide decreases hippocampal levels of serotonin and dopamine in vivo

Nitric oxide (NO) modulates the levels of various neurotransmitters in the CNS. Here we determined whether the specific nitric oxide synthase (NOS) inhibitor 7‐nitroindazole (7‐NI), the non‐selective inhibitor of guanylate cyclase (GC) and NOS, methylene blue (MB), the NO‐precursor L‐arginine (L‐Arg), and the selective soluble GC inhibitor 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ) affect extracellular levels of serotonin (5‐HT), dopamine (DA), 5‐hydroxyindoleacetic acid (5‐HIAA), and homovanillic acid (HVA) in the rat ventral hippocampus by using microdialysis in freely moving animals. Local perfusion of 7‐NI (1 mM) and MB (1 mM) significantly increased extracellular level of 5‐HT, whereas DA was increased by 7‐NI only. Systemic administration of 7‐NI (50 mg kg−1) and MB (30 mg kg−1) increased the extracellular levels of 5‐HT and DA. Extracellular levels of 5‐HIAA was not influenced by local or systemic MB or 7‐NI. In contrast, extracellular level of HVA was decreased by systemic MB and retrodialyzed MB, but was not influenced by 7‐NI. Retrodialysis of L‐Arg (2 mM) decreased the levels of 5‐HT, DA, 5‐HIAA and HVA in the hippocampus. Systemic administration of L‐Arg (250 mg kg−1) decreased the level of 5‐HT, but failed to influence DA, 5‐HIAA and HVA. Local perfusion of ODQ (400 μM) did not affect 5‐HT overflow in the hippocampus. We conclude that NOS inhibitors increased extracellular levels of 5‐HT and DA in the rat ventral hippocampus after local or systemic administration, whereas the NO precursor L‐Arg had the opposite effect. Thus, endogenous NO may exert a negative control over the levels of 5‐HT and DA in the hippocampus. However, this effect is probably not mediated by cyclic GMP.

Antidepressant- and anxiolytic-like effects of selective neuronal NOS inhibitor 1-(2-trifluoromethylphenyl)-imidazole in mice.

Various inhibitors of nitric oxide synthase (NOS) have been shown to possess antidepressant- and anxiolytic-like properties in animal models. The aim of this study was to compare the behavioural effects of NOS inhibitor 7-nitroindazole (7-NI) with the more selective neuronal NOS inhibitor 1-(2-trifluoromethylphenyl)imidazole (TRIM) in animal models predictive of antidepressant- and anxiolytic-like activity in order to clarify the role of distinct isoforms of NOS in the regulation of depression and anxiety. Both TRIM (50 mg/kg) and 7-NI (50 mg/kg) decreased the immobility time in the forced swimming test. The magnitude of the effect was comparable to that of the tricyclic antidepressant imipramine (15 mg/kg). The antidepressant-like effect of TRIM was counteracted by pretreatment with L-arginine (250 mg/kg). The systemic administration of TRIM (50 mg/kg), but not 7-NI (up to 50 mg/kg) increased the time spent in the light side of the apparatus in the light-dark compartment test. The anxiolytic-like effect of TRIM was antagonised by pretreatment with L-arginine. Both TRIM and 7-NI decreased the locomotion of animals in the open field and caused motor incoordination on rotarod. These motor side effects were more pronounced in the case of 7-NI and were not diminished by pretreatment with L-arginine. We conclude that neuronal NOS seems to play the key role in the antidepressant- and anxiolytic-like effects of NOS inhibitors.

Local, but not systemic, administration of serotonergic antidepressants decreases hippocampal nitric oxide synthase activity

Nitric oxide (NO) is an unconventional transmitter molecule in the nervous system, synthesized by nitric oxide synthase (NOS) following activation of the N-methyl-D-aspartate (NMDA) receptor. Several in vivo studies have demonstrated that NO modulates the extracellular levels of various neurotransmitters in the central nervous system, while serotonin (5-HT) re-uptake may be influenced by the NO pathway. Moreover, inhibitors of NOS exhibit antidepressant-like and anxiolytic-like properties in various animal models. Therefore, the aims of the present study were to clarify the involvement of distinct antidepressants acting on the serotonin re-uptake site in the regulation of the activity of hippocampal NOS in vitro, in vivo and ex vivo. We found that citalopram, paroxetine, imipramine and N(G)-nitro-L-arginine dose dependently decreased the hippocampal NOS activity in vitro. Moreover, local administration of citalopram, paroxetine, tianeptine, imipramine and N(G)-nitro-L-arginine significantly decreased the hippocampal NOS activity in vivo at a concentration significantly lower than in vitro. No effect on NOS activity following retrodialysis with 5-HT was observed. Acute (5 mg/kg, s.c.) and chronic (3 weeks, 20 mg/kg/24 h) systemic administration of citalopram did not influence NOS activity ex vivo. The effects on NOS represent a response to structurally dissimilar serotonergic antidepressants. However, since these data reflect effects on basal NOS activity, we believe that serotonergic antidepressants do not directly affect NOS at dosages used clinically, but the findings may reflect a secondary action of antidepressants on the glutamate NMDA receptor following their primary inhibitory action at the 5-HT transporter.

7-Nitroindazole, a nitric oxide synthase inhibitor, has anxiolytic-like properties in exploratory models of anxiety.

Abstract The action of the novel nitric oxide synthase (NOS) inhibitor 7-nitroindazole (7-NI) was studied in different exploratory models of anxiety. In the rat plus-maze test, 7-NI potently increased time spent on open arms and percentage of open arm visits in a dose-dependent manner with the minimal effective dose of 40 mg/kg. 7-NI caused an anxiolytic-like effect in the rat social interaction test. The minimal dose increasing social interaction time was 20 mg/kg. However, the drug also produced a clear sedative effect occurring even at smaller doses (10 mg/kg) in the open field test. 7-NI also showed an anxiolytic-like profile in the mouse light-dark compartment test and in the elevated plus-maze test, but the doses required were higher (80–120 mg/kg) than in rat models. Also, the sedative effect occurred at these doses in open field. We failed to demonstrate any effect of L-arginine either in the rat elevated plus-maze test or in the open field test at doses up to 600 mg/kg IP. These results indicate that there are no major interspecies differences between rats and mice in respect of action of 7-NI. The clear anxiolytic-like action of the nitric oxide synthase inhibitor in four different models shows that nitric oxide is involved in the process of anxiety and that NOS could be a new target in developing anxiolytic drugs.

Inhibition of nitric oxide synthase causes anxiolytic-like behaviour in an elevated plus-maze.

The action of inhibition of nitric oxide (NO) synthase by NG-nitro-L-arginine methyl ester (L-NAME) (1-20 mg kg-1) on the exploratory behaviour of rats in the elevated plus-maze was studied. L-NAME induced an anxiolytic-like effect in the plus-maze test, showing a reverse U-shape action behaviour, with a maximal effect at 10 mg kg-1. This effect was not related to a non-specific increase in motor activity, since in the open field test L-NAME did not affect locomotor activity of rats. Pretreatment of rats with L-NAME (1-10 mg kg-1) also tended to attenuate the anti-exploratory action of CCK agonist caerulein (5 micrograms kg-1), but this action was not significant. In conclusion, it appears that NO may be involved in the process that can lead to anxiety in the rat.

A high-fat diet exacerbates depressive-like behavior in the Flinders Sensitive Line (FSL) rat, a genetic model of depression

Major depressive disorder (MDD) and diabetes mellitus type II (T2DM) are two of the major health challenges of our time. It has been shown that MDD and T2DM are highly co-morbid, and recent work has proposed a bi-directional connection between the diseases. The aim of the current study was to investigate the effect of a high-fat diet (HFD) on behavior and metabolism in a genetic rat model of depression, the Flinders Sensitive and Resistant Line (FSL/FRL) rats. Age and weight matched rats were fed a HFD or control diet for 10 weeks and subjected to behavioral testing and metabolic assessment. We found that HFD exacerbated the depressive-like behavior of the FSL rat in the Forced Swim Test (FST), a depression screening tool, although it did not affect the non-depressed FRL rat despite a higher caloric intake. Moreover, the depressive-like phenotype was associated with reduced anxiety and impairment in novel object recognition memory, while HFD consumption led to diminished object recognition memory as well. In both strains HFD increased insulin levels during an oral glucose tolerance test, although fasting blood glucose levels were only significantly increased by HFD in the FSL rat, suggesting a greater metabolic susceptibility in this rat strain. We conclude that compared with the FRL rat, the FSL rat is more susceptible to developing aberrant behaviors related to depression following metabolic stress induced by HFD. Further studies with a mechanistic focus could potentially lead to a better understanding of a possible pathophysiological link between T2DM and MDD.

The antidepressant action of imipramine and venlafaxine involves suppression of nitric oxide synthesis.

Depressive disorders represent a major public health problem worldwide. The limitations of current antidepressant drugs have warranted on-going research to identify pharmacological agents and strategies that offer a greater therapeutic efficacy. The NMDA/L-arginine nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) cascade is an important signaling pathway that is also implicated in the regulation of depression. In animal models detecting antidepressant activity, distinct NO synthase inhibitors display antidepressant-like action. Therefore, the aim of current study was to evaluate whether pretreatment with L-arginine (precursor of NO) could counteract antidepressant-like effects of distinct antidepressant classes in the mouse forced swimming test (FST), and whether these drugs are able to modulate the nitric oxide synthesis in the brain. We found in the FST that pretreatment with L-arginine (500 mg/kg) counteracted the antidepressant-like effect of imipramine (IMI, 15 mg/kg) and venlafaxine (VENL, 6 mg/kg), but not the effects of bupropion (BUPR, 20mg/kg) or fluoxetine (FLX, 20mg/kg). Increasing the dose of L-Arg to 1000 mg/kg attenuated the antidepressant-like effects of BUPR, but did not modify the action of FLX. L-Arginine was devoid of any locomotor effects on the animals. The effect of antidepressants on brain NO metabolism paralleled their behavioral action in case of IMI and VENL which decreased the nitrite+nitrate concentration in the brain. BUPR and FLX did not have any effect on brain nitrite+nitrate concentration. These results support the idea that some antidepressants are able to inhibit nitric oxide synthesis in the brain, an effect which could be mechanistically related to the ability of L-arginine to counteract their antidepressant-like effects.

Increased stress-evoked nitric oxide signalling in the Flinders sensitive line (FSL) rat: a genetic animal model of depression

Stress engenders the precipitation and progression of affective disorders, while stress-related release of excitatory mediators is implicated in the degenerative pathology observed especially in the hippocampus of patients with severe depression. Nitric oxide (NO) release following stress-evoked N-methyl-d-aspartate (NMDA) receptor activation modulates neurotransmission, cellular memory and neuronal toxicity. We have investigated the Flinders rat (FSL/FRL), a genetic animal model of depression, regarding the response of the hippocampal nitrergic system following exposure to an escapable stress/inescapable stress (ES-IS) paradigm. Hippocampal tissue from naive FSL/FRL rats and those exposed to ES-IS were studied with respect to constitutive nitric oxide synthase (cNOS) activity and neuronal nitric oxide synthase (nNOS) protein levels, as well as transcript expression of upstream regulatory proteins in the NMDA-NO signalling pathway, including NMDAR1, nNOS, CAPON, PIN and PSD95. Within stress-naive animals, no differences in hippocampal cNOS activity and nNOS expression or PIN were evident in FSL and FRL rats, although transcripts for NMDAR1 and CAPON were increased in FSL rats. Within the group of ES-IS animals, we found an increase in total hippocampal cNOS activity, nNOS protein levels and mRNA expression in FSL vs. FRL rats, together with an increase in PSD95 transcripts, and a reduction in PIN. In conclusion, ES-IS enhanced hippocampal cNOS activity in FSL rats, but not FRL rats, confirming the NMDA-NO cascade as an important vulnerability factor in the depressive phenotype of the FSL rat.

Methylene blue inhibits hippocampal nitric oxide synthase activity in vivo.

The aim of the present study was to investigate the effect of methylene blue, a guanylate cyclase inhibitor, on the hippocampal nitric oxide synthase activity in vivo. We used a microdialysis-based technique of measuring conversion of [3H]l-arginine to [3H]l-citrulline in freely moving rats. The administration of methylene blue (0.1 and 1 mM) via the microdialysis probe caused a dose-dependent decrease in [3H]l-citrulline efflux comparable with the effect of unselective NOS inhibitor NG-nitro-L-arginine (2 mM). We conclude that methylene blue inhibits brain NOS activity in vivo and thus interferes with NO-cGMP cascade in different levels.

Nitric Oxide Synthase Inhibitors as Antidepressants

Affective and anxiety disorders are widely distributed disorders with severe social and economic effects. Evidence is emphatic that effective treatment helps to restore function and quality of life. Due to the action of most modern antidepressant drugs, serotonergic mechanisms have traditionally been suggested to play major roles in the pathophysiology of mood and stress-related disorders. However, a few clinical and several pre-clinical studies, strongly suggest involvement of the nitric oxide (NO) signaling pathway in these disorders. Moreover, several of the conventional neurotransmitters, including serotonin, glutamate and GABA, are intimately regulated by NO, and distinct classes of antidepressants have been found to modulate the hippocampal NO level in vivo. The NO system is therefore a potential target for antidepressant and anxiolytic drug action in acute therapy as well as in prophylaxis. This paper reviews the effect of drugs modulating NO synthesis in anxiety and depression.