# BOLD-Perfusion Coupling during Monocular and  Binocular Stimulation

Authors: Claudine Gauthier; Richard D. Hoge
Journal: International Journal of Biomedical Imaging
 (2008)

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## Abstract

Previous studies have suggested that during selective activation of a subset of the zones comprising a columnar system in visual cortex, perfusion increases uniformly in all columns of the system, while increases in oxidative metabolism occur predominantly in the activated columns. This could lead to disproportionately large blood oxygenation level-dependent (BOLD) signal increases for a given flow increase during monocular (relative to binocular) stimulation, due to contributions from columns which undergo large increases in perfusion with little or no change in oxidative metabolism.  In the present study, we sought to test this hypothesis by measuring BOLD-perfusion coupling ratios in spatially averaged signals over V1 during monocular and binocular visual stimulation. It was found that, although withholding input to one eye resulted in statistically significant decreases in BOLD and perfusion signals in primary visual cortex, the ratio between BOLD and perfusion increases did not change significantly. These results do not support a gross mismatch between spatial patterns of flow and metabolism response during monocular stimulation.


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## Body

## 1. Introduction

Although blood oxygenation level-dependent (BOLD) functional MRI
has assumed a role of great importance in systems neuroscience, our
understanding of factors determining the amplitude and spatial extent of the
BOLD effect under different conditions remains incomplete. Relevant parameters
include baseline values and reactive capacity for cerebral perfusion, oxidative
metabolism, and blood volume. An understanding of how these contribute to the
BOLD response is important, since in general they may vary due to age or
disease, and also depending on the nature of the neural system targeted by an
applied stimulus. In particular, the exact nature and extent of the coupling
between changes in oxidative metabolism and perfusion increases during neuronal
activation is still the subject of debate.
While recent studies have focused on quantification of responses during
nonspecific activation of diffuse regions of sensory and motor cortex [1–4], this topic has also arisen in the context of highly localized
responses in cortical columnar systems [5, 6]. In the present study, we sought
to bridge the gap between these two regimes by looking at the effect of
selective activation of only part of a small-scale cortical columnar system on
the apparent BOLD response observed at a spatial resolution typical of studies
used in human subjects.Early optical imaging studies [7–9] suggested that although
evoked changes in oxidative metabolism exhibit a high degree of spatial
specificity, brain perfusion is regulated on a much coarser spatial scale. If
this is true, then there might be profound implications for the BOLD MRI
signal, especially when measured during manipulations such as monocular
stimulation, which preferentially activates the set of ocular dominance columns
projecting to the stimulated eye. In particular, one might expect the spatial pattern
of perfusion response evoked by stimulation of a single eye to be similar to
that seen during binocular stimulation, despite a substantial reduction in the
metabolic response (compared to binocular stimulation) in columns projecting to
the occluded eye. Since the BOLD signal reflects changes in the level of venous
deoxygenated hemoglobin, this gratuitous hyperperfusion in unstimulated ocular
dominance columns could be expected to result in a higher BOLD signal at a
given level of perfusion increase (considering spatial averages over multiple
columns, which would be applicable at commonly used spatial resolutions in
fMRI).The present study examines joint changes in perfusion and BOLD
signals during monocular and binocular stimulation, to test the hypothesis that
spatial decoupling of flow and metabolic responses during stimulation of only a
partial subset of the columnar regions distributed within primary visual cortex
leads to a significant shift in the ratio between spatially averaged BOLD and
perfusion signals (measured using arterial spin-labeling). By combining
quantitative MRI-based measures of these two physiological quantities, we hope
to provide new insight into the spatial precision with which cerebral blood
flow is regulated, as well as factors which determine BOLD contrast amplitude
in cortical tissues exhibiting columnar organization.


## 2. Methods

### 2.1. Subjects

Eight healthy subjects (five males and three females) 24 ± 2.6 years old, one left eye and hand dominant
(male) and seven right eye and hand dominant, participated in the study. The
subjects did not suffer from any known visual deficits except myopia
(MRI-compatible corrective glasses were fitted in these cases). All gave
informed consent and the project was approved by the Comité mixte d’éthique de
la recherche du Regroupement Neuroimagerie/Québec. Data from two of the
subjects was not analyzed due to the poor quality of the arterial spin-labeling
(perfusion) data.


### 2.2. Visual Stimulation

Subjects were fitted with a neoprene rubber mask which allowed
occlusion of one eye by a removable patch. The patch was applied and removed as
needed between the appropriate scans, by an experimenter, from the back of the
scanner bore.Each scanning session included eight six-minute acquisitions,
during which alternating one-minute blocks of baseline (uniform grey screen
with central fixation point) and one-minute blocks of visual stimulation (black
and white checkerboard reversing contrast at a rate producing four white
periods per second within a square) were presented, starting with baseline.
During each scanning run, the subject received either binocular (B) or
monocular (M) stimulation to their nondominant eye with separate scans
conducted in the following order: B-B-M-M-B-B-M-M. Subjects were instructed to
direct their gaze at the central fixation point throughout all scans. The nondominant
eye was selected for monocular stimulation to maximize the difference in
activation between the monocular and binocular trials given that there may presumably
be more extensive activation of V1 for the dominant eye [10].


### 2.3. MRI Data Acquisition

MRI data acquisition was carried out using a
Siemens Trio 3 Tesla MRI
system, at software revision VA25A. Images reflecting relative
perfusion were acquired using a PICORE/Q2TIPS arterial spin-labeling (ASL)
acquisition [11, 12]. The spatial resolution was 3.4 mm × 3.4 mm on a 64 × 64 matrix,
with 10 slices of 5 mm thickness. Other
sequence parameters included TR/TE/alpha = 2 s/19 ms/90° and TI1/TI2 = 700 ms/1400 ms. A slab
thickness of 200 mm was used, with a 10 mm gap between the top of the label
slab and the most inferior image slice.
The Q2TIPS stop time was 1350 ms. MR signals were received using an
eight-channel receive-only head coil, with excitation and labeling performed
using the system body coil.A T1-weighted structural scan was also acquired for later use in
spatial normalization. These scans were
at 1 mm isotropic resolution, acquired using an MPRAGE sequence with
TI/TR/TE/alpha = 900/2300/2.94/9. Voxel size was 1.0 × 1.0 ×
1.2 mm.


### 2.4. Analysis

Flow and BOLD images were generated using the “surround
subtraction” approach described in Wong et al. [13], reducing artifactual flow
signals during periods of BOLD signal transition. The sequence of flow images
was generated by computing the difference between each image and the average of
the previous and subsequent images. The sequence of BOLD images was computed by
adding each image to the average of its two neighbors. For each run, the effect
and standard error maps were then generated by fitting a linear signal model to
each voxel in the flow and BOLD series. The model consisted of a term
representing the three task epochs in the run convolved with a dual gamma
function including positive response plus undershoot [14], plus a third-order
polynomial. Multiple runs for each subject were then combined using a mixed-effects
model, followed by spatial normalization to the MNI 152 brain and combination
of normalized maps for different subjects again using a mixed-effects model (as
described in Worsley et al. [15]). Individual and multisubject maps were then
thresholded at P = .001 significance with correction for multiple
comparisons using the stat_threshold routine of the fMRIstat software package
[15].Regions of interest (ROIs) were generated using the NeuroLens
software package (www.neurolens.org). Average BOLD statistical maps for each
subject were used to make a V1 ROI by thresholding as described above. Voxels
exceeding threshold in the BOLD map but located outside the banks of the
calcarine sulcus as visualized in the T1-weighted structural scan were manually
edited from the ROI, to ensure that the signals extracted were associated with
primary visual cortex and therefore contained tissue organized as ocular
dominance columns. The effect values were then averaged within the ROI for each
functional scan and tabulated as effect size ± standard error. Values were converted to percent change as
needed by dividing the effect size by the constant (DC) term fit during linear
modeling and multiplying by 100.


## 2.1. Subjects

Eight healthy subjects (five males and three females) 24 ± 2.6 years old, one left eye and hand dominant
(male) and seven right eye and hand dominant, participated in the study. The
subjects did not suffer from any known visual deficits except myopia
(MRI-compatible corrective glasses were fitted in these cases). All gave
informed consent and the project was approved by the Comité mixte d’éthique de
la recherche du Regroupement Neuroimagerie/Québec. Data from two of the
subjects was not analyzed due to the poor quality of the arterial spin-labeling
(perfusion) data.


## 2.2. Visual Stimulation

Subjects were fitted with a neoprene rubber mask which allowed
occlusion of one eye by a removable patch. The patch was applied and removed as
needed between the appropriate scans, by an experimenter, from the back of the
scanner bore.Each scanning session included eight six-minute acquisitions,
during which alternating one-minute blocks of baseline (uniform grey screen
with central fixation point) and one-minute blocks of visual stimulation (black
and white checkerboard reversing contrast at a rate producing four white
periods per second within a square) were presented, starting with baseline.
During each scanning run, the subject received either binocular (B) or
monocular (M) stimulation to their nondominant eye with separate scans
conducted in the following order: B-B-M-M-B-B-M-M. Subjects were instructed to
direct their gaze at the central fixation point throughout all scans. The nondominant
eye was selected for monocular stimulation to maximize the difference in
activation between the monocular and binocular trials given that there may presumably
be more extensive activation of V1 for the dominant eye [10].


## 2.3. MRI Data Acquisition

MRI data acquisition was carried out using a
Siemens Trio 3 Tesla MRI
system, at software revision VA25A. Images reflecting relative
perfusion were acquired using a PICORE/Q2TIPS arterial spin-labeling (ASL)
acquisition [11, 12]. The spatial resolution was 3.4 mm × 3.4 mm on a 64 × 64 matrix,
with 10 slices of 5 mm thickness. Other
sequence parameters included TR/TE/alpha = 2 s/19 ms/90° and TI1/TI2 = 700 ms/1400 ms. A slab
thickness of 200 mm was used, with a 10 mm gap between the top of the label
slab and the most inferior image slice.
The Q2TIPS stop time was 1350 ms. MR signals were received using an
eight-channel receive-only head coil, with excitation and labeling performed
using the system body coil.A T1-weighted structural scan was also acquired for later use in
spatial normalization. These scans were
at 1 mm isotropic resolution, acquired using an MPRAGE sequence with
TI/TR/TE/alpha = 900/2300/2.94/9. Voxel size was 1.0 × 1.0 ×
1.2 mm.


## 2.4. Analysis

Flow and BOLD images were generated using the “surround
subtraction” approach described in Wong et al. [13], reducing artifactual flow
signals during periods of BOLD signal transition. The sequence of flow images
was generated by computing the difference between each image and the average of
the previous and subsequent images. The sequence of BOLD images was computed by
adding each image to the average of its two neighbors. For each run, the effect
and standard error maps were then generated by fitting a linear signal model to
each voxel in the flow and BOLD series. The model consisted of a term
representing the three task epochs in the run convolved with a dual gamma
function including positive response plus undershoot [14], plus a third-order
polynomial. Multiple runs for each subject were then combined using a mixed-effects
model, followed by spatial normalization to the MNI 152 brain and combination
of normalized maps for different subjects again using a mixed-effects model (as
described in Worsley et al. [15]). Individual and multisubject maps were then
thresholded at P = .001 significance with correction for multiple
comparisons using the stat_threshold routine of the fMRIstat software package
[15].Regions of interest (ROIs) were generated using the NeuroLens
software package (www.neurolens.org). Average BOLD statistical maps for each
subject were used to make a V1 ROI by thresholding as described above. Voxels
exceeding threshold in the BOLD map but located outside the banks of the
calcarine sulcus as visualized in the T1-weighted structural scan were manually
edited from the ROI, to ensure that the signals extracted were associated with
primary visual cortex and therefore contained tissue organized as ocular
dominance columns. The effect values were then averaged within the ROI for each
functional scan and tabulated as effect size ± standard error. Values were converted to percent change as
needed by dividing the effect size by the constant (DC) term fit during linear
modeling and multiplying by 100.


## 3. Results

Figure1 shows mixed-effects BOLD and flow maps over all subjects
for monocular and binocular stimulation. Occlusion of input to one eye reduced
the amount of activation detected in extrastriate areas. However, the
significance levels observed within primary visual cortex appeared similar
during both monocular and binocular stimulation.Figure 1
Mixed-effects response maps for BOLD and flow changes in response
to monocular and binocular visual stimulation (n = 6). Spatial extent and
intensity are
greater for binocular stimulation than for monocular, for both BOLD and flow
signals. Thresholded activation maps are overlaid on average T1 maps for the
six subjects.Averaged time course signals for flow and
BOLD are shown in Figure2, revealing the initial BOLD signal overshoot and
poststimulus undershoot commonly observed in visual cortex during checkerboard
stimulation (seen here during both monocular and binocular stimulation). The
flow signal illustrates the lower signal-to-noise ratio generally obtained in
arterial spin-labeling measurements.BOLD and flow signals (expressed as percent change; black = binocular,
green = monocular) in response to monocular and binocular visual stimulation,
averaged over all subjects (6 subjects ×
4 averages per subject = 24 averages per signal). Initial overshoot and
poststimulus undershoot are observed in BOLD signal for both monocular and
binocular stimulation, as is typical for checkerboard stimulation.
(a)(b)The bar graphs in Figure3 show average percent changes in BOLD
and flow signals within the V1 ROIs of all subjects. The average percent change
in BOLD signal for monocular stimulation was 0.93±0.04, which was significantly
(P < .05) less than the percent
change of 1.11±0.05 observed during binocular stimulation. The percent flow
increase measured during monocular stimulation was 29 ± 2, also significantly
less than the percent change of 37 ± 2 observed during binocular stimulation. Expressed
as a percent reduction in the response amplitude, withholding input from one of
the two eyes resulted in a 16 ± 6% decrease in the BOLD response and 19 ± 9%
decrease in the perfusion response.Percent change (±SE in lighter shade of gray) in BOLD and CBF
signals in response to binocular and monocular stimulation. Responses evoked by
binocular stimulation are significantly greater than those produced by
monocular stimulation.
(a)(b)The percent changes in BOLD signal per percent signal increase in
flow (i.e., the quotientΔ%BOLD ÷  Δ%flow) during
monocular and binocular stimulation were found, respectively, to be 0.031±0.004
and 0.030±0.004 (Figure 4). The difference between these ratios was not
statistically significant, failing to support any difference in flow-metabolism
coupling during the two forms of stimulation.Figure 4
Percent change ratios (±SE in lighter shade of gray) for BOLD and
CBF during binocular and monocular stimulation. There is no significant
difference between the ratios for the two forms of stimulation, suggesting a
comparable degree of flow-metabolism coupling throughout V1 regardless of the
columnar fraction activated.


## 4. Discussion

In this study, we examined the coupling between BOLD and CBF
responses in primary visual cortex during monocular and binocular stimulation. We
found that the BOLD and CBF responses to monocular visual stimulation were both
significantly reduced in V1 relative to the responses observed during binocular
stimulation (Figure3). The ratio of BOLD to CBF effect sizes did not differ
significantly between the two stimulation conditions (Figure 4), indicating
comparable coupling between flow and oxidative metabolism regardless of the
columnar fraction that was activated.The results obtained in the present study do not support the
“strong” form of the theory that tissue perfusion is regulated only on a coarse
spatial scale, irrespective of the spatial precision with which metabolism
might change. This notion has been described previously as “watering the entire
garden for the sake of one thirsty flower” [9]. To borrow the garden analogy,
the experiments described here could be described as an attempt to measure the
total water intake of the garden, as well as the runoff of unused water, to
test this hypothesis. Our results are consistent with recent MRI studies
showing that there is in fact sufficient spatial contrast in the perfusion
response, as imaged using arterial spin-labeling, to resolve columnar
structures in the visual cortex [6]. The study by Duong et al. [6] found that
the early negative BOLD response (initial dip) also exhibited a high degree of
spatial localization, whereas the late positive BOLD response was more diffuse.
It is important to remember that the apparent resolution of each signal is
dictated both by the underlying physiological regulatory precision and by the degree to which confounding vascular structures are superimposed on the
pattern of parenchymal activation. Based on our results and those from Duong et
al., it appears likely that the lack of precision in the late BOLD signal is
due primarily to obscuring effects from the macrovascular anatomy, rather than
a diffuse parenchymal BOLD effect. It has been suggested by other authors [16]
that the increase in the apparent precision of the initial dip arises because
BOLD signal increases in large draining veins do not arise until after the
initial transit of blood through the local capillary bed postulated to take
approximately one second.Given that functional signals of interest may exhibit bias due to
vascular anatomy, designating regions of interest using objective criteria is
an important part of quantitative neuroimaging studies. In the present study, the use of
phase-encoded retinotopic mapping to identify V1 in a separate mapping
experiment would have been the optimal approach, since this procedure yields a
set of voxels exhibiting a specific spatial trend in the polar angle or
eccentricity of their projection in visual field that is unlikely to appear in
a large vein. This would have led to excessively long scan sessions, however.
Instead, we used the fact that the optic radiations terminate in the calcarine
sulcus, making it very probable that activated regions lying in this anatomical
zone are in fact part of primary visual cortex. It is still possible that the
BOLD activation maps used to create ROIs based on a simple “activation
minus baseline” contrast could contain a disproportionate contribution
from large draining veins. These veins mix venous outflow from multiple visual
areas, including regions which do not exhibit eye-specific columnar
segregation, diluting any shift in flow-BOLD coupling present specifically in
V1. In a pilot study of six subjects
performed at 1.5T using retinotopic mapping to identify V1 (but performed using
single-slice ASL at 1.5 T), we obtained a virtually identical result [17]. We therefore do not feel that the results of
the present study are substantially impacted by our ROI selection procedure. Moreover,
the relatively large voxel size and intense stimulus used in the present study
yielded diffuse regions of robust activation that did not appear to be limited
to macrovascular responses (as can occur at higher spatial resolutions or with
weaker stimuli).By measuring total flow and BOLD responses in V1 during
activation of different columnar fractions, we were able to achieve high
signal-to-noise ratios (SNRs) compared to studies that have used extremely high
spatial resolution to actually resolve the columns. The purpose of the present
study was to provide insight into two questions: the first is whether there is in fact a
fundamental difference in the spatial precision with which perfusion and
oxidative metabolism is measured; the second was to determine the impact of
partial activation of a cortical columnar system on the BOLD signal
characteristics observed at a customary fMRI spatial resolution. If there is
indeed a profound mismatch in the spatial extent of increases in oxidative
metabolism and flow, this should be apparent in the total average signal over
V1. That none was found suggests that similar extents are likely to be found at
higher spatial resolutions.However, it is notable that while removal of input from one of
the two eyes did result in a reduction of both BOLD and flow signals, the
response decreased by much less than one half. This is consistent with detailed
autoradiographic studies showing that pronounced ocular dominance segregation
is mainly limited to cortical layer IV, with layers II and III actually exhibitinghigher activation during binocular
than monocular stimulation [10, 18]. This is consistent with later human
neuroimaging studies, in which some regions showed higher apparent activity
levels during the appropriate monocular stimulation than during binocular input
[16, 19]. The columnar structure associated with ocular dominance is therefore
most appropriately viewed as reflecting a moderate bias in overall selectivity,
associated primarily with a single cortical sublayer, superimposed on numerous
other modulating influences.In light of the issues discussed above, it is clear that the
columnar segregation of brain activation is not “all or nothing”
during selective stimulation such as monocular or single-orientation
conditions. Much of the early research in this area, performed using optical
imaging methods capable of producing compelling maps of the columnar
architecture, examined the perfusion of orientation domains (e.g., Malonek and
Grinvald [9]) and not ocular dominance
columns although a number of authors have imaged ocular dominance using a
variety of other methods [5, 6, 10, 19, 20].
It would therefore be informative to replicate the present study using
different combinations of oriented stimuli. It is also possible that certain
stimuli might achieve a higher degree of selectivity than the ones used in this
and prior studies. Perhaps under such conditions a small difference in
flow-BOLD coupling might become detectable. Future investigation of this topic
might include the use of different stimuli designed to selectively activate
pathways involved in stereopsis.


## 5. Conclusion

Our results do not support the theory of spatially decoupled
responses in blood flow and oxidative metabolism during activation of a subset
of cortical ocular dominance columns. The limited impact of monocular blockade
on flow and BOLD response amplitudes is also demonstrated, and should serve as
a caution that ocular dominance contrast is likely to be faint in hemodynamic
imaging methods.



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*Source: 628718-2008-03-02.xml*