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NeuroImage 21 (2004) 236 – 243
Reduced BOLD response to periodic visual stimulation
Laura M. Parkes, a,* Pascal Fries, a,b Christian M. Kerskens, a and David G. Norris a
a
b
F.C. Donders Centre for Cognitive Neuroimaging, University of Nijmegen, 6525 EK Nijmegen, The Netherlands
Department of Biophysics, University of Nijmegen, 6525 EZ Nijmegen, The Netherlands
Received 12 May 2003; revised 12 August 2003; accepted 14 August 2003
The blood oxygenation level-dependent (BOLD) response to entrained neuronal firing in the human visual cortex and lateral
geniculate nuclei was investigated. Periodic checkerboard flashes at a
range of frequencies (4 – 20 Hz) were used to drive the visual cortex
neurons into entrained oscillatory firing. This is compared to a
checkerboard flashing aperiodically, with the same average number
of flashes per unit time. A magnetoencephalography (MEG)
measurement was made to confirm that the periodic paradigm
elicited entrainment. We found that for frequencies of 10 and 15 Hz,
the periodic stimulus gave a smaller BOLD response than for the
aperiodic stimulus. Detailed investigation at 15 Hz showed that the
aperiodic stimulus gave a similar BOLD increase regardless of the
magnitude of jitter (F17 ms compared to F33 ms), indicating that
flashes need to be precise to at least 17 ms to maintain entrainment.
This is also evidence that for aperiodic stimuli, the amplitude of the
BOLD response ordinarily reflects the total number of flashes per
unit time, irrespective of the precise spacing between them,
suggesting that entrainment is the main cause of the BOLD
reduction in the periodic condition. The results indicate that, during
entrainment, there is a reduction in the neuronal metabolic demand.
We suggest that because of the selective frequency band of this
effect, it could be connected to synchronised reverberations around
an internal feedback loop.
D 2003 Elsevier Inc. All rights reserved.
Keywords: Blood oxygenation level-dependent response; Entrained neuronal firing; Human visual cortex
Introduction
The aim of this work is to establish if there is a difference in
the neuronal metabolic rate during oscillatory neuronal firing
(bursts at a fixed frequency) in comparison to random neuronal
firing (jittered around a central frequency).
Repetitive stimulation by a periodically flickering light causes
visual cortex neurons to become entrained. The neurons synchronise their firing to the frequency of the flickering light leading to
strong EEG responses at that frequency (Herrmann, 2001). Of
* Corresponding author. F.C. Donders Centre for Cognitive Neuroimaging, University of Nijmegen, Trigon 181, NL-6500 HB, Nijmegen,
The Netherlands. Fax: +31-24-3610989.
E-mail address: [email protected] (L.M. Parkes).
Available online on ScienceDirect (www.sciencedirect.com.)
1053-8119/$ - see front matter D 2003 Elsevier Inc. All rights reserved.
doi:10.1016/j.neuroimage.2003.08.025
course, repetitive aperiodic stimulation will also produce synchronous firing with a frequency profile reflecting that of the stimulus.
However, periodic stimulation also produces entrainment, where
the bursts of firing increase in amplitude over the first few hundred
milliseconds of stimulation, and become more tightly locked to the
driving frequency. Multi-unit activity (MUA) recordings in the cat
(Rager and Singer, 1998) detailed the effects of repetitive stimulation for a range of flicker frequencies from 2 to 50 Hz. At 2 Hz,
the response was similar to that of a single flash, with brief bursts
in the first 100 ms (the phasic response), a period of reduced firing
and then large bursts between 200 and 600 ms (the tonic response).
For higher frequencies, the initial 200 – 300 ms of stimulation was
characterised by firing bursts of variable amplitude and an ongoing
sustained response component. Following this, entrainment
emerged with regular, stimulus-locked bursting patterns. During
entrainment, phasic bursts increased in amplitude and the tonic
response was suppressed. The amplitude of the MUA response at
the driving frequency shows peaks between 4 and 8 Hz, 16 and 30
Hz, and 30 and 50 Hz, indicating enhanced entrainment. This is in
accordance with earlier single-cell recordings in the macaque
monkey (Foster et al., 1985), which show a similar peak response
between 4 and 8 Hz to a drifting sine-wave grating. Human EEG
recordings in response to flicker stimuli (Herrmann, 2001) show
increased power in the steady-state potentials for driving frequencies between 6 and 20 Hz, with a weaker peak around 40 Hz.
Differences in neuronal metabolic rate can be measured by
the amplitude of the blood oxygenation level-dependent (BOLD)
response, using MRI (Ogawa et al., 1993). The BOLD signal is
sensitive to changes in local blood flow, blood volume, and
oxygen consumption as a result of neuronal activation. Differences in the BOLD amplitude between regions could reflect
differences in the haemodynamic coupling and/or differences in
neuronal activity. However, modulations of the BOLD amplitude
in a particular region can be assumed to reflect modulations in
the underlying metabolic demand due to changes in neural
activity. A number of previous studies have looked at the
temporal frequency tuning of neurons using BOLD measurements (Ozus et al., 2001; Singh et al., 2003; Thomas and
Menon, 1998) and PET (Fox and Raichle, 1984; Mentis et al.,
1997). The studies all show a general increase in response
amplitude up to around 8 Hz, followed by either a plateau or
a decrease for higher frequencies. Our study differs from these
in that, rather than considering the response to periodic stimuli
with increasing frequency, we focus on the difference in re-
L.M. Parkes et al. / NeuroImage 21 (2004) 236–243
237
jitter is also investigated. A magnetoencephalography (MEG)
recording is included to confirm that the periodic stimulus is
producing entrainment.
Materials and methods
Experiment 1
Fig. 1. Experiment 1: Periodic and aperiodic stimuli over a range of
frequencies. There are two stimuli: the checkerboard with fixation cross (a)
which always appears for a duration of 16.7 ms, and the fixation cross on a
black background (b). The fixation cross randomly changes colour between
red, green, and blue. The presentation scheme for the aperiodic and periodic
conditions is shown in (c), where the black bars represent a 16.7-ms flash of
the checker stimulus. The two conditions consist of the same number of
checkerboard flashes separated by different intervals of a black screen with
a fixation cross. The mean inter-flash time for the aperiodic condition is the
same as for the periodic condition.
sponse between periodic and aperiodic stimuli having a constant
stimulus duration.
In this work, we use a periodically flashing checkerboard
stimulus to produce oscillatory entrainment in the human visual
cortex. This is compared to random bursts of neuronal firing
induced by a checkerboard flashing aperiodically, with the same
average number of flashes per unit time. If the response to each
flash is the same, the two conditions should give the same average
BOLD response. However, it is expected that the periodic condition will produce neuronal entrainment, giving firing bursts of
greater amplitude and reduced tonic inter-flash activity than the
aperiodic condition. The BOLD amplitude change is measured for
both periodic and aperiodic stimuli at a range of frequencies (4 –
20 Hz). The robustness of entrainment to the magnitude of the
Six subjects took part in this experiment on the MR scanner
(21 – 42 years of age, one female), all with normal or correctedto-normal vision. The stimulus (see Fig. 1) consisted of 60 s of
fixation cross on a black background followed by 20 s of flashing
checkerboard pattern. The long rest period of 60 s is required for
the BOLD signal to fully return to baseline between trials, so that
the BOLD signal increase is comparable. There were two options
for the flashing checkerboard: 1: periodic, where the screen
alternated between 16.7 ms (the screen refresh time) of checkerboard and a black screen of a fixed interval, and 2: aperiodic,
where the screen alternated between 16.7 ms of checkerboard and
a black screen of a random interval, but mean interval the same
as the periodic condition. These two conditions were presented
twice each, for a range of seven mean frequencies (4, 6, 7.5, 10,
12, 15, 20 Hz), giving a total time of 2240 s. A fixation cross
remained on throughout the experiment and randomly changed
colour between red, blue, and green. The subject was instructed
to fixate on the cross and respond to the colour changes with a
choice of two button presses. This task was included to maintain
attention at fixation throughout the experiment. Following this
functional scan, an anatomical scan was performed.
The experiment was repeated on one subject (27 years of age,
female) using MEG with a slight change to the stimulus scheme:
Without the slow BOLD signal return to baseline, the long 60-s rest
period is no longer required, so the rest and activation periods were
10 s each. Each condition was repeated 4 times for all seven
frequencies, giving a total time of 1120 s. This MEG measurement
is included to establish the different neuronal firing patterns during
the aperiodic and periodic conditions. The measurement is only
performed on one subject, since the results are unlikely to vary
Fig. 3. Typical regions of interest in the visual cortex and LGN. The three images on the left show a typical region of interest for the visual cortex, in the sagittal
(top left), coronal (top right), and transverse (bottom) planes. The threshold was chosen so that the volume of the activated region was approximately 10 cm3.
The three images on the right show a similar view for the LGN. The threshold was increased to include a total volume of approximately 0.5 cm3 for both right
and left LGN together.
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fundamentally between subjects, and the results are not intended to
be directly compared to the MRI data.
produced using the ‘Presentation’ software package (Neurobehavioral Systems, Inc.).
Experiment 2
MRI scanning procedure
The results of Experiment 1 in the MR scanner showed that the
maximum difference between the two conditions (aperiodic and
periodic) occurs at 15 Hz. This frequency was chosen for further
investigation into the nature of neuronal entrainment. Two subjects
(one male 26 years of age, and one female 27 years of age) took part
in the experiment on the MR scanner. The stimulus scheme is
shown in Fig. 2. Three stimulus conditions are used to establish how
exact the periodicity of the stimulus needs to be to achieve
entrainment. For example, it is possible that entrainment is robust
to a small jitter away from the periodic condition. Fig. 2 shows the
three stimulus conditions. The black bar represents a checkerboard
flash and the numbers represent the length of the inter-flash gap, in
units of screen refresh time. At 15 Hz, the periodic condition has a
constant gap, of 50 ms (3 screen refresh times). The conditions
‘jitter narrow’ and ‘jitter wide’ have a non-standard (i.e. not 50 ms)
gap, every alternate gap. ‘Jitter narrow’ allows every other gap to be
either 2 or 4 screen refresh times and ‘jitter wide’ allows every other
gap to be 1, 2, 4, or 5 screen refresh times. This should establish if
entrainment is destroyed by both a small and large jitter away from
the mean gap length.
Each condition was presented for 10 s followed by a 10-s rest
period showing only the fixation cross. As in Experiment 1, the
fixation cross remained on throughout the experiment and
randomly changed colour between red, blue, and green. The
subject was instructed to fixate on the cross and respond to the
colour changes with a choice of two button presses, to maintain
attention at fixation throughout the experiment. Each condition
was presented 5 times, and the order of the conditions was
randomised. With an initial rest period of 20 s, this took a total
time of 320 s. Following an anatomical scan, the functional scan
was repeated; another 320 s. This procedure was repeated 6
times (on 4 different days) giving a total of 12 runs of 320 s for
each subject.
All experiments were performed using a 3T Trio whole body
scanner (Siemens, Erlangen, Germany). A birdcage resonator was
used for RF transmission and signal reception. The maximal
gradient strength was 40 mT/m. Images were acquired using a
gradient-echo EPI sequence (TR 1 s, TE 40 ms, bandwidth 1860
kHz, flip angle 65j) with 10 slices (22.4 cm FOV, 64 64
matrix, 3.5 mm thickness, giving isotropic voxels of 3.5 mm3)
positioned through the visual cortex and the lateral geniculate
nuclei (LGN). This gives a standard BOLD signal. A 3D
MPRAGE sequence with 1 mm3 isotropic resolution was used
for the anatomical scan.
Stimulus equipment
The stimulus (Fig. 1) was presented on a screen of width 40 cm,
height 30 cm, and a distance 80 cm from the subjects’ eyes. The
projector screen refresh rate was set at 60 Hz. The stimulus was
fMRI data analysis
The fMRI data were processed using the BrainVoyager software package (Brain Voyager 2000, Rainer Goebel and the Max
Planck Society). The first eight images (8 s) of each functional
data set were discarded to allow time for the longitudinal magnetization of the water protons to reach equilibrium. The data were
corrected for the different slice timings and baseline drift. Functional runs showing sudden signal spikes or drops in the whole
brain baseline signal were excluded. This only occurred during
Experiment 2, for 3 out of the 24 data sets (2 from subject one and
1 from subject 2). The functional data were aligned onto the 3D
anatomical data using coordinate references from the header files.
The alignment was checked by eye and slight adjustments were
made where necessary. For each data set, activated voxels were
identified by correlating the BOLD signal amplitude with a
haemodynamic response function over all flashing checkerboard
periods (both periodic and aperiodic). This produced a fairly large
region of activation within the visual cortex (see Fig. 3). The
threshold was chosen so that the volume of the activated region
was approximately 10 cm3. For Experiment 2, a region of interest
over the LGN was also analysed (the signal to noise in the LGN
was not high enough in Experiment 1). The threshold was
increased to include a total volume of approximately 0.5 cm3
for both right and left LGN together. The signal time course over
each activated region was recorded. In future studies, it may be
interesting to consider the response in individual visual areas.
Fig. 2. Experiment 2: Narrow and wide aperiodic stimuli at 15 Hz. In this experiment, the rest period (black screen with fixation cross) and activation period are
10 s each. There are three conditions for the activation blocks as shown above. The black bars indicate a checker stimulus of 16.7 ms (Fig. 1a), with the fixation
stimulus (Fig. 1b) shown at all other times. The numbers indicate the length of the gaps between the checkerboard flashes in number of screen refresh times
(one screen refresh time = 16.7 ms). The periodic condition shows there is a fixed gap of 50 ms between each checker flash. For the aperiodic conditions, ‘jitter
narrow’ and ‘jitter wide’, every alternate gap is a non-standard length. For ‘jitter narrow’, this can be either 2 or 4 screen refresh times, and for ‘jitter wide’, the
choice is wider, from 1, 2, 4, or 5 screen refresh times.
L.M. Parkes et al. / NeuroImage 21 (2004) 236–243
239
Results
Experiment 1
Fig. 4. BOLD signal change averaged over six volunteers for the periodic
and aperiodic stimuli. The error bars represent the standard error of the
mean. The difference in BOLD signal between two conditions was
significant at 10 Hz (P = 0.03) and 15 Hz (P = 0.01).
For Experiment 1, the average percentage BOLD increase over
10 s was calculated for each condition (seven frequencies, periodic
and aperiodic), starting 10 s after the onset of stimulation. Baseline
was taken as the mean signal during 30 s of rest preceding the
stimulus. The results were averaged over the two trials of each
condition. For each subject, the percentage BOLD increase for each
condition was normalised to the maximum increase found for any of
the stimulus conditions. The mean normalised BOLD increase was
found for each stimulation condition, averaging over all six subjects.
To test for significant differences between the periodic and aperiodic
conditions, a paired t test (two-tailed distribution) was performed
using these six normalised values at each frequency.
For Experiment 2, the average percentage BOLD increase over
5 s was calculated for each condition (jitter narrow, jitter wide, and
periodic), starting 9 s after the onset of stimulation. A four-point
baseline was used, taken from 2 s before the start of stimulus
presentation to 1 s after. The results were averaged over the five
trials of each condition, giving a total of 10 (subject 1) or 11
(subject 2) values for each condition (1 from each data set). To test
for significant differences between the response to any two conditions, a paired t test (two-tailed distribution) was performed using
these 10 or 11 values of the conditions under test. The mean signal
difference and 95% confidence intervals were also found. Finally,
the signal curves from each condition were averaged to give a
mean signal response from all of the trials.
MEG procedure and analysis
MEG was recorded with a whole-head CTF system using 151
axial gradiometers (CTF Systems, Port Coquitlam, Canada). Similar stimulus equipment was used as in the fMRI experiment, with
stimulus projection onto a screen roughly 1 m from the subjects’
eyes. Signals were low-pass filtered at 150 Hz and sampled at 600
Hz. Analysis of the MEG data was performed using MatLab
software developed in-house. The first second of each stimulation
episode was omitted to allow build up of the steady-state visual
evoked field. The remaining 9 s were first tapered using discrete
prolate spheroidal sequences (Slepian sequences) (Mitra and
Pesaran, 1999), padded out to 10 s length, Fourier transformed,
and squared to calculate the power spectra. Power spectra were
produced for all 151 sensors for each stimulus condition. For
topographic plots, power was taken at the stimulus flicker frequency (or the equivalent flicker frequency for the jittered stimuli).
Fig. 4 shows the BOLD signal change averaged over all six
subjects for each condition with increasing frequency. For frequencies of 8 Hz and below, the two stimulation conditions give
very similar increases in BOLD amplitude. Between 10 and 15
Hz, the aperiodic stimulus gives a higher BOLD signal change
than for the periodic stimulus. A paired t test (two-tailed
distribution) between the normalised amplitude increases for the
periodic and aperiodic condition over all six subjects showed that
the aperiodic signal increase was significant at 15 Hz ( P = 0.01,
signal increase = 0.13 F 0.07 (95% confidence interval)) and 10
Hz ( P = 0.03, signal increase = 0.09 F 0.05).
Fig. 5 shows a map of the MEG signal over all sensors at 15 Hz
for the periodic (Fig. 5a) and aperiodic (Fig. 5b) conditions during
the 15-Hz stimulation. It can be seen that the maximum power for
the periodic condition is found for sensors over the visual cortex.
Fig. 5c shows a power versus frequency plot for the four sensors
circled in Fig. 5a, for both the periodic (red) and aperiodic (blue)
condition. A clear increase in power at 15 Hz can be seen in the
periodic condition, indicating a strong, entrained response at the
driving frequency. Similar results were found at all stimulation
frequencies.
Experiment 2
Fig. 6 shows the average BOLD response curves in the visual
cortex to the three stimulus conditions as described in Fig. 2.
Both subject 1 (Fig. 6a) and subject 2 (Fig. 6b) show that both a
wide and narrow jitter away from the mean gap length give a
similar increased BOLD response compared to the periodic
condition. The results of the statistical analysis are shown in
Table 1. It can be seen that the ‘jitter narrow’ and ‘jitter wide’
conditions gave increased signal compared to the periodic condition for both subjects, although the differences for subject 2 were
more significant. For subject 2, Fig. 6b also shows that the ‘jitter
wide’ condition gave a larger signal response than the ‘jitter
narrow’ condition. However, the results in Table 1 show this to be
non-significant.
Fig. 7 shows the equivalent results from the LGN. Although the
data are noisier, the periodic condition gives the lowest signal
response for both subjects. The results of the statistical analysis in
Table 1 show that the increases in signal for the jittered conditions
are highly significant for subject 2, but for subject 1, only the ‘jitter
narrow’ condition is on the edge of significance (the 95% confidence interval puts the difference just above zero). There are no
significant differences between the ‘jitter narrow’ and ‘jitter wide’
conditions.
In the course of our investigations, we measured the response to
a number of other jitter conditions and found that the periodic
condition gave consistently reduced signal compared to jittered
conditions for both subjects.
This experiment was conducted at 15 Hz since this frequency
showed the maximum difference in response between the periodic
and aperiodic conditions. However, we expect similar results at 10
and 12 Hz since a difference was also found at these frequencies.
The reaction times and error rates of the colour change task
were not significantly different between any conditions for any of
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Fig. 5. MEG data at 15 Hz. Maps of the power distribution over the brain at 15 Hz are shown for the periodic (a) and aperiodic (b) stimulation. The colour bar
indicates power in fT2. A clear increase in power is seen in the occipital cortex for the periodic condition. The mu rhythm (usually strongest at 10 Hz, but can
extend up to 15 Hz) can also be seen in central brain regions for both conditions. (c) Power frequency plot for the four sensors over the activated region, as
indicated in (a). The response to the periodic stimulus (red) shows an abrupt increase in power at the driving frequency (15 Hz), with harmonics at 30 Hz. The
response is square-shaped over a 2-Hz region as a result of the F1-Hz spectral concentration specified in the multi-taper method. The aperiodic condition
(blue) shows a small, diffuse increase in power compared to the periodic condition, spread over 5 – 30 Hz.
Fig. 6. BOLD response to different aperiodic conditions in the visual cortex. (a) Response curves for subject 1 (male) averaged over 50 trials. (b) Response
curves for subject 2 (female) averaged over 55 trials. The conditions correspond to those described in Fig. 2. Statistical analysis is shown in Table 1.
Fig. 7. BOLD response to different aperiodic conditions in the LGN. (a) Response curves for subject 1 (male) averaged over 50 trials. (b) Response curves for
subject 2 (female) averaged over 55 trials. The conditions correspond to those described in Fig. 2. Statistical analysis is shown in Table 1.
L.M. Parkes et al. / NeuroImage 21 (2004) 236–243
241
Table 1
Statistical signal differences between the ‘jitter narrow’, ‘jitter wide’, and periodic conditions
Conditionsa
Subject 1
Subject 2
Visual cortex
mean F CIb (%)
Narrow—periodic
Wide—periodic
Wide—narrow
0.34 F 0.33
0.32 F 0.36
0.02 F 0.34
LGN
Pc
0.07
0.12
0.91
mean F CI (%)
0.20 F 0.20
0.05 F 0.15
0.16 F 0.27
Visual cortex
LGN
P
mean F CI (%)
P
mean F CI (%)
P
0.08
0.58
0.28
0.32 F 0.25
0.51 F 0.26
0.19 F 0.40
0.03
0.003
0.38
0.19 F 0.10
0.27 F 0.18
0.08 F 0.15
0.005
0.01
0.31
a
The two conditions under comparison. For example, (narrow—periodic) considers the signal increase of the ‘jitter narrow’ condition compared to the periodic
condition.
b
The mean difference in signal between the two conditions (given as the absolute difference in % BOLD signal increase from baseline, over 10 data sets
for subject 1 and 11 for subject 2). CI is the 95% confidence interval of the mean.
c
The P value is the result of a paired t test (two-tailed distribution) between the two data sets under consideration. The significant ( P < 0.05) results are
highlighted in bold.
the subjects. This suggests that attention on the fixation cross was
the same for all conditions.
Discussion
The periodic condition gives entrained oscillatory firing
The MEG response to the periodic stimulus (Fig. 5c, red) shows
a strong power increase in the visual cortex at the driving
frequency of the flickering stimulus, in comparison to the aperiodic
condition. This indicates the presence of entrainment, with strong
bursts of firing locked to the stimulus flashes. The increased firing
rate of the stimulus-locked component creates an increase in
synchronous firing, allowing the magnetic fields of each neuron
to add up in phase, causing the increased MEG power. While Fig. 5
shows the results at 15 Hz, similar strong increases in power at the
driving frequency were seen for all frequencies tested. These
results are in agreement with previous findings using LFP (Rager
and Singer, 1998) and EEG (Herrmann, 2001) recordings using
similar flashing stimuli, in which entrainment was seen for a broad
range of frequencies. For the aperiodic stimulus, the MEG response
shows general increased power over a broad frequency band
compared to the periodic condition (Fig. 5c, blue). This is due to
increased firing for the broad range of frequencies present in the
aperiodic stimulus. Although this MEG measurement was only
performed on a single subject, we assume that the entrained
response will not be fundamentally different between subjects.
The BOLD response to ‘jitter narrow’ and ‘jitter wide’ is the
same
Figs. 6a and b show that the average BOLD responses in the
visual cortex to the ‘jitter narrow’ and ‘jitter wide’ conditions are
very similar for both subjects. Table 1 shows that there are no
significant differences between the two conditions. This is evidence that the amplitude of the BOLD response to aperiodic
stimuli ordinarily reflects the total number of flashes per unit time,
irrespective of the precise spacing between them. It is possible that
different flash timings could produce different BOLD amplitudes
due to the non-linear addition of the haemodynamic response for
short interstimulus intervals (Ogawa et al., 2000). The slightly
increased signal in the ‘jitter wide’ condition for subject 2 (Fig. 6b)
could reflect this effect. However, this signal difference is not
statistically significant (Table 1), and cannot therefore be the only
cause of the differences between the jittered and periodic conditions. Likewise, any masking effects, which depend on the gap
length between flashes (Macknik and Livingstone, 1998), do not
appear to have a significant effect. These data also show that
entrainment is destroyed by both the narrow and the wide jitter
conditions, suggesting that the flashes need to be precise to at least
17 ms (one screen refresh time) to maintain entrainment.
Periodic stimuli give reduced BOLD response compared to
aperiodic
Fig. 4 shows that the periodic stimulation gives a reduced
BOLD signal compared to the aperiodic stimulation for frequencies
from 10 to 15 Hz. The two conditions contain the same number of
flashes with the same mean gap length, indicating that the
difference in the BOLD amplitude is due only to the oscillatory
nature of the periodic stimulation. A possible attention difference
between the two stimuli is controlled by the colour change task,
which maintains attention on the fixation cross.
Another possibility is that the difference in the temporal
frequency spectra between the aperiodic and periodic stimuli is
responsible for the different BOLD response. While containing the
same total power as the periodic stimulus, the aperiodic stimulus
contains a broader spread of frequencies. If there is a non-linear
BOLD response to different flicker frequencies, this could cause a
BOLD amplitude difference. However, as shown in Fig. 4, the
response to the periodic stimulus is fairly flat with frequency,
showing a small increase up to 8 Hz followed by a plateau. This is
in good agreement with previous fMRI data, particularly the results
of Ozus et al. (2001), who used a similar stimulus. For frequencies
between 10 and 15 Hz, the aperiodic response is larger than the
periodic response to any frequency. This increased aperiodic
response cannot therefore be caused by the superposition of
periodic responses over a broad range of frequencies. This leaves
entrainment in the periodic condition as the most likely cause of
the BOLD amplitude difference.
Several studies (Arthurs et al., 2000; Gratton et al., 2001;
Logothetis et al., 2001; Ogawa et al., 2000; Singh et al., 2003)
show that the amplitude of the haemodynamic response is coupled
to some sort of average of the underlying neuronal activity. For
example, in the human brain, Arthurs et al. (2000) found a linear
correlation between the evoked potential amplitude and the BOLD
signal for increases in electrical stimulation of the median nerve;
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and in the monkey, Logothetis et al. (2001) showed that the BOLD
amplitude is most strongly correlated to LFP recordings rather than
the MUA or the spike rate. This suggests that the BOLD signal
reflects the synaptic activity due to incoming signals on the
dendrites and soma, and local processing, rather than the energy
required to propagate an action potential. In energetic terms, the
main cost of cell signalling is in restoring ionic balances, most of
which is associated with the production of post-synaptic potentials;
however, 33% is attributed to axonal action potential propagation
(Lennie, 2003). A reduction in BOLD amplitude from the same
region of interest during entrainment could therefore reflect a
reduction in either the number of neurons actively engaged in
the task, or a reduction in their synaptic activity.
As described in the introduction, during entrainment, cat MUA
recordings show an increase in the phasic, stimulus-locked firing
rate and a decrease in the tonic firing rate (Rager and Singer, 1998).
This reduction in tonic firing could be the cause of the reduced
BOLD signal during the periodic stimulus. However, if this were
the case, the amplitude difference might be expected to increase
with increasing stimulation frequency, since the tonic response
becomes more and more suppressed with increasing frequency.
The lack of a BOLD amplitude difference at 20 Hz does not
support this hypothesis.
One explanation for the selective frequency band (10 – 15 Hz)
of the BOLD amplitude difference is that it is due to the broad
tuning of the visual cortex neurons to certain ‘resonance’ frequencies. EEG recordings in the human visual cortex (Herrmann, 2001)
during periodic stimulation showed increased power of steady-state
visual evoked potentials (ssVEPs) around certain resonance frequencies. In the low-frequency range, an average over 10 subjects
showed strong, broad resonance peaks around 10 Hz, starting at
roughly 6 Hz and falling off before 20 Hz. This is in rough
accordance with the frequency band of our observed BOLD
amplitude differences, although a difference might have been
expected at 8 Hz. The increased ssVEP response at these frequencies is thought to be due to the matching of the flash timing with
the temporal characteristics of the neuronal circuits. At the resonance frequency, each stimulus input could reinforce internal
reverberations from an internal feedback loop, increasing the
synchrony of the spikes and therefore the ssVEP. This is supported
by the fact that, in general, neurons have a tendency to engage in
oscillatory activity in certain frequency bands (e.g. 9 – 11 Hz alpha
oscillations in the occipital cortex (Nunez et al., 2001)).
For this resonance activity to result in a reduced BOLD
amplitude, it must involve either a reduced number of active
neurons, or a reduction in their level of synaptic activity, compared
to the non-resonance states. It is possible that a reduced number of
neurons, for example, those involved in the reverbatory loop,
become selectively active for the resonance frequencies, and other
neurons are suppressed.
There is some support in the literature for the observation of
reduced BOLD signal during increases in synchronous activity
around the alpha frequency. A simultaneous EEG/fMRI study
(Goldman et al., 2002) found power in the alpha band (8 – 12
Hz) to be negatively correlated with the BOLD response in the
occipital cortex. Another study (Singh et al., 2002) found increased
BOLD responses to be spatially coincident with decreases in
cortical synchronisation in the 5- to 25-Hz band as measured with
MEG. However, these changes were due to internally generated,
state-dependent oscillations rather than the driven, steady-state
evoked oscillations in our study.
Our finding of a reduced BOLD signal during periodic stimulation is frequency specific. It would be interesting to repeat
Experiment 1 with a projection system with a faster possible flash
rate, to allow testing at higher frequencies. This would determine if
there is a change in the BOLD signal to periodic stimulation
around 40 Hz, which might be expected since the ssVEP response
showed further resonance behaviour at this frequency (Herrmann,
2001). The phenomenon may also be specific to the visual system,
so tests in the auditory and somatosensory domain could be
interesting. The auditory system, in particular, has also shown
resonance behaviour at 40 Hz (Pastor et al., 2002).
Entrainment in the LGN
MRI offers an advantage over EEG and MEG in that the
response to deep brain structures such as the LGN can also be
studied. Fig. 7 shows that entrainment is also present in the LGN,
with the periodic condition giving a consistently lower BOLD
response than the jittered conditions. However, the response curves
are noisier than in the cortex due to the smaller signal change.
Entrainment in the visual cortex is thought to be due to corticocortical or cortico-thalamo-cortical reverberant loops. Several theoretical models show that propagation delays within different
networks can produce oscillations with frequencies in the range
of human EEG measurements (Nunez et al., 2001). In the LGN,
inhibitory neurons in the perigeniculate nucleus are thought to
produce oscillatory activity (Steriade et al., 1993). So, given that
the visual stimulus drives both LGN and visual cortex neurons, and
both systems have the capacity for endogenous oscillations,
entrainment in the LGN is to be expected. Supporting evidence
comes from intracranial electrode recordings in epileptic patients
(Krolak-Salmon et al., 2003), showing ssVEPs in the LGN during
screen flicker.
This work shows that the entrainment of neurons in the visual
cortex and the lateral geniculate nucleus gives a reduced BOLD
response compared to non-entrained firing for flicker frequencies
of 10 and 15 Hz. In a similar manner to Ogawa et al. (2000), our
approach shows how the amplitude of the BOLD response can be
used to probe very fast neural events, despite the sluggishness of
the haemodynamic response.
Acknowledgments
We thank Paul Gaalman for help with the MRI scanning, Ole
Jensen for help with MEG recordings, and Wolf Singer for helpful
comments on this study.
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