Three experiments were conducted. In Experiments 1 and 3 we studied 11 healthy, right-handed volunteers (mean age = 36.4 years, s.d. = 9.9 years, range = 26–57 years; 8 male and 3 female). In Expt 2 we studied 10 healthy, right-handed volunteers (mean age = 36.2 years, s.d. = 11.8 years, range = 23–57 years; 9 males and 1 female), eight of whom also participated in Experiments 1 and 3. Handedness was confirmed using the Oldfield Handedness Inventory (Oldfield, 1971). Subjects were recruited through advertisements in the community and postings within the hospital. All subjects gave their written informed consent and the protocol was approved by the University Health Network Research Ethics Board in accordance with the declaration of Helsinki on the use of human subjects in experiments. Exclusion criteria included a self-reported comorbid medical illness or a history of drug or alcohol abuse.
Surface EMG was recorded from the right and left first dorsal interosseous (FDI) muscles with disposable disc electrodes placed in a tendon-belly arrangement over the bulk of the FDI muscle and the first metacarpo-phalangeal joint. The subject maintained relaxation throughout the experiment and the EMG was monitored on a computer screen and via speakers at high gain. The signal was amplified (Intronix Technologies Corporation Model 2024F, Bolton, Ontario, Canada), filtered (band-pass 2 Hz to 5 kHz), digitized at 5 kHz (Micro 1401, Cambridge Electronics Design, Cambridge, UK) and stored in a laboratory computer for offline analysis.
This study involved three experiments. The first experiment examined the effects of test MEP size on SICI, LICI, ICF and IHI. The second experiment examined the effects of IHI on SICI and ICF. The third experiment examined the effects of LICI on IHI.
TMS of the left motor cortex was performed with a 7 cm figure-of-eight coil and four Magstim 200 stimulators (The Magstim Company, Dyfed, UK) connected via three Bistim modules in a ‘pyramid’ setup. The output of each of the two pairs of Magstim 200 stimulators was connected to one Bistim module. The output from the two Bistim modules was directed to a third Bistim module that was connected to the TMS coil. This setup allowed us to deliver up to four pulses of different stimulus intensities through the same coil at very short interstimulus intervals. The power attenuation of the pyramid system is about 15 %, similar to a single Bistim system (personal communication, Dr R. Jalinous, Magstim Company). The coil was placed at the optimal position for eliciting motor-evoked potentials (MEPs) from the right FDI muscle. The optimal position was marked on the scalp with a felt pen to ensure identical placement of the coil throughout the experiment. The handle of the coil pointed backwards and was perpendicular to the presumed direction of the central sulcus, about 45 deg to the midsagittal line. The direction of the induced current was from posterior to anterior and was optimal to activate the motor cortex transsynaptically (Werhahn et al. 1994; Kaneko et al. 1996).
TMS of the right motor cortex was performed with a 7 cm figure-of-eight coil and a Magstim Super Rapid stimulator. This stimulator produced bi-phasic current in the coil. The coil was placed at the optimal position for eliciting MEPs from the left FDI muscle. Stimulus intensity was set at 75 % of maximum stimulator output. The handle of the coil pointed forward and laterally about 45 deg to the midsagittal line. This orientation was chosen because in some subjects it was not possible to place both coils at the optimal positions with the handle pointed backwards and laterally due to the size of the coil. Previous studies in 11 normal subjects in our laboratory found no difference in the IHI between 75 % and 90 % of the stimulator output and between four coil orientations 90 deg apart (Yung & Chen, 2001).
This section explains the various parameters used in the experiments. The MT is expressed as a percentage of maximum stimulator output and was defined as the lowest intensity that produced MEPs of > 50 μV in at least five out of ten trials with the muscles relaxed. SICI and ICF were tested using paired-TMS with a subthreshold CS preceding a suprathreshold TS. CS2 denotes a conditioning stimulus that occurred 2 ms prior to a TS and CS10 denotes a conditioning stimulus that occurred 10 ms prior to a TS. CS2 was chosen because it consistently leads to SICI (Kujirai et al. 1993; Chen et al. 1998) and largely avoids the phenomenon of I-wave facilitation (Ziemann et al. 1998; Chen & Garg, 2000), which may obscure SICI (Awiszus et al. 1999). CS10 was chosen because it consistently gives rise to ICF (Kujirai et al. 1993; Ridding et al. 1995). LICI was tested with the suprathreshold CS and TS (Valls-Sole et al. 1992). The CS precedes the TS by 100 ms and is termed CS100. CS100 was used because at this interval direct recording of corticospinal waves demonstrated reduced cortical excitability (Nakamura et al. 1997; Chen et al. 1999) without any change in spinal excitability (Fuhr et al. 1991). IHI was tested with a suprathreshold CS delivered to the left motor cortex followed by a suprathreshold TS delivered to the right motor cortex 10 ms later. This CS will be referred to as CCS10 (contralateral conditioning stimulus). CCS10 was chosen because it consistently leads to IHI (Ferbert et al. 1992).
In all experiments the intensities of the TS were often adjusted to produce a target MEP size. An intensity of ‘TS 1 mV’ indicates a stimulator setting (determined to the nearest 1 % of the maximum stimulator output) that produces a peak-to-peak MEP amplitude of ≥ 1 mV in at least 5 out of 10 trials. Similarly, ‘TS 0.2 mV’ and ‘TS 4 mV’ indicate settings that produce peak-to-peak MEP amplitudes of ≥ 0.2 mV and ≥ 4 mV in at least 5 out of 10 trials, respectively.
In Experiments 2 and 3, we compared the effects two inhibitory mechanisms together to that of one inhibitory mechanism alone. If we used the same test intensity throughout, the first inhibitory mechanism would decrease the test MEP amplitude upon which the second mechanism could operate. In order to match for test MEP amplitude, therefore, in some trials we increased the test stimulus intensity such that it would give a 1 mV test MEP in the presence of the first inhibitory mechanism. We then compared the effects of the second inhibitory mechanism on this 1 mV MEP to a 1 mV MEP that was elicited by a weaker single test pulse. Since both test MEP amplitude and test pulse intensity may be important in determining the degree of inhibition but it is not possible to match them at the same time, we designed our protocols to match for test MEP amplitude and test pulse intensity in different trials.
Experiment 1: effects of test stimulus intensity on SICI, ICF, LICI and IHI
In this experiment we examined the effects of different TS intensities on SICI, ICF, LICI and IHI. For SICI (CS2) and ICF (CS10), the intensity of the CS was set to 80 % of the MT (0.8 MT). For LICI the intensity of the suprathreshold CS100 was adjusted to produce a peak-to-peak MEP amplitude of about 1 mV and for IHI the CCS10 was set at 75 % of stimulator output. Each run consisted of 10 trials each of TS alone and four conditions with the conditioning stimulus preceding the test stimulus at different intervals (CS2, CS10, CS100, CCS10) delivered in random order. The time between trials was five seconds. Three TS intensities (TS 0.2 mV, TS 1 mV, and TS 4 mV) were studied in separate runs.
Experiment 2: effects of IHI on SICI and ICF
Here we investigated whether SICI and ICF are altered by IHI. Ten conditions were tested and are listed in Table 1 as 2A-2J. Each run consisted of 10 trials of each of the 10 conditions delivered in a random order (100 trials). Conditions 2A-2D were used to determine SICI, ICF and IHI for a 1 mV test MEP. Since IHI inhibits the test response, and SICI and ICF may be altered by an attenuated test MEP, for conditions 2E-2J the strength of the test stimulus was adjusted to produce 1 mV MEPs in the presence of an earlier CCS10 pulse. This test stimulus is referred to as ‘TS 1 mVCCS10'. This allowed us to match MEP amplitudes to produce a similar degree of corticospinal activation with and without preceding a CCS10. SICI and ICF in the presence of IHI were studied using three pulses in conditions 2I and 2J. We also measured SICI and ICF with the increased TS strength (TS 1 mVCCS10) in conditions 2F and 2G. Therefore, we designed this experiment to compare SICI and ICF in the presence of IHI (2I/2H and 2J/2H) to SICI and ICF in the absence of IHI matched for test MEP amplitude (i.e. TS 1 mV; 2B/2A and 2C/2A) and TS intensity (i.e. TS 1 mVCCS10; 2F/2E and 2G/2E).
Table 1. Stimulus conditions used in Expts 2 and 3
|3C||1 mV||—||—||—||1 mV|
|3F||1 mV||—||—||—||1 mVCS100|
|3G||1 mV||75%||—||—||1 mVCS100|
Experiment 3: effects of LICI on IHI
In this experiment we investigated the effects of LICI on IHI. Seven conditions were tested and are listed in Table 1 as 3A-3G. Each run consisted of 10 trials of each of the 7 conditions delivered in a random order (70 trials). LICI and IHI for a 1 mV test MEP were determined from conditions 3B and 3C. Since IHI may be affected by test MEP amplitude and CS100 inhibits the test MEP, the strength of the test stimuli was adjusted to produce 1 mV MEPs in the presence of the CS100 pulse in conditions 3D-3G. This test pulse is referred to as ‘TS 1 mVCS100'. The interaction between IHI and LICI were studied using three pulses in condition 3G. Therefore, we designed this experiment to compare IHI in the presence of LICI (3G/3F) to IHI in the absence of LICI matched for test MEP amplitude (i.e. TS 1 mV) (3B/3A) and test stimulus intensity (i.e. TS 1 mVCS100; 3E/3D).
The peak-to-peak MEP amplitude for each trial was measured offline. Inhibition or facilitation was expressed as a ratio of the conditioned to mean unconditioned MEP amplitude for each subject. Ratios less than one indicate inhibition, and ratios greater than one indicate facilitation. Values are expressed as mean ± standard deviation (s.d.).
For Expt 1, the effects of TS intensity on SICI, LICI, ICF and IHI were evaluated by repeated-measures analysis of variance (ANOVA). If the effect of TS intensity was significant, Fisher's Protected Least Significant Difference (PLSD) post hoc test was used to detect differences among different TS intensities. Correlations between SICI and IHI were tested by Pearson product-moment correlation coefficients. In addition, it was found that the distribution for IHI values violated the assumptions of normality and homogeneity of variance and, therefore, was log transformed. For Expt 2, SICI and ICF alone at different test stimulus intensities (TS 1 mV and TS 1 mVCCS10) and in the presence of IHI were compared using repeated-measures ANOVA. For Expt 3, IHI alone at different test stimulus intensities (TS 1 mV and TS 1 mVCS100) and in the presence of LICI was compared using repeated-measures ANOVA. The threshold for significance was set at P < 0.05.