Physiol. Res. 59: 811-819, 2010 Pain Perception in Neurodevelopmental Animal Models of Schizophrenia M. FRANĚK1, S. VACULÍN1, A. YAMAMOTOVÁ1, F. ŠŤASTNÝ2, V. BUBENÍKOVÁ- VALEŠOVÁ2, R. ROKYTA1 1Charles University in Prague, Third Faculty of Medicine, Department of Normal, Pathological and Clinical Physiology, Prague, Czech Republic, 2Prague Psychiatric Center affiliated with the Charles University in Prague, Prague, Czech Republic Received February 17, 2009 Accepted January 18, 2010 On-line April 20, 2010 Summary Corresponding author Animal models are important for the investigation of mechanisms Miloslav Franek, Charles University in Prague, Third Medical and therapeutic approaches in various human diseases, including Faculty, Department of Normal, Pathological and Clinical schizophrenia. Recently, two neurodevelopmental rat models of Physiology, Ke Karlovu 4, 120 00 Prague 2, Czech Republic. this psychosis were developed based upon the use of subunit Fax: +420 224 916 896. E-mail: [email protected] selective N-methyl-D-aspartate receptor agonists – quinolinic acid (QUIN) and N-acetyl-aspartyl-glutamate (NAAG). The aim of this study was to evaluate pain perception in these models. QUIN or Introduction NAAG was infused into lateral cerebral ventricles neonatally. In the adulthood, the pain perception was examined. The rats with Schizophrenia is a neurodevelopmental disorder neonatal brain lesions did not show any significant differences in which afflicts about 1 % of the human population acute mechanical nociception and in formalin test compared to worldwide. The etiology and pathogenesis of this controls. However, the neonatally lesioned rats exhibited psychosis is complex but involves the interplay of significantly higher pain thresholds in thermal nociception. polygenic influences and environmental risk factors Increased levels of mechanical hyperalgesia, accompanying the operating on brain maturation processes. Different sciatic nerve constriction (neuropathic pain), were also observed theories as to the cause of schizophrenia and the in lesioned rats. Although hyperalgesia was more pronounced in heterogeneity of clinical symptoms have made it difficult QUIN-treated animals, the number of c-Fos-immunoreactive to develop a valid animal model. A number of limited neurons of the lumbar spinal cord was similar in experimental animal models have been developed to explore various and control rats. We conclude that neonatal brain lesions theories of etiology, progression and treatment of this attenuated the thermal perception in both nociceptive and psychosis (Marcotte et al. 2001). Neonatal excitotoxic neuropathic pain whereas mechanical pain was increased in the disconnection of the fronto-temporo-hippocampal model of neuropathic pain only. Thus, nociceptive and complex may represent a heuristic model of neuropathic pain belongs – in addition to behavioral changes – schizophrenia-like behavior (Bubeníková-Valešová et al. among the parameters which are affected in described animal 2006, Lipska 2004, Šťastný et al. 2005). Two of these models of schizophrenia. models have been based on the neonatal intracerebroventricular (i.c.v.) infusion of substances, Key words which activate N-methyl-D-aspartate receptor (NMDA- Quinolinic acid • N-acetyl-aspartyl-glutamate • Neuropathic pain • R), when overproduced by activated microglia – Rat quinolinic acid (QUIN) (Heyes et al. 1996, Šťastný et al. 2005) and N-acetyl-aspartyl-glutamate (NAAG) PHYSIOLOGICAL RESEARCH • ISSN 0862-8408 (print) • ISSN 1802-9973 (online) © 2010 Institute of Physiology v.v.i., Academy of Sciences of the Czech Republic, Prague, Czech Republic Fax +420 241 062 164, e-mail: [email protected], www.biomed.cas.cz/physiolres 812 Franěk et al. Vol. 59 (Bubeníková-Valešová et al. 2006, Passani et al. 1998). Neonatal brain lesions Similarities in behavioral changes and changes in social Wistar male pups on postnatal day 12 (PND 12) interactions were observed between these models and a were anesthetized by ethylether, fixed in a stereotaxic genetic model of reduced NMDA-R function (Duncan et apparatus (Stoelting, Wood Dale, IL) using ear and tooth al. 2004). bars. Quinolinic acid (QUIN group) or N-acetyl-aspartyl- It is well known that QUIN and NAAG have glutamate (NAAG group) was slowly infused i.c.v. in a also antinociceptive effects on the pain experience dose of 250 nmol/0.25 μl buffered isotonic (Heyliger et al. 1998, Yamamoto et al. 2004). The effect saline/ventricle; one injection was given into each lateral might be related to a hypofunction of NMDA-R in rat cerebral ventricle by a 1 µl Hamilton syringe equipped lumbar spinal cord (Boyce et al. 1999). The individuals with 26S needle with coordinates previously used with schizophrenia (and their relatives) (Hooley and (Bubeníková-Valešová et al. 2006, Lisý et al. 1994, Delgado 2001) are significantly less sensitive to direct Šťastný et al. 2005). Sham-operated control rats (SHAM painful stimuli (Dworkin 1994, Fishbain 1982, Kudoh et group) were i.c.v. injected with 0.25 µl of saline. al. 2000, Rosenthal et al. 1990), but their subjective Selection of sham-operated or lesioned animals was made evaluation of the stimulus is similar to that of healthy randomly (from each litter 3 pups were treated with control subjects (Blumensohn et al. 2002). It may reflect neuroactive substance, 3 pups were treated with saline a dysfunction of glutamatergic system in these patients, and 2 male pups remained as naïve). The pups were which affects the transmission of the noxious stimuli. weaned on the PND 28 (the day of birth was denoted as To solve this problem we can use animal models PND 0). According to the type of brain lesion three although there are difficulties to evaluate the typical groups of animals were used in this study: QUIN, NAAG psychotic symptoms (hallucinations, lack of motivation, and corresponding SHAM groups (n=4-8 animals per social withdrawal, attention deficits) in animals. group in each pain test). However, it is important to know that the nociceptive profile of these models can help to interpret the clinical Neuropathic pain model results obtained in nociceptive tests owing to a validity of Persistent neuropathic pain in rats was evoked the neurodevelopmental animal models of schizophrenia. by chronic constriction of sciatic nerve (CCI) according The changes of pain perception in animal models are not to the model of Bennett and Xie (1988). For this test, homogenous and depend on used model (Al Amin et al. young adult animals with neonatal brain damage (or 2004, Gao et al. 2004, Schneider and Przewlocki 2005). sham-operated animals) were used on PND 50-55. Rats The purpose of our study was to describe nociceptive were anesthetized with pentobarbital (40 mg /kg b.w., profiles of adult animals (which had been neonatally i.p.). Then the right sciatic nerve was exposed and, treated with icv QUIN or NAAG) with neonatal brain proximally to the trifurcation of the nerve, 5 mm of the lesions induced by QUIN or NAAG. sciatic nerve was freed from the adhering connective tissue. Around the loosened nerve four 4-0 chromic Methods catgut sutures were loosely tied at intervals of approximately 1 mm. Finally, the incisioned muscles and Animals skin were sutured in layers. We have been using Wistar:Hann rats (SPF, BioTest, Konárovice Czech Republic) throughout the Acute nociceptive tests experiments. The litters were rearranged so that each Plantar test and von Frey test were performed contained eight males and two females. Mothers with 2-5 days before the CCI and then 2-3 weeks after this their pups were housed in cages having free access to operation. Formalin test was performed one week before food and water. Animals were kept at 22±2 °C (relative the CCI. Each animal was used in only one nociceptive humidity was 40-70 %) under 12-h light/12-h dark cycle. test; they were not repeatedly exposed to pain. All experiments were approved by the Committee for Pain threshold for the thermal stimulation was Animal Care and Use of the 3rd Faculty of Medicine determined using plantar test equipment (Ugo Basile, (Charles University, Prague) and conducted in Comerio, Italy) as described previously (Procházková et accordance with the ethical guidelines of the International al. 2009). Shortly, in this test the latency (in seconds) of Association for the Study of Pain (Zimmermann 1983). hindlimb withdrawal to the noxious thermal stimulation 2010 Pain in Animal Models of Schizophrenia 813 was measured. Each animal was placed individually in a diaminobenzidine and intensified with nickel solution clear plastic box with a clear glass floor and was allowed (DAB Kit, Vector). to acclimatize for 10 min. Cut-off value was set to 22 s to prevent limb injury. The testing box was cleaned between Statistical analysis each animal. Statistical analysis was carried out by Origin 6 For the testing of mechanical allodynia and (Microcal Software, USA) and by Statistica 6.0 (StatSoft hyperalgesia calibrated nylon von Frey monofilaments Inc., USA). Data are expressed as mean ± S.E.M. (Touch-Test Sensory Evaluator, North Coast Medical Statistical differences were evaluated using paired Inc., Morgan Hill, USA) of different thicknesses were Student´s t-test for comparison of ipsilateral versus used as described elsewhere (Howard et al. 2005). With contralateral sides of the same experimental group. For these filaments the plantar surface of hind limbs was comparison of the ipsilateral versus contralateral sides exposed to varying degrees of pressure. The stimulus was between groups one-way ANOVA test followed by post- applied five times at 2 s intervals for each successive hoc Fisher’s test were used. Differences between means filament until five clear withdrawal responses were were considered statistically significant if p<0.05. For the elicited (100 % response). analysis of von Frey test the stimulus-response curves For chemical nociception, formalin test were constructed. The curves were fitted according to the n (injection of 50 µl of 2.5 % formalin into the plantar equation y=(ymax–ymin)/(1+(x/x0) )+ymin. The value x0 surface of right hindpaw) was used.