Major depressive disorder (MDD) is a common and prevalent psychiatric illness that comprises the most significant contributor to the global burden of disease and disability, which causes a substantial impact on the social function and life quality of individuals [1]. Notably, the conventional antidepressants (i.e., monoamine oxidase inhibitors, tricyclics, and monoamine reuptake inhibitors) have several drawbacks, including a therapeutic response that ranges from weeks to months, incomplete remission, and treatment resistance to a large percentage of the patients, and several bothersome effects that reduce the adhesion to treatment [2,3]. Considering these limitations of antidepressant therapy, research for therapeutic agents with new mechanisms of action is needed [4]. In this regard, the glutamatergic system has received special attention and mounting evidence has shown its involvement in the pathophysiology and treatment of MDD [5,6].

Glutamate is the major excitatory neurotransmitter in the mammalian brain and plays a crucial role in several functions, including mood modulation and neuronal plasticity [7]. Under physiological conditions, glutamate may be synthesized through neuronal aerobic metabolism or recycled from glutamine supplied by astrocytes, packaged into synaptic vesicles, and then released into the extracellular space where it can interact with its receptors, such as N-methyl-d-aspartate (NMDA) receptors [8]. Glutamate undergoes uptake from the synapse cleft by excitatory amino acid transporters activity on astrocytes (i.e., EAAT1/GLAST and EAAT2/GLT-1) and is converted into glutamine by glutamine synthetase enzyme [9]. Disturbances in these events can result in glutamatergic synaptotoxicity via extrasynaptic NMDA receptors, thereby causing neuronal atrophy and impairing emotional processing and behavior [10]. Importantly, dysfunctions in glutamate cycling and transmission have been reported in the hippocampus and prefrontal cortex of MDD patients [11] and rodents subjected to stress models [12].

Reinforcing the role of the glutamatergic system in MDD, overwhelming evidence has shown that ketamine, an NMDA receptor antagonist, effectively produces fast (within hours) and sustained (up to 1 week) antidepressant responses in patients with MDD, even in treatment-resistant individuals at risk of suicide [[13], [14], [15]]. However, ketamine has some knock-on effects that limit its widespread use, and therefore, it would be important to discover fast-acting antidepressants devoid of ketamine's side effects. Within this scenario, remarkable studies have suggested that guanosine, a guanine-based purine, could be a promising candidate to exert fast-acting antidepressant responses [16]. Preclinical studies have reported that acute or chronic administration of guanosine produces antidepressant-like responses in the forced swim test (FST) and tail suspension test (TST) [[17], [18], [19], [20]]. Moreover, guanosine exhibits antidepressant-like effects in animal models that induce depression-like behavior, such as olfactory bulbectomy [21] and dorsolateral striatum lesion by 6-hydroxydopamine [22].

Guanosine has also been shown to potentiate the antidepressant-like and pro-synaptogenic effects of subthreshold doses of ketamine [[23], [24], [25]], by modulating the classical targets of ketamine, such as activation of the mechanistic target of rapamycin protein complex 1 (mTORC1) signaling, brain-derived neurotrophic factor (BDNF) release, and synaptic protein translation [16]. Nevertheless, no previous study has investigated the effect of acute guanosine plus ketamine administration on glutamatergic parameters, as glutamine synthetase activity and GLT-1 levels. Another question is whether guanosine and/or ketamine, at the same protocol schedule that exhibits antidepressant-like effects, is effective in protecting hippocampal and prefrontocortical slices against glutamate-induced damage.

Given this background, this study investigated the antidepressant-like and neuroprotective effects elicited by guanosine, ketamine, and the combination of ketamine and guanosine at sub-effective doses in mice. Furthermore, we assessed whether the antidepressant-like and neuroprotective responses displayed by ketamine plus guanosine could be associated with the modulation of glutamine synthetase activity and GLT-1 levels. In addition, the ability of guanosine to interact with the NMDA receptor at the ketamine or glycine/d-serine binding sites was also evaluated by molecular docking analysis.