SECTION 5 measurement of pain 408 Method of limits (MLI): this method entails the administration of stimuli of varying intens ity until the subject notes a certain sensa- tion and indicates this verbally or behavioura lly. Usually, stimuli are delivered in ascending series, the subject \200\231s response sto ps fur- ther increase in intensity. Sensory detection and pain thresholds are defi ned as t he stimulus intensity required to evoke a given sen- sation (e.g. touch, warmth) or pain, r espectively. Whereas sensory detection thresholds rely on discriminating the presence versu s absence of a specifi c sensation, pain thresholds are not only infl u- enced by perceptua l sensitivity, but also by the given verbal label and the subject \200\231s pain labeling b ehavior and the ensuing response bias (Gracely, 2006 ). In order to reduce expectancy or ha bituation eff ects, it is oft en recommended to use both ascending and descend- ing series. MLI is reaction-time dependent (Yarnitsky and Ochoa, 1990 ). Most likely, this accounts fo r thermal thresholds being about 1.5 C higher when determined by MLI as compared to reac tion- time independent methods (e.g. Defrin et al., 2006 ; children: Meier et al., 2001 ). Despite this limitation, MLI is frequently used because instructions are simple and it is l ess time-consuming than other procedures. Th ere are reaction time-independent MLI variants . Th e method of levels entails the administration of stimuli with pre- defi ned intensity and the subject \200\231s response is recorded. Depending on the subject \200\231s positive or negative response, stimulus intensity is systematically either decreased (following a p ositive response) or increased (following a negative response) until it converges onto the threshold (Meier et al., 2001 ; Yarnitsky et al., 1990 ). Stimulus intensity can either be changed in absolute steps or in steps of modality-specifi c, just noticeable, diff erences (Yarnitsky, 1997 ). Reaction-time independent measures are more reliable than reac- tion-ti me dependent measures, but require more trials (Yarnitsky, 1997 ). is helpful for detecting sensory losses and gains such as altered pain sensitivity (hyperal gesia, allodynia, hyperpathia). Including suprathreshold stimuli is important because hyper sensitivity can be observed if thresholds are reduced, elevated, or unchanged. \200\230Dyn amic \200\231 measures require activation of the pain system such that possible changes in specifi c mechanisms of pain processing become evident. For example, sensitization to nocic eptive input is a key factor in the development of chronic pain. Such sensitization occurs in the peripheral and the central nervous system (Woolf, 2011 ). In humans, temporal summat ion (i.e. the increase in per- ceived pain intensity upon prolonged or repetitive painful s timula- tion), is considered to represent a perceptual correlate primarily of central sensi tization (e.g. Granot et al., 2006 ). Accordingly, some QST protocols include measures of t emporal summation. Th e test- ing of descending pain inhibitory mechanisms using procedures of painful counter-irritation is another example of a dynamic QST measure. Comprehensive QST protocols usually include the testing of diff erent sensory modalities su ch as thermal, touch, pressure, and vibratory stimuli. For obvious reasons of impracticabil ity and problems of standardization, chemical stimulation is not included. Testing diff ere nt sensory modalities allows the assessment of the integrity of diff erent types of recepto rs, peripheral nerve fi bres and information integration within the central nervous system. Mechanical stimuli such as light touch, pressure, or punctate stimuli activate A \237 , A , and C fi bres (see Table 40.1 ; Walk et al., 2009 ). Th e processing of warm and heat stimuli is dependent on C-fi bre activa- tion, cold sensations and noxious cold involve A -fi bre and C-fi bre activity, respectively. Th e processing of painful stimuli is media ted both by modality-specifi c and convergent pathways (Craig, 2003 ). Hence, pain sensatio n is the result of central integration of sensory non-nociceptive information and nocicepto r input (Defrin et al., 2002 ). Accordingly, correlations between (supra-)threshold meas- u res across modalities are typically low (e.g. Bhalang et al., 2005 ). Indeed, factor analyz es of diff erent pain modality measures (e.g. heat, ischaemic, pressure, electrical) reveal ed separate factors each representing a sensory modality (Hastie et al., 2005 ; Neziri et a l., 2011a ). In addition, measures of temporal summation loaded on a separate factor, suppo rting the distinction between static and dynamic measures (Hastie et al., 2005 ). Method of constant stimuli: similar to MLI, the subject is exposed to a set of stimuli with predefi ned intensity above and below the threshold of interest. However, these stimuli ar e presented ran- domly and the subject is asked to indicate whether or not the stimulus evo ked a certain sensation. Th e threshold is defi ned as the intensity that the subject repor ts as the sensation of interest in 50% of the trials. Method of adjustment: unlike MLI, this method requires the sub- ject to directly adjust sti mulus intensity until a certain sensation occurs. Th is procedure has to be repeated many t imes. Aside from the long duration, it is disadvantageous in that some of the stimula- tion devices for certain pain modalities do not allow the subject to control stimulus intensity . Sensory testing site When performing QST, the choice of testing sites is important. Th e ob tained values vary considerably between diff erent body regions (e.g. Defrin et al., 2006 ; children: Hogeweg et al., 1996 ; Meier et al., 2001 ; Meh and Denislic, 1994 ). In one of the most comprehensive studies in healthy adults, QST was performed bilaterally on the face , and at the hand and foot (Rolke et al., 2006b ). Similarly, in children, while no left \200\223right diff erences were observed, sensitivity was highest in the face and lowest in the foot and, with interme- diate values in the hand (Blankenburg et al., 2010 ). Presumab ly, this region- specifi c sensitivity is accounted for by diff erences in innervation and/ or thickness of the epidermis. However, these diff erences, at least for thresholds obtaine d by MLI, may also be a result of reaction-time-dependent threshold determination. If the m ethod of levels rather than MLI was used, diff erences in sensitiv- ity between body region s such as foot versus upper body were no longer observed, at least for thermal stimuli (Def rin et al., 2006 ). Although, in healthy subjects, thermal pain sensitivity does not In most cases, QST protocols exclusively rely on MLI for deter- mining sensory detection an d pain thresholds; mostly for practi- cal reasons, such as keeping the duration of the test ing within reasonable limits. Length of testing is an issue because the subject should be a lert and awake and motivated to cooperate with the experimenter. QST measures Sensory modalities Quantitative sensory testing does not refer to defi ned set of meas- ures. QST measures can broadly be categorized as static or dynamic (Arendt-Nielse n and Yarnitsky, 2009 ). Th reshold measures (e.g. detection, pain) and pain magnitude rati ngs for a given stimulus are typically considered as \200\230static \200\231 and are used for assessing the basal state of the somatosensory system. Th reshold determination CHAPTER 40 sensory processing and neurophysiological evaluation in children 409 Table 40.1 Overview of the QST measures included in the German Research Network on Neuropat hic Pain (DFNS) QST protocol (Rolke et al., 2006 ) as used in a large sample of healthy chi ldren and adolescents for determining normative reference values (Blankenburg et al., 2010 ) Static measures QST parameter Peripheral nerve \201 bre Testing procedure Testing device Th ermal Cold Detection threshold (CDT) Pain threshold (CPT) Detection threshold (WDT) Pain threshold (HP T) Th ermal sensory limen (TSL) Paradoxical heat sensations (during TSL) A (cid:303) C, A (cid:303) C C, A (cid:303) C, A (cid:303) MLI (descending) MLI (descending) MLI (ascending) MLI (ascending) Alternating warm/cold sti muli Verbal report Computer-controlled contact heat probe (e.g. TSA II, MEDOC, Israel, or MSA, SOMEDIC, Sweden ) Warm/heat Cold/warm Mechanical Cutaneous punctate blunt sharp A \237 A (cid:303) , C A (cid:303) , C A \237 Detection threshold (MDT) Pain threshold (MPT) Pain threshold (PPT) Detection threshold (VD T) MLI (ascending + descending) MLI (ascending + descending) MLI (ascending) MLI (descending) von Frey \201 laments Calibrated pinpricks (N=7; 8 \200\223512 mN) Pressure algometer Tun ing fork Deep pressure Vibration Dynamic and other measures Mechanical Sharp Numerical pain intensity ratings for all 7 pinpricks (5 repetitions each in pseudo-randomiz ed order) Mean pain rating of 5 series of repetitive stimuli relative to mean pain rating o f 5 single stimuli Mean numerical pain rating for the 3 touch intensities Pain sensitivity A (cid:303) , C Calibrated pinpricks (N = 7 with intensities 8 \200\223512 mN) Wind-up ratio (temporal summation) A (cid:303) , C A \237 3 levels of force: cotton wisp, cotton wool tip, brush Dynamic light touch Dynamic mechanical allodynia MLI = method of limits. seem to vary between diff erent parts of the hand (i.e. dorsum, fi n- ger, thenar eminence) , its interindividual variability is lowest at the thenar eminence, thus facilitating use o f this site for normative ref- erence data (Hagander et al., 2000 ). data are available for both healthy adults (Rolke et al., 2006b ; for a similar protocol se e Neziri et al., 2011b ) and children (Blankenburg et al., 2010 ). Th ese data for children compare fairly well with previ- ous reports for selected measures in healthy children usin g similar instructions and devices (e.g. Hilz et al., 1996 , 1998a , 1998b ; Meier et al., 2001 ). However, values can diff er when compared to studies using at least slightly diff e rent procedures and smaller samples with a greater age range (e.g. Th ibault et al., 1994 ; Walker et al., 2009 ). By comparing the patient \200\231s profi le of sensory losses and g ains to such normative data, absolute references can be made (Magerl et al., 2010 ). Altern atively, relative references are possible. Neuropathic pain oft en aff ects only one side o f the body. In light of the lack of left \200\223right diff erences in healthy individuals , the unaff ected side can then be used as a within-subject (relative) reference and allows characterization of the pattern of disturbed sensory functioning on the aff ected side (e. g. Kemler et al., 2000 ). Comprehensive standardized QST protocol example QST in research and in clinical practice re quires standardized pro- cedures that promise suffi cient reliability, allow comparisons ac ross studies, and reference to norms (e.g. Walk et al., 2009 ). Most recently, the German R esearch Network on Neuropathic Pain (DFNS) com- piled a battery of tests for sensory loss ( small and large fi bres) and sensory gain (hyperalgesia, allodynia, hyperpathia) (Rolke et al., 2006b ). In two sensory modalities (mechanical, thermal), and bilaterally at three mea surement sites (face, hand, foot), both static (detection and pain thresholds) and dynamic (e.g. temporal sum- mation) QST parameters are assessed in a fi xed order within one sessio n with detection thresholds determined prior to pain thresh- olds and suprathreshold measur es. Table 40.1 gives a more detailed overview of the obtained QST parameters. Th e DFNS QST battery is unique as it is the only QST battery for which data in healthy adults, diff ere nt pain populations and children (Blankenburg et al., 2010 , 2011) have been obtained. Psychometric considerations: reliability of QST Aside from the availability of normative da ta, adequate reliability is another prerequisite for validity and clinical usefulness of QS T. In their review of the literature, Siao and Cros ( 2003 ) concluded that reproducibility of QST fi ndings varied greatly across studies depending on study methodology, sample size , type of study sam- ple, number of examiners, and retest interval. More recently, in healt hy adults and patients, retest reliability both for comprehensive QST batteries or thermal measures was reported as good, at least Psychometric considerations: normative QST data Th e clinical usefulness of QST depends on relating individual QST fi ndings to reference data. For the DFNS QST battery, normative SECTION 5 measurement of pain 410 for retest intervals ranging between 1 to 2 days and up to 3 weeks (e.g. Agostinho et al., 2009 ; Geber et al., 2011 ; Heldestad et al., 2010 ; Moloney et al., 2011 ; Wylde et al., 2 011 ). In children, retest reliability has not been systematically investigated in suffi ci ently large healthy and patient samples. Determination of pressure pain thresholds has exce llent retest reliability in children with orthopae- dic disorders, at least when measured o n the same day (Nikolajsen et al., 2011 ). Similarly, retest reliability for sensory detect ion thresh- olds is good (Hilz et al., 1996 ; Th ibault et al., 1994 ). performed fi rst (Grone et al., 2012 ). For the thermal modality, no order eff ects were ob served. Initial assessment of pain threshold may also exert a confounding infl uence on the determination of detection thresholds, at least for thermal stimuli (Heldestad et al., 201 0 ). Especially for thermal testing using contact heat, starting temperature (e.g. Hilz et al., 1995 ), rate of the temperature increase (e.g. Yarnitsky et al., 1990 ) and size of sk in area (e.g. Khalili et al., 2001 ) infl uence the obtained values due to spatial and temp oral summation (cf. Jorum and Arendt-Nielsen, 2003 ). Furthermore, room and skin temperatur e can have an eff ect (Hilz et al., 1995 ; Strigo et al., 2000 ). In most studies, retest reliability has been determined by com- puting correlations, the co effi cient of variation, or by comparing the means for test \200\223retest measurements. Ho wever, these statisti- cal procedures only determine the strength of association, but not a greement (Bland and Altman, 1986 ). In addition, these tests are based on groups and cannot be applied when deciding on the clinical signifi cance of fi ndings. Derived from the Blan d \200\223Altmann graphical procedure for determining agreement between meas- urement metho ds, the coeffi cient of repeatability is a measure of normal day-to-day variability, thus p roviding limits of normal vari- ation ( Bland et al., 1986 ). For example, in a study in he althy adults, normal limits for heat pain threshold (thenar) ranged between 39.5 C and 5 1.9 C and the coeffi cient of repeatability was 5.6 C, i.e. a diff erence between hea t pain thresholds measured on diff erent occasions in the same subject must exceed this mag nitude in order to be considered a true diff erence (Sand et al., 2010 ). Contextual in \202 uences While it is well established that pain responses are modulated b y the psychosocial context in which they occur, surprisingly little is known about how thes e factors impact on QST fi ndings. We observed that mothers \200\231 presence during QST te sting decreased chil- dren \200\231s pain sensitivity although mothers only observed the te sting from a distance and did not interact with their children (Hohmeister et al., 2009 ; Z ohsel et al., 2006 ). Th is modulating infl uence could be accounted for by perceived socia l support and/or by distraction. Situationally induced anxiety may also exert a modulating infl u- ence. For example, in children undergoing thermal, pressure, and cold pain tasks an ticipatory anxiety accounted for about 30% of variance in pain report across tasks (Tsao et al., 2004 ). By contrast, pain-related coping including catastrophizing does not aff ect Q ST results in children (Blankenburg et al., 2011 ). Biological variables such as circadian rhythms may also modulate QST fi ndings. Very few systematic studies exist, but did not con fi rm robust circadian patterns of heat pain thresholds (e.g. Bachmann et al., 2011 ). Since QST is performed by an examiner, interrater reliability is also important. In adults, for the DFNS QST battery, good interrater reliability has been demonstrated both for compu ter- assisted (thermal) and manually obtained (mechanical) QST parameters (Geber et al., 20 11 ). Notably, the experimenters had undergone extensive training at one center. In other s tudies, interrater reliability was substantially lower (e.g. Peters et al., 2003 ). Similar ly, in healthy children, interrater reliability for pressure pain is satisfactory, but lowe r than retest reliability (Nikolajsen et al., 2011 ). Interindividual age-related di \200 erences In adults, age and gender infl uence QST resul ts (e.g. Neziri et al., 2011b ). Healthy participants older than 40 years of age as com- pa red to younger participants had signifi cantly higher detection and pain thresholds except for mechanical pain sensitivity meas- ures (Rolke et al., 2006 a; see also Hilz et al., 199 8a ; Meh et al., 1994 ). Whether sensory and pain sensitivity undergoes a developmental cha nge during childhood and adolescence is less clear. In a fi rst study using the DFNS QST ba ttery, young children (6 \200\2238 years) had higher thermal and detection thresholds and l ower heat, pressure and mechanical pain thresholds when compared to older children (9 \200 \22312 years) and adolescents (13 \200\22316 years) with the latter two groups not diff eri ng signifi cantly (Blankenburg et al., 2010 ). Yet, in a follow-up study comparing exclusiv ely 7- and 14-year-olds (Blankenburg et al., 2011 ), the age eff ect for warmth and mechani - cal detection thresholds was not confi rmed, an age-related increase in pain thresholds w as again observed and is consistent with previ- ous reports (e.g. Haslam, 1969 ; Hogeweg et al., 1996 ). Earlier stud- ies also found detection thresholds to be age-insensitive; howe ver, most of these studies focused on only one sensory modality, and were limited in sample size and age range (e.g. Hilz et al., 1996 ; Meh et al., 1994 ; Th ibault et al., 1994 ). Undoubtedly, somatosensory and pain processing undergoes functional maturation, especially during the fi rst year of life (Fitzgerald, 2005 ). Whether such developmen- tal changes in somatosensory functioning and pain processing con- tinue during childhood and adolescence, is less clear. Importantly, it cannot fully be ruled out that QST in young children may al so be less reliable due to the reaction-time dependence of many QST measures, and due to th e fact that QST parameters do not only Overall, both retest and interrater reliability of QST in children and adolescents requires systematic empirical evaluation. In order to maximize reliability, examiners should follow a highly standard- ized measurement protocol and be suffi ciently trained. Modulating in \202 uences on QST measurement outcomes Being a psychophysical procedure, QS T relies on the subject \200\231s per- ception and response behavior. Hence, procedural, co ntextual and interindividual factors may infl uence the outcome of QST and fur- ther underl ine the necessity for standardization. Procedural and methodological in \202 uences QST parameters are typically determined by ca lculating the mean of several (N = 3 \200\2237) trials in order to yield more reliable esti mates. In one of the few studies testing this assumption, cold, but not warm detection and cold pain thresholds signifi cantly decreased from the fi rst to the second trial with no f urther change across six additional trials (Agostinho et al., 2009 ). Heat pain threshold s trongly increased from the fi rst to the sixth trial; such habitua- tion to painful heat is mediated by peripheral and central proc- esses (Greff rath et al., 2007 ). Th e order of t he tests may also be relevant. For example, mechanical pain sensitivity was higher and mech anical pain threshold lower when thermal testing preceded mechanical testing as opposed to when mechanical testing was CHAPTER 40 sensory processing and neurophysiological evaluation in children 411 refl ect sensory sensitivity, but also entail a response bias component (Gracely, 2006 ). P ossibly, this response bias, rather than the actual sensitivity, may change throughout chil dhood and adolescence. Moreover, QST can be helpful to determine how generalized such altered pain responding is. For example, children with recurrent abdominal pain did not show altered threshold or sensi tization in response to somatic pain stimuli as opposed to previously demon- strated viscer al pain hypersensitivity (Zohsel et al., 2008b ). QST has been performed in children suff e ring from growing pains. Sensory thresholds for innocuous brush, cold, vibration and deep p ressure stimuli were normal, thus there was no evidence for dysfunctional peripheral nerves (Pathirana et al., 2011 ). Interestingly, children with growing pains perceived these non- painful stimuli as more intense, and had lowered pressure pain thresholds which may refl ec t disturbed somatosensory processing (Hashkes et al., 2004 ). Th us far, neuropathic pain i n children has not been systematically evalu- ated using QST. QST can be useful for early d etection of peripheral neuropathy in children with diabetes even in the absence of clinical symptoms (Blankenburg et al, 2012 ; Heimans et al., 1987 ). Interindividual sex-related di \200 erences In healthy adults undergoing QST, females have signifi cantly lower pain thresholds than men, whereas no eff ect of sex on detection thre sholds was observed (Rolke et al., 2006a ). Th is sex-related diff erence in pain sensitivi ty is consistent with conclusions of an earlier meta-analysis on sex-related diff erences i n pain sensitiv- ity (Riley et al., 1998 ). In youth, few studies have systematically addre ssed this issue. In a large cohort of about 175 healthy chil- dren and adolescents between 6 and 16 years old, girls had signifi - cantly lower detection for thermal, pressure, and v ibratory stimuli and lower thermal and pressure pain thresholds, whereas no sex- related di ff erences with regard to mechanical stimuli was observed (Blankenburg et al., 2010 ). By c ontrast, in a follow-up study of 7- and 14-year olds, no sex diff erences in pain sensitivi ty emerged, girls had only lower warmth detection thresholds (Blankenburg et al., 2011 ). S imilarly, no sex diff erences with regard to pres- sure pain thresholds were observed in ch ildren with orthopedic disorders (Nikolajsen et al., 2011 ). Blankenburg et al. speculated that puberty and the concomitant hormonal changes presumably account for the emerging sex d iff erences in pain sensitivity in adulthood. However, a recent meta-analysis of the experi mental literature concluded that even in healthy adults, there is no clear and consistent p attern of sex diff erences in pain sensitivity (Racine et al., 2012a ). One explanation may be that rather than biological sex diff erences, it is psychosocial factors that determine whether or not sex diff erences are observed (Racine et al., 2012b ). Specifi cally, gende r role stereotypes may be particularly important in determin- ing one \200\231s response to painful stimulation (Alabas et al., 2012 ). Th is could at least partially explain why sex diff erences in pain sensitiv- ity are not consistently found both in adults and in youth. QST measurements have rather consistently revealed that repeated pain experiences during th e neonatal period result in long-term altered functioning of the somatosensory and pain sys - tems (e.g. Hermann et al., 2006 ; Schmelzle-Lubiecki et al., 2007 ; Walker et al., 2009 ) . QST may also be useful for detecting hypo- or hypersensitivity to normal sensory stimulat ion which presumably underlies sensory modulation disorder (Bar-Shalita et al., 2009 ). Neurophysiological evaluation Neurophysiological methods and especially neuroimaging have p layed a pivotal role in unraveling the neural substrates of pain. Due to the complex nature of pain, it is not surprising that acti- vation of a large distributed functional network of brain regions, oft en referred to as the \200\230pain matrix \200\231 underlies the exp erience of pain. According to a meta-analysis of neuroimaging data, experi- mental pain is most commonly associated with activations in the primary and secondary somatosensory cortex , thalamus, insula, anterior cingulate and prefrontral region (Apkarian et al., 2005 ). Dep ending on type of stimulation, task, and other contextual and individual factors, regions s uch as amygdala, hippocampus, pari- etal and temporal areas, the cerebellum, and brainstem areas can also be activated. Th e neural representation of chronic pain diff ers from that of experimental pain, and chronic pain modulates brain responses to acute pain (e.g. Baliki et al., 2010 ). For example, the neuronal signature of chronic pain includes brain areas s uch as the medial prefrontal cortex which is involved in the processing of self-relevant in formation (for a review, see Apkarian et al., 2009 ). Neurophysiological methods have helpe d to identify the neuronal mechanisms of pain modulation by cognitive processes such as att ention or pain-related expectations and by emotions (e.g. Tracey and Mantyh, 2007 ; Wiech a nd Tracey, 2009; Wiech et al., 2008). Moreover, these methods have been crucial for confi r ming neuro- nal plasticity, i.e. changes in the functioning and/or structure of the nervous system, as a core mechanism in chronic pain (Tracey and Bushnell, 2009 ; Woolf, 2011 ). Related to the issue of possible sex diff erence, it has been sug- gested that subjects \200\231 responses especially to painful stimuli may depend on the sex of the experimenter. While such an infl uence of experimenter sex has been documented in some studies, Racine e t al. ( 2012b ) concluded that the available empirical literature in adults does not suppor t a clear and consistent pattern of infl uence. In children and adolescents, the issue of e xperimenter sex infl uenc- ing QST measurements has not been systematically addressed. Use of QST in children and adolescents QST is feasible in children as young as 6 years of a ge (Blankenburg et al., 2010 ; Hogeweg et al., 1996 ). Th ermal detection thresholds have e ven been measured in preschool children at the age of 4 years (Hilz et al., 1996 ). In chil dren, QST studies have focused on three research questions. Specifi cally, altered sensory and pain process- ing has been studied in the context of: \231 Chronic pain problems with or without putative sensory abnormalities. Most recently, in various chronic pain conditions, structural changes such as decreased gra y matter density have been observed which, aside from syndrome-specifi c alterations, seem to commonly aff ect brain regions such as cingulate cortex, orbitofrontal cortex, insula, a nd dorsal pons which are involved in sensory-emotional- motivational integration processes and pain regulation (Apkarian et al., 2009 ; May, 2008 ). Th ese structural changes are lik ely to be consequence of repeated pain and can be reversed by successful \231 Diseases with an increased risk for neuropathic pain. \231 Delineating the impact of neonatal pain. Chronic pediatric pain is associated with enhanced pain sen- sitivity as indicated by lower ed pain thresholds and augmented temporal summation which, however, does not necessarily af f ect all sensory modalities (e.g. Walco et al., 1990 ; Zohsel et al., 2006 ). SECTION 5 measurement of pain 412 treatment (Rodriguez-Raecke et al., 2009 ; Seminowicz et al., 2011 ). While there is a weal th of data in adults, relatively few studies have used these methods to study pain processi ng during childhood. To some extent, this may be explained by not all neurophysiological te chniques being easy to use in children. in the blood and, therefore, is referred to as a blood oxygen lev- el-dependent (BOLD) imag ing technique. Oxygen consumption due to increased neuronal activity is followed by an incr ease in blood fl ow and blood volume that leads to reduced concentration of deoxygenated he moglobin and a concomitant increase in the BOLD signal (e.g. Wager et al., 2007 ). Major ad vantages of fMRI are the relatively high spatial resolution for localizing changes in corti - cal, subcortical, brainstem, and even spinal activity, and non-inva- siveness that permit s its use in children. Nonetheless, fMRI studies in children are challenging because the sc anner environment is likely to be experienced as mildly anxiety-provoking, at least ini- ti ally. Moreover, fMRI recordings are highly subject to movement artifacts and, therefore, re quire the subject to lie still during the scanning. Th is can be a diffi cult task, especia lly for younger chil- dren. Th e processing of fMRI data requires that individual data are related to a standard template of common spatial coordinates in order to anatomically local ize brain activations. Th e human brain not only grows in size, but also undergoes structur al changes from infancy through childhood until adulthood (e.g. Johnson, 2001 ). Only recen tly, MRI atlas templates for diff erent age groups have become available that are based on MRI recordings in larger and more representative samples of children and adolescents (Fonov et al., 2011 ). Neurophysiological methods used for the study of pediatric pain In the past two decades, an unprecedented progress in the devel- opment of neuroimaging techniques occurred. A thoroug h review of these techniques is beyond the scope of this chapter. Here, the focus is on met hods that have been used for studying pediatric pain. Neuroimaging Generally, neuroimaging methods can be subdivided into those assessing structu ral, and those assessing functional, changes in the brain. Structural changes (e.g. gray ma tter density) or changes in structural connectivity between brain regions are measured by v oxel-based morphometry and diff usion tensor imaging, respec- tively (Sava et al., 2009 ). Functional imaging techniques diff er with regard to the parameter that is measured as a pr oxy of neu- ral activity. Similar to fMRI, near-infrared spectroscopy (NIRS) is a non- invasive imaging technique bas ed on measuring shift s in (de-) oxygenated hemoglobin. Whereas fMRI uses the diff erent pa ra- magnetic properties as recording signal, NIRS relies on the diff er- ent optical proper ties in the visible and near-infrared light range (Bunce et al., 2006 ). Light administered to the scalp is absorbed by tissue and scattered back to the surface. Oxygenated and deoxy genated hemoglobin diff er in their absorption sensitivity depending on the wavelength of t he light. By using wavelengths maximizing the absorption by (de)oxygenated hemoglobin, meas uring the light scattered back to the surface can be used to infer brain activity. NIRS is an especially promising technique for use in children and even infants because it is safe, non-invasive, and can even be performed as part of a clinical exam (Ranger et al., 2011 ). Electrophysiological techniques Techniques such as EEG or magnetoencephalography directly m easure electrical activity of neuronal assemblies with high temporal resolution (Pizzagalli , 2007 ). EEG activity recorded on the scalp is presumably generated by excitatory and inhi bi- tory postsynaptic potentials in cortical neurons, hence EEG is not well suited for meas uring subcortical activity, and does not yield whole-brain imaging data. Spatial informatio n about the source of the electrical activity has to be inferred by computational mod- els. Due to the high temporal resolution, it is possible to separate event-related, i.e. time-l ocked brain activity from background EEG oscillation. Th ese event-related potentials (ERPs ) are char- acterized by distinct components, i.e. negative and positive peaks in the EEG w aveform, which refl ect diff erent aspects of stimu- lus processing. For example, in respon se to painful stimulation, somatosensory ERPs typically show an initial negative peak at ab out 150 ms post-stimulus followed by a positive peak at about 260 ms (N150 \200\223P260). T h e absolute value of this negative-positive vertex response correlates positively with sub jective pain intensity (Garcia-Larrea et al., 1997 ). Later ERP components (e.g. P300) refl ect cognitive processes such as attention and memory (Fabiani et al., 2007 ). Typically, E RPs are derived based on averaging many trials such that time-locked EEG changes can be dis criminated from random variations of EEG activity. However, statistical methods (e.g. princ ipal component analysis) can be applied for single trial analysis such as the cortical resp onse to a heel lance (Slater et al., 2010c). Understanding pediatric pain \200\224evidence from neurophysiological studies Evaluation in early life Neurophysiological methods have fundamentally contributed to unraveling the dev elopment of pain processing, especially dur- ing early infancy (e.g. Fabrizi et al., 2011 ; see also Chapter 40 40 Poorun and Slater, Chapter 39, this volume). In 2006, using NIRS, t wo studies were the fi rst to demonstrate that, as early as 25 weeks postmenstrual age, an invasive painful medical procedure (e.g. heel lance, venepuncture) as compared to non-painf ul touch elicits a distinct haemodynamic response in the contralateral somatosensory cortex , hence at this developmental stage thalam- ocortical pathways are suffi ciently mature for cortical processing of nociceptive input (Bartocci et al., 2006 ; Slater et al., 2006 ). A lso, diff erent intensive care procedures are accompanied by signifi cant cerebral haemodyn amic changes (Limperopoulos et al., 2008 ). Extending these fi ndings, time-locked EEG reco rdings in infants undergoing heel lance revealed a distinct component consisting of a negat ive and positive defl ection at about 500 ms (N420 \200\223P560) Indirect measures of neural activity: metabolism, oxygen consumption, and blood \202 ow A nother approach is to record changes in metabolism, oxygen consumption, and blood fl ow bas ed on the assumption that these are closely related to neural activity. FMRI is based on me asuring changes in the ratio of oxygenated to deoxygenated hemoglobin CHAPTER 40 sensory processing and neurophysiological evaluation in children 413 Chronic pain In chronic pediatric pain, EEG has been used to study cognitive aspects of pai n processing and specifi c pathophysiological mech- anisms. In children with migraine and c hildren with recurrent abdominal pain, we recorded somatosensory-evoked potentials in respo nse to painful and non-painful mechanical stimuli which had to be ignored while performing a primary task (Hermann et al., 2008 ; Zohsel et al., 2008a ). Painful stimuli elicited a s ignifi cantly larger vertex potential (N150 \200\223P260) compared to non-painful stimuli. In both pain groups, the P300 in response to both pain- ful and non-painful stimuli was sig nifi cantly greater than in the controls (see Figure 40.2 ). Th is suggests that chronic pa in in chil- dren is associated with automatic attention to painful and poten- tially painfu l stimuli. In migraine, heightened cortical excitability and altered cortical sensory proce ssing have been proposed as pathogenetic mechanisms. Consistent with such concepts, in chil - dren with migraine, both cross-sectional and longitudinal studies entailing the measureme nt of ERPs to sensory stimuli (e.g. visual) and slow cortical potentials suggest altered ma turation of cortical information processing (Oelkers-Ax et al., 2008 ; Siniatchkin et al., 2010 ). Th us far, only one study has used fRMI in chronic pediatric pain. Children with co mplex regional pain syndrome (CRPS) were exposed to non-noxious thermal and mechanical stim ulation during the acute pain phase and aft er recovery (Lebel et al., 2008 ; see also Clin ch, Chapter 24, this volume). In the acute pain phase, patients experienced allodynia in th e aff ected extremity and, consistently, at electrode sites above the somatosensory cortex that is specifi c for nociceptive input a nd is larger in preterm compared to full- term-born infants (Slater et al., 2010b , 2010c). Such direct meas- uring of cortical activity is especially promising for evaluating the ef fi cacy of (non-)pharmacological treatments for infants given that, at this age, pain measu rement usually relies exclusively on behavioral observations and physiological parameters ( e.g. heart rate, oxygen saturation). For example, in infants, oral sucrose as compared to w ater did not reduce the cortical response to a pain- ful heel stick despite resulting in an attenuated behavioral pain response (Slater et al., 2010a ). Using fMRI, we studied the lo ng- term impact of neonatal pain experiences on brain activation in response to heat pain i n school-aged preterm and full-term-born children having undergone treatment in a neonatal intensive care unit (NICU) and control children (Hohmeister et al., 2010 ). In all three gr oups, tonic heat pain led to signifi cant activations of brain regions typically involved i n pain processing (see Figure 40.1 ). Th e preterm, but not the full-term NICU children exh ibited signifi - cant activations in a number of brain regions (thalamus, anterior cingulat e cortex, cerebellum, basal ganglia, and periaqueductal gray) that were not signifi cantly activated in controls. Moreover, the preterms showed signifi cantly higher activations than con- trols in primary somatosensory cortex, anterior cingulate cortex, and insula. Hence, exposure to pain at a vulnerable stage of the developing pain circuitry, may aff ect both s ensory and aff ective processing of pain later in life. Figure 40.1 Signi \201 cant brain activations ( p <0.001, k \211 10) during painful toni c heat stimulation in NICU-preterms (red), NICU-full-terms (blue), and the controls (green) , superimposed on a normalized T 1 -weighted brain. X-coordinates of brain slices (in mm) r efer to the Montreal Neurological Institute standard brain. PFC = prefrontal cortex; PPC = posterior parietal cortex; PMC = premotor cortex; antIns = anterior insula; Cereb = cerebel lum; BG = basal ganglia; pre-SMA = pre-supplementary motor area; ACC = anterior cingulate c ortex; Th al = thalamus; PAG = periaqueductal gray. Reproduced from Johanna Hohmeister et a l., Cerebral processing of pain in school-aged children with neonatal nociceptive input: An exploratory MRI study, PAIN , Volume 150, pp. 257 \200\223267, Copyright 2010. Th is \201 gure has been reproduced with permission of the International Association for the St udy of Pain (IASP). Th e \201 gure may NOT be reproduced for any other purpose without permission. Th is \201 gure is reproduced in color on the back endpaper. SECTION 5 measurement of pain 414 this is most likely accounted for by ethical considerations given the relatively high deman d on children \200\231s compliance with instructions and measurement requirements. References Agostinho , C. M. S. , Scherens , A. , Richter , H. , Schaub , C. , Rolke , R. , Treede , R. D. , et al . ( 2009 ). Habituation and short-term repeatability of thermal tes ting in healthy human subjects and patients with chronic non- neuropathic pain . Eur J Pain , 13 , 779 \200\223785. 204 \200\223216 ms 220 \200\223232 ms 236 \200\223248 ms