Fear-potentiated Startle Response Research Articles

Medial prefrontal cortex stimulation modulates the processing of conditioned fear.

The extinction of conditioned fear depends on an efficient interplay between the amygdala and the medial prefrontal cortex (mPFC). In rats, high-frequency electrical mPFC stimulation has been shown to improve extinction by means of a reduction of amygdala activity. However, so far it is unclear whether stimulation of homologues regions in humans might have similar beneficial effects. Healthy volunteers received one session of either active or sham repetitive transcranial magnetic stimulation (rTMS) covering the mPFC while undergoing a 2-day fear conditioning and extinction paradigm. Repetitive TMS was applied offline after fear acquisition in which one of two faces (CS+ but not CS−) was associated with an aversive scream (UCS). Immediate extinction learning (day 1) and extinction recall (day 2) were conducted without UCS delivery. Conditioned responses (CR) were assessed in a multimodal approach using fear-potentiated startle (FPS), skin conductance responses (SCR), functional near-infrared spectroscopy (fNIRS), and self-report scales. Consistent with the hypothesis of a modulated processing of conditioned fear after high-frequency rTMS, the active group showed a reduced CS+/CS− discrimination during extinction learning as evident in FPS as well as in SCR and arousal ratings. FPS responses to CS+ further showed a linear decrement throughout both extinction sessions. This study describes the first experimental approach of influencing conditioned fear by using rTMS and can thus be a basis for future studies investigating a complementation of mPFC stimulation to cognitive behavioral therapy (CBT).

Conditioned fear is modulated by CRF mechanisms in the periaqueductal gray columns

The periaqueductal gray (PAG) columns have been implicated in controlling stress responses through corticotropin-releasing factor (CRF), which is a neuropeptide with a prominent role in the etiology of fear- and anxiety-related psychopathologies. Several studies have investigated the involvement of dorsal PAG (dPAG) CRF mechanisms in models of unconditioned fear. However, less is known about the role of this neurotransmission in the expression of conditioned fear memories in the dPAG and ventrolateral PAG (vlPAG) columns. We assessed the effects of ovine CRF (oCRF 0.25 and 1.0μg/0.2μL) locally administered into the dPAG and vlPAG on behavioral (fear-potentiated startle and freezing) and autonomic (arterial pressure and heart rate) responses in rats subjected to contextual fear conditioning. The lower dose injected into the columns promoted proaversive effects, enhanced contextual freezing, increased the blood pressure and heart rate and decreased tail temperature. The lower dose of oCRF into the vlPAG, but not into the dPAG, produced a pronounced enhancement of the fear-potentiated startle response. The results imply that the PAG is a heterogeneous structure that is involved in the coordination of distinct behaviors and autonomic control, suggest PAG involvement in the expression of contextual fear memory as well as implicate the CRF as an important modulator of the neural substrates of fear in the PAG.

Fear-potentiated startle response is unrelated to social or emotional functioning in adolescents with autism spectrum disorders.

It has been suggested that atypical amygdala function contributes to the social impairments characteristic of autism spectrum disorders (ASDs). Previous research has demonstrated that adolescents and adults with ASD generate normal response during a fear-potentiated startle paradigm, suggesting this aspect of amygdala function is intact and may not account for the social dysfunction associated with the condition. The amygdala also plays a crucial role in the expression of anxiety and may contribute to high rates of reported anxiety in individuals with ASD. The present study partially replicates prior work by examining the fear-potentiated startle response in adolescents with ASD, and extends this to investigate the relationship between startle response and anxiety. Eyeblink magnitude and latency (electromyographic activity; EMG) were collected from 20 adolescents with ASD and 19 typically developing (TD) age-matched adolescents during a fear-potentiated startle paradigm. Parent-report and self-report of anxiety and additional psychiatric symptoms were collected. Parental reports indicated higher rates of associated psychopathology in adolescents with ASD compared with TD adolescents. Consistent with previous results, both groups showed normal potentiated startle response, and no group differences in EMG were found. Symptoms of anxiety and level of social impairment were unrelated to startle response. These findings held for all levels of anxiety, suggesting that within the context of the fear-potentiated startle paradigm, amygdala response is not associated with degree of atypical social or emotional functioning in ASD.

Fear-Potentiated Startle Response Is Unrelated to Social or Emotional Functioning in Adolescents With Autism Spectrum Disorders: Startle response in adolescents with ASD

Fear-Potentiated Startle Response Is Unrelated to Social or Emotional Functioning in Adolescents With Autism Spectrum Disorders: Startle response in adolescents with ASD

Temporal properties of fear extinction—Does time matter?

Fear extinction can be defined as the weakening of the expression of a conditioned response (CR) by extended experience of nonreinforcement. Conceptually, two distinct models have been invoked to account for extinction. R. A. Rescorla and A. R. Wagner (1972, A theory of Pavlovian conditioning: Variations in the effectiveness of reinforcement and nonreinforcement, in A. H. B. W. F. Prokasy (Ed.), Classical conditioning: II. Current research and theory, pp. 64-99, New York, NY, Appleton-Century-Crofts) postulated that the number of exposure trials is the primary determinant of CR decrement, whereas C. R. Gallistel and J. Gibbon (2000, Time, rate, and conditioning, Psychological Review, Vol. 107, pp. 289-344) proposed that the decisive event is the cumulated exposure time to the nonreinforced conditioned stimulus (CS) elapsed after the last CS reinforcement. We evaluated these two accounts in a human differential fear conditioning study in which CR was measured with the fear-potentiated startle response. Cumulated duration of nonreinforcement fails to explain our findings, whereas the number of trials appeared critical. In fact, many CS trials with a duration shorter than the acquisition CS duration facilitated within-session extinction, but this effect did not predict the recovery of fear.

Reduced neural activation during an inhibition task is associated with impaired fear inhibition in a traumatized civilian sample

IntroductionImpaired inhibition of fear in the presence of safety cues and a deficiency in the extinction of fear cues are increasingly thought to be important biological markers of Posttraumatic stress disorder (PTSD). Other studies have suggested that there may be altered neural activation during behavioral inhibition tasks in subjects with PTSD. The current study aimed to see whether neural activation during inhibition was reduced in a highly traumatized civilian population, and whether atypical activation was associated with impaired fear inhibition. MethodsThe participants were 41 traumatized women (20 PTSD+, 21 PTSD−) recruited from Grady Memorial Hospital in Atlanta, GA. We used a Go/NoGo procedure with functional magnetic resonance imaging (fMRI) in a high-resolution 3T scanner. Participants were instructed to press a button whenever an “X” or “O” appeared on the screen, but not if a red square appeared behind the letter. Participants were assessed for trauma history and PTSD diagnosis, and completed a fear-potentiated startle and extinction paradigm. ResultsWe found stronger activation in the ventromedial prefrontal cortex (vmPFC) in traumatized subjects without PTSD compared to those with PTSD in the NoGo greater than Go contrast condition. Activation in the vmPFC was negatively correlated with fear-potentiated startle responses during safety signal learning (p = .02) and fear extinction (p = .0002). ConclusionsThese results contribute to understanding of how the neural circuitry involved in inhibitory processes may be deficient in PTSD. Furthermore, the same circuits involved in behavioral inhibition appear to be involved in fear inhibition processes during differential fear conditioning and extinction.

Glutamatergic mechanisms of the dorsal periaqueductal gray matter modulate the expression of conditioned freezing and fear-potentiated startle

It is well known that excitatory amino acids induce unconditioned fear responses when locally injected into the dorsal periaqueductal gray matter (dPAG). However, there are only few studies about the involvement of excitatory amino acids mediation in dPAG in the expression of conditioned fear. The present series of experiments evaluates the participation of AMPA/Kainate and NMDA glutamatergic receptors of dPAG in the expression of conditioned fear, assessed by the fear-potentiated startle (FPS) and conditioned freezing responses. Wistar rats were subjected to fear conditioning to light. Twenty-four hours later, they received intra-dPAG injections of kainic acid or NMDA (AMPA/Kainate and NMDA agonists) and 1,2,3,4-Tetrahydro-6-nitro-2,3-dioxo-benzo[f]quinoxaline-7-sulfonamide disodium salt hydrate (NBQX) or d(−)-2-Amino-7-phosphonoheptanoic acid (AP7) (AMPA/Kainate and NMDA antagonists) and were submitted to the FPS test. Conditioned freezing response was simultaneously measured. Effects of drug treatment on motor activity were evaluated in the open-field test. Intra-dPAG injections of glutamatergic agonists enhanced conditioned freezing and promoted pro-aversive effects in the FPS. Lower doses of the agonists had no effect or enhanced FPS whereas higher doses disrupted FPS, indicating a non-monotonic relationship between fear and FPS. The antagonist NBQX had no significant effects while AP7 decreased conditioned freezing but did not affect FPS. Both antagonists reduced the effects of the agonists. The obtained results cannot be attributed to motor deficits. The results suggest an important role of the AMPA/Kainate and NMDA mechanisms of the dPAG in the expression of conditioned freezing and FPS.

Neural reactivity tracks fear generalization gradients

Recent studies on fear generalization have demonstrated that fear-potentiated startle and skin conductance responses to a conditioned stimulus (CS) generalize to similar stimuli, with the strength of the fear response linked to perceptual similarity to the CS. The aim of the present study was to extend this work by examining neural correlates of fear generalization. An initial experiment (N=8) revealed that insula reactivity tracks the conditioned fear gradient. We then replicated this effect in a larger independent sample (N=25). Activation in the insula, anterior cingulate, right supplementary motor cortex and caudate increased reactivity as generalization stimuli (GS) were more similar to the CS, consistent with participants’ overall ratings of perceived shock likelihood and pupillary response to each stimulus.

Favoured genetic background for testing anxiolytics in the fear-potentiated and light-enhanced startle paradigms in the rat

The fear-potentiated startle (FPS) and the light-enhanced startle (LES) paradigms are rodent tests of fear and anxiety, which combine face validity with predictive validity for clinically effective anxiolytic drugs. However, systematic strain comparisons aimed at identifying a rat strain that shows robust and reliable fear and anxiety responses in both models are missing. Here, we investigated four commonly used laboratory rat strains: Wistar, Sprague Dawley, Long-Evans and F344. Following strong cued fear conditioning training [60 conditioned stimulus–unconditioned stimulus (CS–US) pairings], all strains except Wistar exhibited significant FPS responses. F344 rats showed the strongest FPS response. Following milder cued fear conditioning protocols, designed to reduce the underlying component of contextual fear conditioning (by context pre-exposure or less CS–US pairings), also Wistar rats were able to show significant FPS, albeit still to a lesser extent than F344 rats tested under identical conditions. When tested in the LES protocol (light intensity ∼1500 lx), all strains except Long-Evans displayed significant light-enhanced startle responses. F344 and Wistar showed the strongest LES responses, which were of similar magnitude. The most sensitive strain in both paradigms, F344, was chosen for further pharmacological validation. The clinically active anxiolytic alprazolam (0.3, 1, 3 mg/kg p.o.) dose-dependently reduced both fear-like responses in the FPS paradigm and anxiety-like responses in the LES paradigm at non-myorelaxant dosages. We propose that the F344 rat strain is particularly suited for the predictability assessment of novel anxiolytic drugs in both startle paradigms.

Hydrocortisone suppression of the fear-potentiated startle response and posttraumatic stress disorder

This study examined the effects of oral administration of 20mg hydrocortisone on baseline and fear-potentiated startle in 63 male veterans with or without PTSD. The procedure was based on a two-session, within-subject design in which acoustic startle eyeblink responses were recorded during intervals of threat or no threat of electric shock. Results showed that the magnitude of the difference between startle responses recorded during anticipation of imminent shock compared to "safe" periods was reduced after hydrocortisone administration relative to placebo. This effect did not vary as a function of PTSD group nor were there were any significant group differences in other indices startle amplitude. Findings suggest that the acute elevations in systemic cortisol produced by hydrocortisone administration may have fear-inhibiting effects. This finding may have implications for understanding the role of hypothalamic-pituitary-adrenal (HPA)-axis function in vulnerability and resilience to traumatic stress.

Two-way avoidance acquisition is negatively related to conditioned freezing and positively associated with startle reactions: A dissection of anxiety and fear in genetically heterogeneous rats

The double, fear-driven "passive avoidance/active avoidance" conflict appearing during acquisition of two-way active avoidance, involves high levels of anxiety and a dominant tendency for freezing responses, which in turn run against the appearance of active escape/avoidance behavior. In the present study, by using a large sample genetically heterogeneous (N/Nih-HS) rats, we have tested the hypothesis that rats showing relatively higher levels of context-conditioned freezing (during the initial trials of that task) should show lower efficiency to acquire two-way avoidance behavior, i.e. the prediction that the initial context-conditioned freezing in the shuttle box would be negatively related to avoidance acquisition efficiency. In agreement with such a hypothesis, the results from the three rat subsamples used show that context-conditioned freezing (during the first 5 inter-trial intervals of the 40-trial two-way avoidance session) is negatively correlated (r=-0.34 to r=-0.64, p<0.001) with two-way avoidance acquisition, in a way that subgroups of rats with extremely high or low levels of freezing markedly differ in their avoidance performance: "high freezer" rats show much worse avoidance acquisition than "low freezers". Moreover, the relationships of conditioned freezing and avoidance acquisition with baseline and fear-potentiated startle, as well as with unconditioned anxiety (in the elevated zero-maze test), have also been studied. Taken collectively, the results indicate that: (i) context conditioned freezing is a reliable (and negative) predictor of two-way avoidance acquisition; (ii) baseline and fear-potentiated startle responses show positive associations with avoidance responding, and (iii) unconditioned anxiety in the elevated zero-maze is also negatively associated with two-way avoidance acquisition. Such patterns of associations are considered to be very informative in regard to the search for (common or differential) neural and genetic mechanisms of different forms of (unlearned or learned) anxious or fear responses.

Effects of intracerebral ventricular administration of gastrin-releasing peptide and its receptor antagonist RC-3095 on learned fear responses in the rat

Several lines of evidence have implicated bombesin and its mammalian analogue, gastrin-releasing peptide (GRP), in the mediation and/or modulation of the stress response. However, the physiological role of GRP in mediating conditioned fear responses remains to be elucidated. The objective of the present study was to characterize the role(s) of GRP and its receptor antagonist (D-Tpi6, Leu13 psi[CH2NH]-Leu14) BB (6–14) (RC-3095) in fear-related responses using two animal models of conditioned fear. To this end, the effects of intracerebroventricular (i.c.v.) administration of GRP (0.062, 0.30, 3.0 nmol) and RC-3095 (0.3, 3.0 and 9.0 nmol) were assessed in the conditioned emotional response (CER) and the fear-potentiated startle (FPS) paradigms. In the CER paradigm, i.c.v. administration of GRP dose-dependently (all doses) attenuated the expression of both contextual and cued fear as reflected by a reduction in freezing behavior to both the context (cage where shock was received) and cue (tone paired with shock). Conversely, pretreatment with RC-3095 (high dose), blocked the reduction of contextual and cued fear normally observed over time. Further, in the FPS paradigm, i.c.v. administration of GRP significantly attenuated the fear-potentiated startle response at medium and high doses without affecting basal startle amplitude. In contrast, pretreatment with RC-3095 at the highest dose (9.0 nmol) significantly increased the basal startle amplitude without affecting fear-potentiation, suggesting elevated fear at the onset of testing. These data provide further evidence that GRP is involved in conditioned fear responses.

Attachment and emotion in school-aged children.

One of the primary functions of the attachment behavioral system is to regulate emotional experience under conditions of threat. Although research supports this association among infants and adults, few studies examine the relation between emotion and attachment in middle childhood. This study examined the concurrent associations among children's attachment organization and three indices of emotion reactivity/regulation: self- and parent-assessments of emotion, neuroendocrine reactivity, and fear-potentiated startle response. Ninety-seven 8- to 12-year-old children completed the Child Attachment Interview (CAI) and a fear-potentiated startle paradigm on separate occasions, with salivary cortisol assessed before and after each assessment. Greater attachment security was related to greater child-reported positive trait- and state-level emotion, lower pre-CAI cortisol levels, higher initial startle magnitude during threat, and a faster decrease in startle magnitude during threat. The findings provide initial support that attachment security is related to select measures of emotion, though different methods of assessment yielded discrepant findings. The findings are discussed in terms of their contribution to theory and research examining attachment and emotion.

Chronic anabolic androgenic steroid exposure alters corticotropin releasing factor expression and anxiety-like behaviors in the female mouse

In the past several decades, the therapeutic use of anabolic androgenic steroids (AAS) has been overshadowed by illicit use of these drugs by elite athletes and a growing number of adolescents to enhance performance and body image. As with adults, AAS use by adolescents is associated with a range of behavioral effects, including increased anxiety and altered responses to stress. It has been suggested that adolescents, especially adolescent females, may be particularly susceptible to the effects of these steroids, but few experiments in animal models have been performed to test this assertion. Here we show that chronic exposure of adolescent female mice to a mixture of three commonly abused AAS (testosterone cypionate, nandrolone decanoate and methandrostenolone; 7.5 mg/kg/day for 5 days) significantly enhanced anxiety-like behavior as assessed by the acoustic startle response (ASR), but did not augment the fear-potentiated startle response (FPS) or alter sensorimotor gating as assessed by prepulse inhibition of the acoustic startle response (PPI). AAS treatment also significantly increased the levels of corticotropin releasing factor (CRF) mRNA and somal-associated CRF immunoreactivity in the central nucleus of the amygdala (CeA), as well as neuropil-associated immunoreactivity in the dorsal aspect of the anterolateral division of the bed nucleus of the stria terminalis (dBnST). AAS treatment did not alter CRF receptor 1 or 2 mRNA in either the CeA or the dBnST; CRF immunoreactivity in the ventral BnST, the paraventricular nucleus (PVN) or the median eminence (ME); or peripheral levels of corticosterone. These results suggest that chronic AAS treatment of adolescent female mice may enhance generalized anxiety, but not sensorimotor gating or learned fear, via a mechanism that involves increased CRF-mediated signaling from CeA neurons projecting to the dBnST.

Amygdala-dependent fear conditioning in humans is modulated by the BDNFval66met polymorphism.

The brain-derived neurotrophic factor (BDNF) is critically involved in neuroplasticity, as well as the acquisition, consolidation, and retention of hippocampal- and amygdala-dependent learning. A common functional A-->G single nucleotide polymorphism (BDNFval66met) in the prodomain of the human BDNF gene is associated with abnormal intracellular trafficking and reduced activity-dependent BDNF release. We studied the effect of BDNFval66met in an aversive differential fear conditioning, and a delayed extinction paradigm in 57 healthy participants. Pictures of male faces were used as stimuli and fear learning was quantified by fear potentiated startle (FPS) and skin conductance responses (SCR). Aware BDNF met-carriers show a deficit in amygdala-dependent fear conditioning as indicated by an absence of FPS responses in the last acquisition block. This deficit was maintained in the first block of extinction. No genotype differences were found in conditioned SCR discrimination. These data provide evidence for the involvement of BDNF signaling in human amygdala-dependent learning. We suggest that the BDNF met-allele may have a protective effect for the development of affective pathologies that may be mediated via reduced synaptic plasticity induced by negative experience.

Serotonergic mechanisms of the median raphe nucleus–dorsal hippocampus in conditioned fear: Output circuit involves the prefrontal cortex and amygdala

Independent studies have shown that the median raphe nucleus (MRN) and dorsal hippocampus (DH) are involved in the expression of contextual conditioned fear (CFC). However, studies that examine the integrated involvement of serotonergic mechanisms of the MRN–DH are lacking. To address this issue, a CFC paradigm was used to test whether the serotonergic projections from the MRN to DH can influence CFC. Serotoninergic drugs were infused either into the MRN or DH prior to testing sessions in which freezing and startle responses were measured in the same context where 6 h previously rats received footshocks. A reduction of serotonin (5-HT) transmission in the MRN by local infusions of the 5-HT 1A agonist 8-hydroxy-2-(di- n-propylamino)-tetralin (8-OH-DPAT) decreased freezing in response to the context but did not reduce fear-potentiated startle. This pattern of results is consistent with the hypothesis that MRN serotonergic mechanisms selectively modulate the freezing response to the aversive context. As for the DH, a decrease in postsynaptic 5-HT receptor activity at projection areas has been proposed to be the main consequence of 5-HT 1A receptor activation in the MRN. Intra-DH injections of 8-OH-DPAT inhibited both the freezing and fear-potentiated startle response to the context. To reconcile these findings, an inhibitory mechanism may exist between the incoming 5-HT pathway from the MRN to DH and the neurons of the DH output to other structures. The DH–amygdala or medial prefrontal cortex projections could well be this output circuit modulating the expression of CFC as revealed by measurements of Fos immunoreactivity in these areas.

Nesfatin-1 increases anxiety- and fear-related behaviors in the rat

Nesfatin-1, derived from the protein NEFA/nucleobindin2 (NUCB2), is a newly identified peptide that acts as a potent satiety agent. It has been reported that peptides involved in the regulation of ingestive behavior are also involved in the regulation of the stress response. However, the relation between nesfatin-1 and stressor-related behaviors like anxiety and/or fear has not yet been investigated. The effects of intracerebroventricular (ICV) injection of nesfatin-1 (0, 5, and 25 pmol/3 microl) were assessed in several paradigms that are thought to reflect anxiety and/or fear in rats. Consistent with an anxiogenic effect, nesfatin-1 dose-dependently decreased the percentage of time spent on the open arms of the elevated plus maze, increased latency to approach, and decreased consumption of a palatable snack in an anxiogenic (unfamiliar) environment. Moreover, ICV nesfatin-1 increased the fear-potentiated startle response and the time spent freezing to both context and conditioned cues in a conditioned emotional response test. These findings suggest that in addition to its role as a satiety peptide, nesfatin-1 may also be involved in the mediation of anxiety- and/or fear-related responses.

Activation of ERK/MAPK in the Lateral Amygdala of the Mouse is Required for Acquisition of a Fear-Potentiated Startle response

There is considerable interest in examining the genes that may contribute to anxiety. We examined the function of ERK/MAPK in the acquisition of conditioned fear, as measured by fear-potentiated startle (FPS) in mice as a model for anticipatory anxiety in humans. We characterized the following for the first time in the mouse: (1) the expression of the ERK/MAPK signaling pathway components at the protein level in the lateral amygdala (LA); (2) the time course of activation of phospho-activated MAPK in the LA after fear conditioning; (3) if pharmacological inhibition of pMAPK could modulate the acquisition of FPS; (4) the cell-type specificity of pMAPK in the LA after fear conditioning. Using western blot and immunohistochemistry techniques and injecting the MEK inhibitor U0126 in the LA, we showed the following: (1) both MEK1/MEK2 and ERK1/ERK2 were co-expressed in the LA of the adult mouse brain; (2) there is a peak of pMAPK at 60 min after fear conditioning; (3) the ERK/MAPK signaling pathway activation is essential for the acquisition of an FPS response; (4) at 60 min, the pMAPK are exclusively neuronal and not glial. These results emphasize the importance of this signaling pathway in the acquisition of conditioned fear in the mouse. Given the widely held view that conditioned fear models the essential aspects of anxiety disorders, the results confirm the ERK/MAPK signaling pathway as a molecular target for the treatment of anxiety disorders in the clinic.

Selective involvement of GABAergic mechanisms of the dorsal periaqueductal gray and inferior colliculus on the memory of the contextual fear as assessed by the fear potentiated startle test

The inferior colliculus (IC) together with the dorsal periaqueductal gray (dPAG), the amygdala and the medial hypothalamus make part of the brain aversion system, which has mainly been related to the organization of unconditioned fear. However, the involvement of the IC and dPAG in the conditioned fear is still unclear. It is certain that GABA has a regulatory role on the aversive states generated and elaborated in these midbrain structures. In this study, we evaluated the effects of injections of the GABA-A receptor agonist muscimol (1.0 and 2.0 nmol/0.2 μL) into the IC or dPAG on the freezing and fear-potentiated startle (FPS) responses of rats submitted to a context fear conditioning. Intra-IC injections of muscimol did not cause any significant effect on the FPS or conditioned freezing but enhanced the startle reflex in non-conditioned animals. In contrast, intra-dPAG injections of muscimol caused significant reduction in FPS and conditioned freezing without changing the startle reflex in non-conditioned animals. Thus, intra-dPAG injections of muscimol produced the expected inhibitory effects on the anxiety-related responses, the FPS and the freezing whereas these injections into the IC produced quite opposite effects suggesting that descending inhibitory pathways from the IC, probably mediated by GABA-A mechanisms, exert a regulatory role on the lower brainstem circuits responsible for the startle reflex.

Effects of threat of electric shock and diazepam on the N1/P2 auditory-evoked potential elicited by low-intensity auditory stimuli

The acoustic startle response includes rapid muscular contractions elicited by loud sounds; it may be measured in humans as the electromyographic response of the orbicularis oculi muscle. Enhancement of this response during exposure to threat of electric shock (fear- potentiated startle) is a widely used model of human anxiety. A problem with the use of the startle reflex in studies of human anxiety is the aversiveness of startle-eliciting sounds, which may, in some subjects, exceed the aversiveness of the electric shock itself. We have recently found that the long-latency N1/P2 auditory-evoked potential elicited by loud sounds is subject to fear potentiation. However, it is not known whether N1/P2 potentials elicited by low-intensity sounds, which do not elicit the startle response, are also subject to fear potentiation. This study examined the susceptibility of the N1/P2 potential elicited by low-intensity sounds to fear potentiation, and the effect of the anxiolytic diazepam on the N1/P2 potential in the absence and presence of threat of electric shock. Fifteen male volunteers (18-43 years) participated in three sessions in which they received placebo, diazepam 5 mg and diazepam 10 mg according to a double-blind protocol. Sixty minutes after treatment, auditory-evoked potentials were elicited by 40 ms 1 kHz tones 5, 10, 15, 20 and 25 dB[A] above a background of 70 dB[A]. Recording sessions consisted of eight alternating 2 min THREAT and SAFE blocks; unpredictable shocks (1.8 mA, 50 ms) were delivered to the subject's wrist in THREAT blocks (1-4 shocks per block). The amplitude of the N1/P2 potential increased monotonically as a function of stimulus intensity. The responses were significantly greater during THREAT blocks than during SAFE blocks (fear potentiation). Diazepam attenuated the responses in both the SAFE and THREAT conditions. Fear potentiation of the N1/P2 potential was significantly reduced by diazepam. Diazepam reduced subjective alertness and lowered critical flicker fusion frequency, a measure of arousal. The results suggest that fear potentiation of the N1/P2 potential is not simply a manifestation of the fear-potentiated startle response. The use of low-intensity stimuli may be advantageous in studies of fear potentiation in humans.