Andrew M. Poulos

Concussive Brain Injury Enhances Fear Learning and Excitatory Processes in the Amygdala

BACKGROUND Mild traumatic brain injury (cerebral concussion) results in cognitive and emotional dysfunction. These injuries are a significant risk factor for the development of anxiety disorders, including posttraumatic stress disorder. However, because physically traumatic events typically occur in a highly emotional context, it is unknown whether traumatic brain injury itself is a cause of augmented fear and anxiety. METHODS Rats were trained with one of five fear-conditioning procedures (n = 105) 2 days after concussive brain trauma. Fear learning was assessed over subsequent days and chronic changes in fear learning and memory circuitry were assessed by measuring N-methyl-D-aspartate receptor subunits and glutamic acid decarboxylase, 67 kDa isoform protein levels in the hippocampus and basolateral amygdala complex (BLA). RESULTS Injured rats exhibited an overall increase in fear conditioning, regardless of whether fear was retrieved via discrete or contextual-spatial stimuli. Moreover, injured rats appeared to overgeneralize learned fear to both conditioned and novel stimuli. Although no gross histopathology was evident, injury resulted in a significant upregulation of excitatory N-methyl-D-aspartate receptors in the BLA. There was a trend toward decreased γ-aminobutyric acid-related inhibition (glutamic acid decarboxylase, 67 kDa isoform) in the BLA and hippocampus. CONCLUSIONS These results suggest that mild traumatic brain injury predisposes the brain toward heightened fear learning during stressful postinjury events and provides a potential molecular mechanism by which this occurs. Furthermore, these data represent a novel rodent model that can help advance the neurobiological and therapeutic understanding of the comorbidity of posttraumatic stress disorder and traumatic brain injury.

Persistence of fear memory across time requires the basolateral amygdala complex

Mammals evolved a potent fear-motivated defensive system capable of single-trial fear learning that shows no forgetting over the lifespan of the animal. The basolateral amygdala complex (BLA) is considered an essential component of this conditional fear learning system. However, recent studies challenge this view and suggest that plasticity within other brain regions (i.e., central nucleus of the amygdala) may be crucial for fear conditioning. In the present study, we examine the mnemonic limits of contextual fear conditioning in the absence of the BLA using overtraining and by measuring remote fear memories. After excitotoxic lesions of the BLA were created, animals underwent overtraining and were tested at recent and remote memory intervals. Here we show that animals with BLA lesions can learn normal levels of fear. However, this fear memory loses its adaptive features: it is acquired slowly and shows substantial forgetting when remote memory is tested. Collectively, these findings suggest that fear-related plasticity acquired by brain regions outside of the BLA, unlike those acquired in the intact animals, do so for a relatively time-limited period.

Compensation in the neural circuitry of fear conditioning awakens learning circuits in the bed nuclei of the stria terminalis

The basolateral amygdala (BLA) is thought to be essential for fear learning. However, extensive training can overcome the loss of conditional fear evident following lesions and inactivation of the BLA. Such results suggest the existence of a primary BLA-dependent and a compensatory BLA-independent neural circuit. We tested the hypothesis that the bed nuclei of the stria terminalis (BST) provides this compensatory plasticity. Using extensive context-fear conditioning, we demonstrate that combined BLA and BST lesions prevented fear acquisition and expression. Additionally, protein synthesis in the BST was critical only for consolidation of BLA-independent but not BLA-dependent fear. Moreover, fear acquired after BLA lesions resulted in greater activation of BST regions that receive hippocampal efferents. These results suggest that the BST is capable of functioning as a compensatory site in the acquisition and consolidation of context-fear memories. Unlocking such neural compensation holds promise for understanding situations when brain damage impairs normal function or failure to regulate compensatory sites leads to anxiety disorders.

Amnesia for Early Life Stress Does Not Preclude the Adult Development of Posttraumatic Stress Disorder Symptoms in Rats

BACKGROUND Traumatic experience can result in life-long changes in the ability to cope with future stressors and emotionally salient events. These experiences, particularly during early development, are a significant risk factor for later life anxiety disorders such as posttraumatic stress disorder (PTSD). However, because traumatic experience typically results in strong episodic memories, it is not known whether such long-term memories are necessary for particular features of PTSD, such as enhanced fear and anxiety. Here, we used a fear conditioning procedure in juvenile rats before maturation of the neural systems supporting declarative memory to assess the necessity of early memory to the later life development of PTSD-related symptoms. METHODS Nineteen-day old rats were exposed to unpredictable and inescapable footshocks, and fear memory for the shock context was assessed during adulthood. Thereafter, adult animals were either exposed to single-trial fear conditioning or elevated plus maze or sacrificed for basal diurnal corticosterone and quantification of neuronal glucocorticoid and neuropeptide Y receptors. RESULTS Early trauma exposed rats displayed stereotypic footshock reactivity, yet by adulthood, hippocampus-dependent contextual fear-related memory was absent. However, adult rats showed sensitized fear learning, aberrant basal circadian fluctuations of corticosterone, increased amygdalar glucocorticoid receptors, decreased time spent in the open arm of an elevated plus maze, and an odor aversion associated with early-life footshocks. CONCLUSIONS These results suggest that traumatic experience during developmental periods of hippocampal immaturity can promote lifelong changes in symptoms and neuropathology associated with human PTSD, even if there is no explicit memory of the early trauma.

Retrieval and Reconsolidation Accounts of Fear Extinction.

Extinction is the primary mode for the treatment of anxiety disorders. However, extinction memories are prone to relapse. For example, fear is likely to return when a prolonged time period intervenes between extinction and a subsequent encounter with the fear-provoking stimulus (spontaneous recovery). Therefore there is considerable interest in the development of procedures that strengthen extinction and to prevent such recovery of fear. We contrasted two procedures in rats that have been reported to cause such deepened extinction. One where extinction begins before the initial consolidation of fear memory begins (immediate extinction) and another where extinction begins after a brief exposure to the consolidated fear stimulus. The latter is thought to open a period of memory vulnerability similar to that which occurs during initial consolidation (reconsolidation update). We also included a standard extinction treatment and a control procedure that reversed the brief exposure and extinction phases. Spontaneous recovery was only found with the standard extinction treatment. In a separate experiment we tested fear shortly after extinction (i.e., within 6 h). All extinction procedures, except reconsolidation update reduced fear at this short-term test. The findings suggest that strengthened extinction can result from alteration in both retrieval and consolidation processes.

Decremental effects of context exposure following delay eyeblink conditioning in rabbits.

The conditioning context arises from the relatively static features of the training environment. In rabbit eyeblink conditioning, procedures that retard acquisition (conditioned stimulus [CS] preexposure, unconditioned stimulus preexposure, blocking manipulations) are attenuated by context changes. In this article the authors investigate the effect of context exposure after initial delay conditioning. After conditioned responses (CRs) were established, one group received 6 sessions of context exposure, whereas control groups either remained in their home cages or received exposure to handling and a novel context. Thereafter, all groups received CS-alone testing. The expression of CRs was substantially reduced following context exposure relative to any retention loss in the home-cage control. Exposure to handling and a novel context facilitated the CRs rather than reducing them.

Disruption of cerebellar cortical inhibition in the absence of learning promotes sensory-evoked eyeblink responses.

Theories of cerebellar learning propose that alterations in synaptic plasticity resulting in decreases in cerebellar cortical inhibition and increases in sensory activation of interpositus nuclei underlie the development of adaptively timed conditioned motor responses. The authors found that with concurrent pharmacological disconnection of the cerebellar cortex and intense sensory stimulation in the untrained rabbit, eyeblink responses were generated. Neither sensory stimulation nor disconnection alone generated significant eyeblink responses. These results are consistent with dual plasticity models of cerebellar learning and strongly support the general hypothesis that conditioned responses are the result of strengthening of preexisting connections in the nervous system.

Timing of conditioned responses utilizing electrical stimulation in the region of the interpositus nucleus as a CS

A large body of evidence indicates that the cerebellum is essential for the acquisition, retention, and expression of the standard delay conditioned eyeblink response and that the basic memory trace appears to be established in the anterior interpositus nucleus (IP). Adaptive timing of the conditioned response (CR) is a prominent feature of classical conditioning—the CR peaks at the time of onset of the unconditioned stimulus (US) over a wide range of CS-US interstimulus intervals (ISI). A key issue is whether this timing is established by the cerebellar circuitry or prior to the cerebellum. In this study timing of conditioned eyeblink responses established via electrical stimulation of the interpositus nucleus as a conditioned stimulus (CS) was analyzed prior to and following modification of the CS-US interval in well-trained rabbits. Consistent with previous results, learning under these conditions is very rapid and robust. The CR peak eyeblink latencies are initially timed to the US onset and adjust accordingly to lengthening or shortening of the CS-US interval, just as with peripheral CSs. The acquisition of conditioned eyeblink responses by direct electrical stimulation of the IP as a CS thus retains temporal flexibility following shifts in the CS-US delay, as found in standard classical eyeblink conditioning procedures.

Sensitization of fear learning to mild unconditional stimuli in male and female rats.

Stress-enhanced fear learning (SEFL) refers to the long-lasting nonassociative sensitization produced by intense stress (e.g., repeated and unpredictable footshock) that results in increased fear learning to a mild conditioning regimen (e.g., one shock). SEFL experiments suggest that one component of posttraumatic behavior is inappropriately strong fear conditioning occurring to relatively mild stressors. Past reports of SEFL have used the same intensity (1 mA) of footshock to cause both the sensitization and conditioning of new fear. SEFL would be a particularly problematic component of posttrauma behavior if intense stress results in substantial fear conditioning under conditions that would not normally support conditioning. Therefore, we determined if SEFL occurred when the conditioning shock was substantially milder than the SEFL-inducing shock. The results indicate that exposure to a sensitizing regimen of shock can convert a mild footshock that normally does not support measurable levels of fear conditioning into one that causes substantial learned fear. Moreover, as the intensity of single footshock increases, so does the capacity of the prior stressor to contribute to the sensitization of fear responses. Consistent with prior studies, males acquired and retained a greater level of fear conditioning than female rats, however the level of sensitization did not differ between sexes.

Effects of a corneal anesthetic on extinction of the classically conditioned nictitating membrane response in the rabbit (Oryctolagus cuniculus).

Rabbits (Oryctolagus cuniculus) were presented with 7 daily sessions of tone-alone training after conditioning. Before the beginning of each of the first 4 extinction sessions, an artificial tear solution or tetracaine hydrochloride was administered to the cornea of rabbits in the control group (n = 6) and experimental group (n = 7), respectively. There were no between-group differences in the percentage of conditioned responses between both groups. However, the amplitude of the conditioned response was notably reduced in the tetracaine group (M = 0.40, SEM +/- 0.216) relative to the control group (M = 1.32, SEM +/- 0.639) early in extinction. Results seem to suggest that although motor output has been found to play an important role in extinction, corneal sensory feedback is not necessary.