Reduced attention and increased impulsivity in mice lacking NPY Y2 receptors: Relation to anxiolytic-like phenotype

Behavioural Brain Research 169 (2006) 325–334

Research report

Reduced attention and increased impulsivity in mice lacking NPY Y2 receptors: Relation to anxiolytic-like phenotype Barbara Greco, Mirjana Carli ∗ Department of Neuroscience, Istituto di Ricerche Farmacologiche “Mario Negri”, via Eritrea 62, 20157 Milano, Italy Received 12 October 2005; received in revised form 23 January 2006; accepted 2 February 2006

Abstract Neuropeptide (NPY) Y2 receptors play an important role in some anxiety-related and stress-related behaviours in mice. Changes in the level of anxiety can affect some cognitive functions such as memory, attention and inhibitory response control. We investigated the effects of NPY Y2 receptor deletion (Y2−/− ) in mice on visual attention and response control using the five-choice serial reaction time (5-CSRT) task in which accuracy of detection of a brief visual stimulus across five spatial locations may serve as a valid behavioural index of attentional functioning. Anticipatory and perseverative responses provide a measure of inhibitory response control. During training, the Y2−/− mice had lower accuracy (% correct), and made more anticipatory responses. At stimulus durations of 2 and 4 s the Y2−/− were as accurate as the Y2+/+ mice but still more impulsive than Y+/+ . At stimulus durations of 0.25 and 0.5 s both groups performed worse but the Y2−/− mice made significantly fewer correct responses than the Y2+/+ controls. The anxiolytic drug diazepam at 2 mg/kg IP greatly increased the anticipatory responding of Y2−/− mice compared to Y2+/+ . The anxiogenic inverse benzodiazepine agonist, FG 7142, at 10 mg/kg IP reduced the anticipatory responding of Y2−/− but not Y2+/+ mice. These data suggest that NPY Y2 receptors make an important contribution to mechanisms controlling attentional functioning and “impulsivity”. They also show that “impulsivity” of NPY Y2−/− mice may depend on their level of anxiety. These findings may help in understanding the pathophysiology of stress disorders and depression. © 2006 Elsevier B.V. All rights reserved. Keywords: Y2 knockout mice; Attention; Impulsivity; Anxiety; Stress; Diazepam; FG 7142; 5-CSRT task

1. Introduction Neuropeptide Y (NPY) is a highly conserved 36-aminoacid peptide, abundantly expressed in the central and peripheral nervous systems [87]. It plays an important role in various basic physiological processes such as food and water intake, pain, seizure activity, memory and learning [30,39,82,92]. Animal and human studies have suggested that NPY, together with its receptors, might be directly implicated in the pathophysiology of affective disorders such as depression and anxiety [5,6,40,52,65,71,84,89,93,94]. Several studies have reported low levels of NPY in cerebrospinal fluid (CSF) [40,95] and plasma [65] of patients with major depression. In addition, individuals with low levels of NPY appear to be predisposed to anxiety-related depression [40].

∗

Corresponding author. Tel.: +39 0239014466; fax: +39 023546277. E-mail address: [email protected] (M. Carli).

0166-4328/$ – see front matter © 2006 Elsevier B.V. All rights reserved. doi:10.1016/j.bbr.2006.02.002

Loss of concentration, difficulty to direct attention and impairment in executive functioning have been reported in patients suffering from major depression [50,57,72,96]. However, whether the attentional deficits in depressed patients are due to some change in NPY activity is not known. NPY effects are mediated through at least five distinct receptor subtypes [58]. The NPY Y2 receptors are abundant in the hippocampal formation, the lateral septum, amygdala and in the basal ganglia nuclei while the highest concentration of Y1 receptors are in superficial layers of cortex and thalamic nuclei [27,31,36]. The Y1 and Y2 receptors modulate anxiety-related and stress-related effects of NPY, in opposite ways. The Y1 receptor agonist [27] NPY mimics the anxiolytic and antidepressant action of NPY [38,71,84] while Y2 receptor agonists, such as NPY13–36 and C2-NPY, induce anxiogenic-like responses [63,78]. In contrast, the Y1 receptor antagonist BIBP3226 has anxiogenic properties [46] while Y2 receptor knockout mice have an anxiolytic-like phenotype that may explain their superior stress-coping ability [70,89].

326

B. Greco, M. Carli / Behavioural Brain Research 169 (2006) 325–334

Several data support the theory that changes in the level of anxiety can affect some aspects of cognitive functions such as attention, aspects of executive function and memory. In humans, anxiolytic drugs such as the benzodiazepine receptor (BZR) agonist diazepam impairs performance in tests dependent on frontal lobe functions such as planning, extra-dimensional attentional set shifting, spatial working memory and in go/no-go tasks [19–21,23]. In rodents, BZR agonists such as chlordiazepoxide and diazepam affect the acquisition of the Morris water maze task [55], have amnesic effects in short-term memory tests [12,91], and impair sustained attention [53]. Anxiogenic ␤-carbolines such as FG 7142 and ␤CCM enhance learning and memory in several tests such as passive avoidance and delayed-matching-to-position [12,91], and affect sustained attention in vigilance tasks and spatial working memory [43,53,62,83]. Anxiety may either facilitate or impair performance, depending on the nature and the difficulty of the task [28,29] and unpleasant emotional state such as anxiety can alter the level of arousal [66]. These observations suggest a close relationship between anxiety, arousal and performance [29,98]. Anxious and over-aroused mice showed changes in performance in a fivechoice serial reaction time (5-CSRT) task [90], water T-maze, Morris water maze and a hole-board test [67]. Mice with a low level of anxiety also had altered performance in spatial learning, memory and attention [34,73]. These findings suggested that the cognitive performance of Y2−/− mice could be influenced by the level of anxiety [89]. This study investigated the role of Y2 receptor deletion on visual attention and inhibitory response control. Attentional performance was assessed using a murine version of the 5-CSRT task [34,44,51] in which the detection of a brief visual stimulus across five spatial locations may serve as a valid behavioural index of aspects of sustained and divided visual-spatial attentional functioning. Additional measures of performance such as anticipatory “impulsive” and perseverative responses provide a measure of inhibitory response control [75]. In order to assess whether the reduced attentional performance in knockout mice was due to an alteration in anxietyrelated behaviour, we investigated the effects of the anxiolytic drug diazepam and the anxiogenic BZR partial inverse agonist FG 7142 on 5-CSRT task performance by wild-type and Y2 knockout mice. 2. Materials and methods 2.1. Animals One group of nine F2 male Y2 knockout (Y2−/− ) and one group of nine F2 male Y2 wild-type (Y2+/+ ) mice were used. The Y2−/− mice were developed in the laboratory of Dr. Herbert Herzog (Garvan Institute of Medical Research, Sydney, Australia) as previously described [77,89]. The Y2−/− and Y2+/+ were bred as homozygous from littermates on a mixed C57BL/6-129SvJ background. The founders of Y2−/− and Y2+/+ mice used in this study were obtained from Dr. Herbert Herzog and the colony expanded at the Charles River facilities in Italy (Calco, Italy). At the start of the experiments the mice weighed 25–30 g and were housed under temperature-controlled conditions (21 ◦ C) with a 12 h light:12 h dark cycle (light on 7:00 a.m.–7:00 p.m.). Food (Rieper, Italy) was available ad libitum. The animals had 2 h of access to water at the end of each

day’s testing. Over the weekends, from Friday evening until Sunday evening, animals had free access to water. Procedures involving animals and their care were conducted in conformity with the institutional guidelines that are in compliance with the national (Decreto Legislativo n. 116, Gazzetta Ufficiale, suppl., 40, 18 Febbraio 1992, Circolare No. 8, Gazzetta Ufficiale, 14 luglio 1994) and international laws and policies (EEC Council Directive 86/609, OJ L 358,1, Dec. 12, 1987; Guide for the Care and Use of Laboratory Animals, U.S. National Research Council, 1996).

2.2. Behavioural procedures 2.2.1. Apparatus The test apparatus consisted of four 21.6 cm × 17.8 cm × 12.7 cm chambers (Med Associates Inc. USA), as previously described [34]. Stimuli and recording of responses were managed by a SmartCtrlTM Package 8 In/16 Out (Med Associates Inc. USA) with additional interfacing by MED-PC for Windows (Med Associates Inc. USA). The running program for the 5-CSRT task was customwritten. 2.2.2. Habituation to liquid reinforcer and nose-poking in the holes Mice were handled for 1 week and their body weight recorded. They were then water-deprived by allowing them 2 h access to water in the early evening until their body weight had stabilised (8 days). Then, over the next 2 days the mice were habituated in their home cages to the 10% sucrose solution used afterwards as reward in the nose-pocking training and 5-CSRT task. On the following 2 days mice were habituated to the operant boxes. During this stage, 10% sucrose solution was available in a small bowl placed below the receptacle hole of the box. First, mice had to learn that every 5 s the liquid reward was available in a small cup in the receptacle hole. During this period head entries were recorded. During the next period, Y2+/+ and Y2−/− mice were trained to poke their noses into the illuminated holes. Immediately after a poke in the water receptacle a visual stimulus was presented by turning on a light emitting diode (LED) at the rear of one of the holes. A nose-poke in the lighted hole extinguished the light stimulus and the liquid dipper provided a 0.01 mL liquid reward in the receptacle hole. Any response in one of the other four holes had no consequence and was not recorded. The light stimulus was presented in all five holes in random order. A mouse was switched to the 5-CSRT task after it had completed at least 50 rewarded nose-poke trials in one 30 min session. 2.2.3. The five-choice serial reaction time task The start of the session was signalled by illumination of the house-light and the delivery of a 0.01 mL liquid reward. Nose-poking in the receptacle hole began the first trial. After a fixed delay (the inter-trial interval, ITI), the LED at the rear of one of the holes came on for a short period. The LED stimulus was presented the same number of times in each hole during a complete session, with the order of presentation randomised by the computer. While the light was on, and for a short period afterwards (the limited hold), responses in the hole that was illuminated (correct response) resulted in the liquid reward. Responses in the holes that had not been illuminated (incorrect responses) or failure to respond within the limited hold (omissions) caused the house-lights to be turned off for a short period (time out). Responses in the holes while the house-light was off restarted the time out. After the delivery of the liquid reward, or at the end of time out, the mouse started the next trial by poking its nose into the receptacle hole. Responses made in the holes after a correct response (perseverative responses), or after the end of time out before nose-poking into the receptacle hole, resulted in a period of time out. Responses in the holes during the ITI (anticipatory responses) also resulted in a period of time out. After anticipatory responses a nose-poke into the receptacle hole restarted the current trial. Each daily session consisted of 100 trials or 30 min of testing, whichever was completed sooner, after which all lights were turned off and further responses had no effect. In the first session of the test schedule, the stimulus and limited hold each lasted 1 min and, depending on individual performance, they were progressively reduced to 1 s. The stimulus duration was reduced in the following sequence: 60, 30, 10, 5, 2.5, 2, 1.5 and 1 s (baseline). The ITI and time out both lasted 2 s during the first session and the ITI was raised to 5 s in subsequent

B. Greco, M. Carli / Behavioural Brain Research 169 (2006) 325–334 sessions; time out was not changed. Throughout the whole period of training and experiments each mouse had one session per day on a 5-CSRT task. Throughout the whole duration of these studies each mouse had only one session per day of training or testing always at the same time of the day (between 9 a.m. and 1 p.m.). 2.2.4. Manipulation of basic task parameters Once stable performance was achieved at 1.0 s stimulus duration, a series of manipulations were done to the basic task, designed to increase the attentional load. First, we manipulated the attentional load by presenting to the mouse visual stimuli of varying durations. The visual stimuli of 0.25, 0.5, 1.0, 2.0 and 4.0 s durations were mixed randomly during a session of 250 trials. Each stimulus duration was presented on 50 occasions. The session lasted 1 h. Second, the attentional load on performance was increased by disrupting the temporal predictability of the stimulus onset. After a series of sessions under baseline conditions, the presentation of the visual stimuli was rendered unpredictable by randomly mixing four different ITIs (3, 4, 5 and 6 s) during a single session with 200 trials (50 per ITI) lasting 1 h. 2.2.5. Pharmacological challenges of anxiety levels To analyse the relationship between the anxiety phenotype of Y2−/− mice with their performance in the 5-CSRT task the level of anxiety was manipulated by giving anxiolytic (benzodiazepine) and anxiogenic (␤-carboline) drugs. The experimental design employed a counterbalanced order of repeated drug injections. On the first day of testing half of Y2+/+ and half of Y2−/− mice received vehicle (0.5% CMC) the other half received 2 mg/kg diazepam. Three days later the treatments were inverted. During the intervening days mice were always trained to re-establish baseline performance and to check for long-lasting effects of drugs. One month after the experiments with diazepam mice were injected with vehicle or FG 7142 (10 mg/kg) according to the same counterbalanced treatment scheme. During the period between the drug testing mice were trained daily. The doses of diazepam (2 mg/kg) and FG 7142 (10 mg/kg) used for these studies were previously reported for their anxiolytic and anxiogenic activity in a variety of mouse strains and murine models of anxiety such as elevated plus maze, social interaction, hole-board exploration, light–dark, elevated T-maze and T-maze alternation [9,24,35,37,48,68,76,86]. Higher doses of diazepam and FG 7142 were not used as they might affect spontaneous locomotor activity and thus, interfere with the performance of the 5-CSRT task [35,37]. Under baseline task conditions vehicle (0.5% carboxymethylcellulose, CMC) or diazepam 2 mg/kg (Ravizza, Italy), dissolved in 0.5% CMC in a volume of 10 mL/kg was injected to mice intraperitoneally (IP) 15 min before the test. The effects of the anxiogenic drug FG 7142 (Schering, Germany) were examined with a longer ITI (9 s). Vehicle (0.5% CMC) or FG 7142 10 mg/kg (dissolved in 0.5% CMC) were injected IP in a volume of 10 mL/kg 15 min before the test session. The various treatments were administered at least 48 h apart in counterbalanced order. 2.2.6. Statistical analysis The main dependent variables selected for analysis were: (a) the percentage of correct responses (total correct responses/total correct + total incorrect responses × 100); (b) percentage of omissions (total omissions/total correct responses + total incorrect responses + total omissions × 100); (c) the number of anticipatory responses in the holes during the ITI; (d) the number of perseverative responses in the holes after a correct response; (e) the number of trials completed (total correct responses + total incorrect responses + total omissions) and (F) the number of nose-pokes in the receptacle hole. Correct responses and√omissions, as percentages, were transformed according to the formula 2 arcsin % × 100, to normalize the distributions in accordance with the ANOVA model [97]. The numbers of initial nose-poke training sessions for the two groups of mice (Y2+/+ , n = 9; Y2−/− , n = 9), were compared by Student’s t-test. Each variable of attentional performance was analysed by a split-plot ANOVA with a betweensubjects factor Genotype and a within-subjects factor Training Level (60, 30, 10, 5.0, 2.5, 2.0, 1.5 and 1.0 s). Similarly, the data collected during the challenge sessions with variable stimulus durations or unpredictable ITI’s were examined by split-plot ANOVA with between-subjects factor Genotype and within-subjects factor either Stimulus Duration (0.25, 0.5, 1.0, 2.0 and 4.0 s) or ITI (3, 4, 5 and 6 s). Although the effects of pharmacological challenges were examined in the

327

same mice, data of diazepam and FG 7142 were analysed independently by a split-plot ANOVA with a between factor Genotype and within factor Drug.

3. Results 3.1. Acquisition of the 5-CSRT task The Y2−/− mice took twice as many sessions as Y2+/+ to consistently nose-poke into an illuminated hole (Y2+/+ 11.3 ± 1.4; Y2−/− 23.0 ± 3.8; t = 2.86, P = 0.011). Both Y2+/+ and Y2−/− showed a progressive improvement in accuracy during the acquisition phase. The mouse genotype strongly influenced the accuracy of detection throughout the whole period of training as shown by a significant effect of genotype (F1,16 = 41.5 P < 0.0001), Training Level (F7,112 = 19.2 P < 0.0001) and the interaction Genotype × Training Level (F7,112 = 2.9 P = 0.008). At each Training Level the Y2−/− made significantly fewer correct responses than Y2+/+ mice (Student’s t-test, P < 0.05). As shown in Fig. 1A, although the Y2−/− mice improved with

Fig. 1. The attentional performance of Y2+/+ and Y2−/− mice during acquisition of the 5-CSRT task. Accuracy measured as % correct responses is presented in, A and the number of anticipatory responses in, B. The stimulus duration was progressively reduced as follows sequence: 60, 30, 10, 5, 2.5, 2.0, 1.5 and 1.0 s. Mean ± S.E.M. of nine Y2+/+ and nine Y2−/− mice averaged over the last three sessions at any given Training Level. * P < 0.05 vs. Y2+/+ mice (Tukey’s test).

328

B. Greco, M. Carli / Behavioural Brain Research 169 (2006) 325–334

Table 1 The 5-CSRT task performance of Y2+/+ and Y2−/− mice during training Stimulus durations

60

30

10

5

2.5

2.0

1.5

1.0

No. of perseverative responses Y2+/+ 38.4 ± 4.3 Y2−/− 31.7 ± 4.4

39.7 ± 7.2 39.7 ± 6.6

46.0 ± 5.0 35.2 ± 7.0

34.9 ± 3.2 37.9 ± 8.2

41.1 ± 7.8 44.5 ± 8.3

35.7 ± 6.8 46.8 ± 7.8

35.9 ± 7.9 51.3 ± 8.1

30.2 ± 5.7 49.2 ± 7.2

Omissions (%) Y2+/+ Y2−/−

2.0 ± 0.7 6.0 ± 2.7

5.8 ± 1.3 11.9 ± 3.8

14.4 ± 1.8 33.2 ± 5.7*

30.6 ± 5.1 48.3 ± 4.0*

33.0 ± 4.0 51.7 ± 2.8*

35.5 ± 5.8 44.3 ± 5.5*

33.6 ± 5.1 40.2 ± 5.4

35.0 ± 5.4 44.0 ± 5.8

No. of trials completed Y2+/+ Y2−/−

54 ± 4 55 ± 6

73 ± 7 54 ± 7

81 ± 6 61 ± 7

75 ± 8 63 ± 7

78 ± 7 66 ± 6

77 ± 7 59 ± 6

78 ± 6 59 ± 3

78 ± 6 61 ± 5

Mean ± S.E.M. of nine Y2+/+ and nine Y2−/− mice averaged over the last three sessions at any Training Level. * P < 0.05 vs. Y2+/+ mice (Student’s t-test).

training, thus showing a degree of stimulus control, they never reached the accuracy of wild-type mice (Y2+/+ ). At the beginning of training, the accuracy of wild-type mice was about 75% and after only four training levels they reached a stable accuracy of 97%; the accuracy of Y2−/− mice was 50% at the start of training and never got better than 87%. During training when the stimulus duration was shorten to less than 2 s the accuracy of Y2−/− but not of Y2+/+ declined, reaching 70% at 1.0 s. However, with further training at stimulus duration 1.0 s (baseline) the correct responses (accuracy) stabilised at 90–95% for Y2+/+ and at 70–80% for Y2−/− . As illustrated in Fig. 1B, Y2−/− mice made many more anticipatory responses than the wild-type (F1,16 = 13.7 P < 0.002) but the number of anticipatory responses was not affected by the training (Training Level, F7,112 = 1.1 P = 0.4; Genotype × Training Level, F7,112 = 0.9 P = 0.4). The mean number of anticipatory responses by Y2+/+ and Y2−/− mice was about 5 and 12, respectively and was constant throughout all experiments. Table 1 shows that the number of perseverative responses was not affected by Genotype (F1,16 = 0.3 P = 0.5). The mean number of perseverative responses by Y2+/+ remained steady at about 38 whereas those by Y2−/− mice increased with training from 31 to 50 (Training Level, F7,112 = 0.7 P = 0.6; Genotype × Training Level, F7,112 = 2.3 P = 0.03). Omissions shown in Table 1 increased on shortening the stimulus duration (Training Level, F7,112 = 75.5 P < 0.0001) particularly in Y2−/− mice (Genotype, F1,16 = 8.0 P = 0.01; Genotype × Training Level F7,112 = 2.3 P = 0.03). The number of trials completed (Table 1) increased steadily but equally in both groups (Genotype, F1,16 = 4.0 P = 0.06; Training Level, F7,112 = 4.6 P = 0.001; Genotype × Training Level, F7,112 = 1.7 P = 0.12). Compared to Y2−/− mice, the Y2+/+ mice completed a larger number of trials but a difference was not significant.

a significant effect of Genotype (F1,12 = 155 P = 0.002), Stimulus Duration (F4,48 = 21.0 P < 0.0001) and Genotype × Stimulus Duration (F4,48 = 2.7 P = 0.03). As illustrated in Fig. 2A, when the stimulus lasted 1 s, Y2+/+ still made more correct responses

3.2. Manipulation of task parameters 3.2.1. Variable stimulus duration In this experiment the attentional load was manipulated by presenting the mouse with visual stimuli of different duration (0.25, 0.5, 1, 2 and 4 s) in a single session. ANOVA indicated

Fig. 2. The effects of stimulus duration on accuracy, A and anticipatory responses, B of Y2+/+ and Y2−/− mice. Different stimulus durations (0.25, 0.5, 1.0, 2.0 and 4.0 s) were presented randomly during a single session. Mean ± S.E.M. of nine Y2+/+ and nine Y2−/− mice. * P < 0.05 vs. Y2+/+ mice (Tukey’s test).

B. Greco, M. Carli / Behavioural Brain Research 169 (2006) 325–334

329

than Y2−/− mice, although the difference was not significant (Tukey’s test P = 0.08). Cutting the stimulus to 0.25 and 0.5 s impaired the accuracy of both groups. In addition, the accuracy of Y2−/− mice in the task at stimulus durations of 0.25 and 0.5 s was significantly lower than that of Y2+/+ mice (both Student’s t-test, P < 0.05). As shown in Fig. 2B, the stimulus duration had no effect on the number of anticipatory responses (Stimulus Duration, F4,48 = 0.7 P = 0.6) or on the interaction between genotype and stimulus duration (F4,48 = 1.4 P = 0.2). Overall, the Y2−/− group made more anticipatory responses than the Y2+/+ mice (Genotype, F1,12 = 4.8 P = 0.04). The genotype did not affect the number of perseverative responses (Genotype, F1,12 = 0.2 P = 0.6) nor did stimulus duration (Stimulus duration, F4,48 = 0.2 P = 0.9; Genotype × Stimulus Duration, F4,48 = 0.5 P = 0.7) (data not shown). The percentage of omissions increased equally in both groups of mice at short stimulus duration (Stimulus Duration, F4,48 = 37.2 P < 0.0001 equally (Genotype, F1,12 = 1.1 P = 0.3; Genotype × Stimulus Duration F4,48 = 2.1 P = 0.09). The number of trials completed was similar in the two groups (Genotype, F1,12 = 0.5 P = 0.5) and it was not affected by stimulus duration (Stimulus Duration, F4,48 = 0.6 P = 0.7; Genotype × Stimulus Duration F4,48 = 0.3 P = 0.9) (data not shown). 3.2.2. Variable inter-trial interval In this experiment the attentional load was manipulated by making the presentation of visual stimuli temporally unpredictable. This was achieved by random presentation of different ITIs. As illustrated in Fig. 3A, although the Y2−/− were still less accurate than Y2+/+ mice (Genotype, F1,16 = 35.6 P < 0.0001), disrupting temporal predictability had no additional effects on accuracy (ITI, F3,48 = 0.3 P = 0.8; Genotype × ITI, F3,48 = 0.6 P = 0.6). Overall, the Y2−/− mice made significantly more anticipatory responses than the Y2+/+ group (Genotype, F1,16 = 5.1 P = 0.03). The number of anticipatory responses was increased by ITI (ITI, F3,48 = 4.0 P = 0.01) but the interaction between genotype and ITI was not significant (F3,48 = 2.4 P = 0.08). However (Fig. 3B), the Y2−/− mice made more anticipatory responses at longer ITI (Tukey’s test, P < 0.05). ITI (F3,48 = 1.3 P = 0.3), genotype (F1,16 = 0.6 P = 0.4) and their interaction (F3,48 = 0.5 P = 0.7) had no significant effects on perseverative responses. Presenting ITIs of different lengths unpredictably had no systematic effect on omissions (Genotype, F1,16 = 3.5 P = 0.08; ITI, F3,48 = 0.2 P = 0.8; Genotype × ITI, F3,48 = 0.4 P = 0.8) or trials completed (Genotype, F1,16 = 3.5 P = 0.08; ITI, F3,48 = 0.2 P = 0.8; Genotype × ITI, F3,48 = 0.4 P = 0.8) (data not shown). 3.3. Pharmacological manipulation of anxiety level 3.3.1. Effects of diazepam Fig. 4A shows that Y2−/− mice made fewer correct responses than wild-type Y2+/+ mice (Genotype, F1,16 = 18.8 P = 0.0005). The anxiolytic drug diazepam, at 2 mg/kg had no effect in Y2+/+ mice but showed a tendency to further reduce the percentage of correct responses of Y2−/− mice (Genotype × Diazepam,

Fig. 3. Accuracy, A and anticipatory responding, B of Y2+/+ and Y2−/− mice when the visual stimuli were presented unpredictably by varying the ITIs. Four different ITIs (3.0–4.0–5.0–6.0 s) were presented randomly during a single session. Mean ± S.E.M. of nine Y2+/+ and nine Y2−/− mice. * P < 0.05 vs. Y2 +/+ mice (Tukey’s test). # P < 0.05 vs. ITI 3 s (Tukey’s test).

F1,16 = 4.1 P = 0.05). The effects of diazepam in Y2−/− mice was statistically not significant (Tukey’s test P > 0.05). As shown in Fig. 4B, Y2−/− mice made more anticipatory responses than Y2+/+ mice (Genotype, F1,16 = 22.7 P = 0.0002). Diazepam significantly increased the number of anticipatory responses in both groups (Diazepam, F1,16 = 15.5 P = 0.001) but the effect was stronger in Y2−/− mice (Genotype × Diazepam, F1,16 = 5.7 P = 0.03). Comparison of treatment means showed that Y2−/− mice treated with diazepam made more anticipatory responses (Tukey’s test, P < 0.05). The number of perseverative responses was the same in both groups and was not affected by 2 mg/kg diazepam (Genotype, F1,16 = 0.9 P = 0.4; Diazepam, F1,16 = 3.9 P = 0.06; Genotype × Diazepam, F1,16 = 1.2 P = 0.3) (data not shown). Diazepam had no effect on the percentage of omissions (Diazepam, F1,16 = 1.5 P = 0.2; Genotype × Diazepam F1,16 = 0.3 P = 0.6) or the number of trials completed (Diazepam,

330

B. Greco, M. Carli / Behavioural Brain Research 169 (2006) 325–334

Fig. 4. Effects of diazepam on accuracy, A and anticipatory responding, B of Y2+/+ and Y2−/− mice. Stimulus duration was 1 s and ITI was 5 s. Fifteen minutes before the test session mice were injected IP with vehicle (0.5% CMC) or 2 mg/kg diazepam. Drug test sessions were at least 48 h apart. The histograms show the mean ± S.E.M. of nine Y2+/+ and nine Y2−/− mice. White bar: vehicle and grey bar: diazepam. * P < 0.05 vs. Y2 +/+ mice (Student’s t-test). # P < 0.05 vs. vehicle (Tukey’s test).

F1,16 = 1.4 P = 0.2; Genotype × Diazepam: F1,16 = 2.8 P = 0.1) (data not shown). 3.3.2. Effects of FG 7142 To assess the effect of the anxiogenic compound FG 7142 on the attentional performance of wild-type Y2+/+ and knockout Y2−/− mice, the duration of ITI was increased to 9 s while the stimulus duration was kept at 1.0 s. Fig. 5A illustrate the effects of 10 mg/kg FG 7142 on accuracy. The Y2+/+ mice had significantly higher accuracy than Y2−/− mice, as shown by their higher proportion of correct responses (Genotype, F1,16 = 12.1 P = 0.003). However, FG 7142 had no effect on accuracy of either Y2+/+ or Y2−/− mice (FG 7142, F1,16 = 1.7 P = 0.2; Genotype × FG 7142, F1,16 = 0.9 P = 0.3). As shown in Fig. 5B, Y2−/− mice made more anticipatory responses than Y2+/+ (Genotype, F1,16 = 9.8 P = 0.006). FG 7142 significantly reduced the number of anticipatory responses in Y2−/− but had no such effect in Y2+/+ mice (FG 7142, F1,16 = 9.8 P = 0.006; Genotype × FG 7142, F1,16 = 4.6 P = 0.05). The number of perseverative responses was the same in both groups and was not affected by FG 7142 (Genotype, F1,16 = 0.3 P = 0.5; FG 7142, F1,16 = 0.3 P = 0.6; Genotype × FG

Fig. 5. Effects of FG 7142 on accuracy, A and anticipatory responding, B of Y2+/+ and Y2−/− mice. Stimulus duration was 1 s and ITI was 9 s. Fifteen minutes before the test session mice were injected IP with vehicle (0.5% CMC) or 10 mg/kg FG 7142. Drug test sessions were at least 48 h apart. The histograms show the mean ± S.E.M. of nine Y2+/+ and nine Y2−/− mice. White bar: vehicle and grey bar: FG 7142. * P < 0.05 vs. Y2 +/+ mice (Student’s t-test). # P < 0.05 vs. vehicle (Tukey’s test).

7142, F1,16 = 0.2 P = 0.7) (data not shown). The percentage of omissions (FG 7142, F1,16 = 0.9 P = 0.3; Genotype × FG 7142, F1,16 = 1,2 P = 0.3) and the number of trials completed (FG 7142, F1,16 = 0.03 P = 0.9; Genotype × FG 7142, F1,16 = 0.02 P = 0.9) were not affected by FG 7142 in either group of mice (data not shown). 4. Discussion This study found that the deletion of NPY Y2 receptors results in deficits in attentional functioning and impulsivity (Table 2). Table 2 Summary of the effects of behavioural challenges and anxiety level on accuracy and impulsivity of NPY Y2−/− mice

Short ST Long ST Variable ITI Diazepam FG 7142

Accuracy

Impulsivity

⇓ ⇑ 0 0 0

0 0 ⇑ ⇑ ⇓

ST = stimulus duration; ITI = inter-trial interval; ⇑ = increase; ⇓ = decrease; 0 = no effect.

B. Greco, M. Carli / Behavioural Brain Research 169 (2006) 325–334

Both Y2+/+ and Y2−/− mice learned the task but Y2−/− mice were less accurate and made more anticipatory responses than Y2+/+ mice throughout training and testing. The number of perseverative responses, the proportion of omissions and the number of trials completed were similar in both groups. To test the attentional nature of deficits in Y2−/− mice the difficulty of the 5-CSRT task was manipulated in various ways. Reducing the attentional load by prolonging the visual stimulus from 1 to 2 and 4 s improved the accuracy of Y2−/− mice, while shortening the stimulus from 1 s to 0.5 and 0.25 s further impaired their accuracy. These findings suggest that the differences in accuracy between Y2+/+ and Y2−/− mice might have resulted from differences in attentional functioning. The mouse’s ability to sustain attention was challenged by preventing the mice using temporal strategies to cope with the task requirements. Thus, the visual target stimuli were presented unpredictably by randomly intermixing the ITIs of 3, 4, 5 and 6 s. This had no effect on the accuracy of either Y2+/+ or Y2−/− mice. However, whereas the wild-type mice were not affected, the Y2−/− mice became more impulsive, suggesting that they were unable to time their responses efficiently in anticipation of reward. Moreover, the data show that impulsivity and accuracy are relatively independent [75]. It is interesting that the higher number of perseverative responses by Y2−/− mice was only evident during early training. Impulsivity and perseveration might be measuring two independent mechanisms of response control. Recent data show that inhibitory processes of response control can be dissociated at the neural level; lesions of the orbital frontal cortex induce perseverative responses while lesions of the infralimbic cortex cause impulsive responding in the 5-CSRT task [11]. Mice displaying a high level of locomotor activity in home cages were also impulsive in a delayed-reinforcement task, suggesting that impulsivity in some tasks may depend on motor hyper-activity [45]. The Y2−/− mice were more active than Y2+/+ mice in exploration of a new environment in an open field test [89] although their spontaneous motor activity in home cages was similar [89], suggesting that enhanced exploration might be secondary to emotional responses. Although the relationship between exploration, emotionality and locomotion is unclear [74] several studies have reported that drugs that affect emotionality such as anxiolytics increase locomotion in an openfield test whereas spontaneous locomotion measured in activity cages may even decrease [24,37,86]. Conversely anxiogenic agents were shown to decrease exploratory behaviour [86] at doses that have no effect on spontaneous locomotion in activity cages [37]. Employing various well-established behavioural paradigms to test anxiety such as elevated plus maze, open field and light–dark test, it has been demonstrated that the deletion of NPY Y2 receptor subtypes potently suppresses anxiety-related behaviour such as decreased exploration induced by novel or highly exposed and illuminated environments [70,89]. Increase in impulsive-like behaviour has been reported after anxiolytic drugs [8,23,88]. Thus the impulsivity of Y2−/− mice might have not resulted from motor activation and may well have been due to the lower emotional level. It is well known that in cogni-

331

tive tasks the level of emotion can affect performance, with an inverted U-shaped curve [66,98]. Systemically administered axiolytic drug, diazepam, increased and anxiogenic compound, FG 7142, reduced the number of anticipatory responses of Y2−/− mice. In agreement with our data, increases of impulsivity induced by benzodiazepines have been reported in several behavioural tasks [2]. The benzodiazepine receptor agonists increased impulsivity in a delayed T-maze [88], and a delayed-reinforcement task [8], and disrupted the ability to refrain from responding during no-go trials in go/no-go procedures [15,16]. Increased impulsive behaviour has also been reported in the differential reinforcement of low rates of responding (DRL) schedule [64]. It is interesting to note that deletion of Y2 receptors caused a 60% reduction of corticotropin-releasing factor (CRF) mRNA expression [77]. The CRF pathway strongly influences anxiety-related and stress-related behaviour [18]; CRF increases and CRF antagonists block these anxiety-like and stress-like behaviours [56]. Over-expressing CRH-transgenic mice made fewer anticipatory responses than wild-type mice during training and testing on a 5-CSRT task [90]. FG 7142 did not affect accuracy of Y2−/− mice whereas diazepam reduced accuracy but not significantly. These findings indicate that although the level of anxiety might influence impulsivity its role in accuracy is less clear. Both in humans and rodents, benzodiazepine impaired performance in several tasks [54]. However, no effect of benzodiazepines was reported on the accuracy of well-trained rats performing an operant auditory or visual conditional discrimination task [25,26] and divided attention task [54]. In transgenic mice overproducing CRH, diazepam had no effect on accuracy in a 5-CSRT task [90]. The highest density of NPY Y2 receptors [27] is found in the hippocampus an area particularly involved in learning/memory [79] but also in behavioural inhibition and anxiety [33]. A link between hippocampal functions and attention has not been clearly established. The perforant path lesions had no effect on attentional performance in the 5-CSRT task [47] but one study reported that animals with hippocampal lesions were impaired in the acquisition of efficient performance in this task [3]. A high density of Y2 receptors has also been reported in the amygdala [27], which is implicated in the control of emotional behaviour [22] but also attention [32,42]. It is well known that the level of emotion can affect arousal and attention [29,66]. Thus it could not be excluded that the NPY Y2 receptors in the hippocampus and amygdala might play some role in attention and the acquisition of efficient performance. The Y2 receptors have generally been assumed to be presynaptic receptors that negatively modulate NPY release [17]. Their deletion results in increased NPY release on post-synaptic Y1 receptors and therefore anxiolytic-like actions [85]. Y2 receptors also regulate the release of other neurotransmitters such as glutamate, GABA and dopamine [69,81,85]. Intracerebral injection of NPY causes a dose-related increases of dopamine and DOPAC levels [40]. Excessive or low dopamine activity in the prefrontal cortex (PFC) is detrimental to cognitive functioning in rodents and monkeys [1,4,7,80,99]. Additionally, increasing DA function by d-amphetamine, particularly in

332

B. Greco, M. Carli / Behavioural Brain Research 169 (2006) 325–334

the nucleus accumbens, increases impulsivity [13,14]. Autoradiographic studies show particularly high Y2 binding sites in the ventral tegmental area and substantia nigra compacta, areas of origin of DA cells [31]. Thus, the NPY Y2 receptors may influence the attentional performance by interfering with DA mechanism in the PFC and connected structures. Activation of Y2 receptors suppresses glutamatergic transmission through presynaptic mechanisms [69]. Recent evidence suggests that glutamate NMDA receptor antagonists induced attentional deficit and impulsivity in rats [41,49,59] and mice [34]. Behavioural deficits induced by glutamate NMDA receptor antagonists have been associated with an enhanced firing rate of pyramidal neurons and glutamate release in the frontal cortex [10,60,61]. Therefore, the deletion of Y2 receptors by interfering with glutamatergic and dopaminergic neurotransmission may lead to attentional deficits and impulsivity. These findings add to previous work [89] indicating a major role of NPY Y2 receptors in emotional response and behavioural arousal and suggest that the anxiolytic-like behavioural phenotype of these Y2−/− mice may contribute to impulsivity. However, the additional deficits in their attentional functioning might be independent of the emotional level. Acknowledgements We thank Dr. Herbert Herzog, Garvan Institute of Medical Research, Sydney, Australia and Dr. Guenther Sperk, University of Innsbruck, Austria, for kindly providing the NPY Y2 knockout mice and Dr. Annamaria Vezzani, for helpful discussion of these studies. References [1] Arnsten AF, Cai JX, Murphy BL, Goldman-Rakic PS. Dopamine D1 receptor mechanisms in the cognitive performance of young adult and aged monkeys. Psychopharmacology (Berl) 1994;116:143–51. [2] Bizot JC, Thiebot MH. Impulsivity as a confounding factor in certain animal tests of cognitive function. Brain Res Cogn Brain Res 1996;3:243–50. [3] Bratt AM, Stacey K, Chase RM, Mittleman G. Hippocampal lesions and acquisition of a 5-choice selective attention task. Soc Neurosci Abs 21 1995;21:477.2. [4] Brozoski TJ, Brown RM, Rosvold HE, Goldman PS. Cognitive deficit caused by regional depletion of dopamine in prefrontal cortex of rhesus monkey. Science 1979;205:929–32. [5] Caberlotto L, Hurd YL. Reduced neuropeptide Y mRNA expression in the prefrontal cortex of subjects with bipolar disorder. Neuroreport 1999;10:1747–50. [6] Caberlotto L, Hurd YL. Neuropeptide Y Y(1) and Y(2) receptor mRNA expression in the prefrontal cortex of psychiatric subjects. Relationship of Y(2) subtype to suicidal behavior. Neuropsychopharmacology 2001;25:91–7. [7] Cai JX, Arnsten AF. Dose-dependent effects of the dopamine D1 receptor agonists A77636 or SKF81297 on spatial working memory in aged monkeys. J Pharmacol Exp Ther 1997;283:183–9. [8] Cardinal RN, Robbins TW, Everitt BJ. The effects of d-amphetamine, chlordiazepoxide, alpha-flupenthixol and behavioural manipulations on choice of signalled and unsignalled delayed reinforcement in rats. Psychopharmacology (Berl) 2000;152:362–75. [9] Carvalho-Netto EF, Nunes-de-Souza RL. Use of the elevated T-maze to study anxiety in mice. Behav Brain Res 2004;148:119–32.

[10] Ceglia I, Carli M, Baviera M, Renoldi G, Calcagno E, Invernizzi RW. The 5-HT receptor antagonist M100, 907 prevents extracellular glutamate rising in response to NMDA receptor blockade in the mPFC. J Neurochem 2004;91:189–99. [11] Chudasama Y, Passetti F, Rhodes SE, Lopian D, Desai A, Robbins TW. Dissociable aspects of performance on the 5-choice serial reaction time task following lesions of the dorsal anterior cingulate, infralimbic and orbitofrontal cortex in the rat: differential effects on selectivity, impulsivity and compulsivity. Behav Brain Res 2003;146:105–19. [12] Cole BJ, Hillmann M. Effects of benzodiazepine receptor ligands on the performance of an operant delayed matching to position task in rats: opposite effects of FG 7142 and lorazepam. Psychopharmacology (Berl) 1994;115:350–7. [13] Cole BJ, Robbins TW. Amphetamine impairs the discriminative performance of rats with dorsal noradrenergic bundle lesions on a 5choice serial reaction time task: new evidence for central dopaminergicnoradrenergic interactions. Psychopharmacology (Berl) 1987;91:458–66. [14] Cole BJ, Robbins TW. Effects of 6-hydroxydopamine lesions of the nucleus accumbens septi on performance of a 5-choice serial reaction time task in rats: implications for theories of selective attention and arousal. Behav Brain Res 1989;33:165–79. [15] Cole SO. Diazepam-induced impairment of a go-no go successive discrimination. Behav Neural Biol 1990;53:371–7. [16] Cole SO, Michaleski A. Dose-dependent impairment in the performance of a go-no go successive discrimination by chlordiazepoxide. Psychopharmacology (Berl) 1986;88:184–6. [17] Colmers WF, Bleakman D. Effects of neuropeptide Y on the electrical properties of neurons. Trends Neurosci 1994;17:373–9. [18] Contarino A, Gold LH. Targeted mutations of the corticotropin-releasing factor system: effects on physiology and behavior. Neuropeptides 2002;36:103–16. [19] Coull JT, Middleton HC, Robbins TW, Sahakian BJ. Clonidine and diazepam have differential effects on tests of attention and learning. Psychopharmacology (Berl) 1995;120:322–32. [20] Coull JT, Middleton HC, Robbins TW, Sahakian BJ. Contrasting effects of clonidine and diazepam on tests of working memory and planning. Psychopharmacology (Berl) 1995;120:311–21. [21] Curran H, Psychological approaches to human memory. In Gazzaniga, M, editor. The cognitive neurosciences. MIT Press, Cambridge; 1999, p. 797–804. [22] Davis M. The role of the amygdala in conditional fear. In: Aggleton JP, editor. The amygdala: neurobiological aspects of emotion, memory, and mental dysfunction. New York: Wiley; 1992. p. 255–306. [23] Deakin JB, Aitken MR, Dowson JH, Robbins TW, Sahakian BJ. Diazepam produces disinhibitory cognitive effects in male volunteers. Psychopharmacology (Berl) 2004;173:88–97. [24] Dere E, Topic B, De Souza Silva MA, Srejic M, Frisch C, Buddenberg T, et al. The graded anxiety test: a novel test of murine unconditioned anxiety based on the principles of the elevated plus-maze and light-dark test. J Neurosci Methods 2002;122:65–73. [25] Dudchenko P, Paul B, Sarter M. Dissociation between the effects of benzodiazepine receptor agonists on behavioral vigilance and responsitivity. Psychopharmacology (Berl) 1992;109:203–11. [26] Dudchenko P, Sarter M. Behavioral microanalysis of spatial delayed alternation performance: rehearsal through overt behavior, and effects of scopolamine and chlordiazepoxide. Psychopharmacology (Berl) 1992;107:263–70. [27] Dumont Y, Fournier A, St-Pierre S, Quirion R. Autoradiographic distribution of [125I]Leu31,Pro34]PYY and [125I]PYY3-36 binding sites in the rat brain evaluated with two newly developed Y1 and Y2 receptor radioligands. Synapse 1996;22:139–58. [28] Easterbrook J. The effect of emotion on cue utilization and the organization of behaviour. Psychol Rev 1959;66:183–201. [29] Eysenck MW. Attention and arousal. Berlin: Springer-Verlag; 1982. p. 47–54. [30] Flood JF, Baker ML, Hernandez EN, Morley JE. Modulation of memory processing by neuropeptide Y varies with brain injection site. Brain Res 1989;503:73–82.

B. Greco, M. Carli / Behavioural Brain Research 169 (2006) 325–334 [31] Gackenheimer SL, Schober DA, Gehlert DR. Characterization of neuropeptide Y Y1-like and Y2-like receptor subtypes in the mouse brain. Peptides 2001;22:335–41. [32] Gallagher M, Holland PC. The amygdala complex: multiple roles in associative learning and attention. Proc Natl Acad Sci USA 1994;91:11771–6. [33] Gray JA. The neuropsychology of anxiety. New York: Oxford University Press; 1982. [34] Greco B, Invernizzi RW, Carli M. Phencyclidine-induced impairment in attention and response control depends on the background genotype of mice: reversal by the mGLU(2/3) receptor agonist LY379268. Psychopharmacology (Berl) 2005;179:68–76. [35] Gries DA, Condouris GA, Shey Z, Houpt M. Anxiolytic-like action in mice treated with nitrous oxide and oral triazolam or diazepam. Life Sci 2005;76:1667–74. [36] Gustafson EL, Smith KE, Durkin MM, Walker MW, Gerald C, Weinshank R, et al. Distribution of the neuropeptide Y Y2 receptor mRNA in rat central nervous system. Brain Res Mol Brain Res 1997;46:223–35. [37] Hascoet M, Bourin M. A new approach to the light/dark test procedure in mice. Pharmacol Biochem Behav 1998;60:645–53. [38] Heilig M, McLeod S, Brot M, Heinrichs SC, Menzaghi F, Koob GF, et al. Anxiolytic-like action of neuropeptide Y: mediation by Y1 receptors in amygdala, and dissociation from food intake effects. Neuropsychopharmacology 1993;8:357–63. [39] Heilig M, Soderpalm B, Engel JA, Widerlov E. Centrally administered neuropeptide Y (NPY) produces anxiolytic-like effects in animal anxiety models. Psychopharmacology (Berl) 1989;98:524–9. [40] Heilig M, Widerlov E, Neuropeptide. Y: an overview of central distribution, functional aspects, and possible involvement in neuropsychiatric illnesses. Acta Psychiatr Scand 1990;82:95–114. [41] Higgins GA, Enderlin M, Haman M, Fletcher PJ. The 5-HT2A receptor antagonist M100,907 attenuates motor and ‘impulsive-type’ behaviours produced by NMDA receptor antagonism. Psychopharmacology (Berl) 2003;170:309–19. [42] Holland PC, Han JS, Gallagher M. Lesions of the amygdala central nucleus alter performance on a selective attention task. J Neurosci 2000;20:6701–6. [43] Holley LA, Turchi J, Apple C, Sarter M. Dissociation between the attentional effects of infusions of a benzodiazepine receptor agonist and an inverse agonist into the basal forebrain. Psychopharmacology (Berl) 1995;120:99–108. [44] Humby T, Laird FM, Davies W, Wilkinson LS. Visuospatial attentional functioning in mice: interactions between cholinergic manipulations and genotype. Eur J Neurosci 1999;11:2813–23. [45] Isles AR, Humby T, Walters E, Wilkinson LS. Common genetic effects on variation in impulsivity and activity in mice. J Neurosci 2004;24:6733–40. [46] Kask A, Rago L, Harro J. Anxiogenic-like effect of the neuropeptide Y Y1 receptor antagonist BIBP3226: antagonism with diazepam. Eur J Pharmacol 1996;317:R3–4. [47] Kirkby DL, Higgins GA. Characterization of perforant path lesions in rodent models of memory and attention. Eur J Neurosci 1998;10:823– 38. [48] Krazem A, Borde N, Beracochea D. Effects of diazepam and beta-CCM on working memory in mice: relationships with emotional reactivity. Pharmacol Biochem Behav 2001;68:235–44. [49] Le Pen G, Grottick AJ, Higgins GA, Moreau JL. Phencyclidine exacerbates attentional deficits in a neurodevelopmental rat model of schizophrenia. Neuropsychopharmacology 2003;28:1799–809. [50] Lemelin S, Baruch P, Vincent A, Everett J, Vincent P. Distractibility and processing resource deficit in major depression. Evidence for two deficient attentional processing models. J Nerv Ment Dis 1997;185:542–8. [51] Marston HM, Spratt C, Kelly JS. Phenotyping complex behaviours: assessment of circadian control and 5-choice serial reaction learning in the mouse. Behav Brain Res 2001;125:189–93. [52] Mathe AA, Jimenez PA, Theodorsson E, Stenfors C. Neuropeptide Y, neurokinin A and neurotensin in brain regions of Fawn Hooded “depressed”, Wistar, and Sprague Dawley rats. Effects of electroconvul-

[53]

[54]

[55]

[56]

[57] [58]

[59]

[60]

[61]

[62]

[63]

[64]

[65]

[66]

[67]

[68]

[69]

[70]

[71]

[72]

[73]

333

sive stimuli. Prog Neuropsychopharmacol Biol Psychiatry 1998;22: 529–46. McGaughy J, Sarter M. Behavioral vigilance in rats: task validation and effects of age, amphetamine, and benzodiazepine receptor ligands. Psychopharmacology (Berl) 1995;117:340–57. McGaughy J, Turchi J, Sarter M. Crossmodal divided attention in rats: effects of chlordiazepoxide and scopolamine. Psychopharmacology (Berl) 1994;115:213–20. McNaughton N, Morris RG. Chlordiazepoxide an anxiolytic benzodiazepine, impairs place navigation in rats. Behav Brain Res 1987;24:39–46. Menzaghi F, Howard RL, Heinrichs SC, Vale W, Rivier J, Koob GF. Characterization of a novel and potent corticotropin-releasing factor antagonist in rats. J Pharmacol Exp Ther 1994;269:564–72. Mialet JP, Pope HG, Yurgelun-Todd D. Impaired attention in depressive states: a non-specific deficit? Psychol Med 1996;26:1009–20. Michel MC, Beck-Sickinger A, Cox H, Doods HN, Herzog H, Larhammar D, et al. XVI. International Union of Pharmacology recommendations for the nomenclature of neuropeptide Y, peptide YY, and pancreatic polypeptide receptors. Pharmacol Rev 1998;50:143–50. Mirjana C, Baviera M, Invernizzi RW, Balducci C. The serotonin 5HT2A receptors antagonist M100907 prevents impairment in attentional performance by NMDA receptor blockade in the rat prefrontal cortex. Neuropsychopharmacology 2004;29:1637–47. Moghaddam B, Adams B, Verma A, Daly D. Activation of glutamatergic neurotransmission by ketamine: a novel step in the pathway from NMDA receptor blockade to dopaminergic and cognitive disruptions associated with the prefrontal cortex. J Neurosci 1997;17:2921–7. Moghaddam B, Adams BW. Reversal of phencyclidine effects by a group II metabotropic glutamate receptor agonist in rats. Science 1998;281:1349–52. Murphy BL, Roth RH, Arnsten AF. Clozapine reverses the spatial working memory deficits induced by FG 7142 in monkeys. Neuropsychopharmacology 1997;16:433–7. Nakajima M, Inui A, Asakawa A, Momose K, Ueno N, Teranishi A, et al. Neuropeptide Y produces anxiety via Y2-type receptors. Peptides 1998;19:359–63. Nakamura M, Carney JM. Antagonism by CGS 8216 and Ro 15-1788, benzodiazepines antagonists, of the action of chlordiazepoxide on a timing behavior in rats. Pharmacol Biochem Behav 1984;21:381–5. Nilsson C, Karlsson G, Blennow K, Heilig M, Ekman R. Differences in the neuropeptide Y-like immunoreactivity of the plasma and platelets of human volunteers and depressed patients. Peptides 1996;17: 359–62. Noteboom JT, Barnholt KR, Enoka RM. Activation of the arousal response and impairment of performance increase with anxiety and stressor intensity. J Appl Physiol 2001;91:2093–101. Ohl F, Roedel A, Binder E, Holsboer F. Impact of high and low anxiety on cognitive performance in a modified hole board test in C57BL/6 and DBA/2 mice. Eur J Neurosci 2003;17:128–36. Ohl F, Sillaber I, Binder E, Keck ME, Holsboer F. Differential analysis of behavior and diazepam-induced alterations in C57BL/6N and BALB/c mice using the modified hole board test. J Psychiatr Res 2001;35:147–54. Qian J, Colmers WF, Saggau P. Inhibition of synaptic transmission by neuropeptide Y in rat hippocampal area CA1: modulation of presynaptic Ca2+ entry. J Neurosci 1997;17:8169–77. Redrobe JP, Dumont Y, Herzog H, Quirion R. Neuropeptide Y (NPY) Y2 receptors mediate behaviour in two animal models of anxiety: evidence from Y2 receptor knockout mice. Behav Brain Res 2003;141:251–5. Redrobe JP, Dumont Y, Quirion R. Neuropeptide Y (NPY) and depression: from animal studies to the human condition. Life Sci 2002;71:2921–37. Rief W, Hermanutz M. Responses to activation and rest in patients with panic disorder and major depression. Br J Clin Psychol 1996;35(Pt 4):605–16. Rizk A, Curley J, Robertson J, Raber J. Anxiety and cognition in histamine H3 receptor−/− mice. Eur J Neurosci 2004;19:1992–6.

334

B. Greco, M. Carli / Behavioural Brain Research 169 (2006) 325–334

[74] Robbins TW. A critique of the methods available for the measurements of spontaneous motor activity. In: Iversen LL, Iversen SD, Snyder SH, editors. Principles of behavioural pharmacology, 7. New York: Plenum Press; 1977. p. 37–82. [75] Robbins TW. The 5-choice serial reaction time task: behavioural pharmacology and functional neurochemistry. Psychopharmacology (Berl) 2002;163:362–80. [76] Rodgers RJ, Cole JC, Aboualfa K, Stephenson LH. Ethopharmacological analysis of the effects of putative ‘anxiogenic’ agents in the mouse elevated plus-maze. Pharmacol Biochem Behav 1995;52:805–13. [77] Sainsbury A, Schwarzer C, Couzens M, Fetissov S, Furtinger S, Jenkins A, et al. Important role of hypothalamic Y2 receptors in body weight regulation revealed in conditional knockout mice. Proc Natl Acad Sci USA 2002;99:8938–43. [78] Sajdyk TJ, Schober DA, Smiley DL, Gehlert DR. Neuropeptide Y-Y2 receptors mediate anxiety in the amygdala. Pharmacol Biochem Behav 2002;71:419–23. [79] Scoville WB, Milner B. Loss of recent memory after bilateral hippocampal lesions, 1957. J Neuropsychiatry Clin Neurosci 2000;12:103–13. [80] Seamans JK, Floresco SB, Phillips AG. D1 receptor modulation of hippocampal-prefrontal cortical circuits integrating spatial memory with executive functions in the rat. J Neurosci 1998;18:1613–21. [81] Shimizu H, Bray GA. Effects of neuropeptide Y on norepinephrine and serotonin metabolism in rat hypothalamus in vivo. Brain Res Bull 1989;22:945–50. [82] Stanley BG, Leibowitz SF. Neuropeptide Y injected in the paraventricular hypothalamus: a powerful stimulant of feeding behavior. Proc Natl Acad Sci USA 1985;82:3940–3. [83] Stephens DN, Sarter M. Bidirectional nature of benzodiazepine receptor ligands extends to effects on vigilance. Psychopharmacol Ser 1988;6:205–17. [84] Stogner KA, Holmes PV. Neuropeptide-Y exerts antidepressant-like effects in the forced swim test in rats. Eur J Pharmacol 2000;387:R9–10. [85] Sun QQ, Akk G, Huguenard JR, Prince DA. Differential regulation of GABA release and neuronal excitability mediated by neuropeptide Y1 and Y2 receptors in rat thalamic neurons. J Physiol 2001;531:81–94. [86] Takeda H, Tsuji M, Matsumiya T. Changes in head-dipping behavior in the hole-board test reflect the anxiogenic and/or anxiolytic state in mice. Eur J Pharmacol 1998;350:21–9.

[87] Tatemoto K, Carlquist M, Mutt V. Neuropeptide Y—a novel brain peptide with structural similarities to peptide YY and pancreatic polypeptide. Nature 1982;296:659–60. [88] Thiebot MH, Le Bihan C, Soubrie P, Simon P. Benzodiazepines reduce the tolerance to reward delay in rats. Psychopharmacology (Berl) 1985;86:147–52. [89] Tschenett A, Singewald N, Carli M, Balducci C, Salchner P, Vezzani A, et al. Reduced anxiety and improved stress coping ability in mice lacking NPY-Y2 receptors. Eur J Neurosci 2003;18: 143–8. [90] van Gaalen MM, Stenzel-Poore M, Holsboer F, Steckler T. Reduced attention in mice overproducing corticotropin-releasing hormone. Behav Brain Res 2003;142:69–79. [91] Venault P, Chapouthier G, de Carvalho LP, Simiand J, Morre M, Dodd RH, et al. Benzodiazepine impairs and beta-carboline enhances performance in learning and memory tasks. Nature 1986;321: 864–6. [92] Vezzani A, Sperk G, Colmers WF. Neuropeptide Y: emerging evidence for a functional role in seizure modulation. Trends Neurosci 1999;22:25–30. [93] Weiner ED, Mallat AM, Papolos DF, Lachman HM. Acute lithium treatment enhances neuropeptide Y gene expression in rat hippocampus. Brain Res Mol Brain Res 1992;12:209–14. [94] Widdowson PS, Ordway GA, Halaris AE. Reduced neuropeptide Y concentrations in suicide brain. J Neurochem 1992;59:73–80. [95] Widerlov E, Lindstrom LH, Wahlestedt C, Ekman R. Neuropeptide Y and peptide YY as possible cerebrospinal fluid markers for major depression and schizophrenia, respectively. J Psychiatr Res 1988;22: 69–79. [96] Williams RA, Hagerty BM, Cimprich B, Therrien B, Bay E, Oe H. Changes in directed attention and short-term memory in depression. J Psychiatr Res 2000;34:227–38. [97] Winer B. Statistical principles in experimental design. 2nd ed. New York: McGraw-Hill; 1971. [98] Yerkes RM, Dodson JD. The relation of strength of stimulus to rapidity of habit-formation. J Comp Neurol Psychol 1908;18:459–82. [99] Zahrt J, Taylor JR, Mathew RG, Arnsten AF. Supranormal stimulation of D1 dopamine receptors in the rodent prefrontal cortex impairs spatial working memory performance. J Neurosci 1997;17:8528–35.