Vortioxetine, but not escitalopram or duloxetine, reverses memory impairment induced by central 5-HT depletion in rats: Evidence for direct 5-HT receptor modulation

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Abstract

Depressed patients suffer from cognitive dysfunction, including memory deficits. Acute serotonin (5-HT) depletion impairs memory and mood in vulnerable patients. The investigational multimodal acting antidepressant vortioxetine is a 5-HT3, 5-HT7 and 5-HT1D receptor antagonist, 5-HT1B receptor partial agonist, 5-HT1A receptor agonist and 5-HT transporter (SERT) inhibitor that enhances memory in normal rats in novel object recognition (NOR) and conditioned fear (Mørk et al., 2013). We hypothesized that vortioxetine's 5-HT receptor mechanisms are involved in its memory effects, and therefore investigated these effects in 5-HT depleted rats. Four injections of the irreversible tryptophan hydroxylase inhibitor 4-chloro-dl-phenylalanine methyl ester hydrochloride (PCPA, 86 mg/kg, s.c.) induced 5-HT depletion, as measured in hippocampal homogenate and microdialysate. The effects of acute challenge with vortioxetine or the 5-HT releaser fenfluramine on extracellular 5-HT were measured in PCPA-treated and control rats. PCPA's effects on NOR and spontaneous alternation (SA) performance were assessed along with the effects of acute treatment with 5-hydroxy-l-tryptophan (5-HTP), vortioxetine, the selective 5-HT reuptake inhibitor escitalopram, or the 5-HT norepinephrine reuptake inhibitor duloxetine. SERT occupancies were estimated by ex vivo autoradiography. PCPA depleted central 5-HT by >90% in tissue and microdialysate, and impaired NOR and SA performance. Restoring central 5-HT with 5-HTP reversed these deficits. At similar SERT occupancies (>90%) vortioxetine, but not escitalopram or duloxetine, restored memory performance. Acute fenfluramine significantly increased extracellular 5-HT in control and PCPA-treated rats, while vortioxetine did so only in control rats. Thus, vortioxetine restores 5-HT depletion impaired memory performance in rats through one or more of its receptor activities.

Introduction

Patients suffering from major depressive disorder (MDD) often present with clinically significant cognitive dysfunction (Kuny and Stassen, 1995), including impaired executive function, episodic memory, and spatial working memory (Airaksinen et al., 2004, Austin et al., 2001, Lee et al., 2012). Recent data suggest that cognitive dysfunction hinders functional recovery (Jaeger et al., 2006), and that greater severity of cognitive dysfunction predicts a poorer response to selective serotonin (5-HT) reuptake inhibitors (SSRIs; Dunkin et al., 2000, Kampf-Sherf et al., 2004, Kaneda, 2009).

MDD has for decades been associated with decreased central serotonergic tone. Support of the 5-HT hypothesis of depression is partly ascribed to the success of SSRIs and other monoamine-centered therapies, but is also supported by the observation that acute depletion of the 5-HT precursor, tryptophan, induces low mood in vulnerable patients (Booij et al., 2005, Delgado et al., 1990, Delgado et al., 1994, Benkelfat et al., 1994), and causes cognitive dysfunction (Reidel et al., 1999, Booij et al., 2005, Sobczak et al., 2002). Thus, in combination with the observation that cognitive dysfunction predicts responsiveness to SSRI treatment (Dunkin et al., 2000, Kampf-Sherf et al., 2004, Kaneda, 2009), these data suggest that the relationship between MDD and central 5-HT tone may extend beyond mood and also encompass cognitive function.

Consistent with this hypothesis, serotonergic treatments attenuate depression-related dysfunction in cognitive domains such as episodic memory, working memory, attention, and executive function (Cassano et al., 2002, Constant et al., 2005, Herrera-Guzman et al., 2009, Herrera-Guzman et al., 2010a, Levkovitz et al., 2002), although it should be noted that some studies fail to find significant treatment effects (Ferguson et al., 2003, Nebes et al., 2003). Norepinephrine reuptake inhibitors (NRIs) or serotonin norepinephrine reuptake inhibitors (SNRIs), while not as extensively studied, also appear to have a benefit in cognitive domains such as episodic memory and to a lesser extent, working memory, mental processing speed, and motor performance in patients with MDD (Ferguson et al., 2003, Herrera-Guzman et al., 2009, Herrera-Guzman et al., 2010b; but see Levkovitz et al., 2002). These data also suggest that SNRI treatment may be superior to SSRIs at improving a subset of cognitive functions (Herrera-Guzman et al., 2009, Herrera-Guzman et al., 2010b). However, it is important to note that MDD-related cognitive deficits often persist in a substantial proportion of patients even after mood dysfunction has ameliorated (Herrera-Guzman et al., 2010a, Herrera-Guzman et al., 2010b, Kuny and Stassen, 1995, Nebes et al., 2003). Thus, remediation of cognitive dysfunction remains an unmet need for this patient population.

The investigational antidepressant vortioxetine (Lu AA21004) is one of the first antidepressants with a multimodal mechanism of action, so named because of its two separate modes of action: (1) 5-HT reuptake inhibition and (2) direct pharmacological action at serotonergic receptors. Vortioxetine acts as a 5-HT3, 5-HT1D, and 5-HT7 receptor antagonist, 5-HT1B receptor partial agonist, 5-HT1A receptor agonist, and a 5-HT transporter (SERT) inhibitor in recombinant cell lines (Bang-Andersen et al., 2011, Mørk et al., 2012, Westrich et al., 2012). A recent double-blind placebo controlled clinical study in elderly depressed patients found in its exploratory endpoints that vortioxetine significantly improved performance in the Digit Symbol Substitution Test and the Rey Auditory Verbal Learning Task compared to placebo (Katona et al., 2012). In preclinical studies, vortioxetine enhances memory performance in the novel object recognition (NOR) and conditioned fear tests (Mørk et al., 2013). Several of vortioxetine's receptor activities, for example 5-HT3 receptor antagonism (Petkov et al., 1995, Staubli and Xu, 1995) as well as 5-HT1A receptor agonism (Bertrand et al., 2001, Horiguchi and Meltzer, 2012) have been implicated in enhancing memory function in rats. We therefore hypothesized that one or more of vortioxetine's receptor activities are involved in mediating its memory enhancing activities. To reduce the impact of vortioxetine's SERT inhibitory activity we investigated its effects on memory performance in rats with low central 5-HT tone. Thus, the present study investigated the ability of vortioxetine, the SSRI escitalopram and the SNRI duloxetine, to rescue object recognition and spatial working memory deficits induced by the irreversible tryptophan hydroxylase inhibitor 4-chloro-dl-phenylalanine methyl ester HCl (PCPA), which depletes 5-HT by stopping the rate-limiting step in its synthesis. The degree of depletion was measured from hippocampal tissue homogenate and microdialysate. Furthermore, extracellular 5-HT in the ventral hippocampus was measured in PCPA-treated and control rats after dosing with vortioxetine or the 5-HT releaser, fenfluramine. The level of SERT occupancy after vortioxetine, escitalopram and duloxetine were measured by ex vivo autoradiography.

Section snippets

Animals

Female Long-Evans rats (Charles River Laboratories, Wilmington, MA, USA) weighing 225–350 g were used for all experiments. Rats were group-housed, three per cage in plastic cages (Rat IVC Green Line Sealsafe plus cages; Tecniplast USA, Philadelphia, PA) except that rats used in microdialysis experiments were individually housed after surgeries. All animals had ad libitum access to food and water in their home cages, in a temperature (69–71°F) and humidity controlled environment (30–70%) on a 12 h

5-HT levels in hippocampus homogenate

PCPA administration for four consecutive days dose-dependently decreased the average 5-HT concentration in hippocampal tissue homogenate to approximately 3% of control values at 86 mg/kg/day (Table 1), and this dose was used in all subsequent experiments. 5-HT depletion reached statistical significance at 26 mg/kg/day (H(5, n=25)=23.4, p<0.001). The ED50 of PCPA treatment was 17.3 mg/kg/day. The profound depletion of 5-HT observed at this dose (H(3, n=20)=16.58, p<0.001; Figure 1A) remained stable

Discussion

In the present study we investigated the effects of the multimodal acting antidepressant vortioxetine on memory performance in a rat model of 5-HT depletion-induced memory deficits and addressed the hypothesis that vortioxetine's effects might involve direct 5-HT receptor mediated effects.

In line with previous research we show that repeated administration of the irreversible tryptophan hydroxylase inhibitor, PCPA, results in dose-dependent reductions in brain tissue concentration of 5-HT and

Role of the funding source

This study was funded by H. Lundbeck A/S and the Takeda Pharmaceutical Company, Ltd. Employees of Lundbeck played a role in the design of experiments, as well as the collection, analysis and interpretation of data. Lundbeck employees also played a role in the writing of and the decision to submit the present study.

Contributors

Jesper Bornø Jensen and Kristian Gaarn du Jardin participated in all experiments related to behavior, assessment of 5-HT concentration in hippocampal tissue, and ex vivo receptor occupancy. Dekun Song conducted the in vivo phase of all microdialysis experiments. David Budac analyzed 5-HT concentrations from microdialysate and hippocampal tissue homogenate. Gennady Smagin participated in the design of microdialysis experiments. Connie Sanchez participated in the design of all experiments. Alan

Conflict of interest

Jesper Bornø Jensen, Kristian Gaarn du Jardin, Dekun Song, David Budac, Gennady Smagin, Connie Sanchez, and Alan Pehrson are employees of Lundbeck Research USA, Inc.

Acknowledgments

The authors wish to thank Mr. David J. Simpson, who assisted with proofreading this manuscript.

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