Noradrenaline transmission within the ventral bed nucleus of the stria terminalis is critical for fear behavior induced by trimethylthiazoline, a component of fox odor

Abstract

The bed nucleus of the stria terminalis (BNST) is involved in the mediation of fear behavior in rats. A previous study of our laboratory demonstrated that temporary inactivation of the BNST blocks fear behavior induced by exposure to trimethylthiazoline (TMT), a component of fox odor. The present study investigates whether noradrenaline release within the BNST is critical for TMT-induced fear behavior. First, we confirmed previous studies showing that the ventral BNST is the part of the BNST that receives the densest noradrenaline innervation. Second, using in vivo microdialysis, we showed that noradrenaline release within the BNST is strongly increased during TMT exposure, and that this increase can be blocked by local infusions of the alpha2-receptor blocker clonidine. Third, using intracerebral injections, we showed that clonidine injections into the ventral BNST, but not into neighboring brain sites, completely blocked TMT-induced potentiation of freezing behavior. The present data clearly show that the noradrenergic innervation of the ventral BNST is important for the full expression of behavioral signs of fear to the predator odor TMT.

Figure 1.

Photomicrograph demonstrating the localization of the noradrenergic terminals within the BNST (immunohistochemical staining of DBH). ac, Anterior commissure; aca, anterior part of the anterior commissure; lBNST, lateral part of the BNST; LSI, intermediate part of the lateral septum; LV, lateral ventricle; mBNST, medial part of the BNST.Scale bar, 1 mm.

Figure 2.

Reconstructions of the different injection sites of saline and clonidine into the BNST (A, filled circles, test on TMT-induced freezing and on motor activity; diamonds, misplaced injections) and the microdialysis sites (B, stars). The coronal sections were taken from the atlas of Paxinos and Watson (1997); numbers to the right indicate distance (in millimeters) from bregma. ac, Anterior commissure; aca, anterior part of the anterior commissure; CPu, caudate-putamen; LSI, intermediate part of the lateral septum; VP, ventral pallidum.

Figure 3.

NA levels in the dialysates of the BNST. A, Effect of TMT presentation on NA levels (n = 6). B, Effects of local clonidine (100 μm) perfusion and simultaneously presented TMT on NA levels in the BNST (n = 6). The dashed line indicates 100% (i.e., the baseline mean level). The arrow indicates the time point of the TMT presentation. Clonidine (100 μm) was infused via reversed microdialysis as indicated by the black bar. Data are presented as a percentage of the basal mean level ± SEM. Data were analyzed by repeated-measures one-way ANOVA, followed by Fisher's LSD (protected t) test, if appropriate. ^*p < 0.05, ^**p < 0.01; significant differences from basal mean levels.

Figure 4.

Mean percentage (±SEM) of time spent freezing to air and TMT in the pre-odor (min 1-4) and odor (min 5-15) condition. A, Effects of clonidine injections into the ventral part of the BNST. B, Effects of misplaced clonidine injections. ^*p < 0.05 and ^**p < 0.01 compared with the pre-odor condition (dependent t tests after ANOVA).

Figure 5.

Time course of (A) or mean (B) spontaneous motor activity after clonidine injections into the ventral BNST measured by activity counts (±SEM) during 15 min in a cage.