Effects of vagus nerve stimulation on extinction of conditioned fear and post-traumatic stress disorder symptoms in rats

Abstract

Exposure-based therapies help patients with post-traumatic stress disorder (PTSD) to extinguish conditioned fear of trauma reminders. However, controlled laboratory studies indicate that PTSD patients do not extinguish conditioned fear as well as healthy controls, and exposure therapy has high failure and dropout rates. The present study examined whether vagus nerve stimulation (VNS) augments extinction of conditioned fear and attenuates PTSD-like symptoms in an animal model of PTSD. To model PTSD, rats were subjected to a single prolonged stress (SPS) protocol, which consisted of restraint, forced swim, loss of consciousness, and 1 week of social isolation. Like PTSD patients, rats subjected to SPS show impaired extinction of conditioned fear. The SPS procedure was followed, 1 week later, by auditory fear conditioning (AFC) and extinction. VNS or sham stimulation was administered during half of the extinction days, and was paired with presentations of the conditioned stimulus. One week after completion of extinction training, rats were given a battery of behavioral tests to assess anxiety, arousal and avoidance. Results indicated that rats given SPS 1 week prior to AFC (PTSD model) failed to extinguish the freezing response after eleven consecutive days of extinction. Administration of VNS reversed the extinction impairment and attenuated reinstatement of the conditioned fear response. Delivery of VNS during extinction also eliminated the PTSD-like symptoms, such as anxiety, hyperarousal and social avoidance for more than 1 week after VNS treatment. These results provide evidence that extinction paired with VNS treatment can lead to remission of fear and improvements in PTSD-like symptoms. Taken together, these findings suggest that VNS may be an effective adjunct to exposure therapy for the treatment of PTSD.

Figure 1

Timeline for extinction, reinstatement and behavioral assays. (a) Protocol for PTSD model rats. Rats underwent the SPS procedure followed by 2 days of AFC and 11 days of extinction, five were paired with VNS (red) or sham stimulation (blue) on alternate (even) days. Freezing in the presence of the CS was used as a measure of conditioned fear. Conditioned fear was measured on the odd days that fell between VNS or sham stimulation days. Following extinction, some rats underwent a reinstatement trial where they received one unsignaled footshock. The day after the footshock, these rats were given another day of extinction to measure conditioned fear. The remaining rats were tested on a battery of behaviors to measure PTSD symptoms 1 week after the end of extinction. The order of the tests was counterbalanced. (b) Protocol for AFC alone rats. Rats in the AFC alone group were treated exactly like PTSD model rats; however, the AFC alone group did not undergo the SPS procedure. AFC, auditory fear conditioning; PTSD, post-traumatic stress disorder; SPS, single prolonged stress; VNS, vagus nerve stimulation.

Figure 2

Conditioned fear responding across extinction days. (a) VNS treatment reverses maladaptive fear seen in PTSD model rats. Following 11 consecutive days of extinction, rats exposed to the SPS procedure 1 week before AFC (PTSD model rats) did not reach remission of fear. A single, unpaired reminder of the unconditioned stimulus was sufficient to reinstate conditioned fear to the level of freezing measured on the first day of extinction in the PTSD model. PTSD model rats given VNS reached remission of fear by the end of treatment. VNS also attenuated the reinstatement of conditioned fear. Freezing in rats given VNS was significantly reduced compared with freezing on the first day of extinction. PTSD model rats given VNS showed significantly less fear than sham-treated rats on extinction day 5 (**P<0.01) and extinction days 7, 9 and 11 (****P<0.0001). PTSD model rats given VNS showed reduced reinstatement of conditioned fear versus sham-treated rats (****P<0.0001). (b) VNS treatment accelerates extinction in control rats subjected to AFC alone. All rats that underwent AFC alone reached remission of fear by the end of extinction. VNS led to more rapid remission of fear than sham stimulation. VNS-treated rats showed reduced freezing versus sham-treated rats on extinction day 5 (***P<0.001) and extinction day 7 (*P<0.05). AFC alone rats alone did not show complete reinstatement of conditioned fear following a reminder of the unconditioned stimulus. AFC, auditory fear conditioning; PTSD, post-traumatic stress disorder; SPS, single prolonged stress; VNS, vagus nerve stimulation.

Figure 3

AFC alone rats and PTSD model rats respond similarly to footshock. (a) Acquisition of conditioned fear is similar between groups. Prior to VNS- or sham-paired extinction, rats underwent 2 days of AFC. On the first day of AFC, rats were exposed to four pretones to assess baseline freezing to the tone, pretone freezing was similar between groups. Acquisition of conditioned fear was similar between all rats on these days; however, PTSD model rats show a reduction in freezing at tones 9 and 10 versus AFC alone rats (^##P<0.01). This effect is no longer present on subsequent tones. (b) Following reinstatement shocks, rats show similar levels of freezing. Following a reinstatement shock in context A, rats from all groups show comparable levels of freezing (P>0.05). AFC, auditory fear conditioning; PTSD, post-traumatic stress disorder; VNS, vagus nerve stimulation.

Figure 4

VNS treatment reversed PTSD-like symptoms of anxiety and exaggerated startle. (a) VNS administration during extinction reduced anxiety. One week after the completion of extinction, PTSD model rats spent less time in the open arms of the maze than AFC alone rats (^##P<0.01), indicating heightened anxiety. VNS during extinction reversed this effect in PTSD model rats and also decreased anxiety in AFC alone rats. VNS-treated rats spent more time in the open arms versus sham-treated rats in the PTSD model (**P<0.01) and in the control group that underwent AFC alone (*P<0.05). (b) VNS treatment increased entries into the open arms. Similar to time spent in the open arms, PTSD model rats showed a reduced number of entries into the open arms versus AFC alone rats (^##P<0.01). Administration of VNS during extinction reversed this effect and increased entries into the open arms in PTSD model rats (***P<0.001) and in AFC alone rats (*P<0.05). (c) Total locomotion was not affected. Total time spent moving in the EPM was similar between groups (P>0.05). (d) VNS treatment reduced startle responses. Following extinction, there was no difference in PTSD model rats versus AFC alone rats. Administration of VNS during extinction reduced startle amplitudes in PTSD model rats versus sham (*P<0.05) and in rats that underwent AFC alone versus sham (*P<0.05). AFC, auditory fear conditioning; EPM, elevated plus maze; PTSD, post-traumatic stress disorder; VNS, vagus nerve stimulation.

Figure 5

VNS during extinction decreases novel avoidance and social withdrawal behavior. (a) VNS reduced novel object avoidance in PTSD model rats. One week following extinction, PTSD model rats showed increased marble burying versus AFC alone rats (^##P<0.01). VNS during extinction reversed this effect PTSD model rats (*P<0.05 versus sham). There was no difference in AFC alone rats. (b) VNS increased social interaction. PTSD model rats given sham treatment showed diminished social interaction versus AFC alone rats (^####P<0.0001). Administration of VNS during extinction reversed this effect and increased social interaction (****P<0.0001 versus sham). There was no difference between VNS and sham stimulation in AFC alone rats. AFC, auditory fear conditioning; PTSD, post-traumatic stress disorder; VNS, vagus nerve stimulation.