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Vagus Nerve Stimulation Enhances Extinction of Conditioned Fear in Rats and Modulates Arc Protein, CaMKII, and GluN2B-Containing NMDA Receptors in the Basolateral Amygdala

View Article: PubMed Central - PubMed

ABSTRACT

Vagus nerve stimulation (VNS) enhances the consolidation of extinction of conditioned fear. High frequency stimulation of the infralimbic cortex (IL) produces long-term potentiation in the basolateral amygdala (BLA) in rats given VNS-paired extinction training, whereas the same stimulation produces long-term depression in sham-treated rats. The present study investigated the state of synaptic plasticity-associated proteins in the BLA that could be responsible for this shift. Male Sprague-Dawley rats were separated into 4 groups: auditory fear conditioning only (fear-conditioned); fear conditioning + 20 extinction trials (extended-extinction); fear conditioning + 4 extinction trials paired with sham stimulation (sham-extinction); fear conditioning + 4 extinction trials paired with VNS (VNS-extinction). Freezing was significantly reduced in extended-extinction and VNS-extinction rats. Western blots were used to quantify expression and phosphorylation state of synaptic plasticity-associated proteins such as Arc, CaMKII, ERK, PKA, and AMPA and NMDA receptors. Results show significant increases in GluN2B expression and phosphorylated CaMKII in BLA samples from VNS- and extended-extinction rats. Arc expression was significantly reduced in VNS-extinction rats compared to all groups. Administration of the GluN2B antagonist ifenprodil immediately after fear extinction training blocked consolidation of extinction learning. Results indicate a role for BLA CaMKII-induced GluN2B expression and reduced Arc protein in VNS-enhanced extinction.

No MeSH data available.


Ifenprodil blocks extinction learning in VNS-extinction and EE rats. (a) Timeline of behavioral protocol. On days 1 and 2 rats were subjected to auditory fear conditioning (AFC) followed by a conditioned fear response test (CFRT) on day 3. On day 4 rats underwent extinction training (EXT) or extended-extinction (EE) training that was immediately followed by an i.p. injection of ifenprodil or vehicle. On day 5 rats underwent a second CFRT (CFRT 2). (b) VNS-extinction and EE rats given a vehicle injection immediately after extinction training show a significant decrease in freezing levels during the second conditioned fear response test (CFRT 2) compared to sham-vehicle rats (∗). VNS-vehicle rats show significantly lower freezing levels during CFRT 2 compared to VNS-ifenprodil rats (#).
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fig4: Ifenprodil blocks extinction learning in VNS-extinction and EE rats. (a) Timeline of behavioral protocol. On days 1 and 2 rats were subjected to auditory fear conditioning (AFC) followed by a conditioned fear response test (CFRT) on day 3. On day 4 rats underwent extinction training (EXT) or extended-extinction (EE) training that was immediately followed by an i.p. injection of ifenprodil or vehicle. On day 5 rats underwent a second CFRT (CFRT 2). (b) VNS-extinction and EE rats given a vehicle injection immediately after extinction training show a significant decrease in freezing levels during the second conditioned fear response test (CFRT 2) compared to sham-vehicle rats (∗). VNS-vehicle rats show significantly lower freezing levels during CFRT 2 compared to VNS-ifenprodil rats (#).

Mentions: An increase in GluN2B expression was measured in rats that showed significant extinction of conditioned fear (VNS-extinction and extended-extinction rats; Figure 3(c)). This is consistent with previous findings indicating a critical role of GluN2B in the BLA in successful extinction of conditioned fear [21]. To determine whether GluN2B is a mediator of VNS enhancement of extinction learning, we administered the GluN2B specific antagonist ifenprodil. Rats underwent auditory fear conditioning followed by extinction training paired with VNS or sham stimulation or extended extinction. Ifenprodil or vehicle was injected (i.p.) immediately after extinction training, and conditioned fear was measured by a CFRT 24 hours later (Figure 4(a)). The ANOVA revealed a significant effect for group (Figure 4(b); F(5, 35) = 3.799, p = 0.007), with the VNS-vehicle and EE-vehicle rats showing significantly lower freezing levels during the second CFRT compared to sham-vehicle rats (versus VNS p = 0.006; versus EE p = 0.001), as well as significantly lower freezing levels in VNS-vehicle rats compared to VNS-ifenprodil rats (p = 0.018).


Vagus Nerve Stimulation Enhances Extinction of Conditioned Fear in Rats and Modulates Arc Protein, CaMKII, and GluN2B-Containing NMDA Receptors in the Basolateral Amygdala
Ifenprodil blocks extinction learning in VNS-extinction and EE rats. (a) Timeline of behavioral protocol. On days 1 and 2 rats were subjected to auditory fear conditioning (AFC) followed by a conditioned fear response test (CFRT) on day 3. On day 4 rats underwent extinction training (EXT) or extended-extinction (EE) training that was immediately followed by an i.p. injection of ifenprodil or vehicle. On day 5 rats underwent a second CFRT (CFRT 2). (b) VNS-extinction and EE rats given a vehicle injection immediately after extinction training show a significant decrease in freezing levels during the second conditioned fear response test (CFRT 2) compared to sham-vehicle rats (∗). VNS-vehicle rats show significantly lower freezing levels during CFRT 2 compared to VNS-ifenprodil rats (#).
© Copyright Policy - open-access
Related In: Results  -  Collection

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fig4: Ifenprodil blocks extinction learning in VNS-extinction and EE rats. (a) Timeline of behavioral protocol. On days 1 and 2 rats were subjected to auditory fear conditioning (AFC) followed by a conditioned fear response test (CFRT) on day 3. On day 4 rats underwent extinction training (EXT) or extended-extinction (EE) training that was immediately followed by an i.p. injection of ifenprodil or vehicle. On day 5 rats underwent a second CFRT (CFRT 2). (b) VNS-extinction and EE rats given a vehicle injection immediately after extinction training show a significant decrease in freezing levels during the second conditioned fear response test (CFRT 2) compared to sham-vehicle rats (∗). VNS-vehicle rats show significantly lower freezing levels during CFRT 2 compared to VNS-ifenprodil rats (#).
Mentions: An increase in GluN2B expression was measured in rats that showed significant extinction of conditioned fear (VNS-extinction and extended-extinction rats; Figure 3(c)). This is consistent with previous findings indicating a critical role of GluN2B in the BLA in successful extinction of conditioned fear [21]. To determine whether GluN2B is a mediator of VNS enhancement of extinction learning, we administered the GluN2B specific antagonist ifenprodil. Rats underwent auditory fear conditioning followed by extinction training paired with VNS or sham stimulation or extended extinction. Ifenprodil or vehicle was injected (i.p.) immediately after extinction training, and conditioned fear was measured by a CFRT 24 hours later (Figure 4(a)). The ANOVA revealed a significant effect for group (Figure 4(b); F(5, 35) = 3.799, p = 0.007), with the VNS-vehicle and EE-vehicle rats showing significantly lower freezing levels during the second CFRT compared to sham-vehicle rats (versus VNS p = 0.006; versus EE p = 0.001), as well as significantly lower freezing levels in VNS-vehicle rats compared to VNS-ifenprodil rats (p = 0.018).

View Article: PubMed Central - PubMed

ABSTRACT

Vagus nerve stimulation (VNS) enhances the consolidation of extinction of conditioned fear. High frequency stimulation of the infralimbic cortex (IL) produces long-term potentiation in the basolateral amygdala (BLA) in rats given VNS-paired extinction training, whereas the same stimulation produces long-term depression in sham-treated rats. The present study investigated the state of synaptic plasticity-associated proteins in the BLA that could be responsible for this shift. Male Sprague-Dawley rats were separated into 4 groups: auditory fear conditioning only (fear-conditioned); fear conditioning + 20 extinction trials (extended-extinction); fear conditioning + 4 extinction trials paired with sham stimulation (sham-extinction); fear conditioning + 4 extinction trials paired with VNS (VNS-extinction). Freezing was significantly reduced in extended-extinction and VNS-extinction rats. Western blots were used to quantify expression and phosphorylation state of synaptic plasticity-associated proteins such as Arc, CaMKII, ERK, PKA, and AMPA and NMDA receptors. Results show significant increases in GluN2B expression and phosphorylated CaMKII in BLA samples from VNS- and extended-extinction rats. Arc expression was significantly reduced in VNS-extinction rats compared to all groups. Administration of the GluN2B antagonist ifenprodil immediately after fear extinction training blocked consolidation of extinction learning. Results indicate a role for BLA CaMKII-induced GluN2B expression and reduced Arc protein in VNS-enhanced extinction.

No MeSH data available.