Impact of specific serotonin receptor modulation on behavioral flexibility
Serotonin (5-HT) is known to play a critical role in regulation of essential neural processes, whereas more recent research highlights serotonin\xe2\x80\x99s modulatory effects on cognition and executive functioning. Current examinations have identified specific serotonin receptors for their direct impact on behavioral flexibility. Providing definitive evidence for the impact of specific receptor targets on behavioral flexibility is difficult, due to range of behavioral tests used. Due to limited studies and the sheer amount of different serotonin receptor targets, beginning to bring these studies together is important for the field. Our current review of the available literature aims to differentiate how modulation of specific 5-HT receptors affects behavioral flexibility. Although more studies have examined 5-HT2A, 5-HT2C, and 5-HT6 receptors, it is unclear why this is the case. Above all, there are some paradoxical results pertaining to these receptor targets. There is a clear distinction between 5-HT2A and 5-HT2C, which conveys that these two receptor subtypes have inverse effects when compared to each other. In addition, some findings support one another, such as upregulation of 5-HT6 receptors impairs flexibility, while blockade alleviates this impairment in both drug-induced and disease model rodents. Further understanding how modulatory effects of specific 5-HT receptors impact behavioral flexibility is imperative to advance the development of new therapeutics for neuropsychiatric disorders afflicted by behavioral inflexibility.
INTRODUCTION<!>5-HT1A<!>5-HT1B & 5-HT2B<!>5-HT2A & 5-HT2C<!>5-HT5A<!>5-HT6<!>5-HT7<!>CONCLUSION<!>
<p>The indolamine serotonin is critical in regulation of several biological processes including appetite, mood, and sleep (Ray et al., 2011). Although serotonin is important in regulating these functions, there is further evidence highlighting the importance of serotonin in governing cognition and executive function (Berger et al., 2009; Meltzer & Roth, 2013). Specifically, accumulating evidence points to the direct impact of specific serotonin receptor modulation on adaptive behaviors (Carhart-Harris & Nut, 2017). Behavioral flexibility, also discussed as cognitive flexibility, is often described as the ability of adapting behavioral patterns when environmental or reward contingencies change. This requires the ability to inhibit certain established behaviors for more optimal choices, allowing organisms to adapt to the ever-changing environment accordingly (Izquierdo et al., 2017; Lea et al., 2020). Previous studies have tested the impact of overall serotonin levels on behavioral flexibility in both human (Meltzer & Roth, 2013; Gabriele et al., 2014; Skandali et al., 2018) and animal studies (Izquierdo et al., 2013; Zhukovsky et al., 2017). Several studies have demonstrated how overall serotonin depletion can have a negative impact on behavioral flexibility (Wallace et al., 2014; Lapiz-Bluhm et al., 2009), although Thirkettle et al., (2019) found that serotonin depletion only impaired feedback sensitivity, sparing behavioral flexibility in healthy individuals. In addition, genetically modified mice with depleted serotonin levels, do not display impaired behavioral flexibility (Carlson et al., 2016), while 5-HT reductions as a result of lesions or dietary tryptophan removal leads to impaired reversal learning performance (Clarke et al, 2005; Park et al, 1994). On the other hand, increases in extracellular 5-HT levels through pharmacological or genetic 5-HT transporter inhibition, has been shown to facilitate behavioral flexibility (Brigman et al., 2010). These conflicting findings demonstrate that a general increase or decrease in 5-HT is not sufficient to facilitate behavioral flexibility across behavioral measures.</p><p>Although systemic reductions (Thirkettle et al., 2019) and specific alterations in serotonin levels (Barlow et al., 2015; Hamilton & Brigman, 2015) impacts behavioral flexibility, specific serotonin receptor targeting has provided more divergent findings (Nilsson et al., 2015). As of current, 14 different serotonin receptor targets have been identified (McCorvy & Roth, 2015; Berger et al., 2009). These various receptors allow for complex changes in the modulation of behavior through specific serotonin receptor binding, it can also complicate our understanding of serotonin's impact on behavioral flexibility specifically. In rodent studies, a standard test of behavioral flexibility is reversal learning: a test in which rodents must first successfully learn a behavior, then inhibit that behavior in favor of another more optimal behavior. These can be examined in a standard operant chamber or radial arm maze. Tests such as strategy switching or attentional set-shifting, are also used to probe behavioral flexibility, but are different from reversal learning. Different not only in how behavioral patterns must adapt to changing rules, but even anatomically. While reversal learning requires changes in stimulus reward contingencies and an intact orbital frontal cortex (OFC) is needed, set shifting instead requires switching attention between dimensions, requiring an intact lateral prefrontal cortex (Chudasama & Robbins 2006; Nilsson et al. 2015).</p><p>When discussing the effects of specific serotonin receptors on behavioral flexibility, it is important to consider 5-HT release. The dorsal raphe nucleus (DRN) has the greatest number of serotoninergic cell bodies in the central nervous system. The DRN has efferent projections to the cerebral cortex and several limbic structures (Hale & Lowry, 2011). Regarding behavioral flexibility, 5-HT neurons of the DRN are increasingly active during prediction errors when reward contingencies change, common in a reversal learning task (Matias et al., 2017; Peters et al., 2021). Matias et al., (2017) demonstrated that inactivation of 5-HT neurons of the DRN lead to increases in perseverative responding, leading to impaired behavioral flexibility. This DRN activation was not specific to loss or gain reward, but was linked to prediction errors overall, highlighting the impact of 5-HT signaling in modulating behavioral flexibility. Even though systemic changes in serotonin are not very successful in modulating behavioral flexibility, central changes in DRN activation have a direct impact on flexible behavior. This review we will focus on the impact of specific serotonin receptor modulation on behavioral flexibility measured predominantly by reversal learning and set shifting tasks. Due to paucity of studies that employ a repeated drug treatment regimen, these will not be discussed in this review.</p><!><p>The 5-HT1A receptor couples to the Gi protein and activation leads to hyperpolarization of the neuron membrane, resulting in reduced activity (Nichols & Nichols, 2008). These receptors are widely distributed in the central nervous system, being localized in raphe nuclei as somatodendritic receptors (Riad et al., 2000), or in cortical and limbic areas as post-synaptic receptors (Verge et al., 1985; Radja et al., 1991; Miquel et al., 1992). Few studies have examined the impact of 5-HT1A receptor modulation on behavioral flexibility. Depoortère et al. (2010) found that treatment with the highly selective 5-HT1A receptor agonist F15599 (which primarily activates postsynaptic 5-HT1A receptors) restored PCP-induced cognitive deficits. In addition, the 5-HT1A agonist 8-OH-DPAT and F13714 (a chemical congener of F15599) were also tested. Both F13714 and 8-OH-DPAT required higher doses to mimic the therapeutic effects of F15599 administration, although they never reached the same therapeutic levels. Consequently, these higher doses of F13714 and 8-OH-DPAT had more aversive effects. 5-HT1A agonists, which specifically target postsynaptic receptors, improved behavioral flexibility more consistently than 5-HT1A agonists, which do not primarily activate postsynaptic receptors. The 5-HT1A receptor agonists F15599, 8-OH-DPAT, and F13714 dose dependently improved behavioral flexibility. Although there is limited data, because three different 5-HT1A agonists facilitated performance, additional inquiry examining how 5-HT1A modulation affects behavioral flexibility is warranted. Previous studies have also indicated that activation of 5-HT1A receptors can be anxiolytic (Bauer, 2015; Gordon & Hen, 2004), and may therefore facilitate flexible choice behavior. Together, these findings suggest that activation of 5-HT1A receptors facilitates behavioral flexibility.</p><!><p>Similar to 5-HT1A, research examining the effects of 5-HT1B receptor modulation on behavioral flexibility is limited. The Gi coupled 5-HT1B receptor can be found at high densities on afferent globus pallidus and substantia nigra neurons from the striatum and on Purkinjie cells of cerebellar nuclei. Previous studies have focused on the impact of 5-HT1B receptors on flexible behavior (Boddington et al., 2020; Buhot et al., 2003; Wolff et al., 2003). Wolff et al. (2003) conducted an experiment employing the spatial stepwise water maze task with the 5-HT1B KO (knockout) mouse. 5-HT1B KO mice showed selective facilitation during the relocation phase and multiple starting phases, indicating that 5-HT1B receptor downregulation may facilitate behavioral flexibility. Moreover, 5-HT1B KO mice express greater behavioral flexibility during the spatial stepwise water maze, but not other water maze tasks. In addition, Buhot et al. (2003) found that 5-HT1B KO mice were more resistant to age related reversal learning impairments than normal mice, further supporting these findings. Together, 5-HT1B KO mice show facilitated behavioral flexibility, but the mechanisms behind these changes are not well understood.</p><p>Recently, Boddington et al. (2020) examined the serotonin related genetic alterations associated with discriminative and reversal learning performance. Specifically, genetic variations of the 5-HT1B receptor gene were examined in red junglefowl chicks. In this reversal learning task, behavioral flexibility was measured by latency for chicks to switch behavioral patterns. 5-HT1B did not have a significant effect on performance. Also, 5-HT2B seemingly impacted behavioral flexibility, but may have had a lesser impact compared to 5-HT2A receptors. Thus, prefrontal 5-HT2A and 5-HT2B receptors may be jointly mediating aspects of behavioral flexibility. However as previously discussed, 5-HT1B receptor downregulation has a facilitatory impact on behavioral flexibility in 5-HT1B KO mice (Wolff et al., 2003). Thus, decreased expression of 5-HT1B facilitated behavioral flexibility, but only in mice. Another recent publication found that partial down regulation of the 5-HT2B receptor does not seem to impact behavioral flexibility in mice (Radke et al., 2020). In addition, 5-HT2B receptors are generally found in the lateral septum, hypothalamus, medial amygdala and cerebellum, regions associated with flexible behavior. Together these findings suggest that down regulation of 5-HT1A or 5-HT2B receptors can facilitate behavioral flexibility in rodent subjects.</p><!><p>The 5-HT2A receptor can be found in many brain regions including the hippocampus and cerebellum, but most importantly can be found throughout the cerebral cortex (Miner et al., 2003; Xu et al., 2000). This Gq coupled receptor is also the target of psychedelics and atypical antipsychotics. In contrast to the previously discussed receptors, the effects that 5-HT2A receptors have on behavioral flexibility are more widely studied. Boulougouris et al. (2007) found behavioral flexibility differences utilizing an instrumental two-lever spatial discrimination and serial reversal learning task, while treating rats with the selective 5-HT2A receptor antagonist M100907. The high dose of M100907 impaired reversal learning performance during the initial reversal, increasing both trials to criterion and incorrect responses, while producing no significant effect on retention. Even though 5-HT2A and 5-HT2C selective receptor antagonists were tested, only 5-HT2A led to a significant impact. Boulougouris et al. (2007) finds that the 5-HT2A and 5-HT2C selective receptor antagonists have opposing effects on behavioral flexibility during the initial reversal (Boulougouris et al., 2007). The opposing effects of 5-HT2A and 5-HT2C receptors have been reported in previous studies (Alex & Pehek, 2007; Burton et al., 2013; Fletcher et al., 2008). Interestingly, 5-HT2C receptors are found on GABA neurons of the dorsal raphe highlighting a negative-feedback loop impacting flexible choice behaviors (Serrats et al., 2005). Furthermore, this negative-feedback loop may also be driving the opposing effects of 5-HT2A and 5-HT2C activation.</p><p>In follow up experiments, Boulougouris and Robbins (2010) examined the impact of M100907 with intra-OFC, intra-medial prefrontal cortex (mPFC) or intra-nucleus accumbens (NAc) infusions on reversal learning. Unlike Boulougouris et al. (2007), intra-OFC, intra-mPFC, or intra-NAc infusions did not increase trials to criterion during reversal learning. Conversely, the 5-HT2C receptor antagonist displayed opposing effects when compared to the 5-HT2A drug. The potential therapeutic effects of 5-HT2A receptor modulation on behavioral flexibility, must be considered in relation to the density of 5-HT2A receptors. Baker et al. (2011) instead found that the 5-HT2A/2c receptor antagonist ketanserin improves strategy-switching with acute systemic administration, while the 5-HT2C receptor antagonist SB242084 did not impair strategy-switching utilizing a spatial cue response paradigm. These findings, in relation to Boulougouris et al. (2007), further complicate the extent to which 5-HT2A and 5-HT2C receptors mediate behavioral flexibility and the functional role they individually have on behavioral flexibility. Since reversal learning in an operant chamber task (Boulougouris et al., 2007) and a spatial strategy shifting paradigm (Baker et al., 2011) was employed, directly comparing the two is difficult. In addition, reversal learning and strategy-switching may require different neuronal mechanisms and brain regions, which may potentially explain why 5-HT2A and 5-HT2C receptors are affecting behavioral flexibility in opposing ways. While Boulougouris et al. (2007) points to the 5-HT2C receptor, Baker et al. (2011) points to the 5-HT2A receptor, suggesting that both receptor effects may interact. Furthermore, Amodeo et al. (2020) demonstrated that acute treatment with the 5-HT2A receptor agonist 25CN-NBOH impaired spatial probabilistic reversal learning while the 5-HT2C receptor agonist SER-082 was not sufficient at impairing probabilistic reversal learning. Together, these findings suggest that the 5-HT2A and 5-HT2C receptors do have opposing effects on behavioral flexibility. Together these findings suggest 5-HT2A over activation attenuates behavioral flexibility, while 5-HT2C activation alone may not be sufficient to impair flexibility.</p><p>Other studies have investigated the interactions between 5-HT2A receptor modulation and certain selective serotonin reuptake inhibitors (SSRIs). To illustrate, Furr et al. (2012) tested 5-HT2A modulation in OFC during reversal learning and the extent to which 5-HT2A accounts for the therapeutic effects that SSRIs specifically have on flexible behavior. Microinfusing the 5-HT2A antagonist M100907 into the OFC, impaired reversal learning in the attentional set shifting task (ASST), but only during the initial reversal. Next, the extent of which the role that 5-HT2A receptor in the OFC accounts for the SSRI citalopram's beneficial effects during reversal learning were examined. Although citalopram improved behavioral flexibility after chronic intermittent cold stress, intra-OFC injections of M100907 reversed the benefits of citalopram administration on reversal learning. Thus, the 5-HT2A receptor activation resulting from SSRI treatment, is possibly responsible for the enhanced behavioral flexibility that was observed. Although there are contradictory results between these two, both ASST and reversal learning performance depend on different brain regions such as hippocampus and prefrontal regions.</p><p>Although the aforementioned experiments have been conducted in control rodents, many other studies have examined neuropsychiatric models known to express behavioral rigidity. Amodeo et al. (2014) examined the effects of M100907 and the atypical antipsychotic Risperidone on the BTBR T + tf/J mice (BTBR) mouse model of autism spectrum disorder. Despite having multiple targets, risperidone has a high affinity for the 5-HT2A receptor. Both M100907 and risperidone enhanced reversal learning in the BTBR mice but not in the control, C57BL/6J strain. Incidentally, higher doses of risperidone led to reversal learning impairments in C57BL/6J mice. These findings parallel Furr et al. (2012) with cold intermittent stress, suggesting that 5-HT2A differentially mediates behavior for typical and atypical populations. These findings eventually led to a follow-up study that microinfused M100907 in the dorsomedial striatum and the OFC, exploring their effects on behavioral inflexibility during probabilistic reversal learning (Amodeo et al., 2017). Specifically, behavioral flexibility deficits are attenuated in BTBR mice when M100907 is infused in the dorsomedial striatum. Intra-OFC infusions of M100907, did not improve behavioral flexibility. Together, 5-HT2A receptor blockade in the dorsomedial striatum instead of the OFC, alleviated the behavioral inflexibility in BTBR mice. Due to the hallucinogenic properties of 5-HT2A agonists, fewer studies have examined the effects of 5-HT2A activation in humans. Pokorny et al. (2019) examined the effects of LSD and co administration with the 5-HT2A/2C antagonist ketanserin on behavioral flexibility in humans. LSD alone was found to induce deficits in a variety of cognitive processes, including behavioral flexibility. In the Intra/Extra-Dimensional shift task, ketanserin attenuated the LSD-induced cognitive deficits. When ketanserin was pretreated, LSD did not induce behavioral rigidity. Thus, the 5-HT2A components of the LSD affected behavioral rigidity and were prevented by ketanserin. These results coincide with previous research, since acute administration of ketanserin improved both strategy-switching and response strategy in rats (Baker et al., 2011).</p><p>Previous studies have also examined the region specific effects of 5-HT2A modulation on behavioral flexibility in prefrontal cortex (PFC), medial PFC, OFC and NAc (Alsiö et al., 2015; Amodeo et al., 2017; Boddington et al., 2020; Boulougouris et al., 2007; Boulougouris & Robbins, 2010; Furr et al., 2012; Hankosky et al., 2018). Not to mention, such variations have also been measured in both distinct brain regions and some of the individual sub-regions in areas such as the dorsomedial (Amodeo et al., 2017; Eskenazi et al., 2015) and dorsolateral striatum (Eskenazi & Neumaier, 2011; Eskenazi et al., 2015). Hervig et al. (2020) further explains the effects that 5-HT activity in the lateral OFC and the medial OFC has on behavior, through a touch-screen serial visual reversal learning task for rats that were administered M100907. Although infusing the 5-HT2A antagonist in the medial OFC did not impair reversal learning, reversal learning was impaired when it was infused in the lateral OFC. In sum, these findings suggest that 5-HT2A receptor blockade tends to restore or rescue behavioral flexibility deficits. This rescue is found in both pharmacologically induced and genetically induced impairments in behavioral flexibility (Amodeo et al., 2014; Boulougouris et al., 2007; Boulougouris & Robbins 2010; Furr et al., 2012; Alsiö et al., 2015; Barlow et al., 2015).</p><p>Research examining 5-HT2C and 5-HT2A receptor modulation on behavioral flexibility have found opposing results. The selective 5-HT2C receptor antagonists SB2420845, produces effects within the OFC that are in contrast to 5-HT2A blockade (Boulougouris et al., 2007; Boulougouris & Robbins, 2010) in healthy subjects, while systemic 5-HT2A receptor blockade does not impair reversal learning (Boulougouris et al., 2007). Boulougouris et al. (2007) observed that the 5-HT2C receptor antagonist treatment led to enhanced reversal learning, compared to the controls. These results were only significant during the initial reversal, suggesting that either drug tolerance or initial difficulty of the task may have directly impacted the beneficial effects of 5-HT2C blockade. The 5-HT2C receptor antagonist tended to promote perseverative errors but did not affect learning errors. Intra-OFC infusions of 5-HT2C receptor antagonists reduced both trials to criterion and incorrect responses during the initial reversal, which is similar to results with systemic treatment. Furthermore, intracerebral infusions of SB242084 into the mPFC or NAc did not have the same beneficial effects (Boulougouris & Robbins, 2010). Similar to Boulougouris et al. (2007), another study focused on the role of 5-HT2C receptors of the OFC on reversal learning in rats. Alsiö et al. (2015) that SB242084 improved initial reversal learning when infused into the OFC. SB242084 improves reversal learning through specific mechanisms within the OFC, which is similar to the results found in Boulougouris & Robbins (2010).</p><p>In mice, Del'Guidice et al. (2014) found that treatment with the 5-HT2C agonist CP809.101 attenuated behavioral flexibility deficits produced by a human tryptophan hydroxylase 2 variant. The Tph2-KI mutation results in a reduction of 5-HT synthesis by 40%, which mimics the human Tph2 variant. Tph2 knock-in mice with chronically reduced 5-HT synthesis showed deficits in behavioral flexibility, but 5-HT2C agonist treatment rescued this impairment. These findings suggest 5-HT2C receptor agonists may potentially improve behavioral flexibility with systemic 5-HT reductions.</p><p>Drug exposure during adolescence has been shown to result in long lasting cognitive deficits, including behavioral inflexibility. Hankosky et al. (2018) examined both age- and sex-related differences that amphetamine and methamphetamine (METH) exposure have on behavioral flexibility, these differences were examined utilizing a set shifting task. Depending on sex and age of exposure onset, exposure to amphetamine and METH exhibit differing effects on behavioral flexibility and related to 5-HT2C receptor co-localization. Specifically, METH self-administration exerts functional changes on 5-HT2C receptors within the OFC, but only in adolescent female rats, and solely adolescent female rats exhibited deficits during discrimination and reversal learning tasks. Together, age- and sex-related differences in 5-HT2C receptor densities in the OFC demonstrate that 5-HT2C receptor modulation on behavioral flexibility differs with sex and region.</p><p>5-HT2C receptors are associated with feedback sensitivity differences that are observed in depression and its subsequent effects on behavioral flexibility. Phillips et al. (2018) examined the function of 5-HT2C receptors on biased reinforcement learning, through novel valence-probe visual discrimination and probabilistic reversal learning tasks. During these tasks, the 5-HT2C antagonist SB242084 and the 5-HT2C agonist WAY163909 were compared to determine the effects of 5-HT2C receptor modulation on biased reinforcement learning. The 5-HT2C agonist was associated with improved performance in the probabilistic reversal learning task, whereas the 5-HT2C antagonist impaired acquisition in the probabilistic reversal learning task while sparing reversal learning performance. Together these findings in with 5-HT2C receptor modulation contradict each other, again highlighting the complicated impact specific serotonin receptor regulation has on behavioral flexibility. Together these studies do suggest that 5-HT2C receptor agonist treatment tends to facilitate behavioral flexibility while localized blockade of 5-HT2C receptors within the OFC similarly enhance behavioral flexibility.</p><!><p>The 5-HT5A receptor can be found on neurons of the cerebral cortex, hippocampus, thalamus, hypothalamus, habenula, and cerebellum (Thomas et al., 2006). Treatment with 5-HT5A receptor antagonists have shown to rescue behavioral flexibility deficits induced by ketamine exposure. Nikiforuk et al. (2016) examined the effects of 5-HT5A receptor antagonist SB69955 treatment on ketamine-induced behavioral rigidity using the ASST, novel object recognition task, and social interaction and social choice test. SB69955 not only improved behavioral flexibility in drug naive rats, but also restored behavioral flexibility deficits induced by ketamine by reducing the trials needed to reach criterion. This is highlighted by the enhanced behavioral flexibility in both impaired and control rats due to SB69955 treatment. Nikiforuk et al. (2016) demonstrated that 5-HT5A blockades enhanced performance during the ASST for both control and ketamine treated rats, although the rescued behavioral inflexibility was greater in the ketamine treated group. These results suggest that 5-HT5A receptor antagonists may have promising therapeutic effects for alleviating behavioral flexibility deficits that are observed in neuropsychiatric disorders or ketamine abuse. Even though the research examining the effects that 5-HT5A has on behavioral flexibility is relatively new, the findings warrant further investigation of the modulatory effects of 5-HT5A on behavioral flexibility. Due to the lack of studies examining the effects of increased 5-HT5A receptor activation, our understanding of specific 5-HT5A receptor modulation on behavioral flexibility is skewed, more research is required.</p><!><p>The 5-HT6 receptor can be found in the striatum, nucleus accumbens, hippocampus, dentate gyrus, olfactory tubercles, amygdala and cerebral cortex, and like the 5-HT2C receptor, is restricted to the central nervous system (Hirst et al., 2003; Ruat et al., 1993). Both the selective 5-HT6 receptor antagonists SB399885 and SB271046 have been shown to bolster behavioral flexibility, as measured through the ASST in healthy rodents. For instance, Hatcher et al. (2005) described how healthy subjects have more difficulty with the extra-dimensional shift than the intra-dimensional shift, SB399885 treatment facilitated extra-dimensional shift performance. On the other hand, SB271046 did not facilitate the extra-dimensional shift compared to intra-dimensional shift. Thus, SB271046 treatment was not as effective as SB399885 in facilitating behavioral flexibility, although this may be due to differences in receptor affinities. The same selective 5-HT6 receptor antagonist SB399885 inhibited the performance enhancing effects of 5-HT6 receptor agonist WAY181187 treatment during the ASST, suggesting that 5-HT6 activation may enhance flexibility while co administration of a 5-HT6 receptor antagonist blocks these effects. Burnham et al. (2010) similarly found that the selective 5-HT6 receptor agonist WAY181187 improved performance during the extra-dimensional shift. This enhanced flexibility with the 5-HT6 agonist on the extra-dimensional shift was blocked with co-administration of the selective 5-HT6 antagonist SB399885. Together these findings show that treatment with either a 5-HT6 receptor agonist or 5-HT6 receptor antagonist can in general facilitate behavioral flexibility. Not surprisingly, co-administration with the agonist and antagonist negates the effects produced by the single treatment.</p><p>Early neural activation measures, such as FOS, was quantified in the several brain regions that have previously been associated with behavioral flexibility (Burnham et al., 2010). Elevated FOS expression was found in the NAc and posterior parietal cortex with 5-HT6 agonist WAY181187 treatment. In addition, treatment with the antagonist SB399885 had blocked WAY181187 from increasing FOS-positive cells within the mPFC. This co-administration confirms the agonist is necessary to produce the reported results and this was not due to extraneous effects. SB399885 had no effect on FOS-positive counts when administered alone. These studies demonstrate that the 5-HT6 receptor agonist WAY181187 facilitated behavioral flexibility while increasing FOS expression in the nucleus accumbens (core and shell) and posterior parietal cortex. In addition, Mohler et al. (2012) conducted two experiments with the high affinity 5-HT6 receptor antagonist PRX-07034. PRX-07034 at both 1 and 3 mg/kg facilitated switching between a place and response strategy in a spatial learning task. Importantly, PRX-07034 reduced both perseverative and regressive type errors during strategy switches. These studies suggest both upregulation and downregulation of 5-HT6 receptors have a beneficial effect on behavioral flexibility as measured by the ASST. Although these studies apply different tests of behavioral flexibility, 5-HT6 receptor modulation facilitated performance regardless.</p><p>Other studies have focused on 5-HT6 over expression within subregions of the striatum. Eskenazi & Neumaier (2011) found that 5-HT6 over expression in the indirect basal ganglia pathway of the posterior region of the dorsal medial striatum, slowed learning of an operant task. Eskenazi et al. (2015) demonstrated how overexpression of 5-HT6 receptors of the striatum affect habitual behaviors that are governed by medium spiny neurons in dorsal regions of the striatum. Increases in 5-HT6 receptor over expression in indirect pathway medium spiny neurons of the dorsolateral striatum, enabled behavioral flexibility following overtraining on a simple repetitive operant response task. Increasing 5-HT6 receptor expression in medium spiny neurons of the dorsolateral striatum enhances behavioral flexibility in the face of overtraining and habituation. Yet, expression of 5-HT6 receptors within the dorsomedial striatum has opposing effects, slowing instrumental learning. The authors suggest that 5-HT6 receptor modulation is responsible for balanced responding and thus behavioral flexibility overseen by subregions of the dorsal striatum.</p><p>Previous studies have utilized various reversal learning paradigms that are generally deterministic, with certain rewards and losses. To examine the effects of increased 5-HT6 receptor activation on probabilistic reward contingencies in reversal tasks, Amodeo et al. (2018) examined the effects of 5-HT6 receptor agonist EMD386088 on probabilistic reversal learning on mice. The higher dose of EMD386088, 4 mg/kg, impaired probabilistic reversal learning by enhancing retention of the initial spatial discrimination. In addition, treatment with the 5-HT6 agonist BGC20–761 did not have an effect on probabilistic reversal learning. Research probing 5-HT6 receptor modulation showed that 5-HT6 receptor activation can impair behavioral flexibility, when reinforcement is probabilistic. Whereas, 5-HT6 receptor downregulation improves performance during the attentional set shifting task without impairing probabilistic reversal learning. Consequently, the conflicting research may be explained by dose-dependent effects or even the variations among different reversal learning paradigms. Together these findings highlight the importance of further examinations into 5-HT6 receptor antagonist treatment as a therapeutic for disorders that express high rates of behavioral rigidity such as autism spectrum disorder and obsessive-compulsive disorder.</p><!><p>In general, the 5-HT7 receptor has been shown to have an impact on thermoregulation, circadian rhythm, sleep and cognition (Hedlund & Sutcliffe, 2004). The 5-HT7 receptor can be found in higher densities on neurons of the thalamus, dentate gyrus, septum, hypothalamus, substantia nigra, hippocampus and cortex (Hannon & Hoyer, 2008). Although the 5-HT7 is the most recently discovered receptor found in mammals, few studies have already demonstrated its impact on behavioral flexibility. Nikiforuk & Popik (2013) found that stress-induced behavioral inflexibility can be attenuated with the 5-HT7 antagonist amisulpride utilizing the ASST, while also reducing depression measures and increasing cognitive function. Furthermore, Rajagopal et al. (2016) found that behavioral rigidity, due to sub-chronically administered phencyclidine, was attenuated by treatment of the 5-HT7 receptor antagonist SB269970. Rajagopal et al. (2016) also found that upregulation with the 5-HT7 agonist, AS19, did not rescue reversal learning deficits due to repeated phencyclidine exposure. Together these findings suggest 5-HT7 blockade improves behavioral flexibility deficits, this merits further examination into 5-HT7 antagonists.</p><!><p>Our current review of the literature highlights how the bulk of the research seems to focus on select receptors, such as the 5-HT2A, 5-HT2C, and more recently the 5-HT6 receptors. Although more studies have been conducted examining these specific receptors, we cannot determine why this is the case. It could be that these receptors do in fact have a greater impact on behavioral flexibility but determining this is difficult due to the lack of research examining the other, less examined serotonin receptor targets. Alternatively, the increased focus on these receptors may be due to the increased commercially available compounds directly modulating these receptors.</p><p>Although this review has primarily focused on the effects that specific serotonin receptor modulation has on behavioral flexibility, many other studies have been conducted examining compounds with a more diffuse affinity profile. Although there is a lack of research on the effects of 5-HT3 and 5-HT1D receptor effects on behavioral flexibility, tests with drugs like vortioxetine, which has an affinity for these sites and has been effective in rescuing behavioral flexibility, more studies with these types of drugs could give us some critical information regarding their therapeutic efficacy. Although, drugs like vortioxetine may be not be commonly used in basic research due to its lack of specificity, pre-clinical studies are still needed. Alleviating behavioral flexibility impairments may require drugs that target several receptors, although this was not the focus of the current review. Overall, this provides rationale for developing new poly target compounds with the aim of alleviating behavioral inflexibility. This would require greater understanding of specific receptor effects and eventually understanding their interactions.</p><p>After reviewing the available literature in hopes of unraveling the impact of specific serotonin receptor modulation on behavioral flexibility, this review has highlighted some of the opposing findings commonly found with these receptor targets. For instance, downregulation of 5-HT2A facilitates flexibility and so does 5-HT6 receptor facilitation, while upregulation of 5-HT2C facilitates flexibility, although it depends on the behavioral measures adopted. There is a clear distinction and interaction between 5-HT2A and 5-HT2C receptors, suggesting these two receptors have inverse effects. In contrast, some findings did point to a similar narrative, such as upregulation of 5-HT6 receptors impaired flexibility, while blockade alleviated this impairment in both drug-induced and disease model rodents. Together, the hope is that these findings will lead to the development of new therapeutics for neuropsychiatric disorders afflicted by behavioral inflexibility.</p><!><p>Serotonin receptor modulation and its impact on behavioral flexibility.</p><p>↑receptor target = agonist; ↓receptor target = antagonist; ↑ increased behavioral flexibility; ↓ decreased behavioral flexibility; — = no affect on behavioral flexibility</p><p>= localized administration</p><p>GEN = Genetic manipulation</p><p>= probabilistic reversal learning.</p>