Age-related decline of various cognitive functions in well-experienced male rats treated with the putative anti-aging compound (2R)-1-(1-benzofuran-2-yl)-N-propylpentane-2-amine ((-)BPAP).

Aging-associated cognitive disorders lack proper medication. To meet this need translation-wise, modification of the animal models is also required. In the present study, effect of the putative anti-aging compound (2R)-1-(1-benzofuran-2-yl)-N-propylpentane-2-amine ((-)BPAP, a deprenyl derivative) on age-related cognitive decline was investigated in experienced, aged Long-Evans rats. During their lifetime, animals had acquired knowledge in various cognitive assays. Their performance in these tests was then parallel followed from the age of 27 months until their death meanwhile half of them were treated with BPAP. Cognitive performance in various tasks showed different sensitivities/resistances to age-related impairment. Pot jumping performance (motor skill-learning) started to impair first, at 21 months of age, followed by decreasing performance in five-choice serial reaction time task (attention) at 26 months. Navigation performance in Morris water maze (spatial learning) started to decline at 31 months. Performance in a cooperation task (social cognition) started to decline the latest, at 34 months. Our findings suggest that in this process, the primary factor was the level of motivation to be engaged with the task and not losing the acquired knowledge. The average lifespan of the tested rat population was 36 months. BPAP could not improve the cognitive performance; neither could it prolong lifespan. A possible reason might be that dietary restriction and lifelong cognitive engagement had beneficial effects on cognitive capabilities and lifespan creating a "ceiling effect" for further improvement. The results confirmed that experienced animals provide a translationally relevant model to study age-related cognitive decline and measure the effect of putative anti-aging compounds.

Lack of general learning ability factor in a rat test battery measuring a wide spectrum of cognitive domains.

Objective: In the framework of a larger project aiming to test putative cognitive enhancer drugs in a system with improved translational validity, we established a rodent test battery, where different, clinically relevant cognitive domains were investigated in the same animal population. The aim of the current study was to check whether performances in the different tasks representing different cognitive functions are assay-specific or may originate in an underlying general learning ability factor. Methods: In the experiments 36 Long-Evans and 36 Lister Hooded rats were used. The test battery covered the following cognitive domains: attention and impulsivity (measured in the 5-choice serial reaction time task), spatial memory (Morris water-maze), social cognition (cooperation task), cognitive flexibility (attentional set shifting test), recognition memory (novel object recognition) and episodic memory (water-maze based assay). The outcome variables were analyzed by correlation analysis and principal component analysis (PCA). The datasets consisted of variables measuring learning speed and performance in the paradigms. From the raw variables composite variables were created for each assay, then from these variables a composite score was calculated describing the overall performance of each individual in the test battery. Results: Correlations were only found among the raw variables characterizing the same assay but not among variables belonging to different tests or among the composite variables. The PCAs did not reduce the dimensionality of the raw or composite datasets. Graphical analysis showed variable performance of the animals in the applied tests. Conclusions: The results suggests the assay outcomes (learning performance) in the system are based on independent cognitive domains.

Cognitive profiling and proteomic analysis of the modafinil analogue S-CE-123 in experienced aged rats.

The lack of novel cognitive enhancer drugs in the clinic highlights the prediction problems of animal assays. The objective of the current study was to test a putative cognitive enhancer in a rodent cognitive test system with improved translational validity and clinical predictivity. Cognitive profiling was complemented with post mortem proteomic analysis. Twenty-seven male Lister Hooded rats (26 months old) having learned several cognitive tasks were subchronically treated with S-CE-123 (CE-123) in a randomized blind experiment. Rats were sacrificed after the last behavioural procedure and plasma and brains were collected. A label-free quantification approach was used to characterize proteomic changes in the synaptosomal fraction of the prefrontal cortex. CE-123 markedly enhanced motivation which resulted in superior performance in a new-to-learn operant discrimination task and in a cooperation assay of social cognition, and mildly increased impulsivity. The compound did not affect attention, spatial and motor learning. Proteomic quantification revealed 182 protein groups significantly different between treatment groups containing several proteins associated with aging and neurodegeneration. Bioinformatic analysis showed the most relevant clusters delineating synaptic vesicle recycling, synapse organisation and antioxidant activity. The cognitive profile of CE-123 mapped by the test system resembles that of modafinil in the clinic showing the translational validity of the test system. The findings of modulated synaptic systems are paralleling behavioral results and are in line with previous evidence for the role of altered synaptosomal protein groups in mechanisms of cognitive function.

Procognitive profiling of a serotonin 5-HT6 receptor antagonist in a complex model system in rats: A novel translational approach for clinical prediction.

INTRODUCTION: The serial clinical failures of novel cognitive enhancer candidates point out the lack of predictive power in the preceding animal experimentation. For a more predictive profiling of putative procognitive drugs in rodents, we recently elaborated a methodical approach which consists of three fundamental steps: 1. teaching various learning tasks representing different cognitive domains to the same cohort of animals with the aim to create a population with 'widespread knowledge'. 2. Applying a cognitive deficit-inducing intervention to transform this cohort of animals to a 'patient population'. 3. Testing putative procognitive drugs with a 'clinical trial-like' design on the wide spectrum of cognitive (dys)functions in the actual 'patient population'. The present study has been the first trial to test the feasibility and utility of the proposed system. METHODS: The population with 'widespread knowledge' consisted of 2 year old male Long-Evans rats with a learning history in five-choice serial reaction time task (5-CSRTT, attentional paradigm), Morris water maze (MWM, spatial learning), a cooperative task carried out in pairs (social learning), and a skill-learning task, "pot-jumping". For inducing cognitive deficit, thus creating a 'patient population' we increased the difficulty of the tasks. For the cognitive enhancer mechanism to test in the system we chose a serotonin 5-HT6 receptor antagonist compound, RO4368554. Animals were randomly assigned to vehicle- and drug treated groups based on their baseline learning performance and their response in a pilot test of increase in task difficulty. During the 13-day long treatment with 3 mg/kg ip. RO4368554 all the learning paradigms were repeatedly run with increased difficulty supplemented with a novel object recognition test (NOR, episodic memory). RESULTS: In the 5-CSRTT, reducing the stimulus duration from 1 s to 0.25 s caused a significant decrease in the percentage of correct responses (from 52 % to 31 % in the control group) which was not affected by the 5-HT6 receptor antagonist treatment (correct responses decreased from 58 % to 31 %). In the MWM, replacing the escape platform to a new location did not mean a hard challenge for the rats. Members of both groups could find it within a relatively short time: mean escape latencies were 83 s and 65 s at the first replacement trial and 58 s and 74 s at the second one in the control and drug-treated groups, respectively. In the cooperation paradigm, where the rats had to perform simultaneous nose-pokes to get a reward, task difficulty was increased by requiring two consecutive simultaneous nose-poking from the animals. This caused a fall in the percentage of successful trials in both groups (from 48 % to 12 % and from 50 % to 20 % in the saline - and drug-treated group, respectively), however, by the end of the treatment RO4368554-treated animals showed significantly higher performance (29 %) than saline treated rats (2%). The NOR test, carried out with a 5 -h delay, revealed poor recognition memory in both groups (discrimination index (DI) values were 0.13 and 0.06 for saline and RO4368554, respectively). Performance in the pot jumping test was also not improved by the drug-treatment. CONCLUSIONS: The applied study design allowed parallel measurements of the action of the test compound on several cognitive functions and to follow its time course. RO4368554 did not show notable effects on impaired attention and visual recognition; nor did it affect spatial and procedural learning, but it exerted beneficial effect on cooperative behaviour. The revealed activity pattern highlight the cognitive domain most sensitive to the particular drug effect and may give hints for further target validating and clinical studies.

Following of aging process in a new motor skill learning model, "pot jumping" in rats.

Impairment of procedural memory is a frequent and severe symptom in many neurological and psychiatric diseases as well as during aging. Our aim was to establish an assay in rats in which procedural learning and changes in performance can be studied on the long term. The work was done in the frame of a larger project aiming to establish a complex cognitive animal test battery of high translational value. The equipment was a 190-cm-diameter circular water tank where 12 flower pots were placed upside down in a circle with increasing distances (18-46 cm) between the adjacent ones. Male Lister Hooded and Long-Evans rats were allowed to move on the pots for 3 min. The arena was filled with shallow water to make the rats stay on the pots. Animals were obviously motivated to move around on the pots; however, the distance which required jumping (> 26 cm) meant a barrier for some of them. Development of motor skill was measured by the longest distance successfully spanned. A relatively flat bell-shaped age dependence was observed, with a peak at 13 months of age. A gradual decline in performance could be observed after the age of 20 months which preceded the appearance of overt physical weakness. Long-Evans rats showed more homogeneous performance and higher individual stability than Lister Hooded rats. The method is appropriate to study the development of motor learning and to follow its age-dependent changes. It may also serve as an assay for testing potential drugs for improving motor skills and/or procedural memory.

Establishment of a rodent cooperation assay as a model of social cognition.

INTRODUCTION: Impaired cooperative skills form a characteristic symptom in autism, which lacks adequate treatment. The objective of this study was to establish a rat cooperation assay which fits the feasibility and capacity requirements of drug development. METHODS: Long-Evans and Lister Hooded rats were trained in pairs to simultaneously perform nose-pokes (within 1 s) for reward in a Skinner box equipped with two nose-poke modules. Conditioning took place first with naive-naive pairs, then with naive-experienced and finally with experienced-experienced pairs, when the task was familiar for both rats. In a control experiment, experienced Lister-hooded pairs were tested under the learnt schedule but without the possibility to communicate with each other. RESULTS: Rats were able to learn the task in 8-15 sessions. Experienced-experienced Long-Evans pairs completed the training significantly faster than the other pairs Analysis of the nose-poke latency data, sample video-recordings and the significantly decreased performance of rats in the control experiment suggested that the animals solved the task via real cooperation. DISCUSSION: The newly developed rat cooperation model is quick and has sufficiently high throughput, therefore it may be used in the drug development of putative social cognitive enhancer compounds.

Translational Difficulties in Querying Rats on "Orientation".

The aim of this study was to translate the "orientation" query of the ADAS-Cog inventory to rats and to investigate whether they can determine which time of the day they are. For this purpose, we established a modified Morris water-maze navigation task where the escape platform was placed onto various locations at different times of the day: "morning", "noon" and "evening". In each of these sessions rats swam a "query" trial and a "confirmatory" trial, 30 min apart. Lister Hooded rats randomly chose among the three possible target locations, while Long Evans rats partly followed a win-stay strategy by preferring to visit first to the platform position of the previous session. Despite simplifying the task to a morning-evening discrimination, Lister Hooded rats continued searching by chance, while Long Evans rats switched to the mentally less demanding random strategy. We then inserted a board into the pool which required longer swimming path from the animals when they were correcting an initial wrong choice, but this modification did not result in a change in the above strategies. Lastly, in a separate group of Long-Evans rats, the training conditions were modified inasmuch an incorrect choice was definitely punished by impeding the animals to correct it and confining them to a platform-free part of the maze for the whole trial period. However, even these stricter conditions were not sufficient to make the rats distinguish times of the day. The observed lack of time discrimination may source from an evolutionary built in mechanism characteristic for the rat species or this ability may have only been lost in laboratory rats.

Selegiline reduces adiposity induced by high-fat, high-sucrose diet in male rats.

BACKGROUND AND PURPOSE: Incidence and severity of obesity are increasing worldwide, however, efficient and safe pharmacological treatments are not yet available. Certain MAO inhibitors reduce body weight, although their effects on metabolic parameters have not been investigated. Here, we have assessed effects of a widely used, selective MAO-B inhibitor, selegiline, on metabolic parameters in a rat model of diet-induced obesity. EXPERIMENTAL APPROACH: Male Long-Evans rats were given control (CON) or a high-fat (20%), high-sucrose (15%) diet (HFS) for 25 weeks. From week 16, animals were injected s.c. with 0.25 mg·kg-1 selegiline (CON + S and HFS + S) or vehicle (CON, HFS) once daily. Whole body, subcutaneous and visceral fat was measured by CT, and glucose and insulin tolerance were tested. Expression of glucose transporters and chemokines was assessed by quantitative RT-PCR. KEY RESULTS: Selegiline decreased whole body fat, subcutaneous- and visceral adiposity, measured by CT and epididymal fat weight in the HFS group, compared with HFS placebo animals, without influencing body weight. Oral glucose tolerance and insulin tolerance tests showed impaired glucose homeostasis in HFS and HFS + S groups, although insulin levels in plasma and pancreas were unchanged. HFS induced expression of Srebp-1c, Glut1 and Ccl3 in adipose tissue, which were alleviated by selegiline. CONCLUSIONS AND IMPLICATIONS: Selegiline reduced adiposity, changes in adipose tissue energy metabolism and adipose inflammation induced by HFS diet without affecting the increased body weight, impairment of glucose homeostasis, or behaviour. These results suggest that selegiline could mitigate harmful effects of visceral adiposity.

Pharmacological comparison of traditional and non-traditional cannabinoid receptor 1 blockers in rodent models in vivo.

Cannabinoid receptor 1 (CB1R) antagonists have been proven to be effective anti-obesity drugs; however, psychiatric side effects have halted their pharmaceutical development worldwide. Despite the emergence of next generation CB1R blockers, a preclinical head to head comparison of the anti-obesity and psychiatric side effect profiles of the key compounds has not been performed. Here, we compared classical CB1R antagonists (rimonabant, taranabant, otenabant, ibipinabant, and surinabant) and non-traditional CB1R blockers (the partial agonist O-1269, the neutral antagonists VCHSR and LH-21 and the peripherally acting inverse agonist JD-5037) using an in vivo screening cascade. First, the potencies of these compounds to reduce CB1R agonist-induced hypothermia and decrease fasting-induced food intake were determined. Then, equipotent doses of the non-toxic compounds were compared in a diet-induced obesity (DIO) test, which includes measurements of metabolic syndrome markers. Psychiatric side effects were assessed by measuring anxiogenicity in an ultrasonic vocalization test. All classical CB1R blockers were centrally acting appetite suppressants and decreased body weight and food intake in an obesity-dependent manner, with only slight effects on metabolic syndrome markers. In addition, all classical CB1R blockers increased ultrasonic vocalization. Surprisingly, none of the non-classical CB1R blockers was eligible for the DIO comparison and side effect profiling. O-1269 and LH-21 induced convulsive behavior, whereas VCHSR and JD-5037 were devoid of any in vivo activity. The classical CB1R blockers displayed similar therapeutic and side effect profiles in vivo, whereas the available non-traditional CB1R blockers were not appropriate tools for testing the therapeutic potential of alternative CB1R inhibitors.

Effect of a kynurenic acid analog on home-cage activity and body temperature in rats.

BACKGROUND: N-(2-N,N-Dimethylaminoethyl)-4-oxo-1H-quinoline-2-carboxamide hydrochloride (SzR-72) is a kynurenic acid (KYNA) amide analog that displays neuroprotective action. Whereas its brain penetration ability and its solubility limit the therapeutic use of KYNA: the corresponding properties of the analog exceed those of the parent compound. Although SzR-72 has been extensively studied, its exact mechanism of action has not yet been fully clarified. As KYNA induces hypothermia in laboratory rodents, it may be hypothesized that SzR-72 may have a similar effect. This would be of major importance, since the hypothermia generated by external cooling is neuroprotective, thus a putative hypothermic effect of SzR-72 could contribute to its neuroprotective action. METHODS: The effects of SzR-72 on the body temperature and home-cage activity of rats were studied by using a telemetry system. In order to follow the longitudinal changes in the effects of the compound, subchronic drug administration was applied. RESULTS: The initial administration of the compound induced substantial hypothermia and reduced the home-cage activity. During the 5 days of SzR-72 administration, partial tolerance developed to the hypothermic effect, while the inhibition of home-cage activity detected after the acute administration was completely tolerated. CONCLUSIONS: On the basis of these results, it cannot be excluded that the hypothermic effect of SzR-72 contributes to its neuroprotective action.

α7 nicotinic acetylcholine receptors and their role in cognition.

The precise role of nicotinic acetylcholine receptors (nAChRs) in central cognitive processes still remains incompletely understood almost 150 years after its initial discovery. Central nAChRs are activated by acetylcholine, which functions in the extracellular space as a nonsynaptic messenger. Recently, a novel concept in the nAChR mode of operation has been described as a fast-type nonsynaptic transmission. In this review, we attempt to summarise the experimental findings that support the role of one of the most distributed receptor subtypes, the α7 nAChRs, and particularly focus on its procognitive effects following receptor activation. The basic characteristics of α7 nAChRs are discussed, from receptor homology to cellular-level functions. Synaptic plasticity is often implicated with α7 nAChRs on the basis of several diverse studies. Here, we provide a summary of the plastic features of the α7 receptor subtype and its role in higher level cognitive function. Finally, recent clinical evidence is reviewed, which demonstrates with increasing confidence the promise α7 nAChRs as a molecular target in future pharmacotherapy to prevent cognitive decline in various types of dementia, specifically, via the development of positive allosteric modulator compounds.

Interactions of CB1 and mGlu5 receptor antagonists in food intake, anxiety and memory models in rats.

CB(1) receptor antagonists proved to be effective anti-obesity drugs, however, their depressive and anxiogenic effects became also evident. Finding solution to overcome these psychiatric side effects is still in focus of research. Based on the available clinical and preclinical results we hypothesized that the combination of CB(1) and mGlu(5) receptor antagonisms may result in a pharmacological intervention, where the anxiolytic mGlu(5) receptor inhibition may counteract the anxiogenic psychiatric side effects of CB(1) antagonism, while CB(1) antagonism may ameliorate the memory impairing effect of mGlu(5) receptor antagonism. Further, the two components will synergistically interact in blocking food-intake and reducing obesity. For testing the interaction of mGlu(5) and CB(1) receptor antagonism MTEP [3-[(2-methyl-1,3-thiazol-4-yl)ethynyl]pridine; SIB-1757, 6-methyl-2-(phenylazo)-3-pyridinol)] (mGlu(5) antagonist) and rimonabant [(5-(4-Chlorophenyl)-1-(2,4-dichloro-phenyl)-4-methyl-N-(piperidin-1-yl)-1H-pyrazole-3-carboxamide)hydrochloride] (CB(1) antagonist) were used. All experiments were carried out in rats. Effects of the compounds on anxiety were tested in two foot shock induced ultrasonic vocalization paradigms, appetite suppression was assessed in the food intake test, while memory effects were tested in a context conditioned ultrasonic vocalization setup. MTEP abolished the anxiogenic effect of rimonabant, while there was an additive cooperation in suppressing appetite. However, rimonabant did not ameliorate the memory impairing effect of MTEP. By combination of CB(1) and mGluR5 antagonism, anxiety related side effects might be attenuated, appetite suppression maintained, nevertheless, the possible emergence of unwanted memory impairments can overshadow its therapeutic success.

Effect of 5-HT5A antagonists in animal models of schizophrenia, anxiety and depression.

The few available data on the pharmacological effect of 5-HT5A receptors suggest that antagonists may have anxiolytic, antidepressant and antipsychotic activity. The aim of our study was to verify these suggestions in relevant animal models. Two 5-HT5A antagonist ligands, SB-699551-A (N-[2-(dimethylamino)ethyl]-N-[[4'-[[(2-phenylethyl)amino]methyl][1,1'-biphenyl]-4-yl]methyl]cyclopentanepropanamide dihydrochloride) (3-60 mg/kg, intraperitoneally) and A-843277 (N-(2,6-dimethoxybenzyl)-N'[4-(4-fluorophenyl)thiazol-2-yl]guanidine) (3-30 mg/kg, intraperitoneally), were examined in the open-field test, in a foot-shock-induced ultrasonic vocalization test, in the forced swim test (FST) and in the amphetamine-induced and phencyclidine-induced hyperlocomotion tests to examine their effect on general behavioural patterns, and their anxiolytic-like, antidepressant-like and antipsychotic-like properties, respectively. In the open-field test, SB-699551-A induced sedation and A-843277 induced writhing. In the ultrasonic vocalization test, SB-699551-A reduced vocalizations, whereas A-843277 was ineffective. In the FST, SB-699551-A was ineffective and A-843277 reduced immobility, but only at the highest dose. In the amphetamine-induced and phencyclidine-induced hyperlocomotion test, both compounds were ineffective. SB-699551-A showed an anxiolytic-like property in the ultrasonic vocalization test; however, this compound has a sedative effect. A-843277 showed an antidepressant-like property in the FST, but its immobility-reducing effect may also be a consequence of abdominal irritation. Consequently, further investigations are required to define the therapeutic potential of 5-HT5A receptor ligands in anxiety, depression and schizophrenia models.

Shock priming enhances the efficacy of SSRIs in the foot shock-induced ultrasonic vocalization test.

Data on the effect of acutely administered serotonin reuptake inhibitors (SSRIs) in animal anxiety models have been inconsistent. In some of the models these compounds showed anxiolytic properties, while in others they were ineffective or even anxiogenic. In the foot shock-induced ultrasonic vocalization (USV) test in the adult rat, SSRIs were reported to be effective, however, they were only tested with protocols using multiple shocking design. In the present study, anxiolytic effects of various SSRI compounds (sertraline, fluoxetine, paroxetine, escitalopram) were tested in three distinct USV test protocols in comparison with alprazolam and 8-OH-DPAT. In the single shocking protocol, animals were exposed to one shocking session after the drug treatment. In the multiple shocking protocol, rats went through a foot shock priming session before each drug test. On priming days animals received foot shocks without drug treatment. On the test day (the day after), rats received drug treatment and then were shocked again. In the context conditioning protocol animals were exposed to foot shocks on two consecutive days before the drug test. On the third, test day, after drug treatment animals were replaced to the shocking chamber, but this time shocks were not delivered. SSRIs were ineffective using the single shocking protocol. In the context conditioned protocol, all SSRIs showed linear dose-response relationship with ED50 values of 8.5, 2.2, 0.77 and 0.93 mg/kg i.p. for fluoxetine, sertraline, paroxetine and escitalopram, respectively. Using the multiple shocking protocol, SSRIs were only partially effective with maximum inhibitions ranging between 44% and 62%. In contrast to SSRIs, the benzodiazepine anxiolytics, alprazolam showed anxiolytic activity with linear dose-response relationship in all of the test protocols, with ED50 values varying from 1.3 to 4.0 mg/kg i.p. The serotonin 5HT1A receptor antagonist 8-OH-DPAT also showed linear dose-response relationship in all protocols, but this compound was less potent in the single shocking design (ED50 values were 0.27, 0.04 and 0.07 mg/kg i.p. in the single shocking, multiple shocking and context conditioned protocol, respectively). In conclusion, our results show that priming has a major impact on the effectiveness of SSRIs in the USV test, and the three test protocols applied in this study have different predictive and face validity.