Stress-Related Neuronal Clusters in Sublenticular Extended Amygdala of Basal Forebrain Show Individual Differences of Positions

To understand functional neuronal circuits for emotion in the basal forebrain, patterns of neuronal activation were examined in mice by immunohistochemistry of immediate-early gene products (Zif268/Egr1 and c-Fos). In all mice examined, clusters of 30–50 neurons expressing Zif268 were found on both sides in the area between the extended amygdala (EA) and globus pallidus (GP), generally designated as sublenticular extended amygdala (SLEA). The clusters consisted of 79.9 ± 3.0% of GABAergic neurons in GAD65-mCherry mice. The expression of the cholinergic marker choline acetyltransferase and the GP markers parvalbumin, proenkephalin, and FoxP2 indicated that these neurons were different from known types of neurons in the EA and GP; therefore, we named them the sublenticular extended amygdalar Zif268/Egr1-expressing neuronal cluster (SLEA-zNC). Sublenticular extended amygdalar Zif268/Egr1-expressing neuronal clusters participated in stress processing because increasing numbers of cells were observed in SLEA-zNCs after exposure to restraint stress (RS), the induction of which was suppressed by diazepam treatment. Mapping SLEA-zNCs showed that their positions and arrangement varied individually; SLEA-zNCs were distributed asymmetrically and tended to be situated mainly in the middle region between the anterior commissure (AC) and posterior end of the GP. However, the total cell number in SLEA-zNCs was compatible between the right and left hemispheres after activation by RS. Therefore, SLEA-zNCs were distributed asymmetrically but were not lateralized. Because time courses of activation differed between the Zif268 and c-Fos, the sequential dual treatment of RSs enabled us to differentiate SLEA-zNCs activated by the first and second RS. The results supported that the same SLEA-zNCs responded to both the first and second RS, and this also applied for all SLEA-zNCs. Thus, we concluded that the cluster positions were invariable under RS in each mouse but were distributed differently between individual mice. We name these newly identified neuronal clusters as stress-related neuronal clusters, SLEA-zNCs, which are considered to be novel functional units of “islands of activation.” Moreover, SLEA-zNCs were situated at different positions in all mice examined, showing individual differences in their positions.