An organization of visual and auditory fear conditioning in the lateral amygdala

Pavlovian fear conditioning is an evolutionary conserved and extensively studied form of associative learning and memory. In mammals, the lateral amygdala (LA) is an essential locus for Pavlovian fear learning and memory. Despite significant progress unraveling the cellular mechanisms responsible for fear conditioning, very little is known about the anatomical organization of neurons encoding fear conditioning in the LA. One key question is how fear conditioning to different sensory stimuli is organized in LA neuronal ensembles. Here we show that Pavlovian fear conditioning, formed through either the auditory or visual sensory modality, activates a similar density of LA neurons expressing a learning-induced phosphorylated extracellular signal-regulated kinase (p-ERK1/2). While the size of the neuron population specific to either memory was similar, the anatomical distribution differed. Several discrete sites in the LA contained a small but significant number of p-ERK1/2-expressing neurons specific to either sensory modality. The sites were anatomically localized to different levels of the longitudinal plane and were independent of both memory strength and the relative size of the activated neuronal population, suggesting some portion of the memory trace for auditory and visually cued fear conditioning is allocated differently in the LA. Presenting the visual stimulus by itself did not activate the same p-ERK1/2 neuron density or pattern, confirming the novelty of light alone cannot account for the specific pattern of activated neurons after visual fear conditioning. Together, these findings reveal an anatomical distribution of visual and auditory fear conditioning at the level of neuronal ensembles in the LA.

A high and low fear phenotype mouse model

Fear-related illnesses such as post-traumatic stress disorder (PTSD) impose a tremendous burden on individual quality of life, families, and the national economy. In the military population, 17-20% of services members returning from deployment are diagnosed with PTSD. While treatments have improved for PTSD and are helpful for some, many people continue to suffer despite therapy. The aim of this research is to examine fear memory behaviourally and at the cellular level in the amygdala by using a unique inter-cross strain of high and low fear phenotype mice. An extended outcross C57BL/6J x DBA/2J (F8) are selected for high or low Pavlovian fear memory to context and cue. On presentation of either the original learning context or the cue (tone) mice display high or low levels of freezing as a behavioural measure of fear. In order to identify key aspects of the cellular basis of this difference in fear memory behaviour we are making measurements of protein levels and neuron numbers of a known pathway involved in the consolidation of a long term fear memory (pMAPK). Ongoing studies aim to determine if high fear behaviour is associated with differential signalling in the lateral amygdala compared to low fear behaviour. Additionally, by blocking this pathway in the lateral amygdala (LA), we aim to reduce fear behaviour following Pavlovian fear conditioning. This research will help to unravel the cellular mechanisms underlying high fear behaviour and advance the field toward targeted treatment and improved outcomes, ultimately improving human quality of life.

How are falls and fear of falling associated with objectively measured physical activity in a cohort of community-dwelling older men?

BACKGROUND: Falls affect approximately one third of community-dwelling older adults each year and have serious health and social consequences. Fear of falling (FOF) (lack of confidence in maintaining balance during normal activities) affects many older adults, irrespective of whether they have actually experienced falls. Both falls and fear of falls may result in restrictions of physical activity, which in turn have health consequences. To date the relation between (i) falls and (ii) fear of falling with physical activity have not been investigated using objectively measured activity data which permits examination of different intensities of activity and sedentary behaviour. METHODS: Cross-sectional study of 1680 men aged 71-92 years recruited from primary care practices who were part of an on-going population-based cohort. Men reported falls history in previous 12 months, FOF, health status and demographic characteristics. Men wore a GT3x accelerometer over the hip for 7 days. RESULTS: Among the 12% of men who had recurrent falls, daily activity levels were lower than among non-fallers; 942 (95% CI 503, 1381) fewer steps/day, 12(95% CI 2, 22) minutes less in light activity, 10(95% CI 5, 15) minutes less in moderate to vigorous PA [MVPA] and 22(95% CI 9, 35) minutes more in sedentary behaviour. 16% (n = 254) of men reported FOF, of whom 52% (n = 133) had fallen in the past year. Physical activity deficits were even greater in the men who reported that they were fearful of falling than in men who had fallen. Men who were fearful of falling took 1766(95% CI 1391, 2142) fewer steps/day than men who were not fearful, and spent 27(95% CI 18, 36) minutes less in light PA, 18(95% CI 13, 22) minutes less in MVPA, and 45(95% CI 34, 56) minutes more in sedentary behaviour. The significant differences in activity levels between (i) fallers and non-fallers and (ii) men who were fearful of falling or not fearful, were mediated by similar variables; lower exercise self-efficacy, fewer excursions from home and more mobility difficulties. CONCLUSIONS: Falls and in particular fear of falling are important barriers to older people gaining health benefits of walking and MVPA. Future studies should assess the longitudinal associations between falls and physical activity.

High and low fear phenotypes show differences in phosphorylated (p44/42 ERK) MAPK-expressing neurons in the lateral amygdala

Post traumatic stress disorder (PTSD) is a serious medical condition effecting both military and civilian populations. While its etiology remains poorly understood it is characterized by high and prolonged levels of fear responding. One biological unknown is whether individuals expressing high or low conditioned fear memory encode the memory differently and if that difference underlies fear response. In this study we examined cellular mechanisms that underlie high and low conditioned fear behavior by using an advanced intercrossed mouse line (B6D2F1) selected for high and low Pavlovian fear response. A known requirement for consolidation of fear memory, phosphorylated mitogen activated protein kinase (p44/42 (ERK) MAPK (pMAPK)) in the lateral amygdala (LA) is a reliable marker of fear learning-related plasticity. In this study, we asked whether high and low conditioned fear behavior is associated with differential pMAPK expression in the LA and if so, is it due to an increase in neurons expressing pMAPK or increased pMAPK per neuron. To examine this, we quantified pMAPK-expressing neurons in the LA at baseline and following Pavlovian fear conditioning. Results indicate that high fear phenotype mice have more pMAPK-expressing neurons in the LA. This finding suggests that increased endogenous plasticity in the LA may be a component of higher conditioned fear responses and begins to explain at the cellular level how different fear responders encode fear memories. Understanding how high and low fear responders encode fear memory will help identify novel ways in which fear-related illness risk can be better predicted and treated.

Mice behaviorally selected for high Pavlovian fear memory exhibit fear generalization, HPA-axis alterations and increased basal activity in cortico-limbic circuits

Biological factors underlying individual variability in fearfulness and anxiety have important implications for stress-related psychiatric illness including PTSD and major depression. Using an advanced intercross line (AIL) derived from C57BL/6 and DBA/2J mouse strains and behavioral selection over 3 generations, we established two lines exhibiting High or Low fear behavior after fear conditioning. Across the selection generations, the two lines showed clear differences in training and tests for contextual and conditioned fear. Before fear conditioning training, there were no differences between lines in baseline freezing to a novel context. However, after fear conditioning High line mice demonstrated pronounced freezing in a new context suggestive of poor context discrimination. Fear generalization was not restricted to contextual fear. High fear mice froze to a novel acoustic stimulus while freezing in the Low line did not increase over baseline. Enhanced fear learning and generalization are consistent with transgenic and pharmacological disruption of the hypothalamic-pituitary-adrenal axis (HPA-axis) (Brinks, 2009, Thompson, 2004, Kaouane, 2012). To determine whether there were differences in HPA-axis regulation between the lines, morning urine samples were collected to measure basal corticosterone. Levels of secreted corticosterone in the circadian trough were analyzed by corticosterone ELISA. High fear mice were found to have higher basal corticosterone levels than low line animals. Examination of hormonal stress response components by qPCR revealed increased expression of CRH mRNA and decreased mRNA for MR and CRHR1 in hypothalamus of high fear mice. These alterations may contribute to both the behavioral phenotype and higher basal corticosterone in High fear mice. To determine basal brain activity in vivo in High and Low fear mice we used manganese-enhanced magnetic resonance imaging (MEMRI). Analysis revealed a pattern of basal brain activity made up of amygdala, cortical and hippocampal circuits that was elevated in the High line. Ongoing studies also seek to determine the relative balance of excitatory and inhibitory tone in the amygdala and hippocampus and the neuronal structure of its neurons. While these heterogeneous lines are selected on fear memory expression, HPA-axis alterations and differences in hippocampal activity segregate with the behavioral phenotypes. These differences are detectable in a basal state strongly suggesting these are biological traits underlying the behavioral phenotype (Johnson et al, 2011).

GABAA and NMDA receptors contribute to synaptic integration in lateral amygdala neurons

In classical fear conditioning a neutral conditioned stimulus (CS), is paired with an aversive unconditioned stimulus (US). The CS thereby acquires the capacity to elicit a fear response. This type of associative learning is thought to require co-activation of principal neurons in the lateral nucleus of the amygdala (LA) by two sets of synaptic inputs...

Amygdala-dependent learning increases the number of spinophilin-immunoreactive dendritic spines in the lateral amygdala

During Pavlovian auditory fear conditioning a previously neutral auditory stimulus (CS) gains emotional significance through pairing with a noxious unconditioned stimulus (US). These associations are believed to be formed by way of plasticity at auditory input synapses on principal neurons in the lateral nucleus of the amygdala (LA). One proposed form of cellular plasticity involves structural changes in the number and morphology of dendritic spines...

Differences in intrinsic properties of dorsally and ventrally located lateral amygdala neurons of the rat

During Pavlovian auditory fear conditioning a previously neutral auditory stimulus (CS) gains emotional significance through pairing with a noxious unconditioned stimulus (US). These associations are believed to be formed by way of plasticity at auditory input synapses on principal neurons of the lateral nucleus of the amygdala (LA). While the LA has been implicated as a key brain structure for fear learning, how its network of cellular components performs these operations is not yet known...