T-pattern Analysis Research Articles

Brain histamine depletion enhances the behavioural sequences complexity of mice tested in the open-field: Partial reversal effect of the dopamine D2/D3 antagonist sulpiride

Markers of histaminergic dysregulation were found in several neuropsychiatric disorders characterized by repetitive behaviours, thoughts and stereotypies. We analysed the effect of acute histamine depletion by means of i. c.v. injections of alpha-fluoromethylhistidine, a blocker of histidine decarboxylase, on the temporal organization of motor sequences of CD1 mice behaviour in the open-field test. An ethogram encompassing 9 behavioural components was employed. Durations and frequencies were only slightly affected by treatments. However, as revealed by multivariate t-pattern analysis, histamine depletion was associated with a striking increase in the number of behavioural patterns. We found 42 patterns of different composition occurring, on average, 520.90 ± 50.23 times per mouse in the histamine depleted (HD) group, whereas controls showed 12 different patterns occurring on average 223.30 ± 20.64 times. Exploratory and grooming behaviours clustered separately, and the increased pattern complexity involved exclusively exploratory patterns. To test the hypothesis of a histamine-dopamine interplay on behavioural pattern phenotype, non-sedative doses of the D2/D3 antagonist sulpiride (12.5-25-50 mg/kg) were additionally administered to different groups of HD mice. Sulpiride counterbalanced the enhancement of exploratory patterns of different composition, but it did not affect the mean number of patterns at none of the doses used. Our results provide new insights on the role of histamine on repetitive behavioural sequences of freely moving mice. Histamine deficiency is correlated with a general enhancement of pattern complexity. This study supports a putative involvement of histamine in the pathophysiology of tics and related disorders.

The effects of morphine on the temporal structure of Wistar rat behavioral response to pain in hot-plate.

The largest amount of researches on the hot-plate test was carried out using quantitative assessments. However, the evaluation of the relationships among the different elements that compose the behavioral response to pain requires different approaches. Although previous studies have provided clear information on the behavioral structure of the response, no data are available on its temporal structure. The objective of this study was to investigate the temporal structure of the behavioral response to pain in Wistar rat tested in hot-plate and how this structure was influenced by morphine-induced analgesia. The behavior of four groups of subjects tested in hot-plate, one administered saline and three with different doses (3, 6, 12mg/kg) of morphine IP, was analyzed by means of quantitative and t-pattern analyses. The latter is a multivariate technique able to detect the existence of statistically significant temporal relationships among the behavioral events in time. A clear-cut influence of morphine on quantitative parameters of the response to the noxious stimulation was observed. T-pattern analysis evidenced profound structural changes of behavior. Twenty-four different t-patterns were identified following saline, whereas a dose-dependent reduction was observed following morphine. Such a reduction was accompanied by a decrease of the total amount of t-patterns detected. Morphine, by reducing the effects of the noxious stimulation, orients animal behavior prevalently toward exploratory t-patterns. In addition, it is suggested that the temporal structure of the response is very quickly organized and adapted to environmental noxious cues.

Design and evaluation of a geovisual analytics system for uncovering patterns in spatio-temporal event data

ABSTRACTIt remains difficult to develop a clear understanding of geo-located events and their relationships to one another, particularly when it comes to identifying patterns of events in less-structured textual sources, such as news feeds and social media streams. Here we present a geovisualization tool that can leverage computational methods, such as T-pattern analysis, for extracting patterns of interest from event data streams. Our system, STempo, includes coordinated-view geovisualization components designed to support visual exploration and analysis of event data, and patterns extracted from those data, in terms of time, geography, and content. Through a user evaluation, we explore the usability and utility of STempo for understanding patterns of recent political, social, economic, and military events in Syria.

Behavioural assessment of pain in commercial turkeys (Meleagris gallopavo) with foot pad dermatitis

Two experiments were conducted to investigate the differences in susceptibility to foot pad dermatitis (FPD) of two medium-heavy lines of turkeys, and whether FPD is painful, by detailed analysis of behaviour in birds with and without analgesic treatment (betamethasone).Turkeys housed on dry litter in the first experiment generally had more frequent bouts of different behaviours that were of shorter duration than birds on wet litter. T-patterns (behavioural sequences) were more frequent, varied and complex on dry than on wet litter. Betamethasone-injected birds of line B, but not breed A, had shorter resting and longer standing durations on wet litter than saline-injected birds.In the second experiment, turkeys on wet litter given saline stood less and rested more than all other treatment groups, suggesting that they experienced pain that was alleviated in birds receiving betamethasone. Turkeys on dry litter had more frequent, varied and complex patterns of behaviour than turkeys on wet litter and birds kept on intermediate litter wetness. Betamethasone provision increased pattern variety regardless of litter treatment.Turkeys with low FPD scores transferred to wet litter and given saline injections had a longer total duration of resting and shorter duration of standing compared to betamethasone-treated birds. Low FPD birds transferred to wet litter had a similar number of patterns and total pattern occurrence as high FPD birds transferred to dry litter. Betamethasone increased pattern variety and frequency compared to saline injections whereas overall pattern complexity was similar.It was concluded that wet litter affects the behaviour of turkey poults independently of FPD and that betamethasone may also change the behaviour of turkeys. There was some evidence from analgesic treatment and T-pattern analyses that FPD was painful. However, there was no evidence of differences in susceptibility to FPD of the two commercial hybrids.

Temporal patterns of rat behaviour in the central platform of the elevated plus maze. Comparative analysis between male subjects of strains with different basal levels of emotionality

We have analyzed the temporal patterns of behaviour of male rats of the Wistar and DA/Han strains on the central platform of the elevated plus maze. The ethogram encompassed 10 behavioural elements. Durations, frequencies and latencies showed quantitative differences as to walking and sniffing activities. Wistar rats displayed significantly lower latency and significantly higher durations and frequencies of walking activities. DA/Han rats showed a significant increase of sniffing duration. In addition, DA/Han rats showed a significantly higher amount of time spent in the central platform. Multivariate T-pattern analysis revealed differences in the temporal organization of behaviour of the two rat strains. DA/Han rats showed (a) higher behavioural complexity and variability and (b) a significantly higher mean number of T-patterns than Wistar rats.Taken together, T-pattern analysis of behaviour in the centre of the elevated plus maze can noticeably improve the detection of subtle features of anxiety related behaviour. We suggest that T-pattern analysis could be used as sensitive tool to test the action of anxiolytic and anxiogenic manipulations.

Quantifying the offensive sequences that result in goals in elite futsal matches

The aim of this study was to quantify the type of offensive sequences that result in goals in elite futsal. Thirty competitive games in the Spanish Primera Division de Sala were analysed using computerised notation analysis for patterns of play that resulted in goals. More goals were scored in positional attack (42%) and from set pieces (27%) compared to other activities. The number of defence to offense “transitions” (n = 45) and the start of offensive plays due to the rules of the game (n = 45) were the most common type of sequences that resulted in goals compared to other patterns of play. The central offensive zonal areas were the most common for shots on goal, with 73% of all goals scored from these areas of the pitch compared to defensive and wide zones. The foot was the main part of the body involved in scoring (n = 114). T-pattern analysis of offensive sequences revealed regular patterns of play, which are common in goal scoring opportunities in futsal and are typical movement patterns in this sport. The data demonstrate common offensive sequences and movement patterns related to goals in elite futsal and this could provide important information for the development of physical and technical training drills that replicate important game situations.

Acute nicotine induces anxiety and disrupts temporal pattern organization of rat exploratory behavior in hole-board: a potential role for the lateral habenula.

Nicotine is one of the most addictive drugs of abuse. Tobacco smoking is a major cause of many health problems, and is the first preventable cause of death worldwide. Several findings show that nicotine exerts significant aversive as well as the well-known rewarding motivational effects. Less certain is the anatomical substrate that mediates or enables nicotine aversion. Here, we show that acute nicotine induces anxiogenic-like effects in rats at the doses investigated (0.1, 0.5, and 1.0 mg/kg, i.p.), as measured by the hole-board apparatus and manifested in behaviors such as decreased rearing and head-dipping and increased grooming. No changes in locomotor behavior were observed at any of the nicotine doses given. T-pattern analysis of the behavioral outcomes revealed a drastic reduction and disruption of complex behavioral patterns induced by all three nicotine doses, with the maximum effect for 1 mg/kg. Lesion of the lateral habenula (LHb) induced hyperlocomotion and, strikingly, reversed the nicotine-induced anxiety obtained at 1 mg/kg to an anxiolytic-like effect, as shown by T-pattern analysis. We suggest that the LHb is critically involved in emotional behavior states and in nicotine-induced anxiety, most likely through modulation of monoaminergic nuclei.

A method for discovery and analysis of temporal patterns in complex event data

Pattern analysis techniques currently common within geography tend to focus either on characterizing patterns of spatial and/or temporal recurrence of a single event type (e.g., incidence of flu cases) or on comparing sequences of a limited number of event types where relationships between events are already represented in the data (e.g., movement patterns). The availability of large amounts of multivariate spatiotemporal data, however, requires new methods for pattern analysis. Here, we present a technique for finding associations among many different event types where the associations among these varying event types are not explicitly represented in the data or known in advance. This pattern discovery method, known as T-pattern analysis, was first developed within the field of psychology for the purpose of finding patterns in personal interactions. We have adapted and extended the T-pattern method to take the unique characteristics of geographic data into account and implemented it within a geovisualization toolkit for an integrated computational-geovisual environment we call STempo. To demonstrate how T-pattern analysis can be employed in geographic research for discovering patterns in complex spatiotemporal data, we describe a case study featuring events from news reports about Yemen during the Arab Spring of 2011–2012. Using supplementary data from the Global Database of Events, Language, and Tone, we briefly summarize and reference a separate validation study, then evaluate the scalability of the T-pattern approach. We conclude with ideas for further extensions of the T-pattern technique to increase its utility for spatiotemporal analysis.

T-pattern analysis in soccer games: relationship between time and attack actions

Sports performance consists of a multiple series of strategies that tend to follow one another. Performance analysis in team sports is usually focused on primary (fundamental skill execution), secondary (scoring) and tertiary (match result) outcomes. While there is general agreement over measuring secondary and tertiary outcomes, literature does not show a unanimous agreement over a unique measure of the primary level of performance. The aim of this study was to investigate primary performance outcomes through an analysis of temporal patterns. In particular, we were interested in verifying if changes in tertiary performance outcomes may be related to changes in primary ones. We selected three soccer matches played by a top club during the Serie A league over the 2012-2013 season in which there was a change in match result between *rst and second half (tertiary level of performance). The methodological approach was based on observational design, supported by digital recordings and computer analysis. Data were analyzed with theme 6 beta software, which detects the temporal and sequential structure of datasets, revealing repeated patterns that may regularly or irregularly occur within a period of observation (Tpatterns). Striking di+erences were found comparing first and second half temporal patterns, especially when the *nal match outcome showed an improvement of the first half ’s one. Our results suggest that 'eme software and T-pattern enhance research opportunities by identifying a useful tool to study the link between primary and tertiary level of performance, making this an e+ective research and support instrument for sports analysis.

T-pattern analysis for the study of temporal structure of animal and human behavior: a comprehensive review.

A basic tenet in the realm of modern behavioral sciences is that behavior consists of patterns in time. For this reason, investigations of behavior deal with sequences that are not easily perceivable by the unaided observer. This problem calls for improved means of detection, data handling and analysis. This review focuses on the analysis of the temporal structure of behavior carried out by means of a multivariate approach known as T-pattern analysis. Using this technique, recurring sequences of behavioral events, usually hard to detect, can be unveiled and carefully described. T-pattern analysis has been successfully applied in the study of various aspects of human or animal behavior such as behavioral modifications in neuro-psychiatric diseases, route-tracing stereotypy in mice, interaction between human subjects and animal or artificial agents, hormonal-behavioral interactions, patterns of behavior associated with emesis and, in our laboratories, exploration and anxiety-related behaviors in rodents. After describing the theory and concepts of T-pattern analysis, this review will focus on the application of the analysis to the study of the temporal characteristics of behavior in different species from rodents to human beings. This work could represent a useful background for researchers who intend to employ such a refined multivariate approach to the study of behavior.

Complex spike patterns in olfactory bulb neuronal networks.

T-pattern analysis is a procedure developed for detecting non-randomly recurring hierarchical and multiordinal real-time sequential patterns (T-patterns). We have inquired whether such patterns of action potentials (spikes) can be extracted from extracellular activity sampled simultaneously from many neurons across the mitral cell layer of the olfactory bulb (OB). Spikes were sampled from urethane-anaesthetized rats over a 6h recording session, or a period lasting as long as permitted by the physiological condition of the animal. Breathing was recorded to mark peak inhalation and exhalation. Complex T-patterns of up to ∼20 elements were identified with functional connections often spanning the full extent of the array. A considerable proportion of these sequences incorporated breathing. In contrast to sequence detection by synfire, the incidence of sequences detected in our real data is very much greater than in the same data when randomized either by shuffling, or an alternative procedure preserving the interval structure of each spike train, and so more conservative. Further, when recordings were terminated before completion of the full recording session, the relative pattern detection in real and randomized data was a strong indicator of physiological condition-in recordings leading up to the preparation becoming physiologically unstable, the number of patterns detected in real data approached that in the randomized data. We conclude that such sequences are an important physiological property of the neural system studied, and suggest that they may form a basis for encoding sensory information.

Multivariate temporal pattern analysis applied to the study of rat behavior in the elevated plus maze: methodological and conceptual highlights.

Aim of this article is to illustrate the application of a multivariate approach known as t-pattern analysis in the study of rat behavior in elevated plus maze. By means of this multivariate approach, significant relationships among behavioral events in the course of time can be described. Both quantitative and t-pattern analyses were utilized to analyze data obtained from fifteen male Wistar rats following a trial 1-trial 2 protocol. In trial 2, in comparison with the initial exposure, mean occurrences of behavioral elements performed in protected zones of the maze showed a significant increase counterbalanced by a significant decrease of mean occurrences of behavioral elements in unprotected zones. Multivariate t-pattern analysis, in trial 1, revealed the presence of 134 t-patterns of different composition. In trial 2, the temporal structure of behavior become more simple, being present only 32 different t-patterns. Behavioral strings and stripes (i.e. graphical representation of each t-pattern onset) of all t-patterns were presented both for trial 1 and trial 2 as well. Finally, percent distributions in the three zones of the maze show a clear-cut increase of t-patterns in closed arm and a significant reduction in the remaining zones. Results show that previous experience deeply modifies the temporal structure of rat behavior in the elevated plus maze. In addition, this article, by highlighting several conceptual, methodological and illustrative aspects on the utilization of t-pattern analysis, could represent a useful background to employ such a refined approach in the study of rat behavior in elevated plus maze.

Patterns of co-occurring non-verbal behaviour and self-directed speech; a comparison of three methodological approaches

Self-directed speech – the audible or partially whispered self-talk that children engage in during their daily activities, was proposed by Vygotsky to have a mediating role in the emerging self-regulatory behaviour of young children. Studies with correlational findings tend to lend support to this hypothesis but fail to delineate the real-time temporal interactions between self-directed speech and self-regulatory behaviour. The authors propose the use of lag sequential analysis and t-pattern analysis as useful and complementary methods for detecting significantly recurring patterns of co-occurrence of self-directed speech and non-verbal behaviour (that is either self-regulatory or shows a failure of self-regulation). Furthermore, it is argued that the analysis of these co-occurrences is required to establish the functions of self-directed speech, and to determine in what ways these might be self-regulatory. Illustrative analyses are presented of data from a study comparing the patterns of self-directed speech use during a planning task in typically developing children and matched peers with Specific Language Impairment. The results obtained from t-pattern analysis reveal qualitative differences between these two groups of children, in their use of self-directed speech, which were not detected by the other two methods. Implications of examining recurring temporal patterns in behaviour for research investigating aspects of development, particularly self-regulation, are discussed.

The Routineness of Routines: Measuring Rhythms of Media Interaction

The routines of information work are commonplace yet difficult to characterize. Although cognitive models have successfully characterized routine tasks within which there is little variation, a large body of ethnomethodological research has identified the inherent nonroutineness of routines in information work. We argue that work does not fall into discrete classes of routine versus nonroutine; rather, task performance lies on a continuum of routineness, and routineness metrics are important to the understanding of workplace multitasking. In a study of 10 information workers shadowed for 3 whole working days each, we utilize the construct of working sphere to model projects/tasks as a network of humans and artifacts. Employing a statistical technique called T-pattern analysis, we derive measures of routineness from these real-world data. In terms of routineness, we show that information workers experience archetypes of working spheres. The results indicate that T-patterns of interactions with information and computational media are important indicators of facets of routineness and that these measures are correlated with workers' affective states. Our results are some of the first to demonstrate how regular temporal patterns of media interaction in tasks are related to stress. These results suggest that designs of systems to facilitate so-called routine work should consider the degrees to which a person's working spheres fall along varying facets of routineness.

Significant divergences between the temporal structure of the behavior in Wistar and in the spontaneously more anxious DA/Han strain of rats tested in elevated plus maze

The aim of present research is to study the temporal structure of the behavior in two strains of rats with different basal level of emotionality. To this purpose, the temporal profile of the behavior in Wistar rat and in the spontaneously more anxious DA/Han strain was analyzed in the Elevated Plus Maze.Both quantitative and multivariate t-pattern analyses were carried out. In comparison with Wistar, DA/Han subjects showed a significant reduction of the permanence in open arm and a significant increase of the time spent in the central platform of the maze. Mean frequencies of each behavioral element showed significant modifications both in open and in closed arm. Multivariate t-pattern analyses demonstrated a very different temporal profile of behavior in the two strains: Wistar rats presented 197 t-patterns of different composition, whereas DA/Han rats only 26; as to the mean number of t-patterns, Wistar presented a value of 698.90, whereas DA/Han only 92.80. Similar clear-cut differences were detected for the mean number of t-patterns in the two arms and for the time course of such t-patterns.Present study has evidenced that the temporal organization of the bahavior in Elevated Plus Maze is differently structured in two strains of rats with different basal level of emotionality.

Temporal structure of the rat's behavior in elevated plus maze test

Aim of the research was to evaluate, by means of quantitative and multivariate temporal pattern analyses, the behavior of Wistar rat in elevated plus maze (EPM) test. On the basis of an ethogram encompassing 24 behavioral elements, quantitative results showed that 130.14±8.01 behavioral elements occurred in central platform and in closed arms (protected zones), whereas 88.62±6.04 occurred in open arms (unprotected zones). Percent distribution was characterized by a prevalence of sniffing, walking and vertical exploration. Analysis of minute-by-minute duration evidenced a decrease for time spent in open arms and central platform and an increase for time spent in closed arms. As to multivariate t-pattern analysis, 126 different temporal patterns were detected. Behavioral stripes, summarizing distribution of such t-patterns along time, showed that several t-patterns were not homogeneously distributed along the test observational period: t-patterns encompassing behavioral events occurring prevalently in central platform-open arms were observed during the first minutes, whereas t-patterns structured on the basis of events occurring mainly in central platform-closed arms were detected during the last minutes. Therefore, during the observation in elevated plus maze, rat's behavior undergoes significant rearrangements of its temporal features. Present research demonstrates, for the first time, the existence of complex and significantly timed behavioral sequences in the activity of Wistar rats tested in elevated plus maze. Application of t-pattern analysis can provide useful tools to characterize the behavioral dynamics of anxiety-related rodent behavior and differentiate the effect of various anxioselective substances.

Behavioral Patterns Associated with Chemotherapy-Induced Emesis: A Potential Signature for Nausea in Musk Shrews

Nausea and vomiting are common symptoms in patients with many diseases, including cancer and its treatments. Although the neurological basis of vomiting is reasonably well known, an understanding of the physiology of nausea is lacking. The primary barrier to mechanistic research on the nausea system is the lack of an animal model. Indeed investigating the effects of anti-nausea drugs in pre-clinical models is difficult because the primary readout is often emesis. It is known that animals show a behavioral profile of sickness, associated with reduced feeding and movement, and possibly these general measures are signs of nausea. Studies attempting to relate the occurrence of additional behaviors to emesis have produced mixed results. Here we applied a statistical method, temporal pattern (t-pattern) analysis, to determine patterns of behavior associated with emesis. Musk shrews were injected with the chemotherapy agent cisplatin (a gold standard in emesis research) to induce acute (<24 h) and delayed (>24 h) emesis. Emesis and other behaviors were coded and tracked from video files. T-pattern analysis revealed hundreds of non-random patterns of behavior associated with emesis, including sniffing, changes in body contraction, and locomotion. There was little evidence that locomotion was inhibited by the occurrence of emesis. Eating, drinking, and other larger body movements including rearing, grooming, and body rotation, were significantly less common in emesis-related behavioral patterns in real versus randomized data. These results lend preliminary evidence for the expression of emesis-related behavioral patterns, including reduced ingestive behavior, grooming, and exploratory behaviors. In summary, this statistical approach to behavioral analysis in a pre-clinical emesis research model could be used to assess the more global effects and limitations of drugs used to control nausea and its potential correlates, including reduced feeding and activity levels.

T-pattern analysis of diazepam-induced modifications on the temporal organization of rat behavioral response to anxiety in hole board

By means of t-pattern analysis, it has been observed that the different events, characterizing rat behavior in hole board (HB), present close interrelationships which occur sequentially and with significant constraints on the interval lengths separating them. The aim of present research was to study, by means of descriptive and multivariate t-pattern analyses, the effects of the reference anxiolytic drug diazepam (DZP) on temporal structure of a rat's anxiety-related behavior in HB. Fifty-six male Wistar rats were tested for 10 min in HB. Video files, collected for each animal, were coded by means of a software coder, and event log files, generated for each subject, were analyzed by means of a specific software for temporal pattern analysis (t-pattern analysis). Significant diazepam-induced modifications were observed for durations of walking, climbing, edge-sniff, and face grooming. Dose-dependent decreases of t-patterns' total amount, of their mean occurrences and of their mean length for each group were detected. Also, t-patterns' mean occurrences, in terms of different composition, were reduced. Percent distributions showed a significant increase of t-patterns including walking for all administered groups, and significant reductions of t-patterns including climbing, immobile sniffing, and edge-sniff. Front-paw licking and face grooming were reduced at the higher DZP dose. Present study demonstrates, for the first time, that the temporal structure of Wistar rats' behavioral response to anxiety in HB changes following pharmacological manipulation of anxiety condition. Moreover, t-pattern analysis is suggested to represent a useful tool to evaluate and compare different classes of anti-anxiety molecules.

Behavioral pattern analysis and dopamine release in quinpirole-induced repetitive behavior in rats

Obsessive-compulsive disorder (OCD) is a chronic and disabling psychiatric disease with a lifetime prevalence of 2-3%. People with OCD suffer from intrusive, unwanted and recurrent thoughts (obsessions) and/or repetitive ritualistic behaviors (compulsions). The aim of this study is to quantify the dimensions of ritualistic 'compulsive-like' behavior in quinpirole-induced behavior in rats by using T-pattern behavioral analysis. In addition, we investigated whether the behavioral effects elicited by quinpirole sensitization remained after 2 weeks of cessation of treatment. Finally, to study the neurobiological underpinnings of this 'compulsive-like' behavior, we investigated the effect of quinpirole treatment on the extracellular dopamine levels in the nucleus accumbens. Once established, 'compulsive-like' behavior is dependent upon quinpirole administration, as this behavior rapidly normalized after cessation of treatment. After a single dose of quinpirole the dopamine level decreased more in saline pre-treated animals as compared with animals given quinpirole treatment continuously. Furthermore, T-pattern analysis revealed that quinpirole-induced behavior consists, unlike OCD rituals, of a smaller behavioral repertoire. As seen in patients with OCD, quinpirole-treated animals performed these behaviors with a high rate of repetition. These findings suggest that quinpirole-induced behavior mimics only part of the compulsive behavior as shown in OCD patients.

Temporal patterns analysis of rat behavior in hole-board

The aim of present research was to analyze the temporal structure of rodent's anxiety-related behavior in hole-board apparatus (HB). Fifteen male Wistar rats were tested for 10min. Video files, collected for each subject, were coded by means of a software coder and event log files generated for each subject. To assess temporal relationships among behavioral events, log files were processed by means of a t-pattern analysis. 14 two-element t-patterns, four t-patterns encompassing 3 events and 2 t-patterns encompassing 4 and 5 events respectively were revealed. It was demonstrated that rat behavior in HB was mainly structured on the basis of the temporal patterning among exploratory events; these ones were the most structured t-patterns detected and appeared mainly during the first 5min of exploration, while grooming t-patterns were present prevalently after the fifth minute. Specific t-pattern parameters, such as overall occurrences and mean duration of each given t-pattern in each subject, were also studied. Present research: (a) reports for the first time that some behavioral events occur sequentially and with significant constraints on the interval lengths separating them; (b) presents the temporal flows of some behavioral elements through multimodal behavioral vectors; (c) could also be used to improve HB test reliability and its ability to detect even small induced behavioral changes.