Optimization and pharmacological validation of a set-shifting procedure for assessing executive function in rats.

BACKGROUND: Set-shifting tests represent a reliable paradigm to assess executive functions in humans and animals. In the rat, set-shifting in a cross-maze is a recognized method. In this test, rats must learn an egocentric rule to locate food reinforcement. Once acquired, a second rule, based on visual-cue strategy, allows the location of the food. Ability of rats to shift from the first to the second rule is considered to reflect cognitive flexibility. NEW METHOD: This study aimed at optimizing the most currently used set-shifting protocol in a cross-maze for standardized drug testing by modulating the parameters related to caloric restriction, reward preference, and by redefining the notion of turn bias and classification of errors sub-types, i.e. perseverative vs. regressive. The new protocol has then been used to assess rats treated by sub-chronic phencyclidine administration and investigate the potential reversal effect of tolcapone, a brain penetrant catechol-O-methyl transferase inhibitor. RESULTS: The new procedure resulted in a decreased total duration and a re-definition of turn bias and error subtypes. Despite preferences for sweet rewards, caloric restriction had to be maintained to motivate animals. Overall, sub-chronic PCP-treated rats made mostly perseverative errors compared to controls and required more trials to shift between the two rules. Tolcapone partly reversed impairments observed in PCP-treated rats. CONCLUSION: The new protocol has improved the reliability of key parameters and has contributed to the decrease of the test duration. PCP-treated rats submitted to this protocol have been shown to have significant deficits that could be reversed by tolcapone.

Peripheral administration of an anti-TNF-α receptor fusion protein counteracts the amyloid induced elevation of hippocampal TNF-α levels and memory deficits in mice.

Alzheimer's disease has long been associated with increased inflammation in the brain. Activated microglia and increased production of the inflammatory cytokines such as TNF-α, have been proposed to contribute to the onset and progression of the disease. We investigated if systemic administration of an anti-tumor necrosis factor (TNF) biologic medication clinically validated for rheumatoid arthritis (RA), TNF receptor 2 fused to a Fc domain (TNFR2:Fc), could ameliorate the behavioral symptoms and decrease neuroinflammation in a non-transgenic mouse model mimicking some hallmarks of the disease. Seven days after a single intracebroventricular (icv) injection of aggregated amyloid beta25-35 (9nmoles), mice displayed significant cognitive deficit in spontaneous alternation (working memory) and inhibitory avoidance (long-term memory) tasks. Alternation percentage decreased from 72.4%±1.3 to chance level (52.6%±1.7); step-through retention latency decreased from 247s to 144s. Subcutaneous administration of 30mg/kg TNFR2:Fc every second day post amyloid beta25-35 icv administration counteracted the amyloid-induced decrease in alternation percentage (66.4s±1.8) and the decreased step-through retention latency (248s±9). Measurement of hippocampal TNF-α levels by ELISA after behavioral assessment showed significant elevation in animals injected with amyloid beta25-35 relative to animals injected with control peptide. In animals treated with 30mg/kg TNFR2:Fc, TNF-α levels in the hippocampus were reduced and were similar to control animals. These data suggest that peripheral administration of TNFR2:Fc counteracts amyloid-induced memory impairment and normalizes increased TNF-α levels in hippocampus of a non-transgenic mouse model of amyloid induced cognitive deficit.

Evaluation of symptomatic drug effects in Alzheimer's disease: strategies for prediction of efficacy in humans.

In chronic diseases such as Alzheimer's disease (AD), the arsenal of biomarkers available to determine the effectiveness of symptomatic treatment is very limited. Interpretation of the results provided in literature is cumbersome and it becomes difficult to predict their standardization to a larger patient population. Indeed, cognitive assessment alone does not appear to have sufficient predictive value of drug efficacy in early clinical development of AD treatment. In recent years, research has contributed to the emergence of new tools to assess brain activity relying on innovative technologies of imaging and electrophysiology. However, the relevance of the use of these newer markers in treatment response assessment is waiting for validation. This review shows how the early clinical assessment of symptomatic drugs could benefit from the inclusion of suitable pharmacodynamic markers. This review also emphasizes the importance of re-evaluating a step-by-step strategy in drug development.

Effects of dietary resveratrol on the sleep-wake cycle in the non-human primate gray mouse lemur (Microcebus murinus).

Converging evidence shows that the non-human primate gray mouse lemur (Microcebus murinus) is ideal for the study of the aging process and for testing the effects of new therapies and dietary interventions on age-associated pathologies. One such dietary supplement is resveratrol (RSV), a dietary polyphenolic compound with several positive effects on metabolic functions and longevity. However, little is known about the effect of RSV on the lemur sleep-wake cycle, which reflects mammalian brain function and health. In the present study, the authors investigated this effect by comparing sleep-wake cycles in adult lemurs based on electroencephalographic (EEG) rhythms. The effect of short-term RSV supplementation on the sleep-wake cycle of mouse lemurs was evaluated in entrained conditions (long-day photoperiods, light:dark 14:10). After 3 wks of RSV supplementation, the animals exhibited a significantly increased proportion of active-wake time, occurring mainly during the resting phase of the sleep-wake cycle (+163%). The increase in active-wake time with RSV supplementation was accompanied by a significant reduction of both paradoxical sleep (-95%) and slow-wave sleep (-38%). These changes mainly occurred during the resting phase of the sleep-wake cycle (RSV supplementation induced negligible changes in active-wake time during the active phase of the sleep-wake cycle). The present data suggest that RSV may be a potent regulator of sleep-wake rhythms and could be of major interest in the study of sleep perturbations associated with aging and neuropathology.

Brivaracetam does not alter spatial learning and memory in both normal and amygdala-kindled rats.

Several antiepileptic drugs (AEDs) may induce memory deficits when tested in preclinical models at doses that exert significant protection against seizures. Brivaracetam (BRV) is a novel high-affinity SV2A ligand also displaying inhibitory activity at neuronal voltage-gated sodium channels. In the present study we have investigated the effects of BRV, at doses that exerted marked anticonvulsant effects in kindled rats, upon cognitive functioning and memory in both normal and amygdala-kindled rats using place learning version of Morris water maze. In addition the effect of BRV on long-term potentiation (LTP) in rat hippocampal slices has been investigated. BRV (2.1, 6.8 or 21.0mg/kg i.p.) was injected daily, 60min before each session. Results indicated that in both normal and amygdala-kindled rats BRV did not alter the latency to find the hidden platform or swimming speed during the four consecutive days of learning. Similarly, the time spent in the target quadrant, used as a further independent index of spatial memory, was not modified by BRV treatment. Likewise, BRV did not affect the LTP induction in CA1 hippocampal region when tested at 3-30microM concentration range, which had been demonstrated to significantly reduce epileptiform activity in slice models. Based on the results of the present study it can be expected that BRV will not have detrimental effects on hippocampal-dependent cognitive functions in patients with epilepsy.

Absence of negative impact of levetiracetam on cognitive function and memory in normal and amygdala-kindled rats.

The effect of the new antiepileptic drug (AED) levetiracetam (LEV, Keppra) on cognitive function was studied in normal and amygdala-kindled rats by using the Morris water maze test. In addition, we investigated the effect of LEV on long-term potentiation (LTP) in rat hippocampal slices. Sodium valproate (VPA) was used as comparator in all studies. Clonazepam (CZP) and carbamazepine (CBZ) were used in normal rats. The results indicated that doses of LEV known to suppress motor seizures did not alter cognitive performance. In contrast, similar doses of the classic AEDs all decreased learning performance of the rats. Likewise, VPA did alter LTP but LEV was inactive. Amygdala-kindled rats were more sensitive than normal rats to the effects of VPA. These results suggest that LEV may be devoid of negative impact on cognition in epileptic patients.

The mirror chamber test for testing anxiolytics: is there a mirror-induced stimulation?

A new murine model of anxiety, namely the mirror chamber test, is based on the assumption that, like many species, mice show approach-avoidance behavior when they are confronted by a mirror. It has been suggested that the mirror chamber is a specific and a quantitatively/qualitatively different measure of anxiety than that implicated in other behavioral models such as the elevated plus-maze and the head-dipping assays. The aim of the present study was to investigate whether there was indeed a specific mirror effect by replacing the mirrors in the chamber by either white or dark-gray tiles. Balb/c, DBA-derived, and C57 BL/6j mice were tested under these three experimental conditions. The results indicated that Balb/c and DBA-derived mice avoided the mirror, white and gray chambers similarly while C57 BL/6j mice entered more readily and spent more time in the gray chamber than in the mirrored or white chambers. Thus, depending on the strain of mouse studied, a brightness or a position effect in the chamber could explain the avoidance behavior observed. These results suggest that there is no need to invoke a specific mirror effect.

Arm width and brightness modulation of spontaneous behaviour of two strains of mice tested in the elevated plus-maze.

Amongst the nonpharmacological variables that may influence spontaneous behaviour in the elevated plus-maze, at least two (namely, the width and brightness level of the maze arms) have not been evaluated for possible consequences on baseline activity. We therefore investigated the effect of width (5, 7, or 9 cm) and brightness level (grey vs. white) of the maze arms on the activity patterns of NMRI and C57/BL 10j mice in a plus-maze based on that described by Lister. Results indicated a clear strain difference in spontaneous behaviour in the open/closed arms with C57/BL mice making no visits on the open arms of the maze. In the NMRI strain, enlarging the arms resulted in significantly increased running in open arms but brightness level did not affect behavioural parameters. In the C57/BL mice, no significant behavioural changes were detected after increasing the arm width; however, the maze with a white floor significantly increased the latency to move and decreased ambulation. Finally, employing a maze with white closed arms and open grey arms resulted in ambulation by C57/BL mice into the open arms, a behaviour not normally observed in this strain using a uniformly bright maze. The results are discussed in terms of their putative effects in pharmacological testing.

The aged mouse as a model of cognitive decline with special emphasis on studies in NMRI mice.

The use of the aged mouse as an integrated model of age-related cognitive decline is reviewed, with special emphasis on experiments covering the life span of NMRI mice, using different age-groups ranging from 3 through to 22 months. Age-related changes in the sensorimotor profile, spontaneous behaviour and performance in learning and memory tasks are considered. The data provide evidence for cognitive impairment and decreases in spontaneous activity and exploration from middle age onwards. Chronologically, this age depends on the longevity of the strain selected; in NMRI mice, middle age corresponds to 11-12 months. Complex learning tasks, such as the Morris water maze for spatial learning, appear to be the most sensitive to age-related changes, as are tests requiring prolonged retention of acquired information, for example, using passive avoidance. Cued and simple discrimination learning are only impaired in the oldest animals. Age-related changes in non-cognitive variables, including sensorimotor capacity, pain sensitivity, emotionality, or locomotor activity, do not account for the learning impairments, although deficits in visual acuity cannot be excluded in the very old animals. Detailed analysis of the individual data for middle aged and old mice, using discriminant and correlation studies highlight a marked heterogeneity between animals of any given chronological age. Furthermore, individual aged mice do not exhibit similar degrees of impairment across all the behavioural variables, showing that aging is not a uniform process. The possible relationship between age-related behavioural decline and neurochemical changes is an area as yet unexplored apart from a few isolated investigations, including a study on ChAT and AChE in NMRI mice. The studies in the NMRI mice illustrate the value of investigating the full age-range to detect an age group which shows cognitive decline dissociable from physical or emotional changes and which is representative of the population as a whole.

Spatial processing and emotionality in aged NMRI mice: a multivariate analysis.

The goal of this study was to investigate possible relationships between different aspects of behavioural decline in aged (17 months) NMRI mice by a multivariate analysis. These mice presented defects in spatial processing both in place learning in a Morris-type water maze and in spontaneous alternation in a Y-maze, as well as showing changes in their behaviour in the elevated plus maze test of fear/anxiety and open-field ambulation. Data were first factor analysed and then correlated on the basis of variable factor scores. The results showed clearly that age-related behavioural changes were independent of each other except between open-field activity measured over 6 min and spatial learning indices in the Morris-type water maze, in that mice performing poorly in the latter were more active in the open field, and between the ratio open/total arms visited in the elevated plus maze task and locomotor activity during the first 2 min of the open-field test. It is proposed that the notion of an allocentric or locale system cannot be applied unitarily to both spatial learning and spontaneous alternation defects, and that activity levels in the aged mice are task dependent and reflect different underlying factors.

Behavioural, biochemical and histological effects of AF64A following injection into the third ventricle of the mouse.

Behavioural, biochemical and histological effects were assessed following AF64A injected into the third ventricle of female NMRI mice. Doses from 3 to 7 nmol produced significant changes in behaviour, causing hyperactivity, reduced hole-board exploration, rotational behaviour in a symmetrical Y-maze corresponding to a loss of alternation, abnormal behaviour in a plus-maze task of fear/anxiety with markedly increased exploration of the open arms and finally deficits in passive avoidance responding and spatial orientation in a Morris-type water maze. In this latter test, a cue learning deficit was noted for the two highest doses only. No histological changes of consequence were observed up to 5 nmol. Beyond this dose, at 6 and particularly 7 nmol, necrosis of parts of the hippocampus and septum was apparent. ChAT and AChE activity were decreased in the hippocampus but not in the cortex although the decreases were smaller than generally reported for AF64A-treated rats. ChAT and AChE reductions correlated highly with hyperactivity in the open-field and to a lesser extent, with spatial learning deficits. Monoaminergic activity was also affected in the hippocampus, but not in the cortex, at 4 nmol and above. NE and particularly 5-HT and 5-HIAA levels were reduced although the rate of 5-HT turnover was unaltered. A highly significant correlation was obtained between 5-HT effects and the increased open arm exploration in the plus-maze task of fear/anxiety. The behavioural effects and biochemical changes lasted at least 8-9 weeks postop.

Age-related changes in spontaneous behavior and learning in NMRI mice from middle to old age.

Spontaneous behavior, sensorimotor reflexes and learning of 3-, 11-, 17- and 22-month-old virgin female NMRI mice were compared. Sensorimotor abilities decreased significantly from the age of 17 months in proportion to the muscular and equilibrium demand of the test. Open-field activity, hole board exploration as well as activity in the Y maze and plus maze decreased from the age of 11 months. However, in the open-field and Y maze, it was not possible to distinguish between 11-, 17- and 22-month-old mice, whereas in the plus maze, activity was drastically decreased in 17- and 22-month-old mice. In the plus maze, indices of fear-motivated behavior suggest a greater sensitivity to the situation in the age groups of 17 and 22 months. Spontaneous alternation also decreased from the age of 11 months, but at the age of 22 months, a rotational behavior emerged. Spatial learning was markedly impaired from middle age, i.e., 11 months, as indicated by longer latencies and absence of spatial bias in place learning. In cued learning, the 11- and 17-month groups eventually reached the level of the 3-month-old mice, while the 22-month age group remained impaired. Changes in swimming ability or speed did not appear to account for the swimming maze deficits. In contrast, a sensory or perceptual deficit cannot be excluded for the oldest mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Simplifying environmental cues in a Morris-type water maze improves place learning in old NMRI mice.

Old virgin female NMRI mice aged 17 months were compared with mice aged 3 months for their spatial learning abilities in two versions of the Morris water maze. The first one was a simplified version with a salient configuration of cues comparable to a black/white discrimination and the second one was the classical version of the Morris test with many distal cues surrounding the maze. In the simplified version, old mice presented a slower rate of acquisition and a transient poorer retention compared to young mice. However, old mice achieved a final level of performance statistically comparable to their young counterparts as assessed by latencies to escape onto the concealed platform and by the spatial bias measured in probe trials at intervals during testing. When subsequently subjected to classical Morris maze learning, the same old animals showed marked learning deficits and were persistently impaired in their latencies to escape onto the platform. They presented no spatial bias for the location of the platform in the different probe trials. When the goal was cued at the end of the experiment, the performances of old mice rapidly improved, showing that motivation, motor disabilities, or fatigue and ability to use proximal cues cannot explain the place learning deficit. Our results were discussed in terms of cognitive versus sensory/perceptual disabilities in aged rats and mice.

Cholinergic modulation of spatial learning in mice in a Morris-type water maze.

Injection of the centrally active muscarinic antagonist scopolamine i.p. 20 min pre-test at 3 mg/kg but not at 1 mg/kg, impaired spatial learning of a Morris-type water maze adapted for mice. Both doses caused hyperactivity. D-amphetamine (3 mg/kg, i.p.), which also caused hyperactivity, did not impair spatial learning nor did methylscopolamine (3 mg/kg, i.p.). In a cued version of the water maze, apart from a temporary disturbance on day 1, scopolamine (3 mg/kg) and control groups behaved similarly, indicating that scopolamine-induced place learning deficits are not due to changes in swimming ability, motivation or ability to use proximal cues. Physostigmine (0.1 and 0.2 mg/kg, i.p.) and oxotremorine (0.02 mg/kg but not 0.01 mg/kg, i.p.) antagonized the deficits in the swimming maze. Neither drug affected the scopolamine hyperactivity despite causing hypoactivity per se. In contrast, the peripherally acting cholinergic drug neostigmine was inactive against scopolamine in either test at 0.1 mg/kg. THA (2-8 mg/kg, i.p.), RS86 (0.25-1 mg/kg, i.p.) and nicotine (1 and 3 mg/kg, i.p.) were also unable to antagonize the scopolamine effect. These studies show that scopolamine disrupts acquisition of spatial rather than cued learning in mice in a Morris-type water maze and that this effect appears to be mediated centrally and can be dissociated from drug-induced hyperactivity. Moreover, this deficit can be reversed with certain cholinergic agents.

Age-related changes in spontaneous behavior and learning in NMRI mice from maturity to middle age.

Spontaneous behavior and learning and memory of 3-, 6-, 9- and 12-month-old virgin female NMRI mice were compared. Open field activity and spontaneous alternation in a Y-maze decreased in an age-related manner, reaching a statistical level of significance for the groups aged 9 and 12 months. Spatial learning was highly impaired in 9-and 12-month-old mice in the place version of a Morris-type water maze but not in the cued version of this task. Changes in motor activity, swimming ability or speed did not appear to account for these deficits. In a one-trial passive avoidance, performance was more variable, although a deficit in the oldest age group was clearly evident using a cutoff time of 120 sec. The passive avoidance was not attributable to reduced shock sensitivity. Together, these results suggest that the onset of aging in NMRI mice occurs at the age of 9 and particularly 12 months. NMRI mice of this age could, therefore, represent a viable animal model for the study of cognitive impairments in aging.

Difference in scopolamine sensitivity during ageing in rats: dichotomy between behavioural repertoire and learning performances.

The effects of different doses of scopolamine in adult and aged rats were compared in two behavioural experiments. The first experiment involved visual reversal learning, while the second experiment investigated individual and social behavioural patterns. Results showed that the same dose of scopolamine was more effective in impairing the reverse discrimination in aged than in adult rats. In contrast, low doses which were effective in modifying behavioural patterns in adults, were inactive in aged rats. This difference in scopolamine sensitivity (supersensitivity versus hyposensitivity), observed between the two types of behaviour, is discussed in relation to age-related functional receptor adaptation, selective changes in muscarinic receptors in the different brain areas and dopaminergic- cholinergic interactions.

Investigation into sex-related differences in locomotor activity, place learning and passive avoidance responding in NMRI mice.

Learning abilities of young adult (5-6 weeks) male and female NMRI mice were compared in two tasks involving cognitive functions, namely spatial learning in a water maze and passive avoidance responding. Locomotor activity was also monitored as a putative bias for the results obtained in these learning tests. No sex-related difference was observed either in avoidance responding or in spatial learning using a procedure with the same point of departure throughout testing in the water maze. However, in this test, using a procedure with 3 different points of departure in randomized order day after day, female mice performed better than male mice. The difference was statistically significant on the last acquisition day but was not sustained during a retention trial 72 hr later. Since no sex differences in locomotor activity were observed, the learning effects cannot be attributed to a difference in general activity level.

Scopolamine effects on juvenile conspecific recognition in rats: Possible interaction with olfactory sensitivity.

The social recognition of a juvenile conspecific by an adult male rat was evaluated as the decrease in investigation time when the same juvenile individual was reintroduced 30min after the first exposure period. The results showed that scopolamine impaired this transient individual recognition. A drop in investigation time was also observed in both tests (first and second exposure), with the same and with different juvenile individuals, in scopolamine treated animals. A second experiment showed that scopolamine disturbed the chemosensory preference for familiar odour observed in the control group. In the light of these two experiments, and according to the chemosensory mediation of social recognition in the rat, it is impossible to rule out a lack of odour discrimination in the absence of social recognition after scopolamine injection.

Scopolamine disrupts visual reversal without affecting the first discrimination.

The effect of scopolamine (0.5 mg/kg) was determined in a brightness discrimination test (Y maze) motivated by electrical shocks (escape avoidance). Male adult Sprague Dawley rats were used. Results show that scopolamine impairs significantly the visual reversal without affecting the first brightness discrimination. The qualitative analysis reveals that the anticholinergic drug-induced deficit involves both perseveration, i.e. failure to suppress inappropriate response, and a tendency to adopt a position habit. A parallelism with hippocampal and frontal lobe damage symptoms is discussed and an interpretation in terms of disinhibition and incapacity to solve a more difficult problem is proposed.