Novelty and fear conditioning induced gene expression in high and low states of anxiety.

Emotional states influence how stimuli are interpreted. High anxiety states in humans lead to more negative, threatening interpretations of novel information, typically accompanied by activation of the amygdala. We developed a handling protocol that induces long-lasting high and low anxiety-like states in rats to explore the role of state anxiety on brain activation during exposure to a novel environment and fear conditioning. In situ hybridization of the inducible transcription factor Egr-1 found increased gene expression in the lateral nucleus of the amygdala (LA) following exposure to a novel environment and contextual fear conditioning in high anxiety-like rats. In contrast, low state anxiety-like rats did not generate Egr-1 increases in LA when placed in a novel chamber. Egr-1 expression was also examined in the dorsal hippocampus and prefrontal cortex. In CA1 of the hippocampus and medial prefrontal cortex (mPFC), Egr-1 expression increased in response to novel context exposure and fear conditioning, independent of state anxiety level. Furthermore, in mPFC, Egr-1 in low anxiety-like rats was increased more with fear conditioning than novel exposure. The current series of experiments show that brain areas involved in fear and anxiety-like states do not respond uniformly to novelty during high and low states of anxiety.

Chronic corticosterone administration does not potentiate unconditioned freezing to the predator odor, trimethylthiazoline.

Chronic high levels of corticosterone (CORT) are known to facilitate learning and memory of aversive events. Whether this effect of chronic CORT also generalizes to unconditioned or unlearned fear behavior is not known. The present study investigated whether high levels of chronic CORT enhance unconditioned fear to a predator odor, trimethylthiazoline (TMT), an innate fear stimulus to rodents. TMT induces a dose-related freezing response, a prototypical behavior to fearful stimuli, in rats. The first experiment demonstrated that dose-related freezing to repeated exposures of TMT does not habituate, sensitize or produce contextually conditioned fear, and therefore can be used to measure the effects of chronic CORT on unconditioned fear to repeated exposures of TMT. In Experiment 2, 21-day release corticosterone pellets (200mg) were implanted subcutaneously in male, Sprague-Dawley rats. Control rats received sham implantation. On days when TMT was not present, chronic CORT rats froze significantly more than sham rats. However, while TMT-induced freezing in both chronic CORT and sham rats, freezing during exposure to TMT was not further enhanced in chronic CORT rats. Thus, chronic CORT appears to increase fear as measured by freezing, possibly by enhancing vigilance, but does not facilitate fear behavior induced by the innate fear stimulus, TMT.

Not all rat strains are equal: differential unconditioned fear responses to the synthetic fox odor 2,4,5-trimethylthiazoline in three outbred rat strains.

Predator odors induce unconditioned fear in rats; however, the synthetic predator odor 2,4,5-trimethylthiazoline (TMT) either elicits robust fear behavior (e.g., freezing) or no fear responses at all. The authors investigated whether this is due to the use of different outbred rat strains. TMT induced robust freezing in Sprague-Dawley and Long-Evans rats but not in Wistar rats. All 3 strains avoided TMT, but Wistar rats were less sensitive to TMT. Wistar rats are capable of freezing; all 3 strains displayed the same amount of odor-cue conditioned freezing. Thus, TMT is a robust unconditioned fear stimulus in rats, and prior negative results from other laboratories were due to the choice of a rat strain (Wistar) that is less responsive to TMT.

Animal studies of amygdala function in fear and uncertainty: relevance to human research.

This article reviews research in both animals and humans on the considerable progress made in elucidating a brain circuitry of fear, particularly the importance of the amygdala in fear conditioning. While there is considerable agreement about the participation of the amygdala in fear in both animals and humans, there are several issues about the function of the amygdala raised by the human research that have not been addressed by or may be answered by animal research. Three of these are addressed in this article: (1) is the amygdala involved in or necessary for both fear learning and unconditioned fear? (2) Does the amygdala code for intensity of fear? (3) Is the amygdala preferentially involved in fear, or is it also activated when there are no overt fear or aversive stimuli, but where the situation can be described as uncertain? We present evidence indicating that the rodent amygdala is involved in some types of fear (conditioned fear), but not all types (unconditioned fear), and may therefore have significance for a differential neurobiology of certain anxiety disorders in humans. Further, similar to the human amygdala, the rodent amygdala responds to varying intensities of aversive stimulation. Finally, it is suggested that, similar to humans, the rodent amygdala is involved in the evaluation of uncertainty. We conclude that progress on elucidating the role of the amygdala in fear is facilitated by corroboration of findings from both animal and human research.

Glucocorticoid receptor antagonism in the basolateral amygdala and ventral hippocampus interferes with long-term memory of contextual fear.

The binding of glucocorticoids to the type II or glucocorticoid receptor (GR) is known to play a role in memory consolidation and long-term memory. The present series of studies investigated the locus for GR effects on learning and memory of contextual fear conditioning. The GR antagonist RU 38486 was administered peripherally (10, 20, or 30 mg/kg/ml), as well as centrally into the lateral ventricle (75 or 150 ng/2 microL), basolateral amygdala (BLA; 0.3, 3, or 30 ng/0.2 microL), dorsal hippocampus (DH; 30 ng/microL) or ventral hippocampus (VH; 30 ng/microL) prior to contextual fear conditioning. Peripheral administration of RU 38486 did not affect fear-related levels of freezing immediately following a footshock or in a long-term memory test 24 h later. However, administration into the lateral ventricle, BLA, or VH decreased freezing in a 24 h retention test, while leaving post-shock levels of freezing intact. Both post-shock and retention levels of freezing were unaffected in rats that received RU 38486 in the DH compared to vehicle controls; however, vehicle rats also displayed low levels of freezing during retention. The data indicate that GR activation within the BLA and VH is important for the establishment of long-term memory for contextual fear conditioning.

An egr-1 (zif268) antisense oligodeoxynucleotide infused into the amygdala disrupts fear conditioning.

Studies of gene expression following fear conditioning have demonstrated that the inducible transcription factor, egr-1, is increased in the lateral nucleus of the amygdala shortly following fear conditioning. These studies suggest that egr-1 and its protein product Egr-1 in the amygdala are important for learning and memory of fear. To directly test this hypothesis, an egr-1 antisense oligodeoxynucleotide (antisense-ODN) was injected bilaterally into the amygdala prior to contextual fear conditioning. The antisense-ODN reduced Egr-1 protein in the amygdala and interfered with fear conditioning. A 250-pmole dose produced an 11% decrease in Egr-1 protein and reduced long-term memory of fear as measured by freezing in a retention test 24 h after conditioning, but left shock-induced freezing intact. A larger 500-pmole dose produced a 25% reduction in Egr-1 protein and significantly decreased both freezing immediately following conditioning and freezing in the retention test. A nonsense-ODN had no effect on postshock or retention test freezing. In addition, 500 pmole of antisense-ODN infused prior to the retention test in previously trained rats did not reduce freezing, indicating that antisense-ODN did not suppress conditioned fear behavior. Finally, rats infused with 500 pmole of antisense-ODN displayed unconditioned fear to a predator odor, demonstrating that unconditioned freezing was unaffected by the antisense-ODN. The data indicate that the egr-1 antisense-ODN interferes with learning and memory processes of fear without affecting freezing behavior and suggests that the inducible transcription factor Egr-1 within the amygdala plays important functions in long-term learning and memory of fear.