P 3. Dose-dependence of changes in cortical protein expression induced by theta burst stimulation in the rat

Abstract

Introduction Theta Burst stimulation (TBS) applied via transcranial magnetic stimulation (TMS) is an effective tool to modulate human neocortical excitability ( Huang et al., 2005 ). Repeated application of the same TBS protocol or variation of the number of stimuli has been shown to alter the strength and direction of changes in cortical excitability compared to the standard TBS protocols ( Gentner et al., 2008 , Gamboa et al., 2010 ). TBS applied to rat cortex affected the expression of activity-dependent proteins related to the cortical inhibitory systems, suggesting altered cortical inhibition contributing to the TBS after-effects ( Benali et al., 2011 , Funke and Benali, 2011 ). Objectives Our aim was to investigate the impact of varying numbers of TBS-stimuli applied as multiple blocks of intermittend TBS (iTBS) or continuous TBS (cTBS) on cortical protein expression in the rat, to further our insights in physiological mechanisms underlying TBS-induced changes in cortical excitability. Materials and methods Nine groups of anesthetized rats (male Sprague Dawley, 400–600 g) received TMS. Eight groups received a different number of iTBS/cTBS-blocks summing up to either 600, 1200, 1800, or 2400 stimuli, applied with breaks of 15 min between blocks of 600 stimuli. Sham stimulation (coil more distant to the head) was applied to a control group. Rats were sacrificed for immunohistochemical analysis or western blotting focussing on frontal, motor, sensory and visual cortex. Results In general, quite similar effects for iTBS and cTBS were observed. The expression of the 65-kDa isoform of glutamic acid decarboxylase (GAD65) increased, while that of the 67-kDa isoform (GAD67) and that of the calcium-binding proteins Parvalbumin (PV) and Calbindin (CB) progressively decreased. Also the expression of the immediate early gene c-Fos decreased with an increasing number of blocks. A more detailed analysis, however, revealed that the sensitivity of distinct proteins to stimulation varied with the number of stimuli and type of stimulation. Conclusion Our findings show that both iTBS and cTBS affect the activity of inhibitory interneurons and indicate that repeated TBS elicits no simple accumulative dose-dependent effect for all activity-markers but distinct profiles with threshold characteristics and a waxing-and-waning effect especially for two markers of inhibitory activity, CB and GAD67. Thus, our data do not suggest fundamentally different modulation of the cortical inhibitory systems by iTBS and cTBS. Subtle differences in stimulation after-effects on different neuronal subsystems might contribute to the opposite impact on cortical excitability following iTBS and cTBS and the switching sign of effect with repeated stimulation.