Name: radiotracer administration for high temporal resolution positron emission tomography of the human brain: application to fdg-fpet
Uploaded: 2019-10-22T14:00:00
Description: Watch this Scientific Journal Video about Radiotracer Administration for High Temporal Resolution Positron Emission Tomography of the Human Brain: Application to FDG-fPET at JoVE.com

Jamadar is supported by an Australian Council for Research (ARC) Discovery Early Career Researcher Award (DECRA DE150100406). Jamadar, Ward, and Egan are supported by the ARC Centre of Excellence for Integrative Brain Function (CE114100007). Chen and Li are supported by funding from the Reignwood Cultural Foundation.
Jamadar, Ward, Carey, and McIntyre designed the protocol. Carey, McIntyre, Sasan, and Fallon collected the data. Jamadar, Ward, Parkes, and Sasan analyzed the data. Jamadar, Ward, Carey, and McIntyre wrote the first draft of the manuscript. All authors have reviewed and approved the final version.

This protocol has been reviewed and approved by the Monash University Human Research Ethics Committee (approval number CF16/1108 - 2016000590) in accordance with the Australian National Statement on Ethical Conduct in Human Research24. Procedures were developed under the guidance of an accredited Medical Physicist, Nuclear Medicine Technologist, and clinical radiographer. Researchers should refer to their local experts and guidelines for the administration of ionizing radiation in humans.
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FDG-PET is a powerful imaging technology that measures glucose uptake, an index of cerebral glucose metabolism. To date, most neuroscience studies using FDG-PET use a traditional bolus administration approach, with a static image resolution that represents the integral of all metabolic activity over the course of the scan2. This manuscript describes two alternative radiotracer administration protocols: the infusion-only (e.g., Villien et al., Jamadar et al.19,<su...

Positron emission tomography (PET) is a powerful molecular imaging technique that is widely used in both clinical and research settings (see Heurling et al.1 for a recent comprehensive review). The molecular targets that can be imaged using PET are only limited by the availability of radiotracers, and numerous tracers have been developed to image neural metabolism receptors, proteins, and enzymes2,3. In neuroscience, one of the most used radiotracers is 18F-fluorodeoxyglucose (FDG-PET), which measures glucose uptake, usually interpreted as an index of cerebral glucose metabolism. The human brain requires a constant and reliable supply of glucose to satisfy its energy requirements4,5, and 70-80% of cerebral glucose metabolism is used by neurons during synaptic transmission6. Changes to cerebral glucose metabolism are thought to initiate and contribute to numerous conditions, including psychiatric, neurodegenerative, and ischemic conditions7,8,9. Furthermore, as FDG uptake is proportional to synaptic activity10,11,12, it is considered a more direct and less confounded index of neuronal activity compared to the more widely used blood oxygenation level-dependent functional magnetic resonance imaging (BOLD-fMRI) response. BOLD-fMRI is an indirect index of neural activity and measures changes in deoxygenated hemoglobin that occur following a cascade of neurovascular changes following neuronal activity.
Most FDG-PET studies of the human brain acquire static images of cerebral glucose uptake. The participant rests quietly for 10 min with their eyes open in a darkened room. The full radiotracer dose is administered as a bolus over a period of seconds, and the participant then rests for a further 30 min. Following the uptake period, participants are placed in the center of the PET scanner, and a PET image that reflects the cumulative FDG distribution over the course of the uptake and scanning periods is acquired. Thus, the neuronal activity indexed by the PET image represents the cumulative average of all cognitive activity over uptake and scan periods and is not specific to cognitive activity during the scan. This method has provided great insight into the cerebral metabolism of the brain and neuronal function. However, the temporal resolution is equal to the scan duration (often ~45 min, effectively yielding a static measurement of glucose uptake; this compares unfavourably to neuronal response during cognitive processes and common experiments in neuroimaging. Due to the limited temporal resolution, the method provides a non-specific index of glucose uptake (i.e., not locked to a task or cognitive process) and cannot provide measures of within-subject variability, which can lead to erroneous scientific conclusions due to Simpson&#39;s Paradox13. Simpson&#8217;s Paradox is a scenario, where brain-behavior relationships calculated across-subjects are not necessarily indicative of the same relationships tested within-subjects. Furthermore, recent attempts to apply functional connectivity measures to FDG-PET can only measure across-subjects connectivity. Thus, differences in connectivity can only be compared between groups and cannot be calculated for individual subjects. While it is debatable what exactly across-subject connectivity measures14, it is clear that measures calculated across-but not within-subjects cannot be used as a biomarker for disease states or used to examine the source of individual variation.
In the past five years, the development and wider accessibility of clinical-grade simultaneous MRI-PET scanners has sparked renewed research interest in FDG-PET imaging2 in cognitive neuroscience. With these developments, researchers have focused on improving the temporal resolution of FDG-PET to approach the standards of BOLD-fMRI (~0.5&#8722;2.5 s). Note that the spatial resolution of BOLD-fMRI can approach submillimeter resolutions but the spatial resolution of FDG-PET is fundamentally limited to around 0.54 mm full width at half maximum (FWHM) due to the positron range15. Dynamic FDG-PET acquisitions, which are often used clinically, use the bolus administration method and reconstruct the list-mode data into bins. The bolus dynamic FDG-PET method offers a temporal resolution of around 100 s (e.g., Tomasi et al.16). This is clearly much better compared to static FDG-PET imaging but is not comparable to BOLD-fMRI. Additionally, the window in which brain function may be examined is limited, because the blood plasma concentration of FDG diminishes soon after the bolus is administered.
To expand this experimental window, a handful of studies17,18,19,20,21 have adapted the radiotracer infusion method previously proposed by Carson22,23. In this method, sometimes described as &#39;functional FDG-PET&#39; (FDG-fPET, analogous to BOLD-fMRI), the radiotracer is administered as a constant infusion over the course of the entire PET scan (~90 min). The goal of the infusion protocol is to maintain a constant plasma supply of FDG to track dynamic changes in glucose uptake across time. In a proof-of-concept study, Villien et al.21 used a constant infusion protocol and simultaneous MRI/FDG-fPET to show dynamic changes in glucose uptake in response to checkerboard stimulation with a temporal resolution of 60 s. Subsequent studies have used this method to show task-locked FDG-fPET (i.e., time-locked to an external stimulus19) and task-related FDG-fPET (i.e., not time-locked to an external stimulus17,18) glucose uptake. Using these methods, FDG-fPET temporal resolutions of 60 s have been obtained, which is a substantial improvement over bolus methods. Preliminary data show that the infusion method can provide temporal resolutions of 20&#8722;60 s19.
Despite the promising results from the constant infusion method, the plasma radioactivity curves of these studies show that the infusion method is not sufficient to reach a steady-state within the timeframe of a 90 min scan19,21. In addition to the constant infusion procedure, Carson22 also proposed a hybrid bolus/infusion procedure, where the goal is to quickly reach equilibrium at the beginning of the scan, and then sustain plasma radioactivity levels at equilibrium for the duration of the scan. Rischka et al.20 recently applied this technique using a 20% bolus plus 80% infusion. As expected, the arterial input function quickly rose above baseline levels and was sustained at a higher rate for a longer time, compared to results using an infusion-only procedure19,21.
This paper describes the acquisition protocols for acquiring high temporal resolution FDG-fPET scans using infusion-only and bolus/infusion radiotracer administration. These protocols have been developed for use in a simultaneous MRI-PET environment with a 90&#8722;95 min acquisition time19. In the protocol, blood samples are taken to quantify plasma serum radioactivity for subsequent quantification of PET images. While the protocol&#39;s focus is the application of infusion methods for functional neuroimaging using BOLD-fMRI/FDG-fPET, these methods can be applied to any FDG-fPET study regardless of whether simultaneous MRI, BOLD-fMRI, computed tomography (CT), or other neuroimages are acquired. Figure 1 shows the flowchart of procedures in this protocol.

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			<td>Blood Collection Equipment</td>
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			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>--12-15 vacutainers</td>
			<td>Becton Dickinson, NJ USA</td>
			<td>364880</td>
			<td>Remain in sterile packaging until required to put blood in tube</td>
		</tr>
		<tr>
			<td>--12-15 10mL LH blood collecting tubes</td>
			<td>Becton Dickinson</td>
			<td>367526</td>
			<td>Marked with the sample number (e.g., S1, S2&#8230;) and subsequently marked with the sample time (e.g., time 0 + x min [T0+x])</td>
		</tr>
		<tr>
			<td>--2-15 10mL Terumo syringe</td>
			<td>Terumo Tokyo, Japan</td>
			<td>SS+10L</td>
			<td>These are drawn up on the day of the study and capped with the ampoule that contained the saline</td>
		</tr>
		<tr>
			<td>-- pre-drawn 0.9% saline flushes</td>
			<td>Pfizer, NY, USA</td>
			<td>61039117</td>
			<td></td>
		</tr>
		<tr>
			<td>--12-15 5mL Terumo syringes</td>
			<td>Terumo Tokyo, Japan</td>
			<td>SS+05S</td>
			<td>Remain in sterile packaging until ready to withdraw a blood sample</td>
		</tr>
		<tr>
			<td>Safety &#38; Waste Equipment</td>
			<td></td>
			<td></td>
			<td>All objects arranged on a plastic chair inside the scanner room on the same side as the arm from which the blood samples will be taken. Biohazard and non-biohazard waste bags to be used. Gloves and waste bags to be easily accessible when preparing the radioactivity in the dispensing area and when pipetting the plasma samples. Biohazard and non-biohazard waste bags to be used. All waste generated is checked with the Geiger counter to ensure that radioactive contaminated waste is stored until it is safe to be disposed of according to Australian Radiation Protection and Nuclear Safety Agency (APRANSA) guidelines for Radiation protection series No.6 (2017).</td>
		</tr>
		<tr>
			<td>-- Gloves</td>
			<td>Westlab, VIC, Australia</td>
			<td>663-219</td>
			<td></td>
		</tr>
		<tr>
			<td>-- waste bags</td>
			<td>Austar Packaging, VIC, Australia</td>
			<td>YIW6090</td>
			<td></td>
		</tr>
		<tr>
			<td>--cello underpads &#8216;blueys&#8217; Underpads 5 Ply</td>
			<td>Halyard Health, NSW, Australia</td>
			<td>2765A</td>
			<td></td>
		</tr>
		<tr>
			<td>--Blue Sharpie pen</td>
			<td>Sharpie, TN, USA</td>
			<td>S30063</td>
			<td></td>
		</tr>
		<tr>
			<td>Dose Syringes</td>
			<td></td>
			<td></td>
			<td>Remain in sterile packaging until ready for use. All syringes used in this facility have an additional 20% volume capacity above the stated volume on the packaging. This is important for the 50mL syringe where the total capacity of 60mL is used</td>
		</tr>
		<tr>
			<td>--5mL</td>
			<td>Terumo Tokyo, Japan</td>
			<td>SS+05S</td>
			<td></td>
		</tr>
		<tr>
			<td>-- 20mL</td>
			<td>Terumo Tokyo, Japan</td>
			<td>SS+20L</td>
			<td></td>
		</tr>
		<tr>
			<td>--50mL</td>
			<td>Terumo Tokyo, Japan</td>
			<td>SS*50LE</td>
			<td></td>
		</tr>
		<tr>
			<td>--1 Terumo 18-gauge needle</td>
			<td>Terumo Tokyo, Japan</td>
			<td>NN+1838R</td>
			<td>Remain in sterile packaging until ready to inject [18F]FDG into the saline bag</td>
		</tr>
		<tr>
			<td>--100mL 0.9% saline bag</td>
			<td>Baxter Pharmaceutical, IL, USA</td>
			<td>AHB1307</td>
			<td>Remain in sterile packaging until ready to inject [18F]FDG</td>
		</tr>
		<tr>
			<td>Radiochemistry Lab Supplies</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>--Heraeus Megafuge 16 centrifuge; Rotor Bioshield 720</td>
			<td>ThermoScientific MA, USA</td>
			<td>75004230</td>
			<td>Relative Centrifugal Force = 724 Our settings are 2000RPM for 5mins. Acceleration and deceleration curves set to 8</td>
		</tr>
		<tr>
			<td>--Single well counter</td>
			<td>Laboratory Technologies, Inc. IL, USA</td>
			<td>630-365-1000</td>
			<td>Complete daily quality control (includes background count) and protocol set to 18F and 4mins. Cross calibration is performed between the well counter, dose calibrator and scanner on a bi-monthly basis.</td>
		</tr>
		<tr>
			<td>--Pipette</td>
			<td>ISG Xacto, Vienna, Austria</td>
			<td>LI10434</td>
			<td>We use a 100-1000 &#956;L set to 1000&#956;L. It is calibrated annually.</td>
		</tr>
		<tr>
			<td>--12-15 plasma counting tubes</td>
			<td>Techno PLAS; SA Australia</td>
			<td>P10316SU</td>
			<td>Marked in the same manner as the LH blood tubes</td>
		</tr>
		<tr>
			<td>--12-15 pipette tips</td>
			<td>Expell Capp, Denmark</td>
			<td>5130140-1</td>
			<td></td>
		</tr>
		<tr>
			<td>--3 test tube racks</td>
			<td>Generic</td>
			<td></td>
			<td>Checked with a Geiger counter to ensure there is no radiation contamination on them</td>
		</tr>
		<tr>
			<td>--500mL volumetric flask and distilled water</td>
			<td>Generic</td>
			<td></td>
			<td>Need approximately 500mL of distilled water to prepare the reference for gamma counting</td>
		</tr>
		<tr>
			<td>--Synchronised clocks in scanner room, console and radiochemistry lab</td>
			<td>Generic</td>
			<td></td>
			<td>Synchronisation checks are routinely completed in the facility on a weekly basis</td>
		</tr>
		<tr>
			<td>--Haemoglobin Monitor</td>
			<td>EKF Diagnostic Cardiff, UK Haemo Control.</td>
			<td>3000-0810-6801</td>
			<td>Manufacturer recommended quality control performed before testing on participant&#8217;s blood sample.</td>
		</tr>
		<tr>
			<td>--Glucometre</td>
			<td>Roche Accu-Chek</td>
			<td>6870252001</td>
			<td>Accu-Chek Performa is used to measure participant blood sugar levels in mmol/L. Quality control is performed daily using high and low concentration solution control test.</td>
		</tr>
		<tr>
			<td>Cannulating Equipment</td>
			<td></td>
			<td></td>
			<td>Check expiry dates and train NMT to prepare aseptically for cannulation.</td>
		</tr>
		<tr>
			<td>--Regulation tourniquet</td>
			<td>CBC Classic Kimetec GmBH</td>
			<td>K5020</td>
			<td></td>
		</tr>
		<tr>
			<td>--20, 22 and 24 gauge cannulas</td>
			<td>Braun, Melsungen Germany</td>
			<td>4251644-03; 4251628-03; 4251601-03</td>
			<td></td>
		</tr>
		<tr>
			<td>--tegaderm dressings</td>
			<td>3M, MN USA</td>
			<td>1624W</td>
			<td></td>
		</tr>
		<tr>
			<td>--alcohol and chlorhexidine swabs</td>
			<td>Reynard Health Supplies, NSW Australia</td>
			<td>RHS408</td>
			<td></td>
		</tr>
		<tr>
			<td>--0.9% saline 10mL ampoules; for flushes</td>
			<td>Pfizer, NY, USA</td>
			<td>61039117</td>
			<td></td>
		</tr>
		<tr>
			<td>--10mL syringes</td>
			<td>Terumo Tokyo, Japan</td>
			<td>SS+10L</td>
			<td></td>
		</tr>
		<tr>
			<td>--3-way tap</td>
			<td>Becton Dickinson Connecta</td>
			<td>394600</td>
			<td></td>
		</tr>
		<tr>
			<td>--IV bung</td>
			<td>Safsite Braun PA USA</td>
			<td>415068</td>
			<td></td>
		</tr>
		<tr>
			<td>--Optional extension tube, microbore extension set</td>
			<td>M Devices, Denmark</td>
			<td>IV054000</td>
			<td></td>
		</tr>
		<tr>
			<td>Scanner Room Equipment</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>--Siemens Biograph 3T mMR</td>
			<td>Siemens, Erlangen, Germany</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>--Portable lead barrier shield</td>
			<td>Gammasonics</td>
			<td>Custom-built</td>
			<td>MR-conditional lead barrier shield. Positioned at the 2000 Gauss line with the castors locked to provide additional shielding of the radioactivity connected to the infusion pump.</td>
		</tr>
		<tr>
			<td>--Infusion pump BodyGuard 323 MR-conditional infusion pump</td>
			<td>Caesarea Medical Electronics</td>
			<td>300-040XP</td>
			<td>MR-compatible. This model is cleared for use on 1.5 and 3T scanners at 2000 Gauss with castors locked.</td>
		</tr>
		<tr>
			<td>--Infusion pump tubing</td>
			<td>Caesarea Medical Electronics</td>
			<td>100-163X2YNKS</td>
			<td>Tubing is administration set with an anti-siphon valve and male luer lock (REF 100-163X2YNKS).</td>
		</tr>
		<tr>
			<td>--Lead bricks</td>
			<td>Custom built</td>
			<td></td>
			<td>Tested for ferromagnetic translational force</td>
		</tr>
		<tr>
			<td>Other Equipment</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>--Syringe shields</td>
			<td>Biodex, NY USA</td>
			<td>Custom-built</td>
			<td>There is a 5mL tungsten syringe shield that is MR-safe, as well as a 50mL lead shield that has been tested for ferromagnetic attraction prior to use in the MR-PET scanner. It is used to transport the radioactive dose from the radiochemistry lab into the scanner to minimise radiation exposure to the NMT.</td>
		</tr>
		<tr>
			<td>--Geiger counter Model 26-1 Integrated Frisker</td>
			<td>Ludlum Measurements, Inc. TX USA</td>
			<td>48-4007</td>
			<td>This is calibrated annually and used to monitor potential contamination and waste. It is not taken into the MR-PET scanner.</td>
		</tr>
	</tbody>
</table>

radiotracer administration for high temporal resolution positron emission tomography of the human brain: application to fdg-fpet

Functional positron emission tomography (fPET) provides a method to track molecular targets in the human brain. With a radioactively-labelled glucose analogue, 18F-fluordeoxyglucose (FDG-fPET), it is now possible to measure the dynamics of glucose metabolism with temporal resolutions approaching those of functional magnetic resonance imaging (fMRI). This direct measure of glucose uptake has enormous potential for understanding normal and abnormal brain function and probing the effects of metabolic and neurodegenerative diseases. Further, new advances in hybrid MR-PET hardware make it possible to capture fluctuations in glucose and blood oxygenation simultaneously using fMRI and FDG-fPET.
The temporal resolution and signal-to-noise of the FDG-fPET images is critically dependent upon the administration of the radiotracer. This work presents two alternative continuous infusion protocols and compares them to a traditional bolus approach. It presents a method for acquiring blood samples, time-locking PET, MRI, experimental stimulus, and administering the non-traditional tracer delivery. Using a visual stimulus, the protocol results show cortical maps of the glucose-response to external stimuli on an individual level with a temporal resolution of 16 s.

Study-specific methods 
Here, study-specific details for the representative results are reported. These details are not critical to the procedure and will vary across studies.
Participants and task design 
Participants (n = 3, Table 2) underwent a simultaneous BOLD-fMRI/FDG-fPET study. As this manuscript focuses on the PET acquisition protocol, MRI re...

Watch this Scientific Journal Video about Radiotracer Administration for High Temporal Resolution Positron Emission Tomography of the Human Brain: Application to FDG-fPET at JoVE.com

Radiotracer Administration for High Temporal Resolution Positron Emission Tomography of the Human Brain: Application to FDG-fPET

This manuscript describes two radiotracer administration protocols for FDG-PET (constant infusion and bolus plus infusion) and compares them to bolus administration. Temporal resolutions of 16 s are achievable using these protocols.

Functional positron emission tomography (fPET) provides a method to track molecular targets in the human brain. With a radioactively-labelled glucose ...

The advent of simultaneous MR/PET scanners has sparked renewed interest in FDG-PET imaging in cognitive neuroscience. With these developments, researchers have focused on improving the temporal resolution of FDG-PET to approach the standards of BOLD fMRI. Traditional FDG-PET acquisitions using the bolus method provide a static measure of glucose uptake averaged across the scan period.In this protocol, we describe two alternate methods of FDG-PET acquisition:constant infusion and hybrid bolus infusion. These techniques provide a superior temporal resolution of up to 16 seconds which compares very favorably to the traditional bolus approach. The improved temporal resolution achieved with these administration protocols will provide important information about the dynamic use of glucose energy in the brain.To begin, have the nuclear medicine technologist or NMT take into account the estimated start of scanning while preparing the dose. To prepare the dose, aseptically add the radiotracer to the saline bag. Then, prepare the priming dose by withdrawing 20 milliliters from the bag into a syringe and cap it.Calibrate this 20 milliliter syringe and label. Then, to prepare the dose, use a 50 milliliter syringe to withdraw 60 milliliters from the bag and cap with a red combi-stopper. Store both syringes in the radiochemistry lab until ready to scan.Next, prepare the scanner room to ensure that all blood collection equipment is within easy reach of the collection site. Place underpads on any surface that will hold blood containers and place bins for regular waste and biohazardous waste within easy reach of the blood collection site. Set up the infusion pump in the scanner room on the side that will be connected to the participant.Build lead bricks around the base of the pump and place the lead shield in front of the pump. Connect the tubing for the infusion pump that will deliver the infusion to the participant and ensure the correct infusion rate has been entered. Then, connect the 20 milliliter priming dose to the infusion pump.On the end of the tubing that will be connected to the participant, attach a three-way tap and an empty 20 milliliter syringe. Ensure that the tap is positioned to allow the FDG solution to flow from the priming dose through the tubing and collect only into the empty syringe. Pre-set the infusion pump to prime a volume of 15 milliliters and select the prime button on the pump.Follow the prompts to prime the line. Lastly, attach the 50 milliliter dose syringe to the infusion pump in place of the priming dose. Begin by escorting the participant to the testing area.Conduct the informed consent procedure and complete safety screens with the participant. Have the NMT review safety for PET scanning and then have the radiographer review safety for MRI scanning with the participant such as exclusion for pregnancy, medical or nonmedical metallic implants. Then, cannulate the participant with two 22 gauge cannulas, one for dose administration and the other for blood sampling.Collect a 10 milliliter baseline blood sample while cannulating and disconnect all saline flushes under pressure to maintain patency of the line. Next, position the participant in the scanner and cover with a disposable blanket to maintain a comfortable body temperature. Flush the cannula to ensure it is patent with minimal resistance before connecting the infusion line.Tape the tubing to the participant's wrist and instruct them to keep their arm straight during the scan. Check the cannula that will be used for plasma samples to ensure that it is able to withdraw blood with minimal resistance. At the start of the scan, situate the NMT in the scanner room to monitor the infusion equipment.Ensure the NMT wears hearing protection and uses the barrier shield to minimize radiation exposure from the dose where possible. Perform the localizer scan to ensure that the participant is in the correct position and check the details for the PET acquisition. Then, signal the NMT to start the infusion pump 30 seconds after the start of the PET acquisition.Have the NMT observe the pump to ensure it has started to infuse the FDG and ensure there is no immediate occlusion of the line. Initiate any external stimulus such as the start of a functional run and calculate the times for blood samples. Take blood samples at regular time intervals and at the mid collection point, mark this time on the record form as the actual time of sample.Finally, connect the 10 milliliter syringe to the vacutainer and then deposit the blood into the relevant blood tube. Quickly flush the cannula with 10 milliliters of saline disconnected under pressure to minimize any chance of line clotting. In the lab, spin all samples at a relative centrifugal force of 724 times g.After the sample has been spun, place the tube in the pipetting rack. Remove the tube cap to not disturb sample separation and place a labeled counting tube in the rack. Next, ensure the tip is securely fastened to the pipette and have a tissue ready for any drips.Steadily pipette 1, 000 microliters of plasma from the blood tube, transfer to the counting tube, and replace the lids on the counting tube and blood tube. Finally, place the counting tube into the well counter and count for four minutes. Dispose of any blood product waste in biohazard bags.The largest peak plasma radioactivity concentration was obtained using the bolus method and the smallest using the constant infusion method. Critically, plasma radioactivity levels were sustained for the longest duration in the hybrid bolus infusion protocol and was minimally varied for a period of approximately 40 minutes. This shows that the hybrid bolus infusion protocol provides a relatively stable plasma radioactivity until the sensation of the infusion.The PET results indicate that the hybrid bolus infusion participant showed clearer differences between the ROIs compared to bolus only and infusion only participants. In the bolus only protocol, there is a sharp increase in signal following the bolus. In the bolus and infusion protocol, there is a sharp increase in uptake at the start of the scan that is of smaller magnitude than in the bolus only protocol and the uptake continues at a comparatively faster rate for the duration of the scan.Lastly, the signal increased roughly linearly across the recording period for the three protocols. The slope of the line was highest for the infusion only protocol, intermediate for the bolus only, and smallest for the bolus and infusion protocol. This is consistent with the conclusion that the hybrid protocol provides a more stable signal across the recording period.The critical point in the protocol is the start of the acquisition. At this point, the beginning of the PET scan must be timed to the start of the BOLD fMRI sequence as well as to the start of the stimulus presentation so as to facilitate accurate analysis. In order for the procedures to fall correctly within the short time period, communication is important to ensure all staff members are adequately prepared prior to the start of the scan.Simultaneous MRI/PET requires the management of two hazardous scenarios:the presence of the strong magnetic field and the administration of radiation to participants. All staff and participants must comply with the instructions of the supervising radiographer and nuclear medicine technologist.

radiotracer-administration-for-high-temporal-resolution-positron

Research

JoVE Journal

Behavior

Stereotactic Injection of MicroRNA-expressing Lentiviruses to the Mouse Hippocampus CA1 Region and Assessment of the Behavioral Outcome

MicroRNAs (miRNAs) are small regulatory single-stranded RNA molecules around 22 nucleotides long that may each target numerous mRNA transcripts and dim an entire gene expression pathway by inducing destruction and/or inhibiting translation of these targets. Several miRNAs play key roles in maintaining neuronal structure and function and in higher-level brain functions, and methods are sought for manipulating their levels for exploring these functions. Here, we present a direct in vivo method for examining the cognitive consequences of enforced miRNAs excess in mice by stereotactic injection of miRNA-encoding virus particles. Specifically, the current protocol involves injection into the hippocampal CA1 region, which contributes to mammalian memory consolidation, learning, and stress responses, and offers a convenient injection site. The coordinates are measured according to the mouse bregma and virus perfusion is digitally controlled and kept very slow. After injection, the surgery wound is sealed and the animals recover. Lentiviruses encoding silencers of the corresponding mRNA targets serve to implicate the specific miRNA/target interaction responsible for the observed effect, with na&iuml;ve mice, mice injected with saline and mice injected with "empty" lentivirus vectors as controls. One month post-injection, the animals are examined in the Morris Water Maze (MWM) for assessing their navigation learning and memory abilities. The MWM is a round tank filled with colored water with a small platform submerged 1 cm below the water surface. Steady visual cues around the tank allow for spatial navigation (sound and the earth's magnetic field may also assist the animals in navigating). Video camera monitoring enables measuring the route of swim and the time to find and amount the platform. The mouse is first taught that mounting the hidden platform offers an escape from the enforced swimming; it is then tested for using this escape and finally, the platform is removed and probe tests examine if the mouse remembers its previous location. Repeated tests over several consecutive days highlight improved performance of tested mice at shorter latencies to find and mount the platform, and as more direct routes to reach the platform or its location. Failure to show such improvement represents impaired learning and memory and/or anxiety, which may then be tested specifically (e.g. in the elevated plus maze). This approach enables validation of specific miRNAs and target transcripts in the studied cognitive and/or stress-related processes.

MicroRNAs have significant roles in brain structure and function. Here we describe a method to enforce hippocampal miRNA over-expression using stereotactic injection of an engineered miRNA-expressing lentivirus. This approach can serve as a relatively rapid way to assess the in vivo effects of over-expressed miRNAs in specific brain regions.

MicroRNAs (miRNAs) are small regulatory single-stranded RNA molecules around 22 nucleotides long that may each target numerous mRNA transcripts and dim an ...

Uncovering Beat Deafness: Detecting Rhythm Disorders with Synchronized Finger Tapping and Perceptual Timing Tasks

A set of behavioral tasks for assessing perceptual and sensorimotor timing abilities in the general population (i.e., non-musicians) is presented here with the goal of uncovering rhythm disorders, such as beat deafness. Beat deafness is characterized by poor performance in perceiving durations in auditory rhythmic patterns or poor synchronization of movement with auditory rhythms (e.g., with musical beats). These tasks include the synchronization of finger tapping to the beat of simple and complex auditory stimuli and the detection of rhythmic irregularities (anisochrony detection task) embedded in the same stimuli. These tests, which are easy to administer, include an assessment of both perceptual and sensorimotor timing abilities under different conditions (e.g., beat rates and types of auditory material) and are based on the same auditory stimuli, ranging from a simple metronome to a complex musical excerpt. The analysis of synchronized tapping data is performed with circular statistics, which provide reliable measures of synchronization accuracy (e.g., the difference between the timing of the taps and the timing of the pacing stimuli) and consistency. Circular statistics on tapping data are particularly well-suited for detecting individual differences in the general population. Synchronized tapping and anisochrony detection are sensitive measures for identifying profiles of rhythm disorders and have been used with success to uncover cases of poor synchronization with spared perceptual timing. This systematic assessment of perceptual and sensorimotor timing can be extended to populations of patients with brain damage, neurodegenerative diseases (e.g., Parkinson&#8217;s disease), and developmental disorders (e.g., Attention Deficit Hyperactivity Disorder).

Behavioral tasks that allow for the assessment of perceptual and sensorimotor timing abilities in the general population (i.e., non-musicians) are presented. Synchronization of finger tapping to the beat of an auditory stimuli and detecting rhythmic irregularities provides a means of uncovering rhythm disorders.

A set of behavioral tasks for assessing perceptual and sensorimotor timing abilities in the general population (i.e., non-musicians) is presented here ...

Application of MultiColor FlpOut Technique to Study High Resolution Single Cell Morphologies and Cell Interactions of Glia in Drosophila

Cells display different morphologies and complex anatomical relationships. How do cells interact with their neighbors? Do the interactions differ between cell types or even within a given type? What kinds of spatial rules do they follow? The answers to such fundamental questions in vivo have been hampered so far by a lack of tools for high resolution single cell labeling. Here, a detailed protocol to target single cells with a MultiColor FlpOut (MCFO) technique is provided. This method relies on three differently tagged reporters (HA, FLAG and V5) under UAS control that are kept silent by a transcriptional terminator flanked by two FRT sites (FRT-stop-FRT). A heat shock pulse induces the expression of a heat shock-induced Flp recombinase, which randomly removes the FRT-stop-FRT cassettes in individual cells: expression occurs only in cells that also express a GAL4 driver. This leads to an array of differently colored cells of a given cell type that allows the visualization of individual cell morphologies at high resolution. As an example, the MCFO technique can be combined with specific glial GAL4 drivers to visualize the morphologies of the different glial subtypes in the adult Drosophila brain.

Application of MultiColor FlpOut Technique to Study High Resolution Single Cell Morphologies and Cell Interactions of Glia in Drosophila

Cells display different morphologies and establish a variety of interactions with their neighbors. This protocol describes how to reveal the morphology of single cells and to investigate cell-cell interaction by using the well-established Gal4/UAS expression system.

Cells display different morphologies and complex anatomical relationships. How do cells interact with their neighbors? Do the interactions differ between ...

An Automated T-maze Based Apparatus and Protocol for Analyzing Delay- and Effort-based Decision Making in  Free Moving Rodents

Many neurological and psychiatric patients demonstrate difficulties and/or deficits in decision making. Rodent models are helpful to produce a deeper understanding of the neurobiological causes underlying the decision-making problems. A cost-benefit based T-maze task is used for measuring decision making in which rodents choose between a high reward arm (HRA) and a low reward arm (LRA). There are two paradigms of the T-maze decision-making task, one in which the cost is a time delay and the other in which it is physical effort. Both paradigms require a tedious and labor-intensive management of experimental animals, multiple doors, pellet reward, and arm choice recordings. In the current work, we invented an apparatus based on traditional T-maze with full automation for pellet delivery, door management and choice recordings. This automated setup can be used for the evaluation of both delay- and effort-based decision making in rodents. With the protocol described here, our lab investigated the decision-making phenotypes of multiple genetically modified mice. In the representative data, we showed that the mice with ablated medial habenular showed aversions of both delay and effort and tended to choose the immediate and effortless reward. This protocol helps to decrease the variability caused by experimenter intervention and to enhance experiment efficiency. In addition, chronic silicon probe or microelectrode recording, fiber-optic imaging and/or manipulation of neural activity can be easily applied during the decision-making task using the setup described here.

This article introduces an automated T-maze apparatus that we invented, and a protocol based on this apparatus for analyzing delay-based decision making and effort-based decision making in free moving rodents.

Many neurological and psychiatric patients demonstrate difficulties and/or deficits in decision making. Rodent models are helpful to produce a deeper ...

Using Eye Movements Recorded in the Visual World Paradigm to Explore the Online Processing of Spoken Language

In a typical eye tracking study using the visual world paradigm, participants' eye movements to objects or pictures in the visual workspace are recorded via an eye tracker as the participant produces or comprehends a spoken language describing the concurrent visual world. This paradigm has high versatility, as it can be used in a wide range of populations, including those who cannot read and/or who cannot overtly give their behavioral responses, such as preliterate children, elderly adults, and patients. More importantly, the paradigm is extremely sensitive to fine grained manipulations of the speech signal, and it can be used to study the online processing of most topics in language comprehension at multiple levels, such as the fine grained acoustic phonetic features, the properties of words, and the linguistic structures. The protocol described in this article illustrates how a typical visual world eye tracking study is conducted, with an example showing how the online processing of some semantically complex statements can be explored with the visual world paradigm.

The visual world paradigm monitors participants' eye movements in the visual workspace as they are listening to or speaking a spoken language. This paradigm can be used to investigate the online processing of a wide range of psycholinguistic questions, including semantically complex statements, such as disjunctive statements.

In a typical eye tracking study using the visual world paradigm, participants' eye movements to objects or pictures in the visual workspace are recorded ...

Using the Visual World Paradigm to Study Sentence Comprehension in Mandarin-Speaking Children with Autism

Sentence comprehension relies on the ability to rapidly integrate different types of linguistic and non-linguistic information. However, there is currently a paucity of research exploring how preschool children with autism understand sentences using different types of cues. The mechanisms underlying sentence comprehension remains largely unclear. The present study presents a protocol to examine the sentence comprehension abilities of preschool children with autism. More specifically, a visual world paradigm of eye-tracking is used to explore the moment-to-moment sentence comprehension in the children. The paradigm has multiple advantages. First, it is sensitive to the time course of sentence comprehension and thus can provide rich information about how sentence comprehension unfolds over time. Second, it requires minimal task and communication demands, so it is ideal for testing children with autism. To further minimize the computational burden of children, the present study measures eye movements that arise as automatic responses to linguistic input rather than measuring eye movements that accompany conscious responses to spoken instructions.

We present a protocol to examine the use of morphological cues during real-time sentence comprehension by children with autism.

Sentence comprehension relies on the ability to rapidly integrate different types of linguistic and non-linguistic information. However, there is ...

Application of 3D Printing in the Construction of Burr Hole Ring for Deep Brain Stimulation Implants

3D printing has been widely applied in the medical field since the 1980s, especially in surgery, such as preoperative simulation, anatomical learning and surgical training. This raises the possibility of using 3D printing to construct a neurosurgical implant. Our previous works took the construction of the burr hole ring as an example, described the process of using softwares like computer aided design (CAD), Pro/Engineer (Pro/E) and 3D printer to construct physical products. That is, a total of three steps are required, the drawing of 2D-image, the construction of 3D-image of burr hole ring, and using a 3D printer to print the physical model of burr hole ring. This protocol shows that the burr hole ring made of carbon fiber can be rapidly and accurately molded by 3D printing. It indicated that both CAD and Pro/E softwares can be used to construct the burr hole ring via integrating with the clinical imaging data and further applied 3D printing to make the individual consumables.

Here, we present a protocol to demonstrate 3D printing in the construction of deep brain stimulation implants.

3D printing has been widely applied in the medical field since the 1980s, especially in surgery, such as preoperative simulation, anatomical learning and ...

Human Circadian Phenotyping and Diurnal Performance Testing in the Real World

In our continuously developing &#39;around the clock&#39; society, there is a need to increase our understanding of how changes in biology, physiology and psychology influence our health and performance. Embedded within this challenge, is the increasing need to account for individual differences in sleep and circadian rhythms, as well as to explore the impact of time of day on performance in the real world. There are a number of ways to measure sleep and circadian rhythms from subjective questionnaire-based methods to objective sleep/wake monitoring, actigraphy and analysis of biological samples. This paper proposes a protocol that combines multiple techniques to categorize individuals into Early, Intermediate or Late circadian phenotype groups (ECPs/ICPs/LCPs) and recommends how to conduct diurnal performance testing in the field. Representative results show large differences in rest-activity patterns derived from actigraphy, circadian phase (dim light melatonin onset and peak time of cortisol awakening response) between circadian phenotypes. In addition, significant differences in diurnal performance rhythms between ECPs and LCPs emphasizes the need to account for circadian phenotype. In summary, despite the difficulties in controlling influencing factors, this protocol allows a real-world assessment of the impact of circadian phenotype on performance. This paper presents a simple method to assess circadian phenotype in the field and supports the need to consider time of day when designing performance studies.

Here, we present a method to investigate diurnal rhythms in performance following accurate categorization of participants into circadian phenotype groups based on the Munich ChronoType Questionnaire, gold standard circadian phase biomarkers and actigraphic measures.

In our continuously developing &#39;around the clock&#39; society, there is a need to increase our understanding of how changes in biology, physiology and ...

SECONDs Administration Guidelines: A Fast Tool to Assess Consciousness in Brain-injured Patients

Establishing an accurate diagnosis is crucial for patients with disorders of consciousness (DoC) following a severe brain injury. The Coma Recovery Scale-Revised (CRS-R) is the recommended behavioral scale for assessing the level of consciousness among these patients, but its long duration&#160;of administration is a major hurdle in clinical settings. The Simplified Evaluation of CONsciousness Disorders (SECONDs) is a shorter scale that was developed to tackle this issue. It consists of six mandatory items&#160;(observation, command-following, visual pursuit, visual fixation, oriented behaviors, and arousal)&#160;and two conditional items&#160;(communication and localization to pain). The score ranges between 0 and 8 and corresponds to a specific diagnosis (i.e., coma, unresponsive wakefulness syndrome, minimally conscious state minus/plus, or emergence from the minimally conscious state). A first validation study on patients with prolonged DoC showed high concurrent validity and intra- and inter-rater reliability. The SECONDs requires less training than the CRS-R and its administration lasts about 7 minutes (interquartile range: 5-9 minutes). An additional index score allows the more precise tracking of a patient&#39;s behavioral fluctuation or evolution over time. The SECONDs is therefore a fast and valid tool for assessing the level of consciousness in patients with severe brain injury. It can easily be used by healthcare staff and implemented in time-constrained clinical settings, such as intensive care units, to help&#160;decrease misdiagnosis rates and to optimize treatment decisions. These administration guidelines provide detailed instructions for administering the SECONDs in a standardized and reproducible manner, which is an essential requirement for achieving a reliable diagnosis.

These guidelines can be used for the administration of the Simplified Evaluation of CONsciousness Disorders (SECONDs), a short behavioral tool developed to diagnose brain-injured patients in time-constrained settings. This scale examines command-following, communication, visual pursuit, fixation, pain localization, oriented movements and arousal.

Establishing an accurate diagnosis is crucial for patients with disorders of consciousness (DoC) following a severe brain injury. The Coma Recovery ...

The Motivation for Alcohol Reward: Predictors of Progressive-Ratio Intravenous Alcohol Self-Administration in Humans

The Progressive Ratio (PR) self-administration paradigm is a common pre-clinical method used to examine the motivation for a drug attributed to a craving, reward, or the relief of negative affect. The Computer-assisted Alcohol Infusion System (CAIS) enables intravenous alcohol self-administration behavior in humans. This system provides the investigator with control over the trajectory of each incremental breath alcohol concentration (BrAC) reward and the maximum BrAC allowed in a session. This paradigm allows participants to earn these alcohol rewards using a sequence of button presses specified by the investigator. The system employs a physiologically-based pharmacokinetic model-based algorithm to achieve the same incremental BrAC exposure in every participant. Participants (n = 11) took part in two identical sessions to examine test-retest reliability, and an additional group (n = 73) completed a single session. Sessions began with a 25 min priming phase: participants were instructed to press a button an increasing number of times per reward, accumulating four standardized incremental BrAC trajectories. The second phase comprised an ad-lib, PR paradigm lasting 125 min. Each reward required an increasing number of button presses. Measures of self-administration included: average and peak BrAC, total rewards earned, total grams of ethanol consumed per unit of total body water, the total number of button presses, and the average rate of button pressing. Self-administration measures were highly correlated both between and within sessions, demonstrating test-retest reliability and internal consistency. Recent drinking history was strongly associated with self-administration measures; heavier drinkers chose greater alcohol self-administration. These results indicate the reliability and sensitivity of this progressive-ratio intravenous alcohol self-administration method for assessing the motivational properties of alcohol, with the potential for improved testing of the efficacy of new medications thought to reduce consumption of alcohol. This method can be used to understand the genetic and environmental determinants of alcohol self-administration in humans.

This study aims to show that the Progressive-Ratio Computer-assisted Alcohol-Infusion System (CAIS) paradigm is a reliable and sensitive method that can be used to examine the motivating properties associated with alcohol self-administration in humans.

The Progressive Ratio (PR) self-administration paradigm is a common pre-clinical method used to examine the motivation for a drug attributed to a craving, ...