Research report
The T-maze continuous alternation task for assessing the effects of putative cognition enhancers in the mouse

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Abstract

The T-maze continuous alternation task (T-CAT) assesses the spatial exploratory performance in mice. We performed a series of four experiments in order to establish the T-CAT in mice in our laboratory, to replicate published findings, and to investigate the effects of scopolamine and donepezil.

In the first experiment, the task was found to be sensitive to differences between mouse strains, corroborating findings reported by Gerlai. HsdWin:CFW1 mice alternated below chance level, C57BL/6JIco and B6D2F1/JIco mice performed above chance level, and C57BL/6NTac and 129S6/SvEvTac mice performed at chance level.

In the second experiment, donepezil (Aricept, E2020) at the dose of 3 mg/kg p.o. increased the rate of alternations above the level of the vehicle-treated control group in C57BL/6JIco mice, suggesting that this drug can act as cognition enhancer in normal animals. 1 mg/kg scopolamine, administered intraperiteoneally (i.p.), impaired the spontaneous alternation behaviour of the mice. The slightly lower dose of 0.75 mg/kg did not affect alternation performance. The high dose of donepezil (3 mg/kg) was able to antagonise the scopolamine-induced performance deficit. With respect to time to complete a session, the results were inconclusive.

In the third experiment, we found that scopolamine, administered i.p. at the dose of 1 mg/kg, or subcutaneously (s.c.) at the dose of 0.1 mg/kg, decreased the rate of spontaneous alternations in C57BL mice in the T-CAT and increased the time to complete a session. Most likely due to adverse side effects induced by the dose of 1 mg/kg scopolamine, 4 out of 10 animals did not complete at least eight free-choice trials during the maximum session duration of 30 min. No such adverse effects were seen after 0.1 mg/kg scopolamine, administered s.c. Finally, we evaluated whether the T-CAT yields replicable results. We conclude that the T-CAT provides a reliable tool for assessing the effects of cognition-modulating treatments in mice.

Introduction

Spontaneous alternation is a measure of exploratory behaviour, most often evaluated in rodents [27]. Gerlai [11], [13] developed a T-maze continuous alternation task (T-CAT) to assess the spatial exploratory performance in mice. Spontaneous alternation is defined as a visit to the other of the two goal arms of a T-maze than that visited in the previous trial. In order to alternate or avoid a revisit, a mouse must remember the goal arm chosen in the previous trial. The information about the previous visit is stored in the spatial working memory [21]. The T-CAT might be suited to assess the effects of putative cognition enhancing and cognition impairing experimental manipulations.

Major reason for the modification of the usual T-maze alternation task, in which a mouse is put back into the start arm by the experimenter after it had entered one of the two goal arms, was to avoid handling during a session. In a series of experiments, Gerlai showed that this task appears to depend upon normal functioning of the hippocampus, is guided by extra-maze cues, and is sensitive to strain differences [11].

Acetylcholine (ACh) has been examined for its role in spontaneous alternation, with drugs that decrease synaptic transmission. The alternation rate is decreased with scopolamine administration to rats [7], [16] and mice [1]. Scopolamine is a parasympatholytic drug that acts as muscarinic cholinoceptor antagonist and disrupts cognitive processes. An impairment in spontaneous alternation is also observed with the administration of pirenzepine, an M1 muscarinic cholinergic receptor antagonist [28]. Contrary to the effects of cholinergic receptor antagonists, physostigmine, a cholinesterase inhibitor (ChEI), increased spontaneous alternation rates [7].

We performed a series of experiments in order to establish the T-CAT in mice in our laboratory, to replicate findings by Gerlai, and to investigate the effects of scopolamine and donepezil.

In a first experiment, we studied the differences between the inbred C57BL/6NTac and 129S6/SvEvTac (according to the revised nomenclature; [8]) mouse strains, the hybrid B6D2F1/Tac mice and an inbred C57BL strain from a different supplier than Taconic, namely C57BL/6JIco from IFFA CREDO.

In experiment 2, the effects of donepezil (Aricept®, E2020) a ChEI that is used for treating patients suffering from mild to moderate Alzheimer’s disease (AD) [26], was tested in normal mice and in mice that had been given scopolamine. AD, a chronic neurodegenerative disorder, is characterised by a progressive decline in cognitive function, including loss of memory and cognitive function and behaviour. Central cholinergic hypofunction is thought to be a component of the disease [2]. Several lines of evidence support the notion that a loss of cholinergic function contributes to the cognitive deficits seen in AD [3], and consequently, an initial therapeutic approach consisted of enhancing cholinergic functions pharmacologically. The most successful of these strategies at this time is to increase the amount of ACh by reducing its degradation by ChEIs, such as donepezil [20], [26]. Kirby et al. [10] and Ogura et al. [20], for example, previously had found that donepezil is able to antagonise scopolamine-induced deficits in short-term or working memory performance in rats.

In experiment 3, scopolamine again was used to induce cognitive impairments. 1 mg/kg scopolamine, administered intraperiteoneally (i.p.) has been reported to disrupt the spontaneous alternation learning of mice [29]. We assessed the effects of different doses and routes of application (i.p. versus s.c.) of scopolamine on the performance of mice in the T-CAT.

Finally, we explored whether the T-CAT procedure yields reproducible results. We did this by comparing the spontaneous alternation rates of the untreated and vehicle-treated groups of C57BL/6JIco mice across different experiments.

Section snippets

Apparatus

The T-maze was constructed by Sembach (Ratingen, FRG) according to the measures provided by Gerlai [11]. The apparatus was made of transparent Plexiglas. The guillotine doors and the floor of the apparatus were black. The guillotine doors could be operated by the experimenter through a system of pulley strings (see Fig. 1, left panel).

Methods

Training of a mouse consisted of one single session, which started with 1 forced-choice trial, followed by 14 free-choice trials (see Fig. 1, right panel).

Forced-choice trial

In the

Animals

Ten male outbred HsdWin:CWF1 mice were supplied by Harlan Winkelmann, Borchen, Germany. Ten inbred C57BL/6JIco male mice were obtained from IFFA CREDO, l’Arbresle, France. Eight inbred C57BL/6NTac, eight hybrid B6D2F1/Tac (C57BL/6 female×DBA/2 male), and eight inbred 129S6/SvEvTac mice were supplied by Taconic, Germantown, NY, USA. All animals were approximately 10 weeks old at the beginning of the experiment. Their weights ranged from 20 to 25 g. All mice per strain were housed together in one

Animals

Sixty male C57BL/6JIco mice, supplied by IFFA CREDO, France, were used. They were approximately 12 weeks old at the beginning of the experiment. Their weights ranged from 25 to 30 g. The mice were housed in groups of 8–10 in standard Makrolon™ type III cages in which food (rat–mouse chow, Ssniff) and water was available ad libitum. Lights in the laboratory were on between 7.00 a.m. and 7.00 p.m. The mice were housed in the laboratory where the T-maze apparatus was situated.

Procedure

The animals were

Experiment 3: effects of intraperitoneal and subcutaneous injections of scopolamine on the performance of male C57BL mice in a T-maze continuous alternation task

Though scopolamine, administered i.p. at the dose of 1 mg/kg body weight, appeared to induce reliable performance deficits in the previous experiment, the drug also appeared to increase the time needed to complete a session consisting of 1 forced-choice and 14 free-choice trials. Moreover, a number of scopolamine-treated mice did complete less than eight free-choice trials during the 30-min session. Their data were excluded from statistical analyses, because the compound apparently suppressed

Replicability of results

The replicability of results was assessed by plotting the means and standard errors of the means of the control groups from seven different drug-evaluation studies and from the strain comparison study (experiment 1) of the present report (see Fig. 6). In all experiments, C57BL/6JIco mice were used. The mean percent spontaneous alternations across experiments ranged between 62 and 76% and these spontaneous alternation rates reliably deviated from chance level, i.e. 50% alternation, as assessed

Discussion

The experiments corroborate findings by Gerlai that mice are able to alternate above chance levels in the T-CAT. The rate of spontaneous alternations, however, depends upon the genotype tested. The spontaneous alternation was disrupted by 1 mg/kg, but not by 0.75 mg/kg scopolamine, administered i.p., and by 0.1 mg/kg scopolamine, administered s.c., 30 min before a spontaneous alternation session started.

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