Impairments in fear conditioning in mice lacking the nNOS gene

Abstract

The fear conditioning paradigm is used to investigate the roles of various genes, neurotransmitters, and substrates in the formation of fear learning related to contextual and auditory cues. In the brain, nitric oxide (NO) produced by neuronal nitric oxide synthase (nNOS) functions as a retrograde neuronal messenger that facilitates synaptic plasticity, including the late phase of long-term potentiation (LTP) and formation of long-term memory (LTM). Evidence has implicated NO signaling in synaptic plasticity and LTM formation following fear conditioning, yet little is known about the role of the nNOS gene in fear learning. Using knockout (KO) mice with targeted mutation of the nNOS gene and their wild-type (WT) counterparts, the role of NO signaling in fear conditioning was investigated. Plasma levels of the stress hormone corticosterone were measured to determine the relationship between physiological and behavioral response to fear conditioning. Contextual fear learning was severely impaired in male and female nNOS KO mice compared with WT counterparts; cued fear learning was slightly impaired in nNOS KO mice. Sex-dependent differences in both contextual and cued fear learning were not observed in either genotype. Deficits in contextual fear learning in nNOS KO mice were partially overcome by multiple trainings. A relationship between increase in plasma corticosterone levels following footshock administration and the magnitude of contextual, but not cued freezing was also observed. Results suggest that the nNOS gene contributes more to optimal contextual fear learning than to cued fear learning, and therefore, inhibition of the nNOS enzyme may ameliorate context-dependent fear response.Anxiety disorders, such as post-traumatic stress disorder (PTSD), constitute the most prevalent mental illnesses in the United States, costing nearly one-third of the country's total health bill (Greenberg et al. 1999). The treatment of these disorders requires overcoming complications such as reluctance to seek mental health treatment and an extremely high comorbidity rate with other affective disorders, reaching 80% (Brady 1997;Solomon and Davidson 1997). Emerging evidence suggests that dysfunctions underlying acquired anxiety and PTSD include an abnormal reaction to stress, which is mediated by specific neurochemical and neuroanatomical substrates (Yehuda and McFarlane 1995;Adamec 1997). Pharmacotherapies that target neuronal signaling molecules, such as nitric oxide (NO), may play a role in the treatment of these disorders.In the brain, N-methyl-D-aspartate receptor (NMDAR) activation and calcium influx into the cell activates the neuronal nitric oxide synthase (nNOS) enzyme to produce NO, which has the role of retrograde messenger (Snyder 1992). NO is involved in memory formation and synaptic plastic events such as late-phase long-term potentiation (LTP) (Lu et al. 1999;Arancio et al. 2001;Puzzo et al. 2006). Behavioral evidence in invertebrates (Lewin and Walters 1999;Muller 2000;Kemenes et al. 2002;Matsumoto et al. 2006) ...

Neuronal nitric-oxide synthase deficiency impairs the long-term memory of olfactory fear learning and increases odor generalization

“…1E), which are necessary for olfactory learning (McNamara et al 2008). Knockout mice failed to express the nNOS cells in these areas, which is in accordance with previous studies (Kelley et al 2009). …”

“…Regarding fear conditioning, it has been shown that nNOS knockout mice display deficits in fear learning to both contextual and visual cues (Kelley et al 2009(Kelley et al , 2010, and that nNOS blockade leads to decreased fear association (Kelley et al 2010;Overeem and Kokkinidis 2012). Similar to contextual fear, we confirmed that the deficits in olfactory fear learning were due to decreased NO production in the Figure 5.…”

“…It has been shown that nNOS knockout mice, or wild-type mice administered L-NAME, display normal pain response thresholds and vocalization induced by foot shock (Kelley et al 2009;Overeem and Kokkinidis 2012). Furthermore, our findings show that nNOS knockout mice exhibited similar freezing during the training session.…”

“…At concentrations known to significantly reduce cerebral NOS activity (Moreno-Lopez et al 2004), L-NAME treatment during the acquisition significantly reduced freezing the following day. In accordance with these findings, fear conditioning to contextual, visual, or auditory cues has been shown to be impaired with nNOS inhibitors (Resstel et al 2008;Kelley et al 2009Kelley et al , 2011Ota et al 2010;Overeem and Kokkinidis 2012). Furthermore, associative olfactory learning in pups was blocked by L-NAME when given before training (Samama and Boehm 1999), and administration of a nNOS blocker prevented maternal memory formation (Kendrick et al 1997).…”

“…The role nNOS plays in fear learning has been investigated using contextual fear conditioning, as well as visual and auditory cues as the CS (Schafe et al 2005;Kelley et al 2010Kelley et al , 2011Overeem et al 2010). And although the nNOS inhibitor 7-nitroindazole showed no disruption in the acquisition and expression of the contextual fear conditioning (Maren 1998), it was demonstrated that animals lacking the nNOS gene (Nos1or nNOS knockout) have shown deficits in fear conditioning paradigms to both contextual and cued associations (Kelley et al 2009(Kelley et al , 2010. Furthermore, the blockade of nNOS signaling in limbic areas involved with aversive learning decreased the fear responses to the CS (Schafe et al 2005;Resstel et al 2008).…”