Repetitive transcranial magnetic stimulation (rTMS) is a noninvasive brain stimulation technique commonly used in clinical and research applications, and it can affect both inhibitory and excitatory neuronal activities thus modulating cortical excitability (Huerta and Volpe, 2009). Of the conventional rTMS patterns, low-frequency (≤1 Hz) rTMS reduces cortical excitability while high-frequency (≥5 Hz) rTMS increases cortical excitability. The theta burst stimulation (TBS) protocol for rTMS involves high stimulation frequencies (50 Hz bursts of 3 pulses repeated at 5 Hz). Intermittent TBS (iTBS) involves the application of a 2 s train of TBS repeated every 10 s for a total of 192 s, and has been shown to enhance cortical excitability. The application of high frequency bursts (γ) coupled with low frequency rhythm (θ) closely mimics the natural firing pattern of the brain. This enables iTBS to produce greater and longer-lasting effects on cortical excitability than high-frequency rTMS (Huang et al., 2005; Di Lazzaro et al., 2011). Accumulating clinical evidence has shown that high-frequency rTMS or iTBS of the prefrontal cortex improved cognitive performances in healthy controls and patients with various neurological disorders (Bagherzadeh et al., 2016; Hoy et al., 2016; Wang et al., 2022; Chou et al., 2020; Miller et al., 2023; Chen et al., 2024; Chu et al., 2022; Wu et al., 2022a). Studies in rodents consistently revealed the pivotal role of the prefrontal cortex in the beneficial effects of the two modalities on memory function (Wu et al., 2022b; Lin et al., 2021; Yang et al., 2020; Wang et al., 2023a). However, whether iTBS, which induces greater after-effects on cortical excitability (Huang et al., 2005; Di Lazzaro et al., 2011), could exert better cognitive effects than high-frequency rTMS remains largely unknown. To date, researchers have mainly focused on the role of prefrontal synaptic plasticity, brain clearance pathways and neuroinflammation in the cognitive effects induced by the two modalities. Few studies have examined whether rTMS-induced cognitive effects are associated with changes in prefrontal neuronal oscillations and neurotransmitter levels, which are highly involved in modulating memory function.

Neuronal oscillations at specific frequencies in the prefrontal cortex are implicated in cognition and memory (Salinas and Sejnowski, 2001; Buzsáki and Draguhn, 2004; Tanigawa et al., 2022). Frontal γ and θ oscillations are responsible for the interaction between cortical structures and other brain areas for encoding and retrieval of episodic memory (Nyhus and Curran, 2010). Greater frontal γ and θ power have been found during the episodic memory task, and γ oscillation covaried with θ oscillation (Burgess and Ali, 2002; Doppelmayr et al., 2000; Düzel et al., 2003; Hanslmayr et al., 2009). While δ oscillation reflects genuine processing involved in memory consolidation (Todorova and Zugaro, 2019), and decreased δ power is related to enhanced memory performance (Cui et al., 2022; Balconi and Pagani, 2015). Besides, mounting evidence has shown that prefrontal excitatory and inhibitory neurotransmitters participate in regulating memory function, and the improvement of memory performance may result from increased glutamate levels and decreased gamma-aminobutyric acid (GABA) levels in the prefrontal cortex (Dawson et al., 2001; Thielen et al., 2019; Piechal et al., 2023; Jentsch et al., 2009; Davydova et al., 2020; Pistis et al., 2002; Sandini et al., 2019). These studies strongly support the involvement of prefrontal neuronal oscillations and levels of extracellular GABA and glutamate in modulating memory function.

rTMS has the potential to regulate network oscillations in the prefrontal cortex with the “entrainment” after-effects (Thut et al., 2011; Suppa et al., 2016; Huerta and Volpe, 2009; Woźniak-Kwaśniewska et al., 2014; Barr et al., 2009; Wang et al., 2023a). Several studies from humans and rodents suggest that high-frequency rTMS or iTBS can enhance working memory while altering the power and coherence of neuronal oscillations in the prefrontal cortex (Barr et al., 2009; Liu et al., 2021; Bai et al., 2018; Wang et al., 2023a). In addition to neuronal oscillations, changes in prefrontal neurotransmitters potentially contribute to the rTMS's after-effects. (Li et al., 2019; Godfrey et al., 2021; Yang et al., 2014; Michael et al., 2003). For example, a previous study showed that high-frequency rTMS elevated prefrontal glutamate levels in patients with major depressive disorder using magnetic resonance spectroscopy (Yang et al., 2014). Another study in healthy controls found that high-frequency rTMS could alter glutamate/glutamine ratio in the prefrontal cortex, and these changes were dependent on the pre-stimulation glutamate/glutamine ratio (Michael et al., 2003). Besides, reduced GABA/Glx ratio was found in the ACC in healthy controls after receiving the prefrontal iTBS protocol (Iwabuchi et al., 2017). Accordingly, we can hypothesize that high-frequency rTMS or iTBS may exert after-effects on episodic memory performance through modulating neuronal oscillations and neurotransmitter levels in the prefrontal cortex.

Herein, we carried out behavioral tests to compare the effects of high-frequency rTMS and iTBS on episodic memory in healthy rats. To further clarify the underlying mechanism, we collected local field potential (LFP) recordings and single-unit recordings, and performed in vivo microdialysis and neurochemical experiments to investigate the effects of the two stimulation modalities on neuronal oscillations and levels of extracellular GABA and glutamate in the anterior cingulate cortex (ACC) which is a subregion of the prefrontal cortex.