Name: functional transcranial doppler ultrasound for monitoring cerebral blood flow
Uploaded: 2021-03-15T13:30:00.000+00:00
Duration: 9 min 41 s
Description: Watch this Scientific Journal Video about Functional Transcranial Doppler Ultrasound for Monitoring Cerebral Blood Flow at JoVE.com

This project is based on research that was partially supported by the Nebraska Agricultural Experiment Station with funding from the Hatch Act (Accession Number 0223605) through the USDA National Institute of Food and Agriculture.

All human subject research was performed in accordance with the Institutional Review Board of the University of Nebraska-Lincoln, and informed consent was obtained from all subjects.
1. Locating the MCA signal by freehand TCD
NOTE: &#8220;Freehand&#8221; TCD refers to operation of TCD with a handheld transducer to find a CBFV signal before beginning an fTCD experiment.
<ol>
	<li>Setting TCD parameters
	<ol>
		<li>Keep the power at a reasonably high value (e.g., 400 mW) during the initial search for the MCA. Once the MCA signal is located, reduce the power as much as possible while still maintaining a &#8220;good&#8221; signal (see step 2.2.7). 
		NOTE: Using a reasonably high power during the initial search does not violate the &#8220;As Low As Reasonably Achievable&#8221; (ALARA) principle of exposure to acoustic radiation because higher power will allow the MCA signal to be discovered more quickly10.</li>
		<li>Set the sample volume to 8&#8211;12 mm during the initial search for the MCA signal. If the signal is difficult to find, increase the gate size to increase the intensity of the signal, but note that this may incorporate the signal from one or more nearby arteries into the signal from the MCA.</li>
		<li>Set the gain at a medium level, with the goal of &#8220;keeping background noise at a minimum, but present&#8221;10.&#160;</li>
		<li>Set the high-pass filter cutoff (normally termed &#8220;threshold&#8221;) to 50&#8211;150 Hz.&#160;</li>
		<li>If the subject is an adult, set the depth to 50 mm, which is the average mid-point depth of the M1 segment of the MCA10 (Figure 1).&#160; 
		&#8203;NOTE: This setting will be discussed in more detail in subsequent steps. Depth settings for children are given in Table 1.</li>
	</ol>
	</li>
</ol>
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/62048/62048fig1.jpg" /> 
Figure 1: Representation of the circle of Willis and the major arteries of the cerebral circulatory system. The bifurcation of the ICA into the ACA and MCA is marked with a black circle. The M1 segment of the MCA is shown. This figure has been modified from24. Abbreviations: ACA = anterior cerebral artery; Bif. = bifurcation; ICA = internal carotid artery; MCA = middle cerebral artery. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/62048/62048fig1large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<ol start="2">
	<li>Locating the temporal window 
	NOTE: The temporal window, also called the transtemporal acoustic window, is a part of the skull where the bone is thinnest11, thus allowing transmission of low-frequency ultrasound energy through the cranium (Figure 2).
	<ol>
		<li>For infants and small children, locate the temporal window just in front of the ear (the &#8220;intertragal space&#8221;) and above the rostral edge of the zygomatic arch, which can be easily felt under the skin.&#160;</li>
		<li>For teenagers and young adults, locate the temporal window via any of the subwindows.&#160; 
		NOTE: The posterior subwindow usually provides the best signal (Figure 2).</li>
		<li>For adults aged 30 years or older, locate the temporal window just in front of the ear. 
		NOTE: The acoustic window decreases in size as people age due to increasing porosity of the bone of the cranium, causing some older people to have a very limited temporal window12. In such individuals, bilateral insonation of the MCA is sometimes impossible.</li>
	</ol>
	</li>
</ol>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/62048/62048fig2.jpg" /> 
Figure 2: The transtemporal window (marked by the dashed ellipse), zygomatic arch (arrow), and subwindows11. (A) Frontal subwindow. (B) Anterior subwindow. (C) Middle subwindow. (D) Posterior subwindow.&#160;<a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/62048/62048fig2large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<ol start="3">
	<li>Applying the transducer
	<ol>
		<li>Apply enough ultrasound gel to cover the surface of the transducer.&#160; 
		&#8203;NOTE: When placed on the head, the gel should cover sufficient space to maintain a seal between the scalp and the Doppler probe&#8217;s surface, thus preventing signal interruption from air coupling underneath the probe&#8217;s surface.</li>
		<li>Alert the subject that the gel may feel cold (if at room temperature).</li>
		<li>Place the transducer on the temporal window, which was located in section 1.2.</li>
	</ol>
	</li>
	<li>Searching for the MCA
	<ol>
		<li>After placing the transducer on the scalp, search for the MCA signal, which will generally be located slightly anterior (forwards) and rostral (towards the head) from the location of the initial transducer scalp placement10.</li>
		<li>If the TCD spectral signal is not immediately obvious, adjust the angle of the transducer while keeping it in the same location relative to the scalp. Slowly angle the probe from rostral to caudal (towards feet) and posterior to anterior.&#160; 
		NOTE: Figure 3 shows two spectra taken from the same position, but at different angles.</li>
		<li>If a signal is still absent after performing step 1.4.2, check the color M-mode display for flow in the MCA at different depths (indicated by red coloring). Increment or decrement the signal depth in 5 mm steps and search as described in step 1.4.2. If flow is visible in M-mode but not in the Doppler spectrum, increase or decrease the depth until the flow signal is visible in the Doppler spectrum.</li>
		<li>If a satisfactory signal is still not obtained, move the transducer to a nearby position on the scalp, which is slightly more anterior, and repeat steps 1.4.1&#8211;1.4.3.&#160;</li>
		<li>When an optimal MCA signal is obtained, note the depth and maximum velocity.</li>
		<li>Using a washable makeup pen, place a mark on the scalp (trace part of the transducer edge) where the optimal signal was found.&#160;</li>
	</ol>
	</li>
</ol>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/62048/62048fig3.jpg" /> 
Figure 3: Sample Doppler spectra and M-mode images from midpoint of M1 segment of the MCA. (A) Spectrum taken right after applying transducer to the temporal window, just in front of the ear. (B) Sample Doppler spectrum at same location and depth as (A). The only change is that the transducer has been angled upwards (superiorly) slightly. In both (A) and (B), depth = 50 mm, gain = 50, sample volume = 12 mm, power = 420 mW/cm2, and filter = 100 Hz. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/62048/62048fig3large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<ol start="5">
	<li>Searching for the bifurcation 
	NOTE: Finding the bifurcation of the internal carotid artery (ICA) is important to help confirm that the MCA is the artery being monitored. This step should be performed on both sides if bilateral monitoring will be performed, as the bifurcation may not be at the same depth on both sides.
	<ol>
		<li>Increase the depth until the signal from the bifurcation of the ICA into the MCA and ACA is noted (Figure 4), typically at a depth of 51&#8211;65 mm10.</li>
		<li>Search for the optimum bifurcation spectral signal using the procedure described in step 1.4.2. Always strive for the highest-velocity spectral signal possible10.&#160;</li>
		<li>When an optimal bifurcation signal is obtained, note the depth of the bifurcation.</li>
		<li>For bilateral monitoring, repeat sections 1.1&#8211;1.4 and steps 1.5.1&#8211;1.5.3 on the other side of the head.</li>
	</ol>
	</li>
</ol>
<img alt="Figure 4" class="xfigimg" src="/files/ftp_upload/62048/62048fig4.jpg" /> 
Figure 4: Spectral Doppler (top) and M-mode (bottom) image of bifurcation of the ICA into the MCA and ACA. Depth = 65 mm, gain = 50, sample volume = 12 mm, power = 420 mW/cm2, and filter = 100 Hz.&#160;<a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/62048/62048fig4large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
2. Relocating the MCA after placing a fixation device
NOTE: For fTCD experiments, it is necessary to monitor CBFV for 10&#8211;90 min or longer. Therefore, a fixation device (Figure 5) is crucial to provide stability.
<ol>
	<li>Placing the fixation device
	<ol>
		<li>By visual inspection, adjust the fixation device (Figure 5) to the subject&#8217;s approximate head size.</li>
		<li>Alert the subject before placing the headset on his or her head. Place the headset on the subject&#8217;s head. 
		NOTE: If the subject has long or thick hair, it may be necessary to tie the subject&#8217;s hair back, depending on the fixation device being used.</li>
		<li>Adjust the fixation device&#8217;s fit, and ask the subject if the device is too tight. 
		NOTE: The device should be tight enough that it does not move when bumped slightly, but loose enough that the subject is not uncomfortable.</li>
	</ol>
	</li>
</ol>
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/62048/62048fig5.jpg" /> 
Figure 5: Subject wearing custom fixation device. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/62048/62048fig5large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<ol start="2">
	<li>Locating the MCA signal
	<ol>
		<li>Loosen the mechanism of the fixation device holding the transducer in place (e.g., loosen the mechanism, shown in in Figure 5, by turning a knob counterclockwise) so that the transducer can move freely.</li>
		<li>Alert the subject before applying gel to the transducers (which should already be in place from section 2.1), and that the gel may be cold (if it has been stored at room temperature).</li>
		<li>Apply enough ultrasound gel to the transducer to cover the face of the transducer.&#160;</li>
		<li>Adjust the fixation device so that the transducer is located over the top of the mark made in step 1.4.6.</li>
		<li>Search for the optimal MCA spectral signal using the procedure described in steps 1.4.1&#8211;1.4.3. Always strive for the highest-velocity spectral signal possible10.&#160; 
		NOTE: When compared to freehand TCD, the optimal depth at which the MCA is located using the fixation device may differ slightly (at most 1&#8211;2 mm) from the depth for the freehand device. This is because the fixation device may hold the transducer slightly further away from the scalp while still maintaining a coupling gel seal.</li>
		<li>When the optimal MCA spectral signal is found, tighten the mechanism of the fixation device to lock the transducer in place. Note the depth and all other settings.&#160;</li>
		<li>Decrease the power (see step 1.1.1) as much as possible while still maintaining a spectral envelope that traces the maximal velocity accurately.</li>
		<li>For bilateral monitoring, repeat steps 2.2.1&#8211;2.2.7 on the other side.</li>
	</ol>
	</li>
</ol>
3. Performing a breath-hold maneuver
NOTE: This section is given as an example of a functional experiment that may be performed using the experimental setup described in section 1 and section 2.
<ol>
	<li>Perform all steps described in section 1 and section 2.</li>
	<li>Begin recording on the TCD software.</li>
	<li>Breathe normally for 3 min to achieve a good baseline recording, and allow CBFV to stabilize from any previous experiments or stimuli.</li>
	<li>Count down slowly from three. On the count of one, ask the subject to begin breath-holding following a normal inspiration13.&#160; 
	NOTE: The subject should not inhale deeply, as this would decrease carbon dioxide in the lungs and decrease the likelihood of observing the increase in CBFV due to cerebrovascular reactivity. The subject should also avoid performing a Valsalva maneuver, in which intrathoracic pressure is substantially increased against a held inspiration14.&#160;</li>
	<li>Place a marker in the TCD recording to signify the start of breath-holding.</li>
	<li>Have the subject hold their breath for 30 s, or until they are no longer comfortable holding their breath.&#160;</li>
	<li>When the subject inhales, place a marker in the TCD recording to signify the end of breath-holding.</li>
	<li>Continue monitoring CBFV using TCD and recording for at least 30 s following the end of breath-holding to ensure that CBFV returns to baseline values.</li>
</ol>

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The authors declare no conflicts of interest.

Critical steps in the protocol include 1) finding the MCA, 2) placing the headband, and 3) performing the breath-holding maneuver.
Modifications may be necessary depending on the subjects in the study. For example, subjects with Alzheimer&#8217;s disease may have difficulty following instructions, necessitating the use of a capnograph to ensure compliance with breath-holding instructions15. Young children may have difficulty following instructions and may be shy of the experimenter; hence, experimental protocols may need to be simplified for such a population (see Lohmann et al.2). Certain settings on the TCD machine may also need to be changed depending on the population of interest. For example, when insonating infants, who have thin cranial bones, reduce the power as much as possible, especially if TCD monitoring will take place over a period lasting several hours16.
Troubleshooting often centers around difficulties finding a good, stable TCD spectral signal. For example, for people older than 50 years of age, the temporal acoustic window becomes increasingly smaller as the age increases due to increased porosity of the bone of the cranium and tends to localize to the region just ahead of the ear (the &#8220;intertragal space&#8221;)12. In such a population, finding a good MCA spectral signal on both sides of the head may sometimes be impossible, and very slight changes in transducer angle or position may cause the signal to be lost. Because a good-quality signal is essential for experiments that depend on the envelope waveform for analysis, every effort should be made to increase MCA spectral signal intensity and quality. For instance, the gain can be adjusted to optimize the signal, and the sample volume can be increased to get a stronger signal. As a last resort, power may be increased. Finally, it is important to note that in approximately 10% of patients, the temporal acoustic window may be absent11,17. The temporal acoustic window can be readily found in infants and small children and is hardest to find in adults over the age of 50.
Limitations of fTCD include the acquisition of CBFV information at one spatial location17 rather than a wide field of view, albeit with very high temporal resolution. Thus, fTCD is a complement to fMRI, which gives cerebral hemodynamic information (and hence neural activity) with a wide field of view at a low temporal resolution18,19. Indeed, fTCD has a temporal resolution comparable to that of fNIRS20, with the important difference that fTCD measures hemodynamic changes at the level of the major cerebral arteries, whereas fNIRS measures changes in the cortex. Therefore, fTCD can fill in significant details about cerebral hemodynamic changes in response to neural activation, which no other neuroimaging modality is currently capable of measuring.
Potential applications of TCD include monitoring for cerebral embolus formation during cardiac surgery16 and monitoring to detect the outcome of tissue plasminogen activator treatment for stroke21. Potential applications of fTCD include any research question involving the neural response to internal or external stimuli, such as studying the lateralized processing of language in the human brain2,3,4, somatosensory &#8220;touch&#8221; stimulation5, or lateralization of visual processing6. In addition, fTCD can be used to study physiological (with or without neural activity changes) responses to stimuli such as exercise22 and breath-holding13,15,23. Finally, the low cost, portability, and simplicity of fTCD make imaging of large numbers of subjects practical, an advantage over fMRI and other neuroimaging modalities such as PET, e.g., when screening for preclinical Alzheimer&#8217;s disease15.&#160;

In neuroscience research, it is often desirable to monitor real-time brain activity noninvasively in a variety of environments. However, conventional functional neuroimaging modalities have limitations that impede the ability to capture localized and/or rapid activity changes. The true (non-jittered, non-retrospective) temporal resolution of functional magnetic resonance imaging (fMRI) is currently of the order of a few seconds1, which may not capture transient hemodynamic changes linked to transient neural activation. In another example, although functional near-infrared spectroscopy (fNIRS) has high temporal resolution (milliseconds) and reasonable spatial resolution, it can only probe hemodynamic changes within the cerebral cortex and cannot provide information about changes taking place in the larger arteries supplying the brain.
In contrast, fTCD&#8212;classified as a neuroimaging modality&#8212;&#8220;imaging&#8221; refers to the dimensions of time and space, rather than two orthogonal spatial directions that are more familiar in an &#8220;image&#8221;. fTCD provides complementary information to other neuroimaging modalities by measuring high temporal resolution (typically 10 ms) hemodynamic changes at precise locations within vessels of the basal cerebral circulation.&#160;As with other neuroimaging modalities, fTCD may be used for a variety of experiments such as studying lateralization of cerebral activation during language-related tasks2,3,4, studying neural activation in response to various somatosensory stimuli5, and exploring neural activation in various cognitive stimuli such as visual tasks6, mental tasks7, and even tool production8.
Although fTCD offers several advantages for use in functional imaging, including low cost of equipment, portability, and enhanced safety (compared to Wada test3 or positron emission tomography [PET] scans), operation of a TCD machine requires skills obtained by practice. Some of these skills, which must be learned by a TCD operator, include the ability to identify various cerebral arteries and the motor skills necessary to precisely manipulate the ultrasound probe during the search for the relevant artery. The goal of this Methods paper is to present a technique for using fTCD to perform a functional imaging experiment. First, the basic steps for identifying and optimizing the signal from the MCA, which perfuses 80% of the cerebral hemisphere9, will be listed. Next, placement of a fixation device for holding the TCD probe in place during the experiment will be described. Finally, the breath-holding experiment, which is one example of a functional imaging experiment using fTCD, will be described, and representative results will be shown.

<table><tbody><tr><td>Aquasonic</td><td>Parker Laboratories, Inc., Fairfield, NJ, USA</td><td>01-50</td><td>Ultrasound Gel</td></tr><tr><td>Doppler Box X</td><td>DWL Compumedics Gmbh, Singen, Germany</td><td>Model &quot;BoxX&quot;</td><td>Transcranial Doppler with 2-MHz monitoring probes</td></tr><tr><td>Kimwipes</td><td>Kimberly-Clark Professional</td><td>34256</td><td>Delicate Task Wipers</td></tr><tr><td>Transeptic&amp;nbsp;</td><td>Parker Laboratories, Inc., Fairfield, NJ, USA</td><td>09-25</td><td>Cleaning Spray</td></tr></tbody></table>

functional transcranial doppler ultrasound for monitoring cerebral blood flow

Functional transcranial Doppler ultrasound (fTCD) is the use of transcranial Doppler ultrasound (TCD) to study neural activation occurring during stimuli such as physical movement, activation of tactile sensors in the skin, and viewing images. Neural activation is inferred from an increase in the cerebral blood flow velocity (CBFV) supplying the region of the brain involved in processing sensory input. For example, viewing bright light causes increased neural activity in the occipital lobe of the cerebral cortex, leading to increased blood flow in the posterior cerebral artery, which supplies the occipital lobe. In fTCD, changes in CBFV are used to estimate changes in cerebral blood flow (CBF).
With its high temporal resolution measurement of blood flow velocities in the major cerebral arteries, fTCD complements other established functional imaging techniques. The goal of this Methods paper is to give step-by-step instructions for using fTCD to perform a functional imaging experiment. First, the basic steps for identifying the middle cerebral artery (MCA) and optimizing the signal will be described. Next, placement of a fixation device for holding the TCD probe in place during the experiment will be described. Finally, the breath-holding experiment, which is a specific example of a functional imaging experiment using fTCD, will be demonstrated.

Figure 3 shows sample Doppler spectra and color M-modes from the midpoint of the M1 segment of the MCA. Figure 3A,B were taken at the same position on the scalp, but at different angles. Note how a very small change in angle, without changing the contact position on the scalp, can greatly improve Doppler signal strength, as shown by the higher-intensity yellow coloring of the spectrogram in Figure 3B. Note also that the M-mode in Figure 3B shows two arteries (blue and red, corresponding to the ACA and MCA, respectively).
Figure 4 shows a sample Doppler spectrum and M-mode from the bifurcation of the ICA into the ACA and MCA. Note the overlapping red- and blue-shaded regions in the M-mode image denoting the MCA and ACA, respectively. Also note the symmetry of the Doppler spectral waveform when comparing flow towards the transducer (positive) with flow away from the transducer (negative).
Figure 6 shows sample spectra and M-mode images from different time points in the breath-hold maneuver. Figure 6A shows the baseline TCD spectrum and M-mode at the beginning of breath-holding. Note the mean velocity of 56 cm/s. Figure 6B shows the TCD spectrum and M-mode at the end of breath-holding. Note that the mean velocity has now increased to 70 cm/s. Figure 6C shows the TCD spectrum and M-mode after the end of breath-holding. Note the undershoot in velocity below baseline values, with the mean dropping to 47 cm/s. Note that the ACA is visible as flow away from the transducer&#160;in the Doppler spectra.
Figure 7 shows the entire breath-holding experiment. Note that the envelope remains elevated for approximately 15 s after breath-holding ends, falls to values lower than those at the beginning of breath-holding for ~20 s, and then finally recovers to baseline values. Note that the ACA is visible as flow away from the transducer in the Doppler spectrum.&#160;
Figure 6 and Figure 7 display good signal intensity in the MCA portion of the TCD spectrum (the MCA is represented by the positive velocities); note how the white line which represents the envelope follows the TCD spectrum very accurately when the spectrum is bright. The spectra of Figure 6 and Figure 7 could be improved by decreasing the monitoring depth by 5&#8211;10 mm so that the ACA portion of the TCD spectrum would not be visible (the ACA is represented by negative velocities) and by changing the scale of the vertical axis in the TCD spectrum to run from approximately -100 cm/s to 100 cm/s, which would allow maximum velocity sampling of the TCD spectrum in the vertical direction.
Figure 8 shows examples of bilateral TCD spectra and M-modes suitable for bilateral fTCD. Figure 8A and Figure 8B demonstrate acceptable, but not optimal, bilateral spectra and M-modes. Note how the gain is higher in Figure 8A (left MCA) than in Figure 8B (right MCA) to compensate for the weaker signal, and how the envelope quality in Figure 8A is slightly poorer than in Figure 8B. Also note how the maximum velocity at systole in Figure 8A is slightly lower than in Figure 8B. By contrast, note how the two spectra in Figure 8C and Figure 8D are very similar in terms of settings, including depth, gain, power, and sample volume, and how the spectral waveforms on both sides have similar maximum velocities and shapes. To address this, it is recommended that the spectrum from the left MCA be consistently placed in the left window and the spectrum from the right MCA in the right window, especially for experiments involving lateralization of blood flow.&#160;
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/62048/62048fig6.jpg" /> 
Figure 6: Sample Doppler spectra and M-mode images from the MCA during different stages of the breath-hold maneuver. (A) Spectrum and M-mode at the beginning of breath-holding. Vertical yellow line in center denotes the start of breath-holding. (B) Spectrum and M-mode at the end of breath-holding. Vertical yellow line in center denotes the end of breath-holding when the subject inhales. (C) Spectrum and M-mode after the end of breath-holding, showing the decrease in flow velocity that persists for approximately 30 s after breath-holding. In all spectra, depth = 56 mm, gain = 50, sample volume = 8 mm, power = 420 mW/cm2, and filter = 100 Hz.&#160;<a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/62048/62048fig6large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/62048/62048fig7.jpg" /> 
Figure 7: Spectrum and M-mode from the MCA throughout breath-holding. Depth = 56 mm, gain = 50, sample volume = 8 mm, power = 420 mW/cm2, and filter = 100 Hz. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/62048/62048fig7large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 8" class="xfigimg" src="/files/ftp_upload/62048/62048fig8.jpg" /> 
Figure 8: Examples of bilateral spectra and M-mode images from the MCA. (A) Acceptable, but not optimal, spectrum and M-mode of the left MCA, with depth = 62 mm, gain = 69, sample volume = 12 mm, power = 420 mW/cm2, and filter = 100 Hz. (B) Good spectrum and M-mode of right MCA, with depth = 62 mm, gain = 56, sample volume = 12 mm, power = 420 mW/cm2, and filter = 100 Hz. (C) Good spectrum and M-mode of the left MCA. (D) Good spectrum and M-mode of the right MCA. For both (C) and (D), depth = 62 mm, gain = 56, sample volume = 12, power = 420 mW/cm2, and filter = 100 Hz.&#160; <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/62048/62048fig8large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Age</td>
			<td>Middle cerebral artery depth (mm)&#160;</td>
		</tr>
		<tr>
			<td>0&#8211;3 monthsa</td>
			<td>25</td>
		</tr>
		<tr>
			<td>3&#8211;12 monthsa</td>
			<td>30</td>
		</tr>
		<tr>
			<td>1&#8211;3 yearsa</td>
			<td>35&#8211;45</td>
		</tr>
		<tr>
			<td>3&#8211;6 yearsa</td>
			<td>40&#8211;45</td>
		</tr>
		<tr>
			<td>6&#8211;10 yearsa</td>
			<td>45&#8211;50</td>
		</tr>
		<tr>
			<td>10&#8211;18 yearsa</td>
			<td>45&#8211;50</td>
		</tr>
		<tr>
			<td>&#62;18 yearsb</td>
			<td>50</td>
		</tr>
	</tbody>
</table>
Table 1: MCA depths at various ages. Sources: a = Bode25, b = Alexandrov et al.10

Watch this Scientific Journal Video about Functional Transcranial Doppler Ultrasound for Monitoring Cerebral Blood Flow at JoVE.com

Functional Transcranial Doppler Ultrasound for Monitoring Cerebral Blood Flow

Functional transcranial Doppler ultrasound complements other functional imaging modalities, with its high temporal resolution measurement of stimulus-induced changes in cerebral blood flow within the basal cerebral arteries. This Methods paper gives step-by-step instructions for using functional transcranial Doppler ultrasound to perform a functional imaging experiment.&#160;

Functional transcranial Doppler ultrasound (fTCD) is the use of transcranial Doppler ultrasound (TCD) to study neural activation occurring during stimuli ...

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Research

JoVE Journal

Bioengineering

7.4K Views.  University of Nebraska-Lincoln. Dit protocol vormt de basis voor een functioneel TCD-experiment omdat bijna alle functionele TCD-experimenten een fixatieapparaat moeten plaatsen om een stabiel signaal over een langere periode op te nemen. Het belangrijkste voordeel van functionele TCD is de hoge temporele resolutie meting van veranderingen in de cerebrale bloedstroom. Het vinden van de middelste hersenslagader met behulp van TCD vergt oefening.Het is cruciaal om de transducer stabiel te houden en heel langzaam te bew...

Video: Functional Transcranial Doppler Ultrasound for Monitoring Cerebral Blood Flow

Simultaneous EEG Monitoring During Transcranial Direct Current Stimulation

Transcranial direct current stimulation (tDCS) is a technique that delivers weak electric currents through the scalp. This constant electric current induces shifts in neuronal membrane excitability, resulting in secondary changes in cortical activity. Although tDCS has most of its neuromodulatory effects on the underlying cortex, tDCS effects can also be observed in distant neural networks. Therefore, concomitant EEG monitoring of the effects of tDCS can provide valuable information on the mechanisms of tDCS. In addition, EEG findings can be an important surrogate marker for the effects of tDCS and thus can be used to optimize its parameters. This combined EEG-tDCS system can also be used for preventive treatment of neurological conditions characterized by abnormal peaks of cortical excitability, such as seizures. Such a system would be the basis of a non-invasive closed-loop device. In this article, we present a novel device that is capable of utilizing tDCS and EEG simultaneously. For that, we describe in a step-by-step fashion the main procedures of the application of this device using schematic figures, tables and video demonstrations. Additionally, we provide a literature review on clinical uses of tDCS and its cortical effects measured by EEG techniques.

Transcranial direct current stimulation (tDCS) is a non-invasive brain stimulation technique that has shown initial therapeutic effects in several neurological conditions. The main mechanism underlying these therapeutic effects is the modulation of cortical excitability. Therefore, online monitoring of cortical excitability would help guide stimulation parameters and optimize its therapeutic effects. In the present article we review the use of a novel device that combines simultaneous tDCS and EEG monitoring in real time.

Transcranial  direct current stimulation (tDCS) is a technique that delivers weak electric  currents through the scalp. This constant electric current ...

Behavior

Assessment of Cerebral Lateralization in Children using Functional Transcranial Doppler Ultrasound (fTCD)

There are many unanswered questions about cerebral lateralization. In particular, it remains unclear which aspects of language and nonverbal ability are lateralized, whether there are any disadvantages associated with atypical patterns of cerebral lateralization, and whether cerebral lateralization develops with age. In the past, researchers interested in these questions tended to use handedness as a proxy measure for cerebral lateralization, but this is unsatisfactory because handedness is only a weak and indirect indicator of laterality of cognitive functions1. Other methods, such as fMRI, are expensive for large-scale studies, and not always feasible with children2.
Here we will describe the use of functional transcranial Doppler ultrasound (fTCD) as a cost-effective, non-invasive and reliable method for assessing cerebral lateralization. The procedure involves measuring blood flow in the middle cerebral artery via an ultrasound probe placed just in front of the ear. Our work builds on work by Rune Aaslid, who co-introduced TCD in 1982, and Stefan Knecht, Michael Deppe and their colleagues at the University of M&uuml;nster, who pioneered the use of simultaneous measurements of left- and right middle cerebral artery blood flow, and devised a method of correcting for heart beat activity. This made it possible to see a clear increase in left-sided blood flow during language generation, with lateralization agreeing well with that obtained using other methods3.
The middle cerebral artery has a very wide vascular territory (see Figure 1) and the method does not provide useful information about localization within a hemisphere. Our experience suggests it is particularly sensitive to tasks that involve explicit or implicit speech production. The 'gold standard' task is a word generation task (e.g. think of as many words as you can that begin with the letter 'B') 4, but this is not suitable for young children and others with limited literacy skills. Compared with other brain imaging methods, fTCD is relatively unaffected by movement artefacts from speaking, and so we are able to get a reliable result from tasks that involve describing pictures aloud5,6. Accordingly, we have developed a child-friendly task that involves looking at video-clips that tell a story, and then describing what was seen.

Assessment of Cerebral Lateralization in Children using Functional Transcranial Doppler Ultrasound fTCD

Functional transcranial Doppler sonography (fTCD) is a simple and non-invasive ultrasound technique which can be used to assess the lateralization of cognitive functions, especially language, and is suitable for use with children.

There are many unanswered questions about cerebral lateralization.  In particular, it remains unclear which aspects of language and nonverbal ability are ...

Neuroscience

Non-invasive Optical Measurement of Cerebral Metabolism and Hemodynamics in Infants

Perinatal brain injury remains a significant cause of infant mortality and morbidity, but there is not yet an effective bedside tool that can accurately screen for brain injury, monitor injury evolution, or assess response to therapy. The energy used by neurons is derived largely from tissue oxidative metabolism, and neural hyperactivity and cell death are reflected by corresponding changes in cerebral oxygen metabolism (CMRO2). Thus, measures of CMRO2 are reflective of neuronal viability and provide critical diagnostic information, making CMRO2 an ideal target for bedside measurement of brain health.
Brain-imaging techniques such as positron emission tomography (PET) and single-photon emission computed tomography (SPECT) yield measures of cerebral glucose and oxygen metabolism, but these techniques require the administration of radionucleotides, so they are used in only the most acute cases.
Continuous-wave near-infrared spectroscopy (CWNIRS) provides non-invasive and non-ionizing radiation measures of hemoglobin oxygen saturation (SO2) as a surrogate for cerebral oxygen consumption. However, SO2 is less than ideal as a surrogate for cerebral oxygen metabolism as it is influenced by both oxygen delivery and consumption. Furthermore, measurements of SO2 are not sensitive enough to detect brain injury hours after the insult 1,2, because oxygen consumption and delivery reach equilibrium after acute transients 3. We investigated the possibility of using more sophisticated NIRS optical methods to quantify cerebral oxygen metabolism at the bedside in healthy and brain-injured newborns. More specifically, we combined the frequency-domain NIRS (FDNIRS) measure of SO2 with the diffuse correlation spectroscopy (DCS) measure of blood flow index (CBFi) to yield an index of CMRO2 (CMRO2i) 4,5.
With the combined FDNIRS/DCS system we are able to quantify cerebral metabolism and hemodynamics. This represents an improvement over CWNIRS for detecting brain health, brain development, and response to therapy in neonates. Moreover, this method adheres to all neonatal intensive care unit (NICU) policies on infection control and institutional policies on laser safety. Future work will seek to integrate the two instruments to reduce acquisition time at the bedside and to implement real-time feedback on data quality to reduce the rate of data rejection.

We combined frequency-domain near-infrared spectroscopy measures of cerebral hemoglobin oxygenation with diffuse correlation spectroscopy measures of cerebral blood flow index to estimate an index of oxygen metabolism. We tested the utility of this measure as a bedside screening tool to evaluate the health and development of the newborn brain.

Perinatal brain injury remains a significant cause of infant mortality  and morbidity, but there is not yet an effective bedside tool that can  accurately ...

Medicine

Optimized System for Cerebral Perfusion Monitoring in the Rat Stroke Model of Intraluminal Middle Cerebral Artery Occlusion

The translational potential of pre-clinical stroke research depends on the accuracy of experimental modeling. Cerebral perfusion monitoring in animal models of acute ischemic stroke allows to confirm successful arterial occlusion and exclude subarachnoid hemorrhage. Cerebral perfusion monitoring can also be used to study intracranial collateral circulation, which is emerging as a powerful determinant of stroke outcome and a possible therapeutic target. Despite a recognized role of Laser Doppler perfusion monitoring as part of the current guidelines for experimental cerebral ischemia, a number of technical difficulties exist that limit its widespread use. One of the major issues is obtaining a secure and prolonged attachment of a deep-penetration Laser Doppler probe to the animal skull. In this video, we show our optimized system for cerebral perfusion monitoring during transient middle cerebral artery occlusion by intraluminal filament in the rat. We developed in-house a simple method to obtain a custom made holder for twin-fibre (deep-penetration) Laser Doppler probes, which allow multi-site monitoring if needed. A continuous and prolonged monitoring of cerebral perfusion could easily be obtained over the intact skull.

Cerebral perfusion monitoring has been demonstrated to improve accuracy in ischemic stroke models. Technical difficulties often limit the use of this essential tool for cerebrovascular research. In this video, an optimized system is shown to obtain a single or multi-site hemodynamic monitoring during intraluminal middle cerebral artery occlusion in rats.

The translational potential of pre-clinical stroke research depends on the accuracy of experimental modeling. Cerebral perfusion monitoring in animal ...

Non-invasive Parenchymal, Vascular and Metabolic High-frequency Ultrasound and Photoacoustic Rat Deep Brain Imaging

Photoacoustics and high frequency ultrasound stands out as powerful tools for neurobiological applications enabling high-resolution imaging on the central nervous system of small animals. However, transdermal and transcranial neuroimaging is frequently affected by low sensitivity, image aberrations and loss of space resolution, requiring scalp or even skull removal before imaging. To overcome this challenge, a new protocol is presented to gain significant insights in brain hemodynamics by photoacoustic and high-frequency ultrasounds imaging with the animal skin and skull intact. The procedure relies on the passage of ultrasound (US) waves and laser directly through the fissures that are naturally present on the animal cranium. By juxtaposing the imaging transducer device exactly in correspondence to these selected areas where the skull has a reduced thickness or is totally absent, one can acquire high quality deep images and explore internal brain regions that are usually difficult to anatomically or functionally describe without an invasive approach. By applying this experimental procedure, significant data can be collected in both sonic and optoacoustic modalities, enabling to image the parenchymal and the vascular anatomy far below the head surface. Deep brain features such as parenchymal convolutions and fissures separating the lobes were clearly visible. Moreover, the configuration of large and small blood vessels was imaged at several millimeters of depth, and precise information were collected about blood fluxes, vascular stream velocities and the hemoglobin chemical state. This repertoire of data could be crucial in several research contests, ranging from brain vascular disease studies to experimental techniques involving the systemic administration of exogenous chemicals or other objects endowed with imaging contrast enhancement properties. In conclusion, thanks to the presented protocol, the US and PA techniques become an attractive noninvasive performance-competitive means for cortical and internal brain imaging, retaining a significant potential in many neurologic fields.

The present work describes a new protocol to perform non-invasive high-frequency ultrasound and photoacoustic based imaging on rat brain, to efficiently visualize deep subcortical regions and their vascular patterns by directing signals on skull foramina naturally present on animal cranium.

Photoacoustics and high frequency ultrasound stands out as powerful tools for neurobiological applications enabling high-resolution imaging on the central ...

Assessing Cerebral Autoregulation via Oscillatory Lower Body Negative Pressure and Projection Pursuit Regression

The process by which cerebral perfusion is maintained constant over a wide range of systemic pressures is known as &#8220;cerebral autoregulation.&#8221; Effective dampening of flow against pressure changes occurs over periods as short as ~15 sec and becomes progressively greater over longer time periods. Thus, slower changes in blood pressure are effectively blunted and faster changes or fluctuations pass through to cerebral blood flow relatively unaffected. The primary difficulty in characterizing the frequency dependence of cerebral autoregulation is the lack of prominent spontaneous fluctuations in arterial pressure around the frequencies of interest (less than ~0.07 Hz or ~15 sec). Oscillatory lower body negative pressure (OLBNP) can be employed to generate oscillations in central venous return that result in arterial pressure fluctuations at the frequency of OLBNP. Moreover, Projection Pursuit Regression (PPR) provides a nonparametric method to characterize nonlinear relations inherent in the system without a priori assumptions and reveals the characteristic non-linearity of cerebral autoregulation. OLBNP generates larger fluctuations in arterial pressure as the frequency of negative pressure oscillations become slower; however, fluctuations in cerebral blood flow become progressively lesser. Hence, the PPR shows an increasingly more prominent autoregulatory region at OLBNP frequencies of 0.05 Hz and below (20 sec cycles). The goal of this approach it to allow laboratory-based determination of the characteristic nonlinear relationship between pressure and cerebral flow and could provide unique insight to integrated cerebrovascular control as well as to physiological alterations underlying impaired cerebral autoregulation (e.g., after traumatic brain injury, stroke, etc.).

Cerebral perfusion is maintained across a range of pressures via cerebral autoregulation. However, characterizing autoregulation requires prominent pressure fluctuations at regulated frequencies. The described protocol will show how oscillatory lower body negative pressure can generate pressure fluctuations to provide data for projection pursuit regression for quantification of the autoregulatory curve.

The process by which cerebral perfusion is maintained constant over a wide range of systemic pressures is known as &#8220;cerebral autoregulation.&#8221; ...

Biology

Analysis of Cerebral Vasospasm in a Murine Model of Subarachnoid Hemorrhage with High Frequency Transcranial Duplex Ultrasound

Cerebral vasospasm that occurs in the weeks after subarachnoid hemorrhage, a type of hemorrhagic stroke, contributes to delayed cerebral ischemia. A problem encountered in experimental studies using murine models of SAH is that methods for in vivo monitoring of cerebral vasospasm in mice are lacking. Here, we demonstrate the application of high frequency ultrasound to perform transcranial Duplex sonography examinations on mice. Using the method, the internal carotid arteries (ICA) could be identified. The blood flow velocities in the intracranial ICAs were accelerated significantly after induction of SAH, while blood flow velocities in the extracranial ICAs remained low, indicating cerebral vasospasm. In conclusion, the method demonstrated here allows functional, noninvasive in vivo monitoring of cerebral vasospasm in a murine SAH model.

The aim of this manuscript is to present a sonography-based method that allows in vivo imaging of blood flow in cerebral arteries in mice. We demonstrate its application to determine changes in blood flow velocities associated with vasospasm in murine models of subarachnoid hemorrhage (SAH).

Cerebral vasospasm that occurs in the weeks after subarachnoid hemorrhage, a type of hemorrhagic stroke, contributes to delayed cerebral ischemia. A ...

Real-Time Monitoring of Neurocritical Patients with Diffuse Optical Spectroscopies

Neurophysiological monitoring is an important goal in the treatment of neurocritical patients, as it may prevent secondary damage and directly impact morbidity and mortality rates. However, there is currently a lack of suitable non-invasive, real-time technologies for continuous monitoring of cerebral physiology at the bedside. Diffuse optical techniques have been proposed as a potential tool for bedside measurements of cerebral blood flow and cerebral oxygenation in case of neurocritical patients. Diffuse optical spectroscopies have been previously explored to monitor patients in several clinical scenarios ranging from neonatal monitoring to cerebrovascular interventions in adults. However, the feasibility of the technique to aid clinicians by providing real-time information at the bedside remains largely unaddressed. Here, we report the translation of a diffuse optical system for continuous real-time monitoring of cerebral blood flow, cerebral oxygenation, and cerebral oxygen metabolism during intensive care. The real-time feature of the instrument could enable treatment strategies based on patient-specific cerebral physiology rather than relying on surrogate metrics, such as arterial blood pressure. By providing real-time information on the cerebral circulation at different time scales with relatively cheap and portable instrumentation, this approach may be especially useful in low-budget hospitals, in remote areas and for&#160;monitoring in open fields (e.g., defense and sports).

Presented here is a protocol for non-invasively monitoring cerebral hemodynamics of neurocritical patients in real-time and at the bedside using diffuse optics. Specifically, the proposed protocol uses a hybrid diffuse optical systems to detect and display real-time information on cerebral oxygenation, cerebral blood flow and cerebral metabolism.

Neurophysiological monitoring is an important goal in the treatment of neurocritical patients, as it may prevent secondary damage and directly impact ...

Mapping Cerebral Blood Flow and Electrical Fields: Electrode Placement in Mice

Placement of Extracranial Stimulating Electrodes and Measurement of Cerebral Blood Flow and Intracranial Electrical Fields in Anesthetized Mice

The detection of cerebral blood flow (CBF) responses to various forms of neuronal activation is critical for understanding dynamic brain function and variations in the substrate supply to the brain. This paper describes a protocol for measuring CBF responses to transcranial alternating current stimulation (tACS). Dose-response curves are estimated both from the CBF change occurring with tACS (mA) and from the intracranial electric field (mV/mm). We estimate the intracranial electrical field based on the different amplitudes measured by glass microelectrodes within each side of the brain. In this paper, we describe the experimental setup, which involves using either bilateral laser Doppler (LD) probes or laser speckle imaging (LSI) to measure the CBF; as a result, this setup requires anesthesia for the electrode placement and stability. We present a correlation between the CBF response and the current as a function of age, showing a significantly larger response at higher currents (1.5 mA and 2.0 mA) in young control animals (12-14 weeks) compared to older animals (28-32 weeks) (p &lt; 0.005 difference). We also demonstrate a significant CBF response at electrical field strengths &lt;5 mV/mm, which is an important consideration for eventual human studies. These CBF responses are also strongly influenced by the use of anesthesia compared to awake animals, the respiration control (i.e., intubated vs. spontaneous breathing), systemic factors (i.e., CO2), and local conduction within the blood vessels, which is mediated by pericytes and endothelial cells. Likewise, more detailed imaging/recording techniques may limit the field size from the entire brain to only a small region. We describe the use of extracranial electrodes for applying tACS stimulation, including both homemade and commercial electrode designs for rodents, the concurrent measurement of the CBF and intracranial electrical field using bilateral glass DC recording electrodes, and the imaging approaches. We are currently applying these techniques to implement a closed-loop format for augmenting the CBF in animal models of Alzheimer's disease and stroke.

Author Spotlight: Stimulation-Based Approach to Improve Cerebral Blood Flow in Alzheimer's Model 

We describe a protocol for assessing dose-response curves for extracranial stimulation in terms of brain electrical field measurements and a relevant biomarker-cerebral blood flow. Since this protocol involves invasive electrode placement into the brain, general anesthesia is needed, with spontaneous breathing preferred rather than controlled respirations.

The detection of cerebral blood flow (CBF) responses to various forms of neuronal activation is critical for understanding dynamic brain function and ...

Periorbital Laser Doppler Flowmetry for MCAO-Induced Ischemic Stroke in Rodents

Periorbital Placement of a Laser Doppler Probe for Cerebral Blood Flow Monitoring Prior to Middle Cerebral Artery Occlusion in Rodent Models

Middle cerebral artery occlusion (MCAO) is the gold-standard method for preclinical modeling of ischemic stroke in rodents. However, successful occlusion is not guaranteed by even the most skilled surgical hands. Errors primarily occur when the filament is not placed at the correct depth and include instances of either no infarction or vessel perforation, which can cause death. Laser Doppler flowmetry (LDF) is a reliable technique that provides real-time feedback on regional cerebral blood flow (CBF) during the MCAO procedure. Here we demonstrate a rapid technique for periorbital placement of a laser Doppler probe for measurement of CBF in both mice and rats. Our rationale was to simplify LDF implementation, encouraging widespread usage for improved surgical reliability. The technique eliminates the need for skull thinning and specialized equipment, with placement at the periorbital region rather than dorsal placement, promoting efficiency and ease of adoption. The protocol described here encompasses presurgical preparations, periorbital Doppler probe placement, and post-operative care. Representative results include visual depictions of procedural elements along with representative LDF tracings illustrating successful MCAO surgeries, with instances of unsuccessful filament placement leading to complications. The protocol illustrates LDF in confirming proper filament placement and offers a simplified procedure compared to alternative methods.

Author Spotlight: Advancing Stroke Research with Periorbital Doppler Monitoring of Regional Cerebral Blood Flow in Rodent Models

A minimally invasive surgical procedure is shown here, which involves placing the laser Doppler probe onto the skull over the distal region of the middle cerebral artery (MCA), a periorbital location suitable for rats and mice, to assess blood flow during transient MCA occlusion.

Middle cerebral artery occlusion (MCAO) is the gold-standard method for preclinical modeling of ischemic stroke in rodents. However, successful occlusion ...

Functional Transcranial Doppler Ultrasound for Monitoring Cerebral Blood Flow

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