Functional MRI of human amygdala activity during Pavlovian fear conditioning: stimulus processing versus response expression

Although laboratory animal studies have shown that the amygdala plays multiple roles in conditional fear, less is known about the human amygdala. Human subjects were trained in a Pavlovian fear conditioning paradigm during functional magnetic resonance imaging (fMRI). Brain activity maps correlated with reference waveforms representing the temporal pattern of visual conditional stimuli (CSs) and subject-derived autonomic responses were compared. Subjects receiving paired CS-shock presentations showed greater amygdala activity than subjects receiving unpaired CS-shock presentations when their brain activity was correlated with a waveform generated from their behavioral responses. Stimulus-based waveforms revealed learning differences in the visual cortex, but not in the amygdala. These data support the view that the amygdala is important for the expression of learned behavioral responses during Pavlovian fear conditioning.     

Effects of exercise on Pavlovian fear conditioning

Exercise promotes multiple changes in hippocampal morphology and should, as a result, alter behavioral function. The present experiment investigated the effect of exercise on learning using contextual and auditory Pavlovian fear conditioning. Rats remained inactive or voluntarily exercised (VX) for 30 days, after which they received auditory-cued fear conditioning. Twenty-four hours later, rats were tested for learning of the contextual and auditory conditional responses. No differences in freezing behavior to the discrete auditory cue were observed during the training or testing sessions. However, VX rats did freeze significantly more compared to controls when tested in the training context 24 hr after exposure to shock. The enhancement of contextual fear conditioning provides further evidence that exercise alters hippocampal function and learning.     

Effects of Pavlovian fear conditioning on septohippocampal metabolism in rats

The effects of classical fear conditioning in different regions of the limbic system were analysed using cytochrome oxidase (CO) histochemistry. Wistar rats were submitted to different conditions. Rats in the group Paired received tone-shock pairing, to elicit conditioned suppression of lever pressing (i.e., tone will evoke conditioned fear responses). The group Unpaired underwent random presentations of these stimuli and developed no conditioned fear. Untrained animals were also included as a control group. A significant decrease in CO activity was found in the medial septal area and the dorsal hippocampus (CA3 subfield and dentate gyrus) in the group Paired as compared with the group Unpaired. Furthermore there was greater metabolic activity in the control group as compared with the other two groups. No differences in CO labelling of the basolateral amygdala were detected among all groups. These findings suggest that the septohippocampal system plays an important role in controlling conditioned fear behaviour.     

The structure of Pavlovian fear conditioning in the amygdala

Do different brains forming a specific memory allocate the same groups of neurons to encode it? One way to test this question is to map neurons encoding the same memory and quantitatively compare their locations across individual brains. In a previous study, we used this strategy to uncover a common topography of neurons in the dorsolateral amygdala (LAd) that expressed a learning-induced and plasticity-related kinase (p42/44 mitogen-activated protein kinase; pMAPK), following auditory Pavlovian fear conditioning. In this series of experiments, we extend our initial findings to ask to what extent this functional topography depends upon intrinsic neuronal structure. We first showed that the majority (87 %) of pMAPK expression in the lateral amygdala was restricted to principal-type neurons. Next, we verified a neuroanatomical reference point for amygdala alignment using in vivo magnetic resonance imaging and in vitro morphometrics. We then determined that the topography of neurons encoding auditory fear conditioning was not exclusively governed by principal neuron cytoarchitecture. These data suggest that functional patterning of neurons undergoing plasticity in the amygdala following Pavlovian fear conditioning is specific to memory formation itself. Further, the spatial allocation of activated neurons in the LAd was specific to cued (auditory), but not contextual, fear conditioning. Spatial analyses conducted at another coronal plane revealed another spatial map unique to fear conditioning, providing additional evidence that the functional topography of fear memory storing cells in the LAd is non-random and stable. Overall, these data provide evidence for a spatial organizing principle governing the functional allocation of fear memory in the amygdala.     

Opioid receptors regulate the extinction of Pavlovian fear conditioning

Rats received a single pairing of an auditory conditioned stimulus (CS) with a footshock unconditioned stimulus (US). The fear (freezing) that had accrued to the CS was then extinguished. Injection of naloxone prior to this extinction significantly impaired the development of extinction. This impairment was mediated by opioid receptors in the brain and was not observed when naloxone was injected after extinction training. Finally, an injection of naloxone on test failed to reinstate extinguished responding that had already accrued to the CS. These experiments show that opioid receptors regulate the development, but not the expression, of fear extinction and are discussed with reference to the roles of opioid receptors in US processing, memory, and appetitive motivation.     

Resting cerebral metabolism correlates with skin conductance and functional brain activation during fear conditioning

We investigated whether resting brain metabolism can be used to predict autonomic and neuronal responses during fear conditioning in 20 healthy humans. Regional cerebral metabolic rate for glucose was measured via positron emission tomography at rest. During conditioning, autonomic responses were measured via skin conductance, and blood oxygen level dependent signal was measured via functional magnetic resonance imaging. Resting dorsal anterior cingulate metabolism positively predicted differentially conditioned skin conductance responses. Midbrain and insula resting metabolism negatively predicted midbrain and insula functional reactivity, while dorsal anterior cingulate resting metabolism positively predicted midbrain functional reactivity. We conclude that resting metabolism in limbic areas can predict some aspects of psychophysiological and neuronal reactivity during fear learning.     

Amygdalar unit activity during three learning tasks: eyeblink classical conditioning, Pavlovian fear conditioning, and signaled avoidance conditioning

Neural activity in central and basolateral amygdala nuclei (CeA and BLA, respectively) was recorded during delay eyeblink conditioning, Pavlovian fear conditioning, and signaled barpress avoidance. During paired training, the CeA exhibited robust learning-related excitatory activity during all 3 tasks. By contrast, the BLA exhibited minimal activity during eyeblink conditioning, while demonstrating pronounced increases in learning-related excitatory responsiveness during fear conditioning and barpress avoidance. In addition, the relative amount of amygdalar activation observed appeared to be related to the relative intensity of the unconditioned stimulus and somatic requirements of the task. Results suggest the CeA mediates the Pavlovian association between sensory stimuli and the BLA mediates the modulation of instrumental responding through the assignment of motivational value to the unconditioned stimulus.     

Arousal-related associative response characteristics of dorsal lateral geniculate nucleus neurons during acoustic Pavlovian fear conditioning

This research determined whether fear-conditioned, acoustic stimuli induce thalamic arousal reflected in associative responses in dorsal lateral geniculate nucleus (dLGN) neurons. Rabbits received a Pavlovian discriminative fear conditioning procedure in which one tone conditioned stimulus (CS +) was always paired with an aversive unconditioned stimulus (US) and another tone (CS-) was never paired with the US. Responses of single dLGN neurons to random CS+ and CS- presentations were then recorded. Nine of 15 recorded neurons demonstrated significantly greater firing during the CS+ versus the CS-. Their spontaneous activity demonstrated tonic firing during increased neocortical arousal and burst firing during decreased neocortical arousal. The results demonstrate that dLGN neurons show associative responses to fear-conditioned, acoustic stimuli and present a model for investigating the neural circuits by which such stimuli affect sensory processing at the thalamic level.     

Reprint of: Resting cerebral metabolism correlates with skin conductance and functional brain activation during fear conditioning

We investigated whether resting brain metabolism can be used to predict autonomic and neuronal responses during fear conditioning in 20 healthy humans. Regional cerebral metabolic rate for glucose was measured via positron emission tomography at rest. During conditioning, autonomic responses were measured via skin conductance, and blood oxygen level dependent signal was measured via functional magnetic resonance imaging. Resting dorsal anterior cingulate metabolism positively predicted differentially conditioned skin conductance responses. Midbrain and insula resting metabolism negatively predicted midbrain and insula functional reactivity, while dorsal anterior cingulate resting metabolism positively predicted midbrain functional reactivity. We conclude that resting metabolism in limbic areas can predict some aspects of psychophysiological and neuronal reactivity during fear learning.     

Impact of healthy aging on awareness and fear conditioning

Fear conditioning has provided a useful model system for studying associative emotional learning, but the impact of healthy aging has gone relatively unexplored. The present study investigated fear conditioning across the adult life span in humans. A delay discrimination task was employed using visual conditioned stimuli and an auditory unconditioned stimulus. Awareness of the reinforcement contingencies was assessed in a postexperimental interview. Compared with young adult participants, middle-aged and older adults displayed reductions in unconditioned responding, discriminant conditioning, and contingency awareness. When awareness and overall arousability were taken into consideration, there were no residual effects of aging on conditioning. These results highlight the importance of considering the influence of declarative knowledge when interpreting age-associated changes in discriminative conditioned learning.     

The effects of intra-amygdaloid infusions of a D2 dopamine receptor antagonist on Pavlovian fear conditioning

The present study examined the effects of bilateral intra-amygdaloid infusions of the D2 receptor antagonist, eticlopride, on the acquisition and expression of Pavlovian fear conditioning as measured by freezing to acoustic and background contextual stimuli in the rat. Infusions of eticlopride before acquisition or before both acquisition and retention testing significantly attenuated conditioned freezing to tone presentations during the retention test 24 hr later. No effects, however, were observed on freezing that emerged during acquisition. Furthermore, these effects were not attributable to state-dependent learning effects or alterations in baseline activity or shock reactivity. In conclusion, these results suggest that amygdaloid dopamine transmission at D2 receptors contributes to the formation and/or consolidation of fear memories.     

Temporal-difference prediction errors and Pavlovian fear conditioning: role of NMDA and opioid receptors

Three experiments studied temporal-difference (TD) prediction errors during Pavlovian fear conditioning. In Stage I, rats received conditioned stimulus A (CSA) paired with shock. In Stage II, they received pairings of CSA and CSB with shock that blocked learning to CSB. In Stage III, a serial overlapping compound, CSB --> CSA, was followed by shock. The change in intratrial durations supported fear learning to CSB but reduced fear of CSA, revealing the operation of TD prediction errors. N-methyl- D-aspartate (NMDA) receptor antagonism prior to Stage III prevented learning, whereas opioid receptor antagonism selectively affected predictive learning. These findings support a role for TD prediction errors in fear conditioning. They suggest that NMDA receptors contribute to fear learning by acting on the product of predictive error, whereas opioid receptors contribute to predictive error.     

Response characteristics of neurons in the medial component of the medial geniculate nucleus during Pavlovian differential fear conditioning in rabbits

Recent evidence suggests that the amygdaloid central nucleus (ACE) may contribute significantly to Pavlovian fear-conditioned bradycardic responses during the presentation of conditioned emotional stimuli. Because the medial component of the medial geniculate nucleus (MGm) is a major source of input to the region of the ACE, the extracellular single-unit responses of MGm neurons were examined during Pavlovian differentially conditioned bradycardic responding in rabbits. Conditioning involved pairing one tone (CS+) with paraorbital shock and presenting another tone (CS-) in the absence of shock. Two general classes of MGm neurons were identified based on their conditioned-response characteristics. Both groups responded differentially to the CSs. One group responded with greater increases in activity and at a shorter latency to the CS+ compared with the CS-, whereas the other group responded with greater increases in activity and at a shorter latency to the CS- compared with the CS+. Recordings from MGm neurons in naive rabbits prior to conditioning provided evidence that the acoustic stimuli used subsequently as the CS+ and CS- did not evoke differential responses. These results suggest that the MGm along with the ACE may be forebrain components of a neural circuit involved in the acquisition and/or expression of Pavlovian fear-conditioned bradycardic responses.     

Phosphatemic control during acute renal failure: intermittent hemodialysis versus continuous hemodiafiltration

BACKGROUND: Achieving "adequacy of dialysis" includes the maintenance of normal serum phosphate concentrations and is an important therapeutic goal in the treatment of acute renal failure (ARF). It is unknown whether this goal is best achieved with intermittent or continuous renal replacement therapy. METHODS: We compared the effects of continuous veno-venous hemodiafiltration (CVVHDF) and intermittent hemodialysis (IHD) on serum phosphate concentrations using daily morning blood tests in 88 consecutive intensive care patients half of which were treated with IHD and half with CRRT RESULTS: Mean patient age was 54+/-14 years for IHD and 60+/-14 years for CVVHDF (NS). However, patients who received CVVHDF were more critically ill (mean APACHE II scores: 24.4+/-5.1 for IHD vs. 29.2+/-5.7 for CVVHDF, p<0.003). Before treatment, the serum phosphate concentration was 2.04+/-0.16 mmoll L for IHD and 1.96+/-0.17 mmoll L for CVVHDF (NS), with abnormal values in 79.4% of IHD patients and in 64.8% of CVVHDF patients (NS). During treatment, CVVHDF induced a greater reduction in serum phosphate (p=0.02) during the first 48 hours and conferred superior subsequent control of hyperphosphatemia (achieved in 64.6% of observations during CVVHDF vs. 41.8% during IHD; p<0.0001). The serum phosphate concentration was also more likely to be within the normal range during CVVHDF (55.3% vs.36.2%; p<0.0001). There was a trend toward more frequent hypophosphatemia (9.3% vs. 5.6%; P<0.1) during CVVHDF CONCLUSIONS: Abnormal serum phosphate concentrations are frequent in ARF patients before and during renal replacement, however, normalization of phosphatemia is achieved more frequently with CVVHDF.     

Effect of hippocampal lesions on heart-rate during classical fear conditioning

Of 22 male albino rats, eight received bilateral lesions at three levels of the hippocampus, six received bilateral lesions of the amygdaloid complex and eight served as unoperated controls. Each rat received 30 fear conditioning trials in which shock was paired with a CS, either light or a tone. After every tenth trial there was a test trial in which heart-beats were recorded. The typical heart-rate response of controls was deceleration-acceleration while for hippocampal lesioned rats it was short latency acceleration. The results were interpreted in terms of two heart-rate theories.     

Effect of rat soleus muscle overload on neuromuscular transmission efficacy during continuous and intermittent activation

Increased neuromuscular activity is known to provoke morphological and functional adaptations at the neuromuscular synapse. Most of these changes have been documented following endurance exercise training programmes. In this study, the effect of rat soleus muscle overload produced by tenotomy plus voluntary wheel-cage activity on neuromuscular transmission efficacy was investigated. The overload protocol increased miniature endplate potential (MEPP) and endplate potential (EPP) amplitudes by 17 and 19%, respectively (both P < 0.01), and increased MEPP frequency by 86% (P < 0.01). EPP amplitude rundown during continuous trains of activation was attenuated by approximately 10% in the overloaded group (P < 0.01). Also, during intermittent activation, the overload protocol attenuated EPP amplitude rundown, mainly by enhancing EPP amplitude recovery by approximately 10% during the quiescent periods (P < 0.01). Although the present results show that both the degree and direction of adaptation are similar to what has been observed at rat soleus neuromuscular junctions following an endurance training protocol, there are important nuances between the results, suggesting different mechanisms through which these changes may occur.     

On testing current cognitive theories of Pavlovian conditioning in the human Pavlovian autonomic transswitching preparation.

The Rescorla-Wagner theory, which Lachnit and Kimmel's transswitching evidence has apparently contradicted, is fundamentally cognitive, inasmuch as it views Pavlovian conditioning as "the learning of relations among events". Kimmel and Lachnit's criticism that Lovibond's theory-saving defence "does little more than immunize a theory against problematic empirical findings" is analogous to the charge of circularity in explanation that cognitive, Tolmanian researchers used to bring against their S-R, Hullian opponents. A way out of the circle in human autonomic transswitching is to measure the cognitive process of relational conditional stimulus-unconditional stimulus contingency learning throughout the experiment, as has been done in simple Pavlovian human autonomic conditioning, although it may also be desirable to check whether such continuous measurement changes the basic transswitching phenomenon itself. The other issue briefly commented on is the relation between animal and human experiments and theoretical formulations, where the more basic distinction may be not between animals and humans, but between conditional stimulus-instrumental response and conditional stimulus-conditional response preparations. The latter preparations (of which human electrodermal transswitching is an instance) appear to be sensitive to S-R as well as (cognitive) S-S factors.     

Does activation of the baroreceptors reinforce differential Pavlovian conditioning of heart rate responses?

High- and low-pitched tones (CS+ and CS?) signalled baroreceptor stimulation or inhibition (US+ and US-) on 6-s conditioning trials (n= 128). Baroreceptor stimulation was induced by the phase-related external suction (PRES) method of Rau et al. (1992) in which a brief pulse of negative external pressure is applied to the neck at systole and one of positive pressure at diastole within each cardiac cycle (the reverse sequence is used for baroreceptor inhibition). Changes in heart period (R-R intervals) confirmed that PRES manipulated the baroreceptors in the presence of CS+ and CS? without habituation over conditioning trials. However, conditioned heart period responses were not observed on test trials (n= 32) in which CS+ and CS? were presented with the baroreceptor manipulation removed. Subjects were unable to state which CS had signalled baroreceptor stimulation and inhibition when given PRES-alone trials after the conditioning phase (differential attention thus controlled). These results (a) confirm that the differential effect of the two PRES stimuli was specific to the baroreceptors and (b) support earlier studies that have found that differential conditioning is impaired when CS?US relations are not processed in attention. We discuss implications regarding when baroreceptor firing might be discriminable and reinforcing.     

Mapping Pavlovian conditioning effects on the brain: blocking, contiguity, and excitatory effects

Pavlovian conditioning effects on the brain were investigated by mapping rat brain activity with fluorodeoxyglucose (FDG) autoradiography. The goal was to map the effects of the same tone after blocking or eliciting a conditioned emotional response (CER). In the tone-blocked group, previous learning about a light blocked a CER to the tone. In the tone-excitor group, the same pairings of tone with shock US resulted in a CER to the tone in the absence of previous learning about the light. A third group showed no CER after pseudorandom presentations of these stimuli. Brain systems involved in the various associative effects of Pavlovian conditioning were identified, and their functional significance was interpreted in light of previous FDG studies. Three conditioning effects were mapped: 1) blocking effects: FDG uptake was lower in medial prefrontal cortex and higher in spinal trigeminal and cuneate nuclei in the tone-blocked group relative to the tone-excitor group. 2) Contiguity effects: relative to pseudorandom controls, similar FDG uptake increases in the tone-blocked and -excitor groups were found in auditory regions (inferior colliculus and cortex), hippocampus (CA1), cerebellum, caudate putamen, and solitary nucleus. Contiguity effects may be due to tone-shock pairings common to the tone-blocked and -excitor groups rather than their different CER. And 3) excitatory effects: FDG uptake increases limited to the tone-excitor group occurred in a circuit linked to the CER, including insular and anterior cingulate cortex, vertical diagonal band nucleus, anterior hypothalamus, and caudoventral caudate putamen. This study provided the first large-scale map of brain regions underlying the Kamin blocking effect on conditioning. In particular, the results suggest that suppression of prefrontal activity and activation of unconditioned stimulus pathways are important neural substrates of the Kamin blocking effect.