Studies examining the acute effects of antidepressant drugs have used the forced swim test (FST) and/or tail suspension test (TST), the gold standards for assaying antidepressant-like activity in rodents (Cryan and Holmes, 2005; Castagné et al., 2011). While there are some inconsistencies among reports (Table 2A), the most common finding is that SSRIs are less effective in juvenile and adolescent rodents, than in adults (Bourin et al., 1998; Mason et al., 2009; Mitchell et al., 2013, 2017). This is in line with clinical reports (Tsapakis et al., 2008; Hetrick et al., 2010, 2012), and underscores the utility of these preclinical tests as predictors of antidepressant-like efficacy in humans.

Interestingly, Mason et al. (2009) found that age-dependent differences in antidepressant-like response were also dependent on mouse strain and test used. For example, in the TST fluoxetine (20 mg/kg) produced an antidepressant-like effect in juvenile (P35) and adult C57BL/6J and F2 mice, but not in BALB/C mice. However, when using the FST, juvenile and adult C57BL/6 mice were insensitive to fluoxetine, but F2 and BALB/C mice responded. This is not surprising in view of elegant studies from Lucki’s group using adult mice to show clear strain-dependent differences in response to fluoxetine in the FST (Lucki et al., 2001) and to citalopram in the TST (Crowley et al., 2005). Regardless, in the Mason study, when antidepressant effects were detectable they were less in juveniles than in adults when using the FST, and less than or equivalent to those of adults when using the TST.

While the majority of studies have used mice, one study used rats (Reed et al., 2008). Using the FST, these investigators found that the SSRIs fluoxetine and escitalopram produced antidepressant-like effects that were of a similar magnitude in P21 and adult rats (Table 2A). Interestingly, they found that the tricyclic antidepressants, desipramine and imipramine, were ineffective in P21 rats, but produced robust antidepressant-like effects in adults. While the mechanistic underpinnings for these results are yet to be determined, a common view is that delayed maturation of the norepinephrine neurotransmitter system relative to serotonin is a factor (Bylund and Reed, 2007). However, in contrast to the rat study, mouse studies generally found tricyclic antidepressants to be either equally effective in producing antidepressant-like effects in the TST and FST in juveniles, adolescents, and adults (Bourin et al., 1998; Mason et al., 2009; Mitchell et al., 2013), or more efficacious in juveniles and adolescents than in adults (David et al., 2001; Mitchell et al., 2017), suggesting that in mice, the norepinephrine neurotransmitter system matures faster than in the rat. In humans, tricyclics are not prescribed for children and adolescents due to poor tolerability (e.g., vertigo, tremors, low blood pressure, and dry mouth; Andersen et al., 2009; Dell’Osso et al., 2011; Hazell and Mirzaie, 2013). However, preclinical findings from mouse studies indicate that the utility of NET, and/or dual NET and SERT blockers in the treatment of juvenile and adolescent depression warrants further investigation. It is possible that the development of new NET (and NET/SERT) targeting drugs that have fewer side effects than currently available drugs may be better suited to the treatment of pediatric depression (Mitchell et al., 2017).

Clearly, differing rates of development of noradrenergic and serotonin systems, and the development of systems that impinge on them (e.g., histamine, see Munari et al., 2015) may also factor into the magnitude of antidepressant-response as a function of age. Rates of maturation also differ among species, and is a consideration when relating findings from rodents to outcomes in humans. The reader is directed to an excellent review by Murrin et al. (2007) for information on differential rates of development of noradrenergic and serotonergic systems among species. While studies in human are relatively scant, those that exist, together with studies in non-human primates, suggest that the serotonin system matures faster than the noradrenergic system (Murrin et al., 2007). It will be of interest for future studies to precisely map treatment response to different classes of antidepressant on developmental stage of the noradrenergic and serotonergic systems in humans.

Lastly, one study included constitutive SERT heterozygote mice as a murine model of humans carrying a common variant of the SERT gene, which confers reduced SERT functionality, and who respond less well to SSRIs than those without this variant (Kronenberg et al., 2007). Consistent with clinical literature, the maximal effect of escitalopram in juvenile SERT heterozygote mice was less than that in juvenile SERT wildtype mice. Moreover, in both SERT genotypes the maximal effect in juveniles was less than that in adults (Mitchell et al., 2016). These findings again underscore the utility of preclinical models of antidepressant-like activity in probing mechanisms contributing to age- and genotype-dependent responses to antidepressants (but see section “Caveats to Currently Available Behavioral Assays for Identifying Drugs With Antidepressant Potential”).

In sum, the most common finding from studies of the acute antidepressant-like effects of SSRIs in preclinical studies is that they are less effective in juveniles and adolescents than in adults, though some studies do report no difference in response among ages. With proper attention to mouse strain, and behavioral test employed, such preclinical models provide valuable tools to study the mechanistic basis for SSRIs being less effective in treating pediatric depression than in adults, as will be discussed further.

While the TST and FST are currently the best tests available to quickly screen for drugs with antidepressant potential, for drugs to be useful clinically they must be effective after chronic administration. Numerous groups have reported antidepressant-like effects following chronic SSRI treatment in adult mice and rats, using tests such as the FST, TST, and novelty-induced hypophagia (Detke et al., 1997; Dulawa et al., 2004; Cryan et al., 2005; Holick et al., 2008; Miller et al., 2008; Balu et al., 2009; Jiao et al., 2011; Roohi-Azizi et al., 2018), with only one group reporting no effect (Griebel et al., 1999). However, only four studies have examined the antidepressant-like effect of chronic SSRI administration in juveniles and/or adolescents. All have used rats (Table 2A). Three used the FST, and though different SSRIs were used (fluoxetine in two studies [Homberg et al., 2011; Bouet et al., 2012] and paroxetine in the other [Karanges et al., 2011]), the overall findings were in general agreement. In the two studies where there was no washout period, chronic SSRI treatment either had no effect in young rats (Karanges et al., 2011), or increased immobility time (i.e., pro-depressive-like behavior; Homberg et al., 2011), whereas in adults there was either no effect (Homberg et al., 2011), or a robust antidepressant-like effect (Karanges et al., 2011), in agreement with published literature using adult rodents. These results are consistent with clinical literature reporting that children and adolescents respond poorly to SSRIs compared with adults, if at all (Tsapakis et al., 2008; Hetrick et al., 2010, 2012). In the one study where a 1-week washout period was allowed before testing in the FST, no antidepressant-like effect of fluoxetine was observed in either juvenile/adolescent or adult rats (Bouet et al., 2012).

Another study utilized inescapable shock and shuttle box tests to assay antidepressant-like activity following a week of twice daily injections of escitalopram or desipramine in rats. Escitalopram produced similar antidepressant-like effects in juveniles, adolescents, and adults, but desipramine was only effective in adolescent and adult rats, and ineffective in juvenile rats (Reed et al., 2009). This result is consistent with findings reported by the same group following acute administration of these drugs (Reed et al., 2008; see discussion above).

Overall, results from studies examining the chronic effects of SSRIs in rats again support the utility of preclinical assays such as these. However, given that mice seem to more reliably parallel human clinical literature, at least in the acute tests described above, it will be of interest to implement mice in studies of chronic SSRI administration as well.

Though the FST and TST are the current gold standards for assaying antidepressant-like activity in rodents (Cryan and Holmes, 2005), it is important to note that while the vast majority of antidepressant drugs reduce immobility time in these tests, not all drugs that reduce immobility time are antidepressants. Thus, these tests are valuable when comparing the sensitivity to drugs across ages, but they do not necessarily predict therapeutic effectiveness in the treatment of depression. In this realm, the field is in need of new tests (be they behavioral or biochemical) that can reliably predict therapeutic efficacy of novel drugs for the treatment of depression, while screening out false positives.

In addition, to date studies of juvenile and adolescent rodents in tests for antidepressant-like activity of drugs have used only naïve animals. Ideally, such tests should be performed in animal models of depression, such as those afforded by unpredictable chronic mild stress, or social defeat paradigms, to name a few (see O’Leary and Cryan, 2013 for review). While these models have been frequently applied to studies of depression in adult rodents, their application to juveniles (in particular), as well as adolescents, is impractical due to the shortness of these developmental periods (e.g., 7-day juvenile period, 14-day early adolescent, and 14-day late adolescent periods; Spear, 2000; Bylund and Reed, 2007; Hefner and Holmes, 2007; Pechnick et al., 2008; Reed et al., 2008). This is further compounded when the effects of chronic drug treatment need to be studied. While it is not within the scope of this focused review to discuss the pros and cons of existing models for depression research, the reader should be aware that there are limitations to those currently available (see O’Leary and Cryan, 2013 for review). Of course, recapitulating the heterogeneous nature of human depression, without yet fully understanding the etiologies of this complex disorder, makes developing improved animal models a challenging undertaking. However, as we learn more about the complexities of this disorder, new knowledge can be applied to the development of improved models to study depression and to screen novel compounds for therapeutic efficacy in its treatment. Importantly, models that can be applied to the study of juveniles and adolescents are sorely needed.

In adult rodents, chronic treatment with SSRIs downregulates SERT expression (Benmansour et al., 1999; Gould et al., 2003, 2007; Mirza et al., 2007; Lin et al., 2017), which is associated with putatively reduced functionality of SERT, as demonstrated by in vivo chronoamperometry studies measuring clearance of serotonin from extracellular fluid in hippocampus (Benmansour et al., 1999). These decreases were not associated with reduced SERT gene expression or neurotoxicity (Benmansour et al., 1999). Several lines of evidence suggest that SSRI-induced downregulation of SERT function is attributable, at least in part, to internalization of SERT to the cytosolic compartment. For example, in vitro studies using Caco-2 cells transfected with human (h) SERT show that long-term exposure to fluoxetine causes internalization of hSERT, leaving less hSERT on the plasma membrane (Iceta et al., 2007). These studies showed no effect of fluoxetine treatment on either total hSERT protein or mRNA. Studies in rats found that chronic, but not acute fluoxetine treatment causes internalization of SERT in both cell bodies and terminals (Descarries and Riad, 2012). Similarly, translational approaches using stem cell-derived serotonergic neurons and a transgenic mouse expressing hSERT found that citalopram dose-dependently causes internalization of hSERT in both models (Matthäus et al., 2016). Such studies underscore the utility of complementary/translational approaches to understanding antidepressant response on a cellular and molecular level. While it remains to be determined if internalization of SERT following chronic SSRI treatment occurs in humans, and is temporally synced with therapeutic benefit, studies in adult rodents, and in vitro, suggests that downregulation of SERT, putatively by trafficking away from the plasma membrane, is at least in part required for ultimate therapeutic benefit following chronic treatment with SSRIs. Moreover, that it takes chronic exposure to SSRIs for this downregulation of SERT to occur, provides a potential mechanistic basis for the delay to onset of therapeutic benefit.

Currently there is no literature reporting effects of chronic SSRI treatment in juvenile rodents. This is almost certainly due to the very narrow juvenile window in rodents (i.e., P21–27). There is remarkably sparse literature on the effect of chronic SSRI treatment on SERT expression in adolescent rodents (reviewed in Daws and Gould, 2011). However, a consistent finding is that chronic treatment with SSRIs, beginning at either P25 or P28 in rodents, increases SERT expression in a number of brain regions (Wegerer et al., 1999; Karanges et al., 2011; Bouet et al., 2012) (Table 2B). These findings in rodents are further supported by a study in juvenile rhesus macaque monkeys, which found that chronic treatment with fluoxetine increased SERT expression in several brain regions, including neocortex and hippocampus (Shrestha et al., 2014) (Table 2B). With more SERT putatively becoming available to take up serotonin as SSRI treatment continues, the increases in extracellular serotonin that are thought to be needed to trigger the downstream cascade of events leading to ultimate therapeutic benefit could be greatly diminished. Thus, the clinical implications of increased SERT expression in children and adolescents could include a need to increase dose of SSRI in order to fully occupy SERT. It is interesting to note that initiating dosing of SSRIs for pediatric depression is generally less than that for adults. For example, initiating low doses of fluoxetine and citalopram to 5- to 12-year olds is less than 10 mg/d, whereas in adults (18–64 years) the dose is less than 20 mg/d (Bushnell et al., 2016). Before the black-box warning on all antidepressants was issued by the FDA in 2004, the percent of patients initiating low dose SSRI treatment was similar across ages (∼15%), with the exception of 10- to 12-year olds where the percent was much lower (5%). In contrast, following the black-box warning the percent of patients initiating low dose SSRI (relevant to their age) increased markedly (5–9 years to ∼28%, 13–17 years to ∼38%, 18–24 years to ∼25%, 25–64 years to ∼20%, and 10–12 years to ∼11%; see Bushnell et al., 2016). In the context of the present review, this raises the possibility that increasing initial dosing in pediatric depression may be therapeutically beneficial, though clearly with this comes the risk of increased, potentially life-threatening, side-effects. It must also be kept in mind that of the clinical studies summarized in Table 1, SSRI doses much higher than these initiating low doses (e.g., 60 mg/day for fluoxetine; 200 mg/d sertraline) yielded mixed outcomes, some reporting therapeutic efficacy, and others no difference from placebo.

Clearly it is difficult to draw conclusions from existing literature. The important take home message here is that increased SERT expression following chronic SSRI treatment in children and adolescents could account, at least in part, for their therapeutic efficacy being less than that in adults. Though there are no studies of SERT in brain of children and adolescents, one study examining platelet SERT expression before and after treatment with the SSRI sertraline, provides support for this premise. Sertraline treatment was found to decrease [³H]paroxetine binding affinity. Notably increased maximal binding of [³H]paroxetine to SERT was associated with non-response (Sallee et al., 1998). Further study into the benefits of more rapidly ramping SSRI dosing in pediatric depression may therefore be warranted.

To stay within the scope of this focused review, we have discussed only the consequences of chronic SSRI treatment for their primary target, SERT. Of course this is an overly simplistic view of the neural consequences of chronic SSRI treatment in juveniles and adolescents, which has also been shown to impact glycogen synthase kinase-3 (GSK3; Beurel et al., 2012), extracellular signal-regulated protein kinase 1/2 (ERK) and cAMP response element binding protein (CREB; Alcantara et al., 2014), brain derived neurotrophic factor (BDNF) and TrkB receptors (Kozisek et al., 2008), genes associated with cell cycle and survival (Tsapakis et al., 2014), and neuroplasticity (Bastos et al., 1999; Norrholm and Ouimet, 2000; Bock et al., 2013; Klomp et al., 2014). Some of the effects of chronic SSRI treatment on these factors in juveniles and adolescents are in parallel with those reported in adults (Bastos et al., 1999), while others are not (Kozisek et al., 2008; Beurel et al., 2012; Bock et al., 2013; Alcantara et al., 2014; Klomp et al., 2014). A deeper focus on SSRI-induced changes that are specific to juveniles and adolescents will help guide future research to develop improved antidepressants for these young populations.

Prior to 2011, remarkably little was known about the ontogeny of SERT expression during juvenile and adolescent periods (reviewed in Daws and Gould, 2011). The first paper to describe postnatal ontogeny of SERT found that in terminal regions, including amygdala and hypothalamus, expression was maximal around P21, decreased at P28, before rising again to stabilize through P70, the oldest age tested (Galineau et al., 2004; Daws and Gould, 2011). Subsequent studies of SERT expression from P21 through adulthood reported either no change across those ages (Karanges et al., 2011; Mitchell et al., 2013), or a steady increase from P21 through adulthood in terminal fields (Bouet et al., 2012; Ulloa et al., 2014; Mitchell et al., 2016), and a decrease from P21 in cell body regions (Bouet et al., 2012; Mitchell et al., 2016) (Table 2C). Of particular interest, Mitchell et al. (2013) using [³H]citalopram saturation binding assays in hippocampal homogenates, provided evidence that SERT in juveniles may be functionally less able to transport serotonin than in adults. The modest variation in results among these studies is likely due to nuanced differences in the assays used, ages studies, and species used. However, taken together current evidence suggests that SERT expression and/or function in terminal fields is lower in juveniles than in adults, whereas expression in cell bodies is greater (Table 2C).

It is difficult to speculate on how these postnatal shifts in SERT expression and function ultimately play into therapeutic responsiveness to SSRIs, but this is clearly a line of investigation that warrants further research. Likewise, it will be important to determine the relationship between receptor systems that are well-known to be influenced by SSRIs (e.g., Artigas, 2013; Kulikov et al., 2018), SERT, and therapeutic response in juveniles and adolescents.