Research ReportHippocampal neuronal nitric oxide synthase (nNOS) is regulated by nicotine and stress in female but not in male rats
Research Highlights
► Basal nNOS levels of the three brain regions examined did not show sex differences. ► Forced swim stress and nicotine increased nNOS expression in female hippocampus. ► When stress and nicotine were applied together, there was no additional increment. ► Stress and nicotine did not regulate nNOS expression in the amygdala of either sex. ► Stress and nicotine did not regulate nNOS expression in frontal cortex of either sex.
Introduction
Nitric oxide (NO) is a highly diffusible gas that functions as a signaling molecule in the central and peripheral nervous system. It has been implicated in the regulation of cognitive (Kanit et al., 2003, Koylu et al., 2005), emotional and behavioral processes such as long-term memory formation (Arancio et al., 1996), fear learning (Overeem et al., 2010), and anxiety (Spolidorio et al., 2007). NO also appears to mediate stress responses by regulating autonomic functions (Krukoff, 1999) and hypothalamo-pituitary–adrenal (HPA) axis activity (Rivier, 2001). In fact, recent studies suggest a role for NO in the pathophysiology of depression through the suppression of neurogenesis in the hippocampus (Packer et al., 2003, Zhou et al., 2007).
NO is synthesized from l-arginine in a reaction catalyzed by nitric oxide synthase (NOS). Three isoforms of NOS are expressed in the brain: neuronal (nNOS), endothelial (eNOS), and inducible (iNOS) NOS. Nerve cells constitutively synthesize nNOS, which is activated by the binding of Ca2+/calmodulin complex (Roman and Masters, 2006). nNOS expressing neurons are widely distributed within the central nervous system including the limbic structures amygdala, hippocampus, and prefrontal cortex (Blackshaw et al., 2003, Kubota et al., 1994, McDonald et al., 1993). NO produced in these limbic regions has important roles in the regulation of autonomic nervous system (Resstel and Correa, 2006, Yao et al., 2007) and HPA axis activity (Seo and Rivier, 2001), anxiety (Forestiero et al., 2006, Spolidorio et al., 2007), fear learning (Overeem et al., 2010), and long-term memory formation (Arancio al., 1996).
Various stress procedures (novelty, restraint, osmotic stress) increase nNOS mRNA, nNOS immunoreactivity, and enzyme activity in the amygdala and hippocampus (de Oliveira et al., 2000, Echeverry et al., 2004, Krukoff and Khalili, 1997, Leza et al., 1998, Yao et al., 2007). Additionally, mild intermittent footshock stress and cytokine administration increase nitric oxide metabolites in the prefrontal cortex (Ishizuka et al., 2000, Ishizuka et al., 2008). Enhancement of nNOS synthesis and activity during the stress response will increase the production of NO, which regulates various neuroendocrine, cognitive, emotional, and behavioral processes in these limbic regions.
Studies report that nicotine administration regulates nNOS expression (Jang et al., 2002, Nakamura et al., 1998, Toborek et al., 2000, Yoo et al., 2005) and activity (Tonnessen et al., 2000) and NO release in the nervous system (Pogun et al., 2000, Smith et al., 1998). Nicotinic receptors are expressed at high levels in the hippocampus, amygdala, and frontal cortex (Tribollet et al., 2004). Additionally, nNOS and α7-nicotinic acetylcholine receptor subunits are co-expressed in the rat hippocampal interneurons (Adams and Freedman, 1997) and primate prefrontal cortex (Csillik et al., 1998). Regulation of NO production in these limbic regions through nicotinic receptors may affect learning and memory (Yilmaz et al., 2000) and HPA axis activity (Raber et al., 1995, Zhu et al., 2001).
Signaling pathways induced by the stress response and nicotinic receptors may converge on various signal transduction molecules to regulate nNOS expression in the brain. Studies report increased glutamatergic signaling in the frontal cortex, amygdala, and hippocampus during the stress response (Musazzi et al., 2010, Reznikov et al., 2007, Sunanda et al., 2000). Similarly, cholinergic signaling through nicotinic receptors enhances glutamate release and glutamatergic neurotransmission in these brain regions (Dani, 2001, Girod et al., 2000, Lambe et al., 2003). Glutamatergic signaling through NMDA, AMPA, and metabotropic receptors activate cAMP response element binding protein (CREB), a transcription factor that is reported to regulate nNOS gene transcription (Mao et al., 2004, Sasaki et al., 2000, Boulware et al., 2005). Additionally, stress also increases acetylcholine release and the expression of α-7 nicotinic receptor mRNA in hippocampus (Tajima et al., 1996, Hunter et al., 2010).
In view of the reported findings above, the aim of this study is to assess the effect of forced swim stress, chronic nicotine administration, and the combined effect of both procedures on nNOS expression in the hippocampus, amygdala and frontal cortex of the rat brain. Considering the sex differences observed in the regulation of the stress response (Solomon and Herman, 2009), central effects of nicotine (Pogun and Yararbas, 2009) and NO (Panzica et al., 2006, Pogun, 2001), sex was included as a factor.
Section snippets
Results
Basal nNOS immunoreactivity levels in control animals that received saline injections (Fig. 1) were subjected to a multifactorial ANOVA with nNOS/β-actin immunoreactivity as the dependent variable and sex (male, female) and brain regions (hippocampus, frontal cortex, and amygdala) as factors. No significant differences were depicted, suggesting that basal levels of nNOS immunoreactivity did not show significant variability between brain regions under the current experimental conditions in
Discussion
The results reported in the present study showed that forced swim stress and chronic nicotine administration increased nNOS expression in the hippocampus of female rats. When stress and nicotine were applied together, no additional increment was observed. These data may suggest an increase in the transcription or translation of hippocampal nNOS. Although the effect of the first day of the forced swim stress (a 2-day procedure) would be more prominent, an additional modulation by the second day
Experimental animals
Adult male (250–400 g) and female (200–230 g) Sprague-Dawley rats were used in the study. Rats were housed (3–4 rats/cage) in standard plastic cages with food and water provided ad libitum during the habituation period for 2 weeks. Male and female rats were housed in the same room. However, they were housed in separate cages placed on different racks so that female and male rats were kept far from each other. Animals were maintained on 12:12-h light:dark cycle (lights on 07:00–19:00).
Acknowledgments
This study was supported by Ege University Research Fund grant 03-BAM-001.
References (84)
- et al.
Nicotinic antagonist alpha-bungarotoxin binding to rat hippocampal neurons containing nitric oxide synthase
Brain Res.
(1997) - et al.
Nitric oxide acts directly in the presynaptic neuron to produce long-term potentiation in cultured hippocampal neurons
Cell
(1996) - et al.
Species, strain and developmental variations in hippocampal neuronal and endothelial nitric oxide synthase clarify discrepancies in nitric oxide-dependent synaptic plasticity
Neuroscience
(2003) Overview of nicotinic receptors and their roles in the central nervous system
Biol. Psychiatry
(2001)- et al.
Expression of neuronal nitric oxide synthase mRNA in stress-related brain areas after restraint in rats
Neurosci. Lett.
(2000) - et al.
Acute and delayed restraint stress-induced changes in nitric oxide producing neurons in limbic regions
Neuroscience
(2004) - et al.
Estrogen enhances potassium-stimulated acetylcholine release in the rat hippocampus
Brain Res.
(2003) - et al.
Facilitation of glutamatergic neurotransmission by presynaptic nicotinic acetylcholine receptors
Neuropharmacology
(2000) - et al.
Regulation of the nicotinic receptor alpha7 subunit by chronic stress and corticosteroids
Brain Res.
(2010) - et al.
Differential profiles of nitric oxide and norepinephrine releases in the paraventricular nucleus region in response to mild footshock in rats
Brain Res.
(2000)
Different response between production of free radicals induced by central and peripheral administration of interleukin-1beta in conscious rats
Neurosci. Res.
Nicotine administration decreases nitric oxide synthase expression in the hypothalamus of food-deprived rats
Neurosci. Lett.
The effect of nitric oxide synthase inhibition on cognitive ability and strategies employed for place learning in the water maze: sex differences
Brain Res. Bull.
Effects of nitric oxide synthase inhibition on spatial discrimination learning and central DA2 and mACh receptors
Pharmacol. Biochem. Behav.
Central actions of nitric oxide in regulation of autonomic functions
Brain Res. Brain Res. Rev.
Three distinct subpopulations of GABAergic neurons in rat frontal agranular cortex
Brain Res.
Regulation of nitric oxide synthase messenger RNA expression in the rat hippocampus by glucocorticoids
Neuroscience
Estradiol increases choline acetyltransferase activity in specific basal forebrain nuclei and projection areas of female rats
Exp. Neurol.
Identification of putative nitric oxide producing neurons in the rat amygdala using NADPH-diaphorase histochemistry
Neuroscience
Central cholinergic pathways in the rat: an overview based on an alternative nomenclature (Ch1–Ch6)
Neuroscience
Effects of gonadal hormones on central nitric oxide producing systems
Neuroscience
Sex differences in brain and behavior: emphasis on nicotine, nitric oxide and place learning
Int. J. Psychophysiol.
Nicotine modulates nitric oxide in rat brain
Eur. Neuropsychopharmacol.
Pretreatment with a single estradiol-17beta bolus activates cyclic-AMP response element binding protein and protects CA1 neurons against global cerebral ischemia
Neuroscience
Electron transfer by neuronal nitric-oxide synthase is regulated by concerted interaction of calmodulin and two intrinsic regulatory elements
J. Biol. Chem.
Evidence for novel estrogen binding sites in the rat hippocampus
Neuroscience
Estrogen binding and estrogen receptor characterization (ERalpha and ERbeta) in the cholinergic neurons of the rat basal forebrain
Neuroscience
Nicotine-evoked nitric oxide release in the rat hippocampal slice
Neurosci. Lett.
Sex differences in psychopathology: of gonads, adrenals and mental illness
Physiol. Behav.
Immobilization stress-induced increase of hippocampal acetylcholine and of plasma epinephrine, norepinephrine and glucose in rats
Brain Res.
Nicotine attenuates arachidonic acid-induced overexpression of nitric oxide synthase in cultured spinal cord neurons
Exp. Neurol.
Comparative distribution of nicotinic receptor subtypes during development, adulthood and aging: an autoradiographic study in the rat brain
Neuroscience
Inhibition of MPEP on the development of morphine antinociceptive tolerance and the biosynthesis of neuronal nitric oxide synthase in rat spinal cord
Neurosci. Lett.
Reduced nNOS expression induced by repeated nicotine treatment in mu-opioid receptor knockout mice
Neurosci. Lett.
The elevation of plasma adrenocorticotrophic hormone and expression of c-Fos in hypothalamic paraventricular nucleus by microinjection of neostigmine into the hippocampus in rats: comparison with acute stress responses
Brain Res.
Mammalian nicotinic acetylcholine receptors: from structure to function
Physiol. Rev.
A novel eIF2B-dependent mechanism of translational control in yeast as a response to fusel alcohols
EMBO J.
Estradiol activates group I and II metabotropic glutamate receptor signaling, leading to opposing influences on cAMP response element-binding protein
J. Neurosci.
Nitric oxide synthase and the acetylcholine receptor in the prefrontal cortex: metasynaptic organization of the brain
Neurobiology (Bp).
Acetylcholine mediates the estrogen-induced increase in NMDA receptor binding in CA1 of the hippocampus and the associated improvement in working memory
J. Neurosci.
Anxiolytic-like effects induced by nitric oxide synthase inhibitors microinjected into the medial amygdala of rats
Psychopharmacology (Berl).
1-Aminocyclopropanecarboxylic acid, an antagonist of N-methyl- d-aspartate receptors, causes hypotensive and antioxidant effects with upregulation of heme oxygenase-1 in stroke-prone spontaneously hypertensive rats
Hypertens. Res.
Cited by (16)
Sex differences in the anticompulsive-like effect of memantine: Involvement of nitric oxide pathway but not AMPA receptors
2024, Behavioural Brain ResearchNitric oxide synthase genotype interacts with stressful life events to increase aggression in male subjects in a population-representative sample
2020, European NeuropsychopharmacologyCitation Excerpt :Similarly to Reif et al. (2011), all genotype main effects were only present in the male subjects. Animal studies have shown sex differences in the brain expression patterns and activity of NOS1 (Díaz et al., 2015; Keser et al., 2011) which is influenced by gonadal hormones: testosterone in the male and estrogens in female rats (Viglietti-Panzica et al., 2006). Also, the NOS1-deficient mice were described to display sexual dimorphism in aggressive behaviours – while knockout males engaged in more territorial aggression than wild-type littermates, female knockouts were in turn characterized by the absence of maternal aggression (Chiavegatto and Nelson, 2003).
Neuronal nitric oxide synthase and affective disorders
2018, IBRO ReportsStriatal NOS1 has dimorphic expression and activity under stress and nicotine sensitization
2015, European NeuropsychopharmacologyHippocampus and nitric oxide
2014, Vitamins and HormonesCitation Excerpt :This report is in line with the viewpoints that hippocampal NO is involved in the effects of typical neurotransmitters, especially monoamine. Interestingly, there are several evidences indicate that hippocampal NO plays a role in the sex difference of behaviors (Kant et al., 2000; Keser, Balkan, Gozen, Kanit, & Pogun, 2011; Taskiran, Kutay, Sozmen, & Pogun, 1997). It is commonly suggested that a female preponderance in depression and anxiety is universal and substantial.
Possible role of adrenomedullin and nitric oxide in major depression
2013, Progress in Neuro-Psychopharmacology and Biological Psychiatry