This research was supported by the National Institute of Mental Health MH 086960-01A1 (Christa K. McIntyre).

All procedures described in this protocol are carried out in accordance with the NIH Guide for the Care and Use of Laboratory Animals, and they were approved by the Institutional Animal Care and Use Committee of The University of Texas at Dallas.
1. Construction of VNS Cuffs
<ol>
	<li>Create a drilling tool by sawing off the sharp end of a 22 &#189; G needle.</li>
	<li>Run the now blunt end of the 22 &#189; G needle over a metal file several times to flatten it. Hold the needle at a 45&#186;...

The authors have no competing interests or conflicts.

We present here a protocol that is used to facilitate extinction of conditioned fear during a single session of exposure to conditioned cues19 and to modulate plasticity in the pathway between the infralimbic cortex and the basolateral amygdala that may mediate extinction learning20. A crucial step for the success of this protocol is the proper delivery of VNS during extinction training. Therefore, special care should be given to the construction of the cuff electrodes and the placement of the cuff ...

Classical fear conditioning provides a widely used animal model to study the biological basis of anxiety disorders. During fear conditioning, an aversive stimulus (the unconditioned stimulus, US, e.g., a footshock) is presented in conjunction with a neutral stimulus, such as a tone and/or a context (the conditioned stimulus; CS). During fear conditioning, associations between the CS and the US are formed. Eventually the presentation of the CS alone elicits a fear response (the conditioned response; CR). In fear extinction, the CS is presented repeatedly in the absence of the US, causing the CR to gradually diminish1. Thus, extinction of conditioned fear is an active process in which fearful behavioral responses to neutral stimuli are attenuated when they no longer predict aversive outcomes. Extinction of conditioned responses requires consolidation of new memories that compete with learned associations. A hallmark of anxiety disorders is impaired extinction2-4. Thus, extinction of conditioned fear in animal models serves as an important paradigm both for inhibitory learning and as a model of behavior therapy for human anxiety disorders5,6.
Because there is close correspondence between fear and human anxiety, it is thought that these studies can provide insight into the biological nature of anxiety-related disorders such as post-traumatic stress disorder and will help to develop strategies to treat them. An important goal of preclinical research is to aid extinction learning and to induce targeted plasticity in extinction circuits to consolidate extinction learning. Vagus nerve stimulation (VNS) is a minimally invasive neuroprosthetic approach that might be used to provide tight temporal and circuit-specific modulation of brain areas and synapses engaged in an ongoing task. A series of recent studies from Michael Kilgard&#39;s group at The University of Texas at Dallas have shown that pairing VNS with discrete sensory or motor stimuli (e.g., a tone or a lever pull) is highly effective in promoting cortical plasticity to treat tinnitus7, or to overcome motor deficits following stroke8-10. In addition, non-contingent VNS that occurs within a short time-window after learning similarly promotes cortical plasticity and enhances memory consolidation in rats and in humans11-13.
Considering the role of the vagus nerve in the parasympathetic pathway, it is not surprising that it could participate in modulating memories and synaptic plasticity. Highly emotional events tend to produce stronger memories than non-emotional memories. This is likely due to the influence of stress hormones on memory consolidation. Posttraining administration of the stress hormone adrenaline enhances memory consolidation in human and non-human animals, but adrenaline does not cross the blood-brain-barrier14, 15. Therefore, stress-induced adrenaline release must impact the brain indirectly to enhance memory consolidation. Strong evidence suggests that the vagus nerve may be the link between circulating adrenaline and the brain. Miyashita and Williams16 found that systemic administration of adrenaline increased vagal nerve firing, and increased levels of norepinephrine in the amygdala17. Systemic administration of adrenaline does not enhance memory consolidation when &#946;-adrenergic receptors are blocked in the amygdala18 suggesting that the vagus nerve plays a role in the pathway that turns emotionally arousing experiences into long-term memories.
Thus, pairing VNS with training has the potential to enhance the brain changes that support memory consolidation and exposure to conditioned cues in the absence of reinforcement enhances the consolidation of extinction learning in rats19,20. Here we describe the use of VNS as a tool to promote cortical plasticity and facilitate extinction of a conditioned fear response.

<table border="0" cellpadding="0" cellspacing="0">
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			<td>Alcohol</td>
			<td></td>
			<td></td>
			<td></td>
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			<td>Atropine</td>
			<td>Fisher</td>
			<td>A0132-5G</td>
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			<td>Betadine</td>
			<td>Henry Schein</td>
			<td>69066950</td>
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			<td>Hydrogen peroxide&#160;</td>
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			<td>209478</td>
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			<td>Henry Schein&#160;</td>
			<td>1129300</td>
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			<td>Marcaine</td>
			<td>Henry Schein</td>
			<td>6312615</td>
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			<td>Oxygen</td>
			<td>Home Depot</td>
			<td>304179</td>
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			<td>Pennicillin</td>
			<td>Fisher</td>
			<td>PENNA-10MU</td>
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			<td>Propane</td>
			<td>Home Depot</td>
			<td>304182</td>
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			<td>Xylazine</td>
			<td>Henry Schein</td>
			<td>4019308</td>
			<td></td>
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			<td></td>
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			<td></td>
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			<td>Tools</td>
			<td></td>
			<td></td>
			<td></td>
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			<td>Jewelery Torch</td>
			<td>Smith Equipment</td>
			<td>23-1001D</td>
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			<td>Sewing Needle</td>
			<td>Walgreens</td>
			<td>441831</td>
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			<td>#5 Forceps (2)</td>
			<td>Fine Science Tools</td>
			<td>11254-20</td>
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			<td>Soldering Iron</td>
			<td>Home Depot&#160;</td>
			<td>203525863</td>
			<td></td>
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			<td>AmScope SM-4TX-144A 3.5X-45X Circuit Board Boom Stereo Microscope + 144 LED</td>
			<td>AmScope</td>
			<td>SM-4TX-144A</td>
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			<td>Helping Hands</td>
			<td>A-M Systems&#160;</td>
			<td>726200</td>
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			<td>Scalpel Blade Holder</td>
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			<td>10003-12</td>
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			<td>Metal File</td>
			<td>Home Depot</td>
			<td>6601</td>
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			<td>Ruler</td>
			<td>Home Deopt</td>
			<td>202035324</td>
			<td></td>
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			<td>Curved Hemostats&#160;</td>
			<td>Fine Science Tools</td>
			<td>130009-12</td>
			<td></td>
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		<tr>
			<td>Fine Scissors</td>
			<td>Fine Science Tools</td>
			<td>14058-09</td>
			<td></td>
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			<td>Spatula</td>
			<td>Fine Science Tools</td>
			<td></td>
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			<td>Small Screwdriver</td>
			<td>Home Depot</td>
			<td>646507</td>
			<td></td>
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			<td>Magnetic Fixator Retraction System</td>
			<td>Fine Science Tools</td>
			<td>18200-04, 18200-01, 18200-05</td>
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			<td>Heating Pad</td>
			<td>Walgreens</td>
			<td>30294</td>
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			<td>Clippers</td>
			<td>Walgreens</td>
			<td>277966</td>
			<td></td>
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			<td>Sharpie</td>
			<td>Staples</td>
			<td>125328</td>
			<td></td>
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		<tr>
			<td>Ring Forceps</td>
			<td>Fine Science Tools</td>
			<td>11103-09</td>
			<td></td>
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		<tr>
			<td>Custom Micropipette Glass Tools (J shape and Straight) - Borosilicate glass</td>
			<td>Sutter Instrument</td>
			<td>B150-110-10</td>
			<td></td>
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		<tr>
			<td>Adson Forceps</td>
			<td>Fine Science Tools</td>
			<td>11006-12</td>
			<td></td>
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		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
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		<tr>
			<td>Cuffs</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
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			<td>Tubing</td>
			<td>Braintree Scientific Inc</td>
			<td>MRE-065</td>
			<td></td>
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			<td>Platinum Iridium Wire</td>
			<td>Medwire</td>
			<td>10IR9/49T</td>
			<td></td>
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			<td>Gold Pins</td>
			<td>Mill-Max</td>
			<td>1001-0-15-15-30-27-04-0</td>
			<td></td>
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		<tr>
			<td>Suture Thread</td>
			<td>Henry Schein</td>
			<td>100-5797</td>
			<td></td>
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		<tr>
			<td>22 G Needles</td>
			<td>Fisher&#160;</td>
			<td>14-815-525</td>
			<td></td>
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		<tr>
			<td>Paper Tape</td>
			<td>Fisher&#160;</td>
			<td>03-411-602</td>
			<td></td>
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		<tr>
			<td>Solder</td>
			<td>Home Deopt</td>
			<td>327793</td>
			<td></td>
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		<tr>
			<td>Flux&#160;</td>
			<td>Home Deopt</td>
			<td>300142</td>
			<td></td>
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			<td>Scalpel Blade, 10 or 15</td>
			<td>Stoelting</td>
			<td>52173-10</td>
			<td></td>
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		<tr>
			<td>Silastic Laboratory Tubing .51 mm ID x .94 mm OD</td>
			<td>Fisher&#160;</td>
			<td>508-002</td>
			<td></td>
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		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
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		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
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		<tr>
			<td>Headcaps</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Connector Pieces (male)</td>
			<td>Omnetics Connector Corporation</td>
			<td>A25001-004</td>
			<td></td>
		</tr>
		<tr>
			<td>Headcap pieces (female)</td>
			<td>Omnetics Connector Corporation</td>
			<td>A24001-004</td>
			<td></td>
		</tr>
		<tr>
			<td>Teets Dental Acrylic, Liquid and Powder</td>
			<td>A-M Systems</td>
			<td>525000, 526000</td>
			<td></td>
		</tr>
		<tr>
			<td>26 Gauge Solid Copper Wire</td>
			<td>Staples</td>
			<td>1016882 &#160;</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
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			<td>Surgery</td>
			<td></td>
			<td></td>
			<td></td>
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			<td>Bone Screws</td>
			<td>Stoelting+CB33:C61</td>
			<td>51457</td>
			<td></td>
		</tr>
		<tr>
			<td>Scalpel Blades, 10 or 15</td>
			<td>Stoelting</td>
			<td>52173-10</td>
			<td></td>
		</tr>
		<tr>
			<td>1 ml syringes</td>
			<td>Fisher</td>
			<td>14-826-261</td>
			<td></td>
		</tr>
		<tr>
			<td>22 G Needles</td>
			<td>Fisher&#160;</td>
			<td>14-815-525</td>
			<td></td>
		</tr>
		<tr>
			<td>27 G Needles</td>
			<td>Fisher</td>
			<td>14-826-48</td>
			<td></td>
		</tr>
		<tr>
			<td>2&#34; x 2&#34; Gauze</td>
			<td>Fisher</td>
			<td>22-362-178</td>
			<td></td>
		</tr>
		<tr>
			<td>Swabs</td>
			<td>Fisher</td>
			<td>19-120-472</td>
			<td></td>
		</tr>
		<tr>
			<td>Puppy Pads</td>
			<td>PetCo</td>
			<td>1310747</td>
			<td></td>
		</tr>
		<tr>
			<td>Kim Wipes</td>
			<td>Fisher</td>
			<td>06-666-A</td>
			<td></td>
		</tr>
		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
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		<tr>
			<td></td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td colspan="2">Chamber and Behavioral Setting&#160;</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Husky Metal Front Base Cabinet (30WX19DX34H)</td>
			<td>Home Depot</td>
			<td>100607961</td>
			<td></td>
		</tr>
		<tr>
			<td>Quiet Barrier&#173; HD Soundproofing Material (Sheet) (PSA)</td>
			<td>soundproofcow.com</td>
			<td>10203041</td>
			<td></td>
		</tr>
		<tr>
			<td>Convoluted Acoustic Foam Panel</td>
			<td>soundproofcow.com</td>
			<td>10432400</td>
			<td></td>
		</tr>
		<tr>
			<td>Isolated Pulse Stimulator Model 2100</td>
			<td>A-M Systems</td>
			<td>720000</td>
			<td></td>
		</tr>
		<tr>
			<td>Digital Camera - Logitech Webcam C210</td>
			<td>Logitech</td>
			<td>B003LVZO88</td>
			<td></td>
		</tr>
		<tr>
			<td>MatLab</td>
			<td>Mathworks.com</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Sinometer 10MHz Single Channel Oscilloscope</td>
			<td>Sinometer</td>
			<td>CQ5010C</td>
			<td></td>
		</tr>
		<tr>
			<td>OxyLED T-01 DIY Stick-on Anywhere 4-LED Touch Tap Light</td>
			<td>OXYLED</td>
			<td>B00GD8OKY0</td>
			<td></td>
		</tr>
		<tr>
			<td>5k ohm potentiomter</td>
			<td>Alpha Electronics</td>
			<td>B00CTWDHIO</td>
			<td></td>
		</tr>
		<tr>
			<td>Extech 407730 40-to-130-Decibel Digital Sound Level Meter</td>
			<td>Extech Instruments</td>
			<td>B000EWY67W</td>
			<td></td>
		</tr>
		<tr>
			<td>DSCK-C Dual Output, scrambled shocker</td>
			<td>Kinder Scientific Co</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

vagus nerve stimulation as a tool to induce plasticity in pathways relevant for extinction learning

Extinction describes the process of attenuating behavioral responses to neutral stimuli when they no longer provide the reinforcement that has been maintaining the behavior. There is close correspondence between fear and human anxiety, and therefore studies of extinction learning might provide insight into the biological nature of anxiety-related disorders such as post-traumatic stress disorder, and they might help to develop strategies to treat them. Preclinical research aims to aid extinction learning and to induce targeted plasticity in extinction circuits to consolidate the newly formed memory. Vagus nerve stimulation (VNS) is a powerful approach that provides tight temporal and circuit-specific release of neurotransmitters, resulting in modulation of neuronal networks engaged in an ongoing task. VNS enhances memory consolidation in both rats and humans, and pairing VNS with exposure to conditioned cues enhances the consolidation of extinction learning in rats. Here, we provide a detailed protocol for the preparation of custom-made parts and the surgical procedures required for VNS in rats. Using this protocol we show how VNS can facilitate the extinction of conditioned fear responses in an auditory fear conditioning task. In addition, we provide evidence that VNS modulates synaptic plasticity in the pathway between the infralimbic (IL) medial prefrontal cortex and the basolateral complex of the amygdala (BLA), which is involved in the expression and modulation of extinction memory.

This section illustrates examples of results that can be obtained by using VNS in combination with extinction learning to reduce the expression of the conditioned fear response in rats. For Days 1 and 2 (Auditory Fear Conditioning), rats were trained on an auditory fear conditioning task in which footshocks were paired with a tone. On Day 3 (Pre Treatment Test), tones were presented in the absence of footshocks to measure freezing levels and infer conditioned fear response acquisition. On Day 4 (Treatment) rats received ...

Watch this Scientific Journal Video about Vagus Nerve Stimulation as a Tool to Induce Plasticity in Pathways Relevant for Extinction Learning at JoVE.com

Vagus Nerve Stimulation as a Tool to Induce Plasticity in Pathways Relevant for Extinction Learning

Vagus nerve stimulation (VNS) has emerged as a tool to induce targeted synaptic plasticity in the forebrain to modify a range of behaviors. This protocol describes how to implement VNS to facilitate the consolidation of fear extinction memory.

Extinction describes the process of attenuating behavioral responses to neutral stimuli when they no longer provide the reinforcement that has been ...

vagus-nerve-stimulation-as-tool-to-induce-plasticity-pathways

Research

JoVE Journal

Behavior

22.9K Görüntüleme.  The University of Texas at Dallas.  Vagus sinir stimülasyonu (VNS) davranışları bir dizi değiştirmek için ön beyinde hedeflenen sinaptik plastisite ikna etmek için bir araç olarak ortaya çıkmıştır. Bu protokol korku söndürme bellek konsolidasyonu kolaylaştırmak için VNS uygulamak açıklamaktadır. 