GABAergic antagonism of the central nucleus of the amygdala attenuates reductions in rapid eye movement sleep after inescapable footshock stress.

RAPID EYE MOVEMENT SLEEP (REM) APPEARS TO BE ESPECIALLY SUSCEPTIBLE TO THE EFFECTS OF STRESS.1,2 FOOTSHOCK IS A WIDELY USED STRESSOR in studies involving rodents, and presentation of inescapable footshock stress (IS) to rats and mice can produce subsequent significant decreases in REM after the stressor is removed.2–4 The decreases in REM may be relatively long lasting and can vary with the reactivity and emotionality of rat4 and mouse2,3 strains. By comparison, the effects on NREM have greater variability with decreases, no significant alterations, or increases in NREM amounts, depending on strain.2–4 The pons contains regions critical for the generation and regulation of REM.5 The locus coeruleus (LC) is activated by footshock,6,7 and it is thought to be a major REM regulating region.5 The firing of LC noradrenergic neurons has been linked to the suppression of REM, whereas their silence (along with the silence of serotonergic neurons in the dorsal raphe nucleus) has been hypothesized to permit REM.5,8 Activation of LC neurons also occurs in response to stressful stimuli.9,10 Forebrain regions can also have significant influences on the generation of REM. The central nucleus of the amygdala (CNA), in particular, has been linked to the regulation of REM.11–15 The amygdala has also been prominently linked to fear behavior that occurs in response to IS16,17 and to the regulation of arousal.13,15,18 CNA output is tightly regulated by GABA.19 Microinjection into CNA of the GABAA agonist, muscimol (MUS), selectively decreases REM; whereas microinjection of the GABAA antagonist, bicuculline (BIC), selectively increases REM.12 These findings suggest that GABAergic inhibition of CNA suppresses REM, whereas blocking GABAergic inhibition of CNA promotes REM. CNA has direct projections to the LC20,21 that may be important for certain stress responses.22 In a previous study in mice, we found that training with IS enhanced Fos expression in the LC and in some regions of the amygdala.23 In the amygdala, the CNA failed to show enhanced Fos expression after IS,23 a finding that complements other studies reporting that CNA does not show enhanced Fos expression after certain types of stress.24,25 Intense Fos activation in LC and lack of significant Fos expression in CNA after IS are compatible with the hypothesis that activation of the noradrenergic LC and inhibition of CNA may be factors in the IS-induced suppression of REM. Thus, the purpose of this study was to determine whether CNA and LC participate in reducing REM after IS. To determine whether reduced REM following IS could involve regulation by CNA, we microinjected MUS and BIC into CNA prior to IS and compared post-shock sleep to sleep exhibited by non-shocked rats receiving control microinjections alone and to shocked rats receiving control microinjections. Similar IS-induced alterations in REM after microinjections of MUS and vehicle and attenuation of the reduction after microinjections of BIC would suggest that inhibition of CNA is a significant factor in the effects of IS on REM. In a second experiment, we conducted the same manipulations in different groups of rats but killed the animals after 2 h of sleep recording to examine Fos protein expression as a marker of neuronal activity in the LC. Enhanced Fos expression in the LC associated with reduced REM would suggest a suppressing role for noradrenergic mechanisms in LC. Attenuation of the REM reduction and Fos expression in the LC after BIC microinjected into CNA would suggest that the CNA-LC pathway is important in the regulation of REM in the aftermath of IS.