Attention deficit hyperactivity disorder (ADHD) is the most common disorder in children and adolescents (Posner et al., 2020; Shrestha et al., 2020). ADHD leads not only to poor attention but also to a decrease in motivated behavior and learning abilities (Volkow et al., 2011). In adulthood, certain ADHD symptoms persisted in up to 50% of cases. The patients are less hyperactive but have an impairment of working memory and selectivity of attention (Asherson et al., 2016; Luo et al., 2019). There are several approaches to ADHD diagnoses and treatment, but precise neurobiological mechanisms underlying ADHD pathology are insufficiently investigated so far. Generally, it is believed that ADHD is connected with the dopamine (DA) system dysfunction (Viggiano et al., 2004; Cinque et al., 2018). Beyond ADHD, DA dysregulation may contribute to numerous neurological and psychiatric disorders, such as Parkinson disease, Huntington’s disease, and schizophrenia (Russell et al., 2005; Tripp and Wickens, 2012; Ko et al., 2013; Ng et al., 2015; Grace, 2016; Creed and Goldberg, 2018; Konnova and Swanberg, 2018; Natsheh and Shiflett, 2018; Adinolfi et al., 2019). It is not surprising since the DA system is involved in the control of cognition, locomotion, reward evaluation, and formation of memories for reward–cue associations (Wise, 2004; Keiflin and Janak, 2015; Morrens et al., 2020).

It has been suggested that the development of ADHD and associated behavioral disorders might be caused by abnormalities in the functioning of the plasma membrane DA transporter (DAT). A significant association between ADHD and the gene encoding the DAT was demonstrated (Mill et al., 2005; Yang et al., 2007), and altered DAT expression in the striatum of patients with ADHD symptoms was shown (Madras et al., 2005). Furthermore, it is well known that patients with ADHD react favorably to therapy with Ritalin (methylphenidate) and Adderall (amphetamine), which target DAT (Dresel et al., 2000).

Dopamine transporter, by controlling re-uptake of released DA, plays a critical role in the regulation of both the intraneuronal and extracellular DA homeostasis (Vaughan and Foster, 2013). Mice lacking the DAT (DAT-KO mice) were shown to have high extracellular DA levels, which results in spontaneous hyperlocomotion and certain cognitive dysfunctions (Giros et al., 1996; Spielewoy et al., 2000; Gainetdinov et al., 2002; Efimova et al., 2016; Ide et al., 2018). In mutant mice, long-term alterations in DA signaling lead to profound disturbances in the cortico-striatal connections and functions (Surmeier et al., 2007). In general, numerous observations indicate that the DA system plays an important role in the consolidation of memory traces (Moncada and Viola, 2007; Beeler et al., 2010; Pezze and Bast, 2012; Duszkiewicz et al., 2019). Investigations in knockout mice allowed researchers to clarify many questions concerning the cognitive and behavioral processes that are critical for modeling neuropsychiatric disorders. The recent opportunity to translate these studies to transgenic rats significantly increased the capacity of such investigations. Rats are known to have a wider repertoire of behavioral reactions and can learn complex cognitive tasks (Abbott, 2004); thus, they are a more convenient model for investigating the cognitive functions.

The strain of rats with deletion of DAT gene (DAT-KO rats) was developed (Leo et al., 2018a) to investigate various aspects of DA system dysfunctions with a particular emphasis on cognitive disorders (Vengeliene et al., 2017; Leo et al., 2018b; Sukhanov et al., 2019). DAT-KO rats were described to have increased locomotor activity, an impaired sensorimotor gating, a deficit in operant nose-poke responding reinforced by food pellets, and decreased Y-maze spontaneous alternation. Furthermore, they have significant dysregulation in the fronto-striatal brain-derived neurotrophic factor function (Adinolfi et al., 2018; Cinque et al., 2018; Leo et al., 2018b). Recently, the impairment of spatial working memory was also demonstrated in DAT-KO rats. It was shown that they can learn the behavioral task in an eight-arm radial maze but perform it much less effectively than the control wild-type (WT) rats (Kurzina et al., 2020). Independently developed DAT deficient rats were also hyperactive, failed to show conditioned fear responses, were unable to learn stimulus–reward associations, and showed anhedonia and impaired cognition and social behaviors (Vengeliene et al., 2017; Sukhanov et al., 2019).

Novelty is a factor of the new environment that allows evaluation of the adaptation processes. Recognition memory is connected with the discrimination of novel information. In experiments in rodents, this type of memory is necessary to recognize previously encountered events or objects using combined information about stimuli and the environmental context where animals have encountered the objects including information about the location of the animal (Ergorul and Eichenbaum, 2004; Aggleton and Brown, 2006). Novel stimuli are known to excite DA neurons, which are involved in the exploratory activity in a novel environment (Costa et al., 2014). DAT-KO rats can recognize a novel testing chamber slightly different from the previous one (Adinolfi et al., 2018). Novel object recognition was found to depend on D2 DA receptor activity (França et al., 2016). In DAT knockdown mice, the impairment of novel object recognition was observed (Chang et al., 2020).

There are some new models to study long-term memory (Genzel et al., 2019) and new approaches for investigating memory consolidation (Hardt and Nadel, 2018) in normal rats, but DAT-KO rats were not evaluated yet in these tasks. So far, long-term memory in DAT-KO animals has not been investigated. Since memory improvement in ADHD patients was shown in experiments with novel stimulus presentation (Baumann et al., 2020), this approach is attracting attention to overcome ADHD problems without any medical treatment. The present study focuses on the evaluation of the ability of DAT-KO rats to learn the paradigm of object recognition with rewarded familiar objects and non-rewarded novel objects and store it in long-term memory.

Here, we present the data obtained in the experiments with DAT-KO rats that display behavioral abnormalities relevant to ADHD endophenotypes. These rats have elevated extracellular DA levels and spontaneous hyperactivity. We investigated the influence of the task paradigm on the learning and storage in the memory motor skills by DAT-KO rats. Despite the difficulties with task acquisition, they have demonstrated an ability to learn a novel object recognition task and store the learned paradigm in memory for 3 months.

Hyperactivity of DAT-KO rats and mice and perseverative pattern of motor activity was demonstrated in numerous investigations and are described as their distinctive traits (Pogorelov et al., 2005; Vengeliene et al., 2017; Adinolfi et al., 2018; Cinque et al., 2018; Ide et al., 2018; Leo et al., 2018b). DAT-KO rats spent much longer time and covered longer distances to perform a task, which might be due to their hyperactivity. DAT-KO rats also spent a longer time to reach the first target to be moved. It might be suggested that their object recognition abilities are inferior and not as good as those of WT rats. This fact might be also explained by their hyperactivity and perseverations. The inability of DAT-KO rats to immediately focus their attention on the objects presented may be also related to their attention deficit described earlier (Sukhanov et al., 2019).

In our previous study, we found that DAT-KO rats can learn the spatial cognitive task in the eight-arm radial maze, but the task fulfillment was significantly worse than in control rats. In the spatial task, DAT-KO rats also spent significantly more time and covered longer distances during training than WT rats. DAT-KO rats also demonstrated an increased perseverative pattern during the spatial task. Analysis of repeated entry sequences showed that DAT-KO rats had a sustainable statistically significant tendency to increase the arms revisits (Kurzina et al., 2020). Highly perseverative patterns of locomotor activity during eight-arm radial maze task were previously reported in DAT-KO mice (Gainetdinov et al., 2002; Efimova et al., 2016).

The recognition memory processes, underlying the ability to choose between the stimuli that have occurred previously and the novel ones, are fundamental for recording events and controlling the target of perspective behaviors. Recognition memory is based on various types of information that allows establishing whether a stimulus has been previously encountered and how it might affect subsequent behaviors (Kinnavane et al., 2015). In a place recognition test with a long (24 h) delay period, a 20-min familiarization was sufficient for rats to discriminate between novel and familiar objects (Ozawa et al., 2011). In associative recognition memory tests, rats would make the association between objects, places, and contexts (Antunes and Biala, 2012). The significant differences in time of the novel and familiar object exploration reflect the ability of animals to react to novelty factors. The main difference in our training paradigm is in using unrewarded novel objects and rewarded familiar ones. As a rule, no reward is used in the recognition memory tasks. We found that DAT-KO rats spent long periods of their time to explore novel objects than WT rats. It is known that blocking the DAT leads to an increased preference for novel options in probabilistic decisions: for instance, DAT blockade in monkeys does not modulate the rate of the task learning (Costa et al., 2014). Hyperdopaminergic DAT-KO mice are easily aroused by novelty and always respond with a hyperlocomotion (Spielewoy et al., 2000). We believe that prolonged exploration observed in our task provides an opportunity for better encoding the distinctions between novel and familiar objects.

During learning, DAT-KO rats were making fewer erroneous trials than were WT rats. It might indicate that protracted novel object exploration leads to the imprinting in memory the learned rule that a novel object has never been rewarded. In the present study, the task had two aspects – motor reactions and object recognition. The rats had to run to retrieve food and at the same time to differentiate the objects to be moved. It should be pointed out again that unlike in other recognition tasks, rats received food reinforcement that helped them to memorize the task rules. The task specificity likely affects the DAT-KO rats’ reactions to the rewarded objects. However, in earlier studies that employed other tasks, it was found that DAT-KO rats are less sensitive to the rewarded stimuli than WT rats (Cinque et al., 2018). Some investigations describe the reward deficiency syndrome, which includes ADHD as a behavioral subtype. The patients with dysfunction of the brain reward cascade in the DA system have a high risk of addictive and compulsive behaviors (Blum et al., 2008). It is possible that the dysfunction of the brain reward system may also exist in DAT-KO rats. When we use an attractive reward, we improve the reward system functioning. Thus, reward may play the role of a specific trigger for storing the information of the experimental paradigm in long-term memory. Our experimental paradigm included the new objects that were never repeated, and therefore, the rat’s curiosity and motivation level might have increased. Moreover, regular rewarding of the familiar objects also contributes to task learning.

Change in DA neurotransmission does not necessarily result in alterations of motor skill acquisition and novel stimulus discrimination. Investigations in DAT knockdown (DAT-KD) mice showed that the elevated DA level did not affect instrumental learning but reduced the selectivity of the stimulus control in the Pavlovian conditional training (Yin et al., 2006). In our studies, DAT-KO rats learn and perform motor tasks easier than spatial tasks (Kurzina et al., 2020).

Examining long-term storage of motor skills in the conditions of the novel object presentations, we found that after 3 months, both groups of rats were able to correctly perform behavioral tasks starting from the first day of the retesting. A comparison of almost all behavioral parameters measured showed that DAT-KO rats performed the task better than WT rats. It was only the number of erroneous trials that did not vary significantly across the groups. There are data to indicate that the extension of a period of familiarization with the objects improved the long-term complex associative recognition memory (Shimoda et al., 2019). Acquisition of the task rules in both groups of rats may likely proceed similarly and thus help memory consolidation.

The most interesting findings were obtained when the results of the first and second experimental blocks were compared. DAT-KO rats have improved their results significantly – they performed the task faster, covered shorter distances, and spent shorter periods of their time to reach the first object when familiar objects were presented. We believe that motor task in a stable environment with an additional stimulus with rewarded objects contributes to a task acquisition for a long period. This task based on learning the motor skills is closely connected with the cortico-basal ganglia and the ventral tegmental area (VTA)–hippocampus connections (Hosp et al., 2009). Long-term potentiation (LTP) is one of the cellular mechanisms of long-term memory. It was shown that LTP/long-term depression (LTD) in the rat prefrontal cortex is modulated by DA (Otani et al., 2015), and altered cortical and nucleus accumbens LTP is found in DAT-KO mice (Yao et al., 2004; Xu et al., 2009). Rats that were placed into a novel environment that resembled an environment that they had previously encountered showed activation of D1/D5 DA receptors and synthesis of plasticity-related proteins leading to formation of long-term memory traces (Moncada and Viola, 2007). Likely, novel experiences that have some features in common with those of the past may activate the DA system and help memory consolidation (Duszkiewicz et al., 2019). In our case, an increased DA signaling may also lead to a better consolidation of memory traces.

In general, the data obtained allow us to suggest that DAT-KO rats learn and perform motor tasks easier in comparison with spatial tasks. We have found previously that DAT-KO rats are unable to form an optimal strategy for food search in comparison with WT rats. They also have a pronounced impairment in working memory (Kurzina et al., 2020). Intriguingly, in children with ADHD, the impairment of working memory components related to attention was described (Rodríguez-Martínez et al., 2020). At the same time, children with ADHD have intact long-term memory but a mild deficit in visual-spatial memory (Kibby and Cohen, 2008).

Furthermore, since DA is involved in various aspects of motivational behaviors and psychotic states (Efimova et al., 2016; Howe et al., 2018), the abnormal behaviors of DAT-KO animals may have relevance also for endophenotypes of other psychiatric disorders such as schizophrenia, bipolar disorder, autism, mania, obsessive–compulsive disorder, and addiction disorders (Arime et al., 2012; Wong et al., 2012; Jones et al., 2015; Mereu et al., 2017; Weinstein et al., 2017; Chang et al., 2018; Cinque et al., 2018; Chao et al., 2020). The DAT-KO rats might have good translational value for developing new treatment principles for mental disorders. As regards ADHD, it was reported that in children and adolescents with ADHD, exploration of a novel environment led to significantly better memory consolidation (Baumann et al., 2020). Data indicate that cognitive training of executive functions is more effective in preschooler children with symptoms of ADHD than in children without any developmental risks (Scionti et al., 2020). Children with ADHD were trained to fulfill a working memory task with individual feedback; and a year after training, they showed a pronounced improvement of working memory (Dobrakowski and Łebecka, 2020). Our studies in the animal model based on a goal-directed task may suggest development of new techniques improving learning in hyperactive children.

The data obtained indicate that DAT-KO rats can learn and retrieve the object recognition task from memory following a 3-month interval. Their task performance differs from that of WT rats, as reflected by a longer time of task fulfillment and the distances covered. Also, in a novel object exploration, DAT-KO rats took a longer time. However, they made fewer erroneous trials than WT rats. After a 3-month interval, both DAT-KO and WT rats correctly performed the behavioral task. During the retesting, DAT-KO rats showed remarkable progress: they fulfilled the task faster and traveled shorter distances when the familiar objects were presented than during learning. It means that under the condition of the novel object presentation, hyperactive DAT-KO rats can learn and store motor tasks in memory. Consequently, the data obtained suggest the development of innovative strategies for teaching children with ADHD. A novel stimulus included in the training material may improve the quality of learning. Switching from the routine materials to something new is likely to provide the basis for better information consolidation.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

The animal study was reviewed and approved by the Ethics Committee of Saint Petersburg State University, No. 131-03-4 of September 24, 2018.

NK, AV, and IA performed the experiments and data analysis. NK, AV, and RG designed the study and contributed to the writing and editing of the manuscript. RG contributed to the conceptualization, supervision, and funding acquisition for the project. All authors contributed to the article and approved the submitted version.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.