Nitric oxide synthase inhibition attenuates phencyclidine-induced disruption of cognitive flexibility

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Abstract

Schizophrenia encompasses, amongst other symptoms, a heavy load of cognitive dysfunctionality. Using the psychotomimetic agent, phencyclidine (PCP), we have previously found that PCP-induced disruptions of cognitive function in translational rodent models of schizophrenia are dependent on nitric oxide (NO) production. In the present study, male Sprague–Dawley rats were subjected to a Morris water maze task designed to assess cognitive flexibility (i.e. the ability to cope with an increasingly demanding cognitive task) by means of a “constant reversal learning paradigm”. Experiments were conducted to evaluate the effects of the NO synthase inhibitor, L-NAME (10 mg/kg), on PCP-induced (2 mg/kg) impairments. Control animals significantly improved their learning over the first 3 consecutive days, whereas PCP-treated animals failed to show any significant learning. Pretreatment with L-NAME normalized the PCP-induced disruption of learning to control levels. These findings suggest that PCP-induced disruptions of cognitive flexibility (i.e. ability to modify behaviour according to an increasingly demanding cognitive task) are dependent upon NO production. These observations, together with accumulated clinical findings, suggest that the NO system is a potential treatment target for cognitive dysfunctions in schizophrenia.

Introduction

Dementia praecox was first described by Kraepelin as a disorder with a heavy cognitive dysfunctional burden and later termed schizophrenia by Bleuler in 1911, signifying a “split-mind” (Adityanjee et al., 1999). The cognitive impairments prevalent in schizophrenia are predictive of disease outcome (Green, 1996, Green et al., 2004, Helldin et al., 2006) and treatment response and thus constitute important targets for the development of new treatment strategies. Cognitive functions such as planning, executive functioning and working memory depend to a large extent upon the prefrontal cortex (PFC) and malfunctions within these domains are included in the abnormalities observed in schizophrenic patients (Mahurin et al., 1998, Harvey et al., 2005). Some of these cognitive dysfunctions are also observed in the first degree relatives of patients with schizophrenia (Toulopoulou et al., 2003). Therefore, these cognitive dysfunctions are not only part of the clinical syndrome but also suggested to be endophenotypic markers of the disease. The above-mentioned aspects of cognition may reflect cognitive flexibility, and malfunction in these domains may be manifested as the cognitive stereotypy (i.e. perseveration) observed in schizophrenia.

Several translational animal studies using the phencyclidine (PCP) model of schizophrenia have shown PCP to induce disruptions of cognitive functions that corresponds to dysfunctions observed in the human condition (Jentsch and R.H., 1999, Palsson et al., 2005). Some of these PCP-induced disruptions can be reduced by atypical antipsychotics (Didriksen et al., 2007, Fejgin et al., 2007, Jentsch et al., 1997). However, available antipsychotics do not have a satisfactory effect on cognitive dysfunctions in patients with schizophrenia, hence the need for novel cognitive enhancing treatments. A putative novel treatment approach is targeting the nitric oxide (NO) signalling pathway. Several experimental studies along with clinical findings indicate that the NO system of the brain is involved in the pathophysiology of schizophrenia (Bernstein et al., 2005). In previous animal studies, our research group has shown that PCP-induced disruption of functions such as pre-attentive information processing, non-associative learning, selective attention, working and long-term memory, can be reduced by pretreatment with a NO synthase (NOS) inhibitor (Johansson et al., 1997, Klamer et al., 2001, Klamer et al., 2004a, Klamer et al., 2004b, Klamer et al., 2004c, Klamer et al., 2005a, Klamer et al., 2005b, Klamer et al., 2005c, Wass et al., 2006a, Wass et al., 2006b). Thus, the NO system may be a suitable treatment target for alleviating cognitive dysfunctions in patients with schizophrenia (Palsson, 2006).

NO is formed in a two-step oxidation reaction catalyzed by NOS between the amino acid, l-arginine, and molecular oxygen. In the brain NO is proposed to be a key link between N-methyl-d-aspartate (NMDA) receptor mediated increases in cytoplasmic Ca2+ and activity-dependent long-term changes such as neuronal differentiation and synaptic plasticity (Karatinos et al., 1995, Snyder and Ferris, 2000). Moreover, NO seems to be involved in several aspects of cognition, amongst which learning and memory are implicated, as NO may act as a retrograde messenger during long-term potentiation (O'dell et al., 1991, Zhuo et al., 1994, Lu et al., 1999). Interestingly, the psychotomimetic effect of PCP has been attributed to, e.g. its action at the glutamatergic NMDA receptor. PCP acts as a non-competitive antagonist of this receptor via a binding site inside the channel complex (Javitt and Zukin, 1991, Lodge and Anis, 1982). Somewhat paradoxically, PCP has been shown to increase glutamate release in regions such as the PFC (Adams and Moghaddam, 1998). This glutamatergic hyperactivity may explain some of the behavioural effects of PCP as well as indications of an increased NO production (Fejgin et al., in press). Thus, the PCP-induced behavioural changes could be caused by a loss of inhibitory control via a blockade of NMDA receptors on GABAergic interneurons. This in turn could result in disinhibition of primary corticolimbic neurons leading to a complex neurofunctional imbalance including several neurotransmission systems (Farber, 2003, Olney et al., 1999, Thornberg and Saklad, 1996). By these complex effects, PCP may mimic unique properties of schizophrenia rendering the PCP-model heuristic potential for identifying new treatment rationales (Lipska and Weinberger, 2000) possibly including cognitive dysfunctions. Furthermore, it has been suggested that NO is involved in several aspects of cognition, e.g. learning and memory (Bernstein et al., 2005, Zhuo et al., 1994). Behavioural studies show that inhibitors of NOS disrupt spatial learning (Chapman et al., 1992, Zou et al., 1998) and furthermore that NOS is expressed in e.g. pyramidal neurons of the hippocampus cornu ammonis 1 (CA1), (Pepicelli et al., 2004).

A behavioural paradigm for testing declarative memory in rodents by utilising these animals' well-developed spatial navigation ability was established decades ago (see, e.g., Morris, 1984). The Morris water maze (MWM) model has since been used widely to assess learning and memory function. Acute treatment with PCP has been shown to induce deficits in both spatial learning, working memory, and long-term memory (Wass et al., 2006a, Wass et al., 2006b). Furthermore, (Hanlon et al., 2006) demonstrated that patients with schizophrenia display deficits in learning and memory function using a computerized, virtual version of the MWM. Thus, the MWM has translational value as a cognitive test, with at least face validity, in both preclinical and clinical schizophrenia research. Recent studies by the present authors (Wass et al., 2006a, Wass et al., 2006b) have shown that NOS inhibition reduces PCP-induced disruption of spatial working and long-term memory using different versions of the MWM model. The present study was designed to further investigate the ability of NOS inhibition to counteract the effects of PCP on cognitive function. To this end, the capacity of the NOS inhibitor, NG-nitro-l-arginine methyl ester (L-NAME), to reduce the effect of PCP in rats was studied using a model in which increasing complexity of the learning task was applied with each testing day (“constant reversal learning” i.e. learning a new platform position each training day). In this model, control rats showed an increase in performance over the first three days of training sessions followed by a decline in performance over the last day. One advantage with this model is that it estimates the limit of the rats' cognitive ability.

Section snippets

Animals

Male Sprague–Dawley rats with a body weight of 250 g were purchased from B&K Universal AB, Sollentuna, Sweden. Upon arrival to the animal facility the animals were housed three to four per cage in a colony room for two weeks in order to get used to the new environment and human handling before testing started. Food (B&K Feeds) and water were available ad libitum, and room temperature (20 ± 1 °C) and humidity (55%) were kept constant. The daylight cycle was maintained artificially, lights off

Results

The acquisition sessions were analysed using a repeated measures three-way ANOVA with pretreatment (saline or L-NAME) and treatment (saline or PCP) as independent factors and acquisition session as dependent factor with four different levels (Days 1–4) (Fig. 2).

Discussion

A Morris water maze model designed to assess cognitive flexibility by increasing the cognitive load was used in the present study. The results indicate that rats treated with saline, L-NAME, or L-NAME + PCP, significantly increased their performance in the maze as measured by time to find the platform between Day 1 and Day 2, and Day 1 and Day 3. When the platform was moved to a new position a fourth time (Day 4) there was a decline in performance. Thus, these rats signalled a certain learned

Acknowledgements

Supported by Swedish Research Council (4247), Tornspiran foundation, Lindhés Lawfirm: In memory of Irma and Arvid Larsson-Röst, Wilhelm and Martina Lundgren Forskningsstiftelse, Adlerbertska Forskningsstiftelse, the Theodore and Vada Stanley Stiftelse, Magnus Bergvalls Stiftelse, Stiftelsen Clas Groschinskys Minnesfond, Göteborgs Läkaresällskap, Stiftelsen Bengt Dahréns fond, Svenska Stiftelsen för Medicinsk Forskning, the Swedish Society of Medicine, Åke Wibergs Stiftelse, Fredrik och Ingrid

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