Frequency-specific effects of low-intensity rTMS can persist for up to 2 weeks post-stimulation: A longitudinal rs-fMRI/MRS study in rats
Introduction
Repetitive transcranial magnetic stimulation (rTMS) is a non-invasive neuromodulation technique widely applied in therapeutic and investigative studies of neuropsychiatric conditions including depression [1,2], schizophrenia [3], and Parkinson's disease [4]. Despite increasing use of rTMS to modulate dysfunctional brain networks in humans, the mechanisms underlying its therapeutic effects, particularly in cortical circuits, still require elucidation. Furthermore, patients report that therapeutic benefits of rTMS wear off after treatment finishes [e.g., Refs. [5,6]], suggesting an urgent need to characterise the persistence of rTMS effects.
Therapeutic rTMS application utilises different stimulation frequencies to elicit different cortical changes: low-frequency (<5 Hz) stimulation has inhibitory and high-frequency (>5 Hz) has excitatory effects on the cortex, albeit with some individual variability [7,8]. Preclinical models have identified gene and protein expression changes associated with these frequency-specific changes in excitatory and inhibitory circuit activity [[9], [10], [11], [12]]. In addition, changes in neurotransmitters glutamate (Glu), γ-aminobutyric acid (GABA), and glutamine (Gln), the major precursor for neuronal glutamate and GABA [13], are central to excitability changes observed following stimulation [9,14,15]. High-frequency stimulation increases excitatory neuronal activity [16,17] and induces a long-lasting increase in glutamatergic synaptic strength consistent with long-term potentiation (LTP) of excitatory circuits [9], while simultaneously reducing the strength of GABAergic synapses [11,18]. Conversely, 1 Hz stimulation increases inhibitory circuit activity by increasing GABA levels [19] and modulates expression of calcium-binding proteins in inhibitory interneurons [10,12].
Although animal models have revealed some key mechanisms of rTMS, in most preclinical studies, changes in the brain are investigated post-mortem, providing only a snapshot of rTMS-induced changes. Additionally, most animal and human studies focus on events occurring minutes to hours after single or multiple stimulations. Consequently, the development of rTMS effects over time, and the stability of its after-effects, have not been explored.
Non-invasive magnetic resonance (MR) based techniques are ideal for longitudinal studies of brain function and neuroplasticity [20]. Resting-state functional magnetic resonance imaging (rs-fMRI) is the technique of choice for examining long-term changes in functional networks [[21], [22], [23], [24], [25]], while proton (1H) magnetic resonance spectroscopy (MRS) is one of only few methods that can non-invasively assay neurometabolic changes. Rs-fMRI identifies brain regions showing synchronised resting activity which form organised networks called resting-state networks (RSNs) [26]. RSN dysregulations, and accompanying neurometabolic changes, have been identified in patients with neuropsychiatric disorders [27], e.g., schizophrenia and depression [28]. Both RSN functional connectivity (FC) and neurometabolite levels in humans are sensitive to rTMS and clinical improvements are associated with changes in RSNs [for review, see 25]. However, there have been no reports of longitudinal studies to characterise the timecourse of changes in RSNs or neurometabolites during and after rTMS treatment.
A single session of low-intensity rTMS (LI-rTMS) in rats has frequency-specific effects on RSNs similar to those described in humans following rTMS [21]. This evidence of translational validity suggests that LI-rTMS in rodents can be a useful model in a translational pipeline to inform and guide clinical application of rTMS, particularly given the cost and logistical challenges of longitudinal human studies. For example, recent active fMRI/rTMS studies in animal models of traumatic brain injury have shown that repeated delivery of high-frequency rTMS increased primary somatosensory activity in response to forelimb stimulation, indicating restored cortical function [29,30]. The present study used rs-fMRI and MRS in rats to investigate the emergence of LI-rTMS-induced changes in FC and neurometabolite levels respectively and their maintenance for up to three weeks. Two weeks of daily stimulation resulted in significant changes in FC and neurometabolites that outlasted the duration of stimulation. A better understanding of rTMS effects on RSNs and neurometabolites may prove helpful in the development of long-lasting treatment options to modify dysfunctional connectivity detected in several neuropsychiatric disorders.
Section snippets
Animals
Experimental procedures were approved by the UWA Animal Ethics Committee (RA/3/100/1430) in accordance with the Australian code for the care and use of animals for scientific purposes. 18 adult male Sprague-Dawley rats (6–8 weeks old, 150–250 g) from the Animal Resources Centre (Canning Vale, Western Australia) were maintained in a temperature-controlled animal care facility on a 12-h light-dark cycle with food and water ad libitum. Animals were euthanised after the last imaging session using CO
Template RSNs identified by group-ICA
Template rodent RSNs were identified from baseline data to avoid including LI-rTMS-related RSN alterations in the template components and thus increase the sensitivity of dual regression in detecting between-group differences [39]. Based on visual inspection of the 15 components identified by group-ICA, four components (Fig. 3) were identified as classical rodent RSNs [e.g., Refs. [37,[43], [44], [45]]]. The remaining eleven components were classified as noise (see examples in Fig. A1).
Discussion
rTMS therapy is normally delivered to patients as a course of treatment over several weeks to induce lasting plastic changes. However, the stability of these changes remains unknown. This study examined both the emergence and maintenance of changes in FC and neurometabolite levels, assessed with rs-fMRI and MRS respectively, following repeated LI-rTMS in rats. Our study confirms the frequency-specific effects of LI-rTMS and further suggests that effects of 1 Hz stimulation, although milder, may
Conclusion
Information about the duration of the after-effects of rTMS therapy is vital for the development and improvement of rTMS use as a treatment in a clinical setting. Here, we present the first longitudinal rs-fMRI/MRS investigation of the duration of FC and neurometabolic changes induced by repeated LI-rTMS delivery. Our work confirms the frequency-specific effects of LI-rTMS and further suggests that effects of 1 Hz stimulation, although milder, may persist longer after cessation of treatment
Acknowledgements
The authors thank Ms Marissa Penrose-Menz, Dr Alexander Joos, Ms Katherine Fisher and Ms Michelle Carey for their assistance with the experiments and Dr Alexander Tang for critical review of the manuscript. The authors acknowledge the facilities and scientific and technical assistance of the National Imaging Facility, a National Collaborative Research Infrastructure Strategy (NCRIS) capability, at the Centre for Microscopy, Characterisation and Analysis, The University of Western Australia.
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2021, Neuroscience ResearchCitation Excerpt :The group differences found based on the all-data template were then quantitatively compared to the results of the baseline-only template. For this comparison, only the interoceptive network (C1) was used, since the default mode network (C2) in the all-data template overlapped with another network, and for the salience network (C3), no significant changes were found in the previous publication (Seewoo et al., 2019). Since the visual comparison of identified RSNs is somewhat subjective, the quantitative comparison of group differences is a more meaningful approach for assessing the quality of different analysis pipelines.