Prefrontal/amygdalar system determines stress coping behavior through 5-HT/GABA connection

Abstract

Coping is defined as the behavioral and physiological effort made to master stressful situations. The ability to cope with stress leads either to healthy or to pathogenic outcomes. The medial prefrontal cortex (mpFC) and amygdala are acknowledged as having a major role in stress-related behaviors, and mpFC has a critical role in the regulation of amygdala-mediated arousal in response to emotionally salient stimuli. Prefrontal cortical serotonin (5-hydroxytryptamine (5-HT)) is involved in corticolimbic circuitry, and GABA has a major role in amygdala functioning. Here, using mice, it was assessed whether amygdalar GABA regulation by prefrontal 5-HT is involved in processing stressful experiences and in determining coping outcomes. First (experiment 1), bilateral selective 5-HT depletion in mpFC of mice reduced GABA release induced by stress in basolateral amygdala (BLA) and passive coping in the Forced Swimming Test (FST) (experiment 2). Moreover, prefrontal-amygdala disconnection procedure that combined a selective unilateral 5-HT depletion of mpFC and infusion of an inhibitor of GABA synthesis into the contralateral BLA, thereby to disrupt prefrontal-amygdalar serial connectivity bilaterally, showed that disconnection selectively decreases immobility in the FST. These results point to prefrontal/amygdala connectivity mediated by 5-HT and GABA transmission as a critical neural mechanism in stress-induced behavior.

Figure 1

Effects of bilateral medial prefrontal cortex (mpFC) 5-hydroxytryptamine (5-HT) depletion and L-allyglycine infusion in basolateral amygdala (BLA) on GABA output in BLA exposed to 10 min of Forced Swimming Test (FST). Results are expressed as percent changes (means±SE) from basal values preceding L-allyglycine infusion in BLA and FST. Statistical analyses were performed on raw data. *P<0.05 from basal values. §P<0.05 in comparison with the corresponding time point of mpFC Sham groups.

Figure 2

Schematic representation of disconnection of medial prefrontal cortex (mpFC) and basolateral amygdala (BLA) (a–d). ‘X' represents the location of the unilateral inactivation to the mpFC or BLA. Green arrows represent intact pathways. Red lines and X-symbols represent blocked, nonfunctional pathways. In unilateral mpFC-5-hydroxytryptamine (5-HT)-depleted mice (red-shaded area), the mpFC relay of the pathway is lost on one side of the brain. However, on the non-depleted side (green area and arrows), the circuitry is intact and functional. When L-allyglycine (red-shaded area) is infused in the BLA contralateral to depletion, it blocks GABA output, hence impairing BLA GABA functioning in the non-5-HT-depleted side of the brain. Therefore, after unilateral L-allyglycine infusions in the contralateral BLA of mpFC-depleted animals, functional activity within the mpFC/BLA system is disrupted bilaterally. (a) This group received unilateral vehicle in mpFC (Sham) and cerebrospinal fluid (CSF) in the controlateral BLA during Forced Swimming Test (FST). (b) This group received unilateral toxin 5,7-dihydroxytryptamine (5,7-DHT) in mpFC to deplete 5-HT and CSF in the controlateral BLA during FST. (c) This group received unilateral vehicle in mpFC (Sham) and L-allyglycine (to block GABA synthesis) in the controlateral BLA during FST. (d) This group received unilateral toxin 5,7-DHT in mpFC to deplete 5-HT and L-allyglycine (to block GABA synthesis) in the controlateral BLA during FST (disconnection).

Figure 3

Effects of selective prefrontal cortical 5-hydroxytryptamine (5-HT) depletion on 5-HT outflow in the medial prefrontal cortex (mpFC) (a) and GABA outflow in the basolateral amygdala (BLA) (b) of mice exposed to 120 min of restraint. Results are expressed as percent changes (means±SE) from basal values. Statistical analyses were performed on raw data. §P<0.05 from the basal values. *P<0.05 in comparison with the corresponding time point of sham group. Samples of chromatograms of basal and stress 5-HT outflow in the mpFC (c) and GABA outflow in the BLA (d). Representative positions of the probe in the mpFC (e) and BLA (f). The segment of the membrane probe is also shown.

Figure 4

(a) Effects of selective prefrontal cortical 5-hydroxytryptamine (5-HT) depletion on Forced Swimming Test (FST)-induced c-fos immunoreactive cell nuclei (mean number of nuclei/0.1 mm²±SEM) in central amygdala (CeA) and basolateral amygdala (BLA). *P<0.05. (b) Representative Nissl-stained section illustrating CeA and BLA boundaries as revealed by cytoarchitecture. Scale bar=1 mm. Representative photomicrographs showing FST-induced c-fos immunoreactive cell nuclei in CeA and BLA of 5-HT-depleted group (c) and mpFC 5-HT sham group (d) (according to the atlas of Franklin and Paxinos, 1997). Scale bar=100 μm.

Figure 5

Effects of 5-hydroxytryptamine (5-HT) in medial prefrontal cortex (mpFC) and GABA in basolateral amygdala (BLA) disconnection on immobility in the Forced Swimming Test (FST). Results are expressed as mean±SE duration (second) of immobility. *P<0.05 in comparision with 5-HT sham+ally, 5-HT depl+cerebrospinal fluid (CSF), and 5-HT sham+CSF groups.