Name: acquisition of resting-state functional magnetic resonance imaging data in the rat
Uploaded: 2021-08-28T15:00:00
Description: Watch this Scientific Journal Video about Acquisition of Resting-State Functional Magnetic Resonance Imaging Data in the Rat at JoVE.com

This work was supported by funding from the National Institute of Health (NIH)&#39;s National Institute on Drug Abuse (NIDA) [DJW, EDKS, and EMB were supported by Grant R21DA044501 awarded to Alan I. Green and DJW was supported by Grant T32DA037202 to Alan J. Budney] and the National Institute on Alcohol Abuse and Alcoholism (NIAAA) [Grant F31AA028413 to Emily D. K. Sullivan]. Additional support was provided through Alan I. Green&#39;s endowed fund as the Raymond Sobel Professor of Psychiatry at Dartmouth.
Hanbing Lu is supported by the National Institute on Drug Abuse Intramural Research Program, NIH.
The authors wish to acknowledge and thank the late Alan I. Green. His unwavering dedication to the field of co-occurring disorders helped to establish collaboration among the authors. We thank him for his mentorship and guidance, which will be greatly missed.

All experiments were performed on a 9.4 T MRI scanner, and were approved by the Institutional Animal Care and Use Committee at Dartmouth College. Additional approval was obtained to record and show the animals used in the video and figures below.
1. Preparations before scanning
<ol>
	<li>Subcutaneous infusion line
	<ol>
		<li>Partially remove a 23 G needle from its package so that the needle point remains sterile.</li>
		<li>Securely hold the hub of the needle and use a razor blade to score ...

<ol>
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Stability of the animal, both physically and physiologically, is key to obtaining high-quality resting-state data. This protocol achieves stability by moving through four distinct phases of anesthesia. It is imperative that the animal has met the set physiological thresholds before moving to the next phase of anesthesia; since this method relies on physiological autoregulatory mechanisms, individual animals may require slightly different amounts of time at each anesthesia phase. It is our experience that taking more time...

Resting-state functional magnetic resonance imaging (rs-fMRI) is a measure of the blood-oxygen-level-dependent (BOLD) signal when the brain is at rest and not engaged in any particular task. These signals can be used to measure correlations between brain regions to determine the functional connectivity within neural networks. rs-fMRI is widely used in clinical studies due to its non-invasiveness and the low amount of effort required of patients (as compared to task-based fMRI) making it optimal for diverse patient populations1.
Technological advances have allowed rs-fMRI to be adapted for use in rodent models to uncover mechanisms underlying disease states (see reference2 for review). Preclinical animal models, including disease or knockout models, allow a wide range of experimental manipulations not applicable in humans, and studies can also make use of post-mortem samples to further enhance experiments2. Nevertheless, due to the difficulty in both limiting motion and mitigating stress, MRI acquisition in rodents is traditionally performed under anesthesia. Anesthetic agents, depending on their pharmacokinetics, pharmacodynamics, and molecular targets, influence brain blood flow, brain metabolism, and potentially affect neurovascular coupling pathways.
There have been numerous efforts to develop anesthetic protocols that preserve neurovascular coupling and brain network function3,4,5,6,7,8. We previously reported an anesthetic regime that applied a low dose of isoflurane along with a low dose of the &#945;2 adrenergic receptor agonist dexmedetomidine9. Rats under this method of anesthesia exhibited robust BOLD responses to whisker stimulation in regions consistent with established projection pathways (ventrolateral and ventromedial thalamic nuclei, primary and secondary somatosensory cortex); large-scale resting-state brain networks, including the default mode network10,11 and salience network12 have also been consistently detected. Furthermore, this anesthetic protocol allows for repeated imaging on the same animal, which is important for monitoring the disease progression and the effect of experimental manipulations longitudinally.
In the present study, we detail the experimental setup, animal preparation, and physiological monitoring procedures involved. In particular, we describe the specific anesthetic phases and acquisition of scans during each phase. Data quality is assessed following each resting-state scan. A brief summary of post-scan analysis is also included in the discussion. Laboratories interested in uncovering the potential of using rs-fMRI in rats will find this protocol useful.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>9.4T MRI</td>
			<td>Varian/Bruker</td>
			<td></td>
			<td>Varian upgraded with Bruker console running Paravision 6.0.1 software</td>
		</tr>
		<tr>
			<td>Air-Oxygen Mixer</td>
			<td>Sechrist</td>
			<td>Model 3500CP-G</td>
			<td></td>
		</tr>
		<tr>
			<td>Analysis of Functional NeuroImages (AFNI)</td>
			<td>NIMH/NIH</td>
			<td>Version AFNI_18.3.03</td>
			<td>Freely available at: https://afni.nimh.nih.gov/</td>
		</tr>
		<tr>
			<td>Animal Cradle</td>
			<td>RAPID Biomedical</td>
			<td>LHRXGS-00563</td>
			<td>rat holder with bite bar, nose cone and ear bars</td>
		</tr>
		<tr>
			<td>Animal Physiology Monitoring &#38; Gating System</td>
			<td>SAII</td>
			<td>Model 1025</td>
			<td>MR-compatible system including oxygen saturation, temperature, respiration and fiber optic pulse oximetry add-on</td>
		</tr>
		<tr>
			<td>Antisedan (atipamezole hydrochloride)</td>
			<td>Patterson Veterinary</td>
			<td>07-867-7097</td>
			<td>Zoetis, Manufacturer Item #10000449</td>
		</tr>
		<tr>
			<td>Ceramic MRI-Safe Scissors</td>
			<td>MRIequip.com</td>
			<td>MT-6003</td>
			<td></td>
		</tr>
		<tr>
			<td>Clippers</td>
			<td>Patterson Veterinary</td>
			<td>07-882-1032</td>
			<td>Wahl touch-up trimmer combo kit, Manufacturer Item #09990-1201</td>
		</tr>
		<tr>
			<td>Dexmedesed (dexmedetomidine hydrochloride)</td>
			<td>Patterson Veterinary</td>
			<td>07-893-1801</td>
			<td>Dechra Veterinary Products, Manufacturer Item#17033-005-10</td>
		</tr>
		<tr>
			<td>Digital Rectal Thermometer Covers</td>
			<td>Medline</td>
			<td>MDS9608</td>
			<td></td>
		</tr>
		<tr>
			<td>FMRIB Software Library</td>
			<td>FMRIB</td>
			<td>MELODIC Version 3.15</td>
			<td>Freely available at: https://fsl.fmrib.ox.ac.uk/fsl/fslwiki</td>
		</tr>
		<tr>
			<td>Heating Pad</td>
			<td>Cara Inc.</td>
			<td>Model 50</td>
			<td></td>
		</tr>
		<tr>
			<td>Hemostat forceps, straight</td>
			<td>Kent Scientific</td>
			<td>INS750451-2</td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Patterson Veterinary</td>
			<td>07-893-1389</td>
			<td>Patterson Private Label, Manufacturer Item #14043-0704-06</td>
		</tr>
		<tr>
			<td>Isoflurane Vaporizer</td>
			<td>VetEquip Inc.</td>
			<td>911103</td>
			<td></td>
		</tr>
		<tr>
			<td>Lab Tape, 3/4&#34;</td>
			<td>VWR International</td>
			<td>89097-990</td>
			<td></td>
		</tr>
		<tr>
			<td>Needles, 23 gauge</td>
			<td>BD</td>
			<td>305145</td>
			<td>plastic hub removed</td>
		</tr>
		<tr>
			<td>Parafilm Laboratory Film</td>
			<td>Patterson Veterinary</td>
			<td>07-893-0260</td>
			<td>Medline Industries Inc., Manufacturer Item #HSFHS234526A</td>
		</tr>
		<tr>
			<td>Planar Surface Coil</td>
			<td>Bruker</td>
			<td>T12609</td>
			<td>2cm</td>
		</tr>
		<tr>
			<td>Polyethylene Tubing</td>
			<td>Braintree Scientific</td>
			<td>PE50 50FT</td>
			<td>0.023&#34; (inner diameter), 0.038&#34; (outer diameter)</td>
		</tr>
		<tr>
			<td>Puralube Ophthalmic Ointment</td>
			<td>Patterson Veterinary</td>
			<td>07-888-2572</td>
			<td>Dechra Veterinary Products, Manufacturer Item #211-38</td>
		</tr>
		<tr>
			<td>Sprague Dawley Rats</td>
			<td>Charles River</td>
			<td>400 SAS SD</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile 0.9% Saline Solution</td>
			<td>Patterson Veterinary</td>
			<td>07-892-4348</td>
			<td>Aspen Vet, Manufacturer Item #14208186</td>
		</tr>
		<tr>
			<td>Sterile Alcohol Prep Pads</td>
			<td>Medline</td>
			<td>MDS090735</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical Tape, 1&#34; (3M Durapore)</td>
			<td>Medline</td>
			<td>MMM15381Z</td>
			<td>3M Healthcare, &#34;wide medical tape&#34;</td>
		</tr>
		<tr>
			<td>Surgical White Paper Tape, 1/2&#34; (3M Micropore)</td>
			<td>Medline</td>
			<td>MMM15300</td>
			<td>3M Healthcare</td>
		</tr>
		<tr>
			<td>Syringes, 1 mL w/ 25 gauge needle</td>
			<td>BD</td>
			<td>309626</td>
			<td></td>
		</tr>
		<tr>
			<td>Syringes, 3 mL</td>
			<td>BD</td>
			<td>309657</td>
			<td></td>
		</tr>
		<tr>
			<td>Vented induction and scavenging system</td>
			<td>VetEquip Inc.</td>
			<td>942102</td>
			<td>2 liter induction chamber with active scavenging</td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td>411724</td>
			<td>omega flowmeter</td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td>931600</td>
			<td>scavenging cube, &#34;vacuum&#34;</td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td>921616</td>
			<td>nose cone, non-rebreathing</td>
		</tr>
	</tbody>
</table>

acquisition of resting-state functional magnetic resonance imaging data in the rat

Resting-state functional magnetic resonance imaging (rs-fMRI) has become an increasingly popular method to study brain function in a resting, non-task state. This protocol describes a preclinical survival method for obtaining rs-fMRI data. Combining low dose isoflurane with continuous infusion of the &#945;2 adrenergic receptor agonist dexmedetomidine provides a robust option for stable, high-quality data acquisition while preserving brain network function. Furthermore, this procedure allows for spontaneous breathing and near-normal physiology in the rat. Additional imaging sequences can be combined with resting-state acquisition creating experimental protocols with anesthetic stability of up to 5 h using this method. This protocol describes the setup of equipment, monitoring of rat physiology during four distinct phases of anesthesia, acquisition of resting-state scans, quality assessment of data, recovery of the animal, and a brief discussion of post-processing data analysis. This protocol can be used across a wide variety of preclinical rodent models to help reveal the resulting brain network changes that occur at rest.

Following each resting-state scan, stability is assessed using an independent component analysis (ICA; example script included in Supplementary Files). Figure 6 shows examples of component outputs from resting-state scans. Figure 6a shows a signal component from a scan with high stability. Note that spatially, the component has high regionality. Within the time course below the spatial component, the signal is stable and not predictable, indicat...

Watch this Scientific Journal Video about Acquisition of Resting-State Functional Magnetic Resonance Imaging Data in the Rat at JoVE.com

Acquisition of Resting-State Functional Magnetic Resonance Imaging Data in the Rat

This protocol describes a method for obtaining stable resting-state functional magnetic resonance imaging (rs-fMRI) data from a rat using low dose isoflurane in combination with low dose dexmedetomidine.

Resting-state functional magnetic resonance imaging (rs-fMRI) has become an increasingly popular method to study brain function in a resting, non-task ...

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