Transcranial direct current stimulation of the mouse prefrontal cortex modulates serotonergic neural activity of the dorsal raphe nucleus

Transcranial direct current stimulation (tDCS) is being increasingly considered as a non-pharmacological alternative for various psychiatric and neurological disorders. In major depressive disorder (MDD), anodal tDCS applied over the left dorsolateral prefrontal cortex (DL-PFC) improves symptoms in drug-resistant patients [1]. However, the molecular mechanisms underlying the efficacy of tDCS in MDD are still unknown. One of the main neurotransmitters involved in the pathophysiology and psychopharmacology of MDD is serotonin (5-HT), released by neurons of the dorsal raphe nucleus (DRN), which have widespread projections to cortical, subcortical and brainstem regions. In rodents, the prefrontal cortex directly innervates the DRN [2], and the activation of this pathway by deep brain stimulation (DBS) was shown to have antidepressant activity [3]. We investigated whether tDCS over the PFC in mice affects the activity of DRN 5-HT neurons using single-unit in-vivo electrophysiology [4] and analysis of cFos levels (as ameasure of cell activation) in the prelimbic cortex (PrL) and the dorsal portion of the DRN (DRD) of adult (2e3 months) anesthetized male C57BL/6 mice. For in-vivo electrophysiology, once a spontaneous and stable DRN 5-HT neuronwas identified and isolated [4], we recorded the activity for 4-min at rest, for 10 min during 250 mA anodal or cathodal left PFC tDCS and for further 10min after stimulation (1 neuron/ mouse; 8 neurons for both anodal and cathodal stimulations). tDCS was applied through a plastic tube filled with saline solution just prior to stimulation. The centre of the active electrode (0.045 cm2) was positioned over the left frontal areas (1.5 mm lateral and 1.8 mm anterior to bregma). The counter electrode was a saline soaked sponge (5.2 cm2) adhering to the ventral thorax [5]. Mean firing activity during and after stimulation was measured and calculated as percentage of baseline. For cell activation analysis [6], cFos expression was quantified in 15 mice (5/group: sham, anodal, cathodal). Briefly, 90 minutes after tDCS anesthetized mice were transcardially perfused with PBS followed by PBS-4%