Measuring anxious responses to predictable and unpredictable threat in children and adolescents

Abstract

Research has highlighted the need for new methods to assess emotions in children on multiple levels to gain better insight into the complex processes of emotional development. The startle reflex is a unique translational tool that has been used to study physiological processes during fear and anxiety in rodents and in human participants. However, it has been challenging to implement developmentally appropriate startle experiments in children. This article describes a procedure that uses predictable and unpredictable aversive events to distinguish between phasic fear and sustained anxiety in children and adolescents. We investigated anxious responses, as measured with the startle reflex, in youths (N = 36, mean age = 12.63 years, range = 7–17) across three conditions: no aversive events (N), predictable aversive events (P), and unpredictable aversive events (U). Short-duration cues were presented several times in each condition. Aversive events were signaled by the cues in the P condition but were presented randomly in the U condition. Participants showed fear-potentiated startle to the threat cue in the P condition. Startle responses were also elevated between cues in the U condition compared with the N condition, suggesting that unpredictable aversive events can evoke a sustained state of anxiety in youths. This latter effect was influenced by sex, being greater in girls than in boys. These findings indicate the feasibility of this experimental induction of the startle reflex in response to predictable and unpredictable events in children and adolescents, enabling future research on interindividual differences in fear and anxiety and their development in youths.

Introduction

To understand the development of normal and pathological emotional processing, research on all components of emotional experiences in children and adolescents is crucial (Zeman, Klimes-Dougan, Cassano, & Adrian, 2007). Research on emotion faces at least two major methodological challenges: the induction of emotions and the measurement of emotion. Emotion induction in children can be problematic due to ethical considerations, especially if negative emotions are investigated. Regarding measurement, it has been argued that emotions should be assessed across several domains, including verbal report, physiological activity, and overt behavior. Currently, the majority of studies use only a single measurement instrument, and many studies assessing emotions still rely solely on self-reports or other-reports (e.g., parent reports and teacher reports) as the sole source of information on the emotional experiences of children (Zeman et al., 2007). This is problematic because these different response systems (i.e., verbal–cognitive, behavioral, and physiological) can be activated independent of each other (Lang, 1993). Furthermore, self-report measures are vulnerable to voluntary and involuntary distortions such as cognitive appraisal, social desirability, and recall bias (Dadds et al., 1998, Matt et al., 1992). Measuring physiological responses to emotional experiences can overcome some of these problems and add additional information that might not be consciously accessible. The goal of this study was to develop an experimental procedure to measure physiological responses in children and adolescents during two different emotional states: fear and anxiety.

Fear and anxiety are frequently used interchangeably, but although they are closely related concepts, several lines of research suggest that they are two functionally different defense mechanism mediated by distinct brain structures (Davis, Walker, Miles, & Grillon, 2010). Fear is elicited by an imminent threat and leads to a phasic fight-or-flight reaction, whereas anxiety is characterized by a sustained state of heightened vigilance and apprehension due to temporally uncertain danger. Research in rodents, healthy adults, and clinical populations shows that these differences are apparent across several levels of analysis. For instance, the structure of internalizing disorders has been described as consisting of a fear factor and an anxious misery factor (Cox et al., 2002, Krueger, 1999). The fear factor usually consists of phobias such as social phobia, specific phobias, and agoraphobia as well as panic disorder, whereas the anxious misery factor is composed of generalized anxiety disorder, posttraumatic stress disorder, dysphoria, and major depression. Parallel to the distinction between human fear and anxiety, these emotions can also be distinguished in rodent defensive behaviors and their alteration by anxiolytic drugs. In a series of very elegant experiments, Blanchard and colleagues distinguished between defensive behaviors evoked by a Fear/Defense Test Battery that involved the actual presence of a threat (e.g., a cat) and defensive behaviors provoked by an Anxiety/Defense Test Battery in which rodents were presented with a potential threat (e.g., cat odor) (Blanchard, Yudko, Rodgers, & Blanchard, 1993). These authors showed that defensive behaviors to a potential threat were systematically altered by anxiolytic drugs, but defensive behaviors to a clearly identifiable and imminent threat were not. Distinct anatomical structures have been implicated in the mediation of these responses (Davis et al., 2010; Walker, Toufexis, & Davis, 2003). Although the amygdala plays a crucial role in fear processing, the bed nucleus of the stria terminalis mediates responses to sustained states of anxiety, suggesting a functional differentiation between fear and anxiety. Research on this topic has also been conducted in humans. Specifically, clinical and psychopharmacological observations have provided support for an empirical distinction between fear and anxiety (Davis et al., 2010).

Prior studies have been conducted solely on adults, so it has been difficult to identify factors related to the emergence of this distinction. An extension of such work to youths would enhance our understanding of the normal and pathological development of these emotions. Therefore, the primary goal of the current study was to examine the feasibility of a psychophysiological experiment designed to distinguish fear and anxiety in adults for use in children and adolescents.

The startle reflex, a protective response to abrupt and intense stimuli (Landis & Hunt, 1939), is an attractive tool to examine fear and anxiety mechanisms for various reasons. Because startle is an automatic/reflexive response, it is not primarily influenced by intentional control and is resistant to demand effects and response biases that can interfere with verbal reports and voluntary motor responses (e.g., reaction time). Furthermore, the amplitude of the startle reflex can be modulated by emotional states. In contrast to many other psychophysiological measures such as skin conductance, the modulation of the startle reflex depends on the valence of a participant’s emotional state and is not a mere index of emotional arousal. Finally, similar experiments can be conducted in animals and in humans, making this methodology especially valuable for a translational approach (Davis et al., 2010).

The startle reflex is strongly potentiated (fear-potentiated startle) by a short-duration threat cue that predicts an imminent aversive event (e.g., shock). This effect is mediated by the central nucleus of the amygdala (Hitchcock & Davis, 1986). More recently, it has been shown that lesions of the amygdala fail to eliminate startle potentiation that is evoked in a more sustained manner (e.g., by administering shocks unpredictably). Rather, startle potentiation is suppressed by lesions of the bed nucleus of the stria terminalis (BNST). In fact, there is a double dissociation between the effect of lesions of the amygdala and the BNST on startle potentiation to short- and long-duration threats, with lesions of the former affecting response to short-duration threat and lesions of the latter affecting response to long-duration threat (Davis, 1998). Given that a short-duration threat cue is associated with a highly predictive danger that evokes a phasic aversive response and that a long-duration threat cue is associated with a sustained aversive state, it has been proposed that these two manipulations provide laboratory analogs of fear and anxiety, respectively (Davis et al., 2010).

To extend research on phasic fear and sustained anxiety to humans, Grillon and colleagues developed an experiment that assesses startle potentiation during alternating periods of predictable and unpredictable aversive events (Grillon, Baas, Lissek, Smith, & Milstein, 2004). A typical experiment consists of three conditions: neutral (N), where participants are informed that they will not receive any unpleasant events; predictable (P), where unpleasant events occur only during a specific cue; and unpredictable (U), where unpleasant events occur randomly. In a series of studies, Grillon et al., 2008, Grillon et al., 2009 showed that adult patients with panic disorder or posttraumatic stress disorder (PTSD) were selectively more sensitive to unpredictability than healthy controls free of any past or current psychiatric disorder. Patients showed stronger differences in startle potentiation between no-cue phases of the U and N conditions but showed similar potentiation as healthy controls to predictable threat. These results support theories that unpredictability of potential threat is a key element in the development of anxiety disorders (Mineka & Zinbarg, 2006).

Applying this experiment in youths would be a significant methodological advance because it enables us to study normal and pathological developmental trajectories of fear and anxiety that might be distinct from each other. The cross-species nature of the experiment and the increasing data available in animals will then allow us to draw inferences from the neurobiological mechanisms behind these trajectories. From a clinical perspective, a stronger sensitivity for unpredictability may serve as a vulnerability factor for the development of anxiety disorders in children and adolescents. Therefore, identifying these differences in youths provides a developmental approach to fear and anxiety so as to uncover possible pathological mechanisms, risk factors, and potential targets for prevention.

To date, no age-appropriate startle experiment has been developed to assess these fear- and anxiety-related processes in youths. Adapting this experiment for use with children and adolescents is especially challenging because the applied aversive stimulus must be sufficiently unpleasant to elicit enhanced startle potentiation due to unpredictability (Grillon et al., 2004). For example, we found that in adults, although startle is reliably potentiated by a threat cue predicting a shock or a blast of air to the neck, startle is potentiated in a sustained manner only during long periods of unpredictable shocks but not unpredictable blasts of air (Grillon et al., 2004). Because electric shocks could not be employed in children due to ethical considerations, we investigated whether less aversive stimuli that were developed in our laboratory and have been used in earlier studies in adults and youths (Grillon et al., 1999, Lau et al., 2008, Lissek et al., 2005) would be adequate to potentiate startle during unpredictability.

As a secondary aim, we explored potential sex differences in fear-potentiated and anxiety-potentiated startle. It is well established that normal anxiety as well as pathological anxiety is generally higher in girls compared with boys (Costa et al., 2001, Kessler et al., 2005, Mount et al., 2010, Van Oort et al., 2009). Sex differences in anxious behavior have been observed starting at a very young age. Mount and colleagues (2010), for example, reported that mothers observed more anxious behavior in 2½-year-old daughters than sons, and higher levels of trait anxiety among girls have been found to persist into adulthood (Costa et al., 2001). The higher prevalence of anxiety disorders in women and girls (Kessler et al., 2005, Van Oort et al., 2009) suggests that sex may be a key factor in the development of pathological fear and anxiety. Previous analog studies of humans and rodents using startle modulation have reported increased anxiety to uncertain threat but equivalent levels of fear to imminent threat in adult females compared with males (Grillon, 2008, Toufexis, 2007), consistent with the observation that the bed nucleus of the stria terminalis shows a strong sexual dimorphism (Allen & Gorski, 1990). Therefore, we hypothesized greater anxiety but not fear in girls compared with boys.

In summary, the main goal of the current study was to investigate the feasibility of the proposed experimental modulation of the startle reflex in response to predictable and unpredictable events in children and adolescents and to explore whether the sex differences found in adults can be replicated in children. A successful adaptation would enable future psychophysiological research on interindividual differences in fear and anxiety and their development in youths.