N = 28). Based on measured anatomical landmarks from X-ray and MRI, knee joint templates were selected from the atlas library. The cartilage stresses and strains of the X-ray-based model were then compared with the MRI-based model during the stance phase of the gait. The biomechanical responses were statistically not different between MRI- vs. X-ray-based models when the template obtained from X-ray imaging was the same as the MRI template. However, if this was not the case, the peak values of biomechanical responses were statistically different between X-ray and MRI models. The developed X-ray-based framework may pave the way for a clinically feasible approach for knee joint FE modeling.Finite element (FE) modeling is becoming an increasingly popular method for analyzing knee joint mechanics and biomechanical mechanisms leading to osteoarthritis (OA). The most common and widely available imaging method for knee OA diagnostics is planar X-ray imaging, while more sophisticated imaging methods, e.g., magnetic resonance imaging (MRI) and computed tomography (CT), are seldom used. Hence, the capability to produce accurate biomechanical knee joint models directly from X-ray imaging would bring FE modeling closer to clinical use. Here, we extend our atlas-based framework by generating FE knee models from X-ray images (The online version contains supplementary material available at 10.1007/s10439-022-02941-0. The potential of X-ray imaging under a similar workflow has not been evaluated so far. As in primary healthcare, planar radiographs are the primary imaging modality for knee OA diagnostics,16 their compatibility in the modeling workflow would be a valuable asset. The capability to generate 3-D knee joint models from 2-D radiographs could also increase the scalability of the FE modeling to large patient groups.Osteoarthritis (OA) is one of the major causes of disability in elderly people.19 These methods include conventional image segmentation methods, e.g., contour matching, edge detection and statistical shape modeling.48 Methods that include only radiographs as input images are challenging to use in soft tissue segmentation due to very limited soft tissue contrast. Moreover, region-based algorithms are prone to noise and over-segmentation of the X-ray images.45 Statistical shape model (SSM)-based geometry generation has been shown to produce a good accuracy in generation of knee geometries but it requires sufficiently high-quality and large datasets to capture all possible shape variations.12 Though there are several publicly available clinical datasets for digital health research,21 the available datasets are often noisy, incomplete and contain artifacts as clinical imaging is tailored to physicians’ needs and to minimize harm to patients.27Various methods for the 3-D reconstruction of shape and geometry from X-ray images have been developed.43 However, issues related to the image quality and artifacts, as well as the substantial amount of manual labor required for the generation and simulation of FE knee joint models, present challenges when using these techniques. An approach introduced by Rodriguez-Vila et al.37 for the rapid generation of FE meshes is quite promising but has not been tested for a large group of subjects. Recently, we developed an atlas-based FE knee joint modeling workflow utilizing knee CT images for rapid knee FE model construction and simulation without cartilage segmentation.32 We observed that CT-based knee models produce similar mechanical responses in knee joint cartilage as MRI-based models.32 This was an important step and introduces the possibility of using native X-ray imaging, where only bones are visible, for FE model generation.In any clinical application, FE model generation must be fast. Various methods have been applied to reduce model generation times utilizing different semiautomatic and automatic segmentation and meshing techniques.32 We hypothesized that the previously presented approach can generate 3-D knee joint FE models based on 2-D knee radiographs and that these models can simulate similar cartilage mechanical responses when compared to the models generated from MR images.Thus, the key purpose of this study is to investigate whether an automated knee joint FE modeling approach based on knee radiographs can simulate mechanical responses of knee joint cartilage with similar accuracy as our previously verified MRI-based knee joint FE modeling approach.18 Written consent of each patient was taken. The data was collected in compliance with the Declaration of Helsinki, and the ethical permission was granted by the Ethical Committee of Northern Ostrobothnia Hospital District, Oulu University Hospital (number 7/2016).18Previously collected data from 28 patients with ages between 48 and 65 years were used in this study.18 Bilateral posterior–anterior  and lateral  knee radiographs were acquired according to normal clinical routine in a weight-bearing and semi-flexed position .18 The posterior–anterior X-ray images were taken using the SynaFlexer™ X-ray positioning frame  while no knee positioning frame was used for the lateral images. Therefore, there was some variation in the flexion of the knee in the lateral images of X rays. The rotation of the knee was minimized so that the femoral condyles would be superimposed. MRI scans of knee joints of all subjects were also taken on the same day using a sagittal T2-weighted dual-echo steady-state sequence . In our study, the right knees of the patients were analyzed. An experienced radiologist scored these radiographs using the Kellgren–Lawrence grading, and patients had no evidence of clinical cartilage erosion .18The magnetic resonance (MR) and X-ray images of the knee joint of each patient were acquired using a clinical 3 T MRI scanner  and X-ray device .The workflow of the atlas-based method is shown in Fig. 32 Briefly, 3D knee joint geometries (N = 21) from the MR images obtained from the OAI database (https://nda.nih.gov/oai/) were created in earlier studies32 using Mimics v15.01 . Final geometries were imported in Abaqus software , in which the FE models were constructed. Subsequently, to enhance the computational convergence and efficiency, only medial compartment FE models were considered in further simulations. Finally, anatomical landmarks were measured from clinical MR images (OAI database) for each knee joint geometry using our in-house build script in MATLAB, and this information was linked with a corresponding medial compartment model. The five measured anatomical dimensions included the maximum anterior–posterior (AP) distance of the medial and lateral condyles of distal femur, tibiofemoral joint space width (JSW) of the medial and lateral compartments, and lastly, the maximum medial–lateral (ML) width of distal femur. The AP dimensions from MR images were determined as the maximum AP distance of ellipsoidal shaped medial and lateral condyles (major axis of the ellipsoid). Tibiofemoral JSWs were measured at the central location of the tibiofemoral contact region in medial and lateral compartments. ML width was the maximum distance between the medial and lateral epicondyles, i.e., clinical transepicondylar axis , the previously described atlas-based approach was utilized  and tibiofemoral JSWs were measured. To mimic these measurements in X-ray images as taken from MR images, both AP dimensions were determined from the sagittal plane, whereas JSWs and ML dimensions were obtained from the coronal plane  was used to measure these dimensions. Although the sagittal images of X rays were differently oriented than MRIs, the approach of taking the maximum AP length of ellipsoidal shaped medial and lateral condyles was kept the same in both MR and X-ray images. To account for slight image magnification in radiographs, each posterior–anterior and lateral X-ray image contained a calibration disc (diameter = 30 mm) placed in a precise position in the field of view and the disc was used to calibrate the image dimensions. Afterward, these measurements were repeated for the MR images of the same knees. Anatomical dimensions were measured three times from both X-ray and MR images of each patient by the same rater to evaluate the intra-rater reliability14 of the measurements and if they affect the template selection. An experienced musculoskeletal radiologist checked the quality of the dimension measurements. In total, 84 knee joint models for both imaging modalities were generated.To develop X-ray- and MRI-based knee joint FE models for the study subjects  were calculated between the normalized dimensions of the subject and FE models found in the atlas library. The best-matched FE model atlas was determined based on the minimum sum of the RMSE values, and this FE model atlas was selected to represent the subject  values of the best-matched atlas with the percentage difference in AP, ML and tibiofemoral JSW dimensions  between the subject data and best-matched atlas .33More detailed information on boundary conditions and contact definitions can be found from our previous studies.50 The fibrillar matrix was modeled with 4 organized primary and 13 randomly oriented secondary fibrils.50 The total stress tensor of the material is of the form22:σt is the total stress tensor, σnf is the nonfibrillar matrix stress tensor, ith fibril, tot f is the total number of fibrils, p is the fluid pressure. Cartilage material properties were obtained from previous studies  script in Abaqus. FE model simulations were performed in Abaqus/Standard (v6.18).The cartilage in the FE model was considered as a fibril reinforced poroviscoelastic (FRPVE) material involving a porous fluid-filled hyperelastic nonfibrillar matrix and a viscoelastic fibrillar matrix. Thus, the FRPVE material model can determine the contribution of the main tissue constituents  on the biomechanical response of cartilage.49 The mean (over cartilage–cartilage contact area) of the aforementioned parameters were obtained from the element centroids of the tibial cartilage as a function of the stance. Additionally, for determining the maximum values over stance, the element with the peak value of biomechanical parameter was identified. This value was averaged with the values of its connecting (neighboring) elements to eliminate abnormally high parameter values that originate from numerical contact discretization.The following biomechanical cartilage responses were analyzed from the FE models: the maximum principal stress, maximum principal strain, minimum principal strain, collagen fibril strain, and fluid pressure. These mechanical parameters were determined as they are known to be associated with the load-bearing, degeneration and possible failure properties of cartilage.t-test with 95% confidence intervals (CIs). The significance level was α = 0.05. Correlation analysis of anatomical dimensions measured from both imaging modalities is presented separately in the Supplementary Material . These analyses were conducted in IBM SPSS Statistics . Using MATLAB , we compared the 1-D trajectories of the averaged and peak values (over the contact area) of the biomechanical parameters from MRI- and X-ray-based models of each subject through the stance using the pairwise 1-D SPM,36 implemented via 1-D SPM toolbox (https://spm1d.org/). The normality of this data was tested via Shapiro–Wilk test prior to applying the parametric or nonparametric version of SPM. SPM method has an advantage over traditional zero-dimensional (0-D) parametric or nonparametric statistical tests (t-test or Wilcoxon signed-rank test) as it can consider various comparisons on smooth and random 1-D trajectories36 as in the case of the biomechanical FE model results.The normality of anatomical dimension measurement data was verified via the quantile–quantile plot, and the Shapiro–Wilk test. The anatomical dimensions obtained from both modalities were statistically compared using the parametric paired samples p > 0.05) between the mean values of maximum medial  and lateral  JSWs between X-ray and MR images. On the other hand, the mean values of maximum AP dimensions of both medial  and lateral  elliptical-shaped condyles were greater in X-ray compared to MRI (p < 0.001). The mean value of the maximum ML distance of the distal femur was slightly greater  in MR than in X-ray images (p < 0.001). See Supplementary Table ST1 for the 95% CIs of measured parameters.There was no significant difference  at 20, 50 and 80% of stance are shown in Fig. During the analyses, we observed that some of the model templates generated using radiographs matched with MRI generated templates , but not in all models , thus these cases are presented separately in the following sections.Case (1) Average and peak biomechanical responses considering subjects (8) for whom the atlas-based approach determined the same best-matched templates from MRI and X-ray images (number of both MRI- and X-ray-based models = 24).In general, both peak and averaged biomechanical responses over the cartilage–cartilage contact region exhibited no statistical differences between the X-ray- and MRI-based FE models at the loading response and terminal stance phases of the gait cycle Figs.  and 4. YCase (2) Average and peak biomechanical responses considering all subjects (28) for whom the atlas-based approach determined the same or different best-matched templates from MRI and X-ray images (number of both MRI- and X-ray-based models = 84).In general, the peak biomechanical responses over the cartilage–cartilage contact region exhibited statistical differences between the X-ray- and MRI-based FE models at the loading response and terminal stance phases of the gait cycle Fig.  while th32 to incorporate knee radiographs for automated knee FE model generation and analysis of cartilage biomechanical responses. The model results were then compared with MRI-based models of the same subjects. Consistent with our hypothesis, we observed that peak and averaged cartilage stresses, strains and fluid pressures were not statistically different between the X-ray- and MRI-based models at loading response, terminal extension or along the majority of the stance when the same template was employed. However, the peak responses were statistically different along extended periods of stance between the X-ray- and MRI-based models that utilized varied templates.In our study, we expanded our previously presented atlas-based FE knee joint modeling approach14 of the measurements and if their difference affect the template selection. Femoral condyles in radiographic images are visible in the lateral view. However, in this view, it becomes difficult to distinguish between both medial and lateral superimposed condyles . This causes uncertainties in the estimation of AP distance of femoral condyles from X-ray images (Table ST1) leading to sometimes different template selection among both modalities. Moreover, there was no knee positioning frame used when taking the lateral images. Therefore, there is variation in the flexion of the knee in the lateral images of X rays. As lateral views of MRIs and X rays are differently oriented, this may cause differences in condyles measurements from both imaging modalities. One of the limitations in this study was that the measurement accuracy of anatomical dimensions from sagittal and coronal views can be influenced by variations in internal-external positioning of the knee joint in X-ray images.34 Thus, in our presented biomechanical results obtained from X-ray- and MRI-based models, the only possible source of differences in mean and peak values of mechanical parameters arise presumably due to differences in anatomical dimensions and subsequent template selection for model generation. All other inputs in a model generation were the same in both approaches. Furthermore, different pixel size  causes uncertainties in these analyses. These differences may be affected by X-ray image magnification which varies with patient positioning and size even with strict imaging protocols.Medial and lateral JSWs showed no significant difference between X-ray and MRI analyses. The maximum AP lengths of medial and lateral condyles were greater in X rays compared to MR images. Especially, the measured AP  length was almost 8 mm greater in X rays than MRIs. Contrarily, the maximum ML length was slightly greater in MR images compared to that in X-ray images. Anatomical dimensions from both modalities were measured three times for each subject to evaluate the intra-rater reliability28 The subject-specific (one subject) comparisons for the mean values of biomechanical are presented separately for each parameter in Supplementary Fig. S6. Mean values of biomechanical parameters were obtained by averaging over cartilage–cartilage contact area over the whole stance phase, while maximum values were obtained by averaging the biomechanical parameter values from the peak value element and its neighboring elements over the contact region during stance. A high stress or strain concentration even in one node can lead to overestimated peak results in certain periods of the stance phase. Taken together, the results emphasize that utilization of radiographs in knee FE modeling requires careful planning of knee X-ray imaging so that AP and ML dimension measurements would correspond to those of MRI.Statistical analysis focusing only on subjects (8) for whom the atlas-based approach determined the same best-matched templates based on all three anatomical dimensions from MRI and X-ray images showed that the peak responses were not statistically different at the loading response and terminal extension among both models Fig. . HoweverAll models were modeled with 8-node porous linear hexahedron C3D8P element type. Although mesh type was the same among all atlases, cartilage thickness and shape vary due to different knee joint sizes . Hence, different atlases have different elements and nodes distribution. As a result, local stresses and strains may differ among different FE atlas models . However, undesirable mesh collapsing due to secondary scaling was not the issue in this study. We observed that the relative difference between the greatest values of anatomical dimensions i.e., ML, AP medial, AP lateral, JSW medial and JSW lateral distances among the X-ray measurements and corresponding dimensions for atlases in the atlas-library was 0.4, 8, 5, 11 and 13% respectively. Similarly, the relative difference between the smallest values of anatomical dimensions i.e., ML, AP medial, AP lateral, JSW medial and JSW lateral distances among the X-ray measurements and corresponding dimensions for in atlas-library is 0.2, 7.9, 4, 8 and 10% respectively.42 although deep learning-based techniques also look promising.35 In terms of generation of the model geometry, we compared earlier the results of the FE models obtained from the atlas-based approach and manually segmented knee joint models with experimental data.32 In that study, we showed that the models based on the atlas-based approach were able to predict the progression of OA similar to the models based on manual segmentation.Atlas-based segmentation approaches are one of the most robust image-based segmentation techniques in the field of medical simulations, which perform classification and segmentation in one go,44 Specifically, the excessive minimum principal strain has been linked to PG loss via cell death.41 These mechanical parameters can be evaluated computationally along the gait from easily accessible X-ray imaging modality using our presented approach. The results were validated against MRI-based model responses (except peak responses in the case of varied templates selection). Yet, no prediction for OA progression was done in this study as there was no clinical follow-up information of disease progression for these subjects.18 However, it is important to note that our current methodology can be applied for the prediction of disease progression as our previously developed prediction algorithm32 is directly related to analyzed biomechanical parameters estimated in the current study. The algorithm was controlled by the accumulation of maximum principal stresses within cartilage during physiological loading which is suggested to reflect collagen failure in cartilage.32 The predictive FE model was generated from baseline (mainly KL0) and then it was used to predict OA progression, as seen experimentally by increased KL grades. To date, there have been no tools for evaluating cartilage degeneration and joint health, hence, the clinical applicability of the current methodology to predict cartilage failure and OA progression from X rays should be evaluated against a clinical follow-up study.Biomechanical parameters like internal tissue strains have been linked with proteoglycan degeneration and subsequently fixed charge density (FCD) loss, while the collagen fibril strain and maximum principal stress have been suggested to be associated with collagen failure and degeneration.5 This may be partially due to the characteristics of joint load distributions between lateral and medial compartments during walking.32 However, load distribution between lateral and medial compartments can affect the biomechanical responses and to tackle this we are currently working on adding tibiofemoral lateral compartment in our atlas-based workflow. Moreover, the addition of other knee structures like the patellofemoral compartment, including tendons, ligaments and muscles, will drastically increase computational time. To circumvent this, in the present study effects of these biological structures were included in the total force applied in the knee FE model. This can be justified since adding those tissues properly into the FE model needs information from subject-specific motion analysis data.In the current study, we simulated biomechanical responses only from medial tibial cartilage and the lateral compartment of the knee was not considered in this study. The motivation for this was based on the literature as knee OA is known to occur particularly in the medial compartment of the tibiofemoral joint.32 In that approach,32 MRI-based atlas knee models with generic gait loading were able to estimate volumetric cartilage degeneration which was indicative of OA progression. Moreover, clinicians widely use imaging modalities data to diagnose knee joint disorders and OA, whereas it is not viable to obtain motion data of every subject in the clinical setup. However, subject-specific motion data could further enhance and personalize our framework, hence, its role should be investigated.In our knee FE models, the subject-specific knee motion was not considered since the gait data was not collected from the patients. Furthermore, to reduce computational burden of simulations, the contribution of the medial meniscus was considered by subtracting the average contribution of meniscus from the total joint contact forces of the medial compartment. Hence, the current way to apply joint loading is not personalized. Nevertheless, it needs to be emphasized that the aim of this paper was not to produce subject-specific tissue responses during walking, but to investigate can this radiograph-based approach produce similar mechanical responses in cartilage as produced previously by the verified MRI-based approach.30 is applicable for knee joints that don’t show X-ray-based cartilage degeneration.We have currently only 21 atlases in our atlas library and we can simulate biomechanical parameters of only intact cartilages from this approach and cannot account for a plethora of knee cartilage pathologies such as cartilage lesions or defects. Hence, the inclusion of templates with cartilage lesions will lead to a more optimal representation of subject geometry. Additionally, only subjects with KL0 and KL1 grades were modeled. One of the reasons to include subjects based on KL0 and KL1 grade was to demonstrate that atlas-based approach32 JSW has the most effective role on the predicted peak stress levels , but among the models with the same and varied templates, peak responses are more sensitive to template selection. Due to statistical differences in anatomical dimensions among both imaging modalities , the same template was not selected in all subjects. Different  templates produce different peak values Below is the link to the electronic supplementary material.


Psychopathy is a personality disorder characterized by lack of empathy, grandiosity, an impulsive lifestyle and antisociality. Anti-social personality disorder (ASPD) and psychopathy are distinct concepts presenting different criteria. Most people with a diagnosis of psychopathy also meet criteria for ASPD while the reverse is not true. Along the years there has been an increasing interest in investigating genetic and neurobiological factors.To analyze the neurobiological factors involved in psychopathy and anti-social personality disorder according to the scientific knowledge available.Review of scientific literature via PubMed search, using the terms “anti-social personality disorder”, “biology or etiology or pathophysiology and psychopathy”.The strongest evidence base for a genetic pathway is associated with the low-expression variant of the Monoamine Oxidase-A (MAO-A) which is linked to the X chromosome. Other genetic factors involve the 5-HTT gene, dopamine receptor genes (DRD4 and DRD2) and genetic polimorfisms at SNAP25 t-snare protein, OXT gene and the CNR1 and FAAH cannabinoid receptor gene. Structural differences in the brain have been noticed such as reduced gray matter volume in the orbitofrontal cortex, gray matter volume reductions in the mid-anterior insula and left anterior temporal cortex, subtle reductions in gray matter volume across several paralimbic and limbic areas.There is considerable evidence regarding various possible underlying neurobiological processes in psychopathy although it is insufficient to suggest a single biological etiology and environmental influences cannot be excluded from a complete understanding of this disorder. The neurobiological correlates found hold promise for new research and treatment.


Sensory deprivation, following a total loss of one sensory modality e.g. vision, has been demonstrated to result in compensatory plasticity. It is yet not known to which extent neural changes, e.g. higher resting-state activity in visual areas  as a consequence of blindness, reverse, when sight is restored. Here, we used functional MRI to acquire blood oxygen level-dependent resting-state activity during an eyes open and an eyes closed state in congenital cataract-reversal individuals, developmental cataract-reversal individuals, congenitally permanently blind individuals and sighted controls. The amplitude of low frequency fluctuation of the blood oxygen level-dependent signal—a neural marker of spontaneous brain activity during rest—was analyzed. In accordance with previous reports, in normally sighted controls we observed an increase in amplitude of low-frequency fluctuation during rest with the eyes open compared with rest with eyes closed in visual association areas and in parietal cortex but a decrease in auditory and sensorimotor regions. In congenital cataract-reversal individuals, we found an increase of the amplitude of slow blood oxygen level-dependent fluctuations in visual cortex during rest with eyes open compared with rest with eyes closed too but this increase was larger in amplitude than in normally sighted controls. In contrast, congenital cataract-reversal individuals lagged a similar increase in parietal regions and did not show the typical decrease of amplitude of low-frequency fluctuation in auditory cortex. Congenitally blind individuals displayed an overall higher amplitude in slow blood oxygen level-dependent fluctuations in visual cortex compared with sighted individuals and compared with congenital cataract-reversal individuals in the eyes closed condition. Higher amplitude of low-frequency fluctuation in visual cortex of congenital cataract-reversal individuals than in normally sighted controls during eyes open might indicate an altered excitatory–inhibitory balance of visual neural circuits. By contrast, the lower parietal increase and the missing downregulation in auditory regions suggest a reduced influence of the visual system on multisensory and the other sensory systems after restoring sight in congenitally blind individuals. These results demonstrate a crucial dependence of visual and multisensory neural system functioning on visual experience during a sensitive phase in human brain development. Rączy et al. report altered slow blood oxygen level-dependent fluctuations during rest in sight recovery individuals indicating that the typical excitatory–inhibitory balance of visual cortex, as well as the interaction of the visual system with other sensory and multisensory systems require visual input during a sensitive period.  A higher activation of visual cortex in permanently blind humans during rest7 has been interpreted as a higher excitatory–inhibitory (E/I) balance allowing for cross-modal activation.8 Both intra- and cross-modal plasticity have been linked to compensatory performance in blind humans.9A number of studies have demonstrated that sensory deprivation, due to e.g. blindness, results in functional and structural changes of the brain related to intra- and cross-modal plasticity.10 Individuals, who due to the presence of bilateral dense congenital cataracts did not experience any patterned vision after birth, have been shown to suffer persisting visual deficits in multiple visual functions.11–16 Yet it is unknown whether cross-modal plasticity retracts following sight restoration and whether typical visual processing circuits emerge.17 There is some evidence suggesting a larger influence of the auditory system on the visual system in sight recovery individuals with short deprivation epochs using brain imaging18–21 and behavioural measures,22 whereas electrophysiological studies have not yet found analogous evidence.15 Recent results in the deaf cat have argued against cross-modal plasticity limiting auditory recovery.24 In two brain imaging studies in short deprived congenital cataract-reversal individuals cross-modal activation of their visual cortex elicited by auditory stimulation19 was rather weak compared with the typical cross-modal activation observed in permanently blind humans.25 However, longer deprivation periods might cause more extensive and persisting cross-modal plasticity. It has been suggested that what causes persisting visual deficits in visual functions27 is the lack of an experience dependent functional and structural tuning29 of genuine visual processing. Based on this hypothesis we would predict altered resting-state activity in visual brain regions after restoring sight in individuals born without pattern vision. Such a lack of reversibility of typical resting-state activity profiles would be evidence for a sensitive period for typical brain development.30The question has been raised whether adaptations to blindness, in particular cross-modal plasticity interfere with functional recovery when vision is restored by preventing the functional tuning of visual areas for visual processing.31 In humans, Biswal et al.32 demonstrated that low frequency blood oxygen level-dependent (BOLD) fluctuations (0.01–0.08 Hz) within the sensorimotor network during rest mirrored typical sensorimotor activity during a finger tapping task indicating that resting-state BOLD fluctuations correlate across functionally related, even spatially remote brain regions similarly as during task performance.33 Resting-state BOLD connectivity often reflects structural connectivity as assessed with diffusion spectrum imaging in humans34 and as demonstrated by combining non-invasive functional magnetic resonance imaging (fMRI) and invasive anatomical retrograde tracing methods in monkeys.35 Resting-state connectivity, does not, however, provide information about the level of BOLD signal change. In contrast, the amplitude of low-frequency fluctuation (ALFF)36 indicates activity in the low frequency range (0.01–0.08 Hz) of the BOLD response. ALFF reflects the strength of low frequency oscillations (LFOs) and is considered as a good estimate of the overall activity in neural circuits.37Resting brain activity is considered as crucial scaffold for task-related processing. For example, it has been found that resting-state connectivity in the visual cortex of awake ferrets mimics the typical visual elicited brain activity.37–43 and in blind humans44 as compared with the control group. In healthy sighted subjects, ALFF has been compared between eyes open (EO) and eyes closed (EC) resting states. Typically, during EO ALFF is higher than during EC in higher visual cortices  and in orbital frontal cortex, whereas ALFF was found to be decreased in the bilateral pre- and postcentral gyrus, as well as in temporal and insula regions, the thalamus44–47 and in most studies in primary visual cortex.47ALFF has been shown to indicate changes in overall neural activity in psychiatric and neurological populations49 During rest, alpha activity is higher with EC than with EO.51 Alpha band activity has been suggested to reflect a neural mechanism important for the control of the E/I balance of neural circuits to guarantee efficient processing: During task processing alpha band power is high for neural circuits not engaged in task processing but low for task relevant neural systems52 resulting in an improved signal-to-noise ratio in neural networks.Simultaneous EEG-fMRI studies reported a negative correlation of the ALFF and EEG alpha oscillatory activity.53 The authors speculated that the reduced alpha activity in congenital cataract-reversal individuals was due to a reduction of inhibitory mechanisms which seem to be elaborated during sensitive periods.29 In fact, reduced posterior alpha activity is a typical characteristic of the EEG of permanently blind humans.54–57 Moreover, two magnetoencephalography studies later reported enhanced gamma activity in congenitally permanently blind individuals compared with typically sighted controls.59 Recent studies have suggested that alpha and gamma oscillatory activity indicate antagonistic mechanisms with alpha oscillations controlling gamma oscillatory activity.61 Interestingly, in monkeys, it has been observed that slow BOLD fluctuations positively correlate with high frequency local field potentials (LFPs) in the gamma range particularly in an EC condition.62Posterior alpha oscillatory activity was found to be significantly reduced in congenital cataract-reversal individuals as compared with developmental cataract-reversal individuals  and typically sighted controls.63 assessing the amplitude of low frequency oscillations of the BOLD signal in congenital cataract-reversal individuals compared on the one hand to congenitally permanently blind humans and on the other hand, to typically sighted controls allows investigating the degree of functional recovery of visual neural circuit functioning following sight restoration. Hence, the present study investigated the question of whether there is a sensitive period for the development of typical resting-state activity in humans and thus a retraction of cross-modal plasticity as observed in congenitally blind humans.Based on the group differences in oscillatory brain activity , we hypothesized that ALFF is enhanced in congenital cataract-reversal individuals, as well as in congenitally blind individuals compared with typically sighted controls in occipital brain regions. However, resting-state brain activity has not yet been investigated in congenital cataract-reversal individuals. Since resting-state activity builds the foundation of stimulus driven activity and might reflect a reinforcing process for the existing functional connectivity,Resting-state fMRI (rs-fMRI) was recorded both during EO and EC in four groups: (i) congenital cataract-reversal individuals (CC group) who had experienced visual deprivation since birth for up to 18 years due to bilateral dense congenital cataracts, (ii) congenitally permanently blind individuals (CB group) to indicate the adaptation of resting-state activity to congenital blindness, (iii) developmental cataract-reversal individuals (DC group) who had developed cataracts later during childhood to serve as a control for visual impairments and other effects related to a history of cataracts and cataract removal surgery and (iv) typical sighted controls (SC group).36 As aforementioned, we hypothesized to find overall higher ALFF in visual areas of both CC and CB individuals compared with the SC group. As previous studies have revealed retracted cross-modal activity in the CC group64 compared with what has been typically found in CB individuals,65 we predicted on the one hand that ALFF is overall lower in the CC group than in the CB group in early visual areas (including striate and extrastriate brain regions) and on the other hand that ALFF is modulated by EO versus EC in the CC group similarly as seen in normally sighted individuals. In particular, we hypothesized to replicate the typical decrease of ALFF in the EO compared with the EC condition in early visual cortex (striate cortex) and an increase in extrastriate and higher visual areas in both the CC and the SC group. Previous research on resting-state functional connectivity in CB individuals indicated lower connectivity between visual and both sensorimotor and auditory areas66 in particular when compared with sighted individuals with EC.67 Moreover, several studies have demonstrated a positive correlation between ALFF and functional resting-state connectivity.69 Thus, assuming incomplete recovery in CC individuals, we predicted a lower decrease of ALFF for the EO compared with the EC condition in auditory and sensorimotor areas compared with the SC group. We expected the CC versus SC group difference to be specific for the CC group, that is, we predicted to not finding the same pattern of results for the DC versus SC group comparison.Low-frequency oscillations (range: 0.01–0.08 Hz) of the BOLD signal were assessed as described in Yu-Feng et al.All participants were recruited at the LV Prasad Eye Institute (LVPEI) and from the local community of Hyderabad (India). Four groups were included: congenital cataract-reversal individuals (CC group), developmental cataract-reversal individuals (DC group), congenitally permanently blind individuals (CB group) and sighted controls (SC group).The original CC group consisted of 22 individuals. Three of them were excluded from the final analysis; one because of insufficient data due to premature termination of the scanning session (C7), a second due to a deprivation period of <3 months (C12) and in a third participant (C13) it turned out later that cataracts had not been dense. The final CC group consisted of 19 individuals . Mean Logarithm of the Minimum Angle of Resolution (logMAR) visual acuity in the better eye  was 0.86 (range: 0.30–1.80). One participant’s visual acuity could not be measured with the letter charts. Thus visual acuity was assessed with the maximal distance at which the participant was able to count fingers, which in this particular case was 1 m (which was translated to 1.80 logMAR). We only included participants with a history of bilateral dense congenital cataracts confirmed based on the information available in the medical records. In the classification process, additionally to the clinical diagnosis, we considered factors such as presence of sensory nystagmus and strabismus, absence of fundus view before surgery, and a positive family history. For detailed information about the participants, see The original DC group consisted of 16 individuals. Five participants (D12–16) were excluded from the final analysis. We were not able to come to an unambiguous classification and thus refrained from including these participants in either the DC group or the CC group. The final DC group consisted of 11 individuals  due to the lack of the EO condition, a second (CB11) due to a central cause of blindness and a third (CB12) due to non-congenital blindness. The final CB group consisted of nine individuals . For detailed information about the participants, see The SC group comprised 28 individuals . Nineteen of 28 sighted controls were matched in age and sex with the CC group . Eleven of 28 sighted controls were matched in age and sex with the DC group . Nine of 28 sighted controls were matched in age and sex with the CB group . All sighted control participants had normal or corrected to normal vision.All participants or their legal guardians (in case of minors) provided written informed consent and an assent (in case of minors) prior to taking part in the study. The subjects’ consent was obtained according to the Declaration of Helsinki. Participants and in case of minors, their legal guardians received a small compensation for the time of participation (e.g. lost wages) and for other expenses such as travel costs.The study was approved by the Local Ethics Board of the Faculty of Psychology and Movement Sciences, University of Hamburg, Germany, the Ethics Board of the German Psychological Society, as well as the Institutional Ethical Review Board of the LVPEI.2, interslice gap = 4 mm) in ascending order were acquired. Anatomical T1-weighted images, using 3D-spoiled gradient recalled (3D-SPGR) sequence  were additionally acquired for each subject.Data were acquired at a radiology clinic , with a 1.5 T GE Signa HDxt scanner. Resting-state fMRI scans were collected employing a gradient-echo echo-planar imaging (EP/GR) sequence. An 8-channel head coil was used . TRs varied slightly among the participants (range: 1950–2300) due to the participant's head size, body weight and height. This TR adjustment was built into the available GE protocol. Thirty-two  interleaved axial slices .Data preprocessing and analysis of ALFF was performed using the DPARSF software of DPABI V4.3.71 In the next step, the time series for each voxel were filtered (band-pass filtering: 0.01–0.08 Hz),46 to remove the effects of very-low-frequency drifts and high frequency noise. For a given voxel, a fast Fourier transform (FFT) 72 was used to convert the filtered time series to the frequency domain to obtain the power spectrum. The power spectrum was then square-rooted and averaged across the frequency band of 0.01–0.08 Hz at each voxel, which represents ALFF. Finally, the amplitudes (beta scores) of subject-level maps were transformed into Z-scores to create standardized subject-level maps for each participant in the EO and in the EC condition for the statistical analysis.First, basic screening of images  for each participant was performed to check for image quality. Exclusion criteria were signal loss due to susceptibility artefacts and cut off slices. No images were excluded due to this procedure. Then, all time points were transformed from echo-planar imaging (EPI) Digital Imaging and Communications in Medicine (DICOM) to Neuroimaging Informatics Technology Initiative (NIFTI). The first 10 time points were removed for signal stability. Next, a standard preprocessing pipeline was applied: All the acquired functional volumes were aligned to the first slice for EPI distortion and slice acquisition time (slice timing). The functional volumes were subsequently spatially realigned (using rigid body transformations to correct for head movements), spatially normalized to the standard adult East Asian template (MNI space), and smoothed with a 4 mm full width at half maximum Gaussian kernel. In the next step, linear trends were removed from the time series (detrend) using polynomial regressors. Structural images were segmented into grey matter, white matter (WM) and CSF. Subsequently, nuisance covariates were regressed out: Friston 24 head motion parameters, CSF and WM signals. The use of full nuisance regression including polynomial detrending in ALFF data optimizes the group-level analysis.73 a voxel-wise paired t-test was carried out in the group of 28 typically sighted individuals comparing the EO and the EC condition .74To determine whether we were able to replicate previous results assessed with ALFF in the EO compared with the EC condition in typically sighted individuals,ondition . The stat-tests were separately calculated for: (i) the SC group matched in sex and age to the CC group, (ii) the CC group, (iii) the CB group and (iv) the DC group .74To determine the brain regions with significantly higher and lower ALFF between the EO compared with the EC condition in each of the tested groups, voxel-wise paired DC group . The staZ-scores) was carried out. Three group models were run: (i) CC group versus SC group; (ii) DC group versus SC group; (iii) CB group versus SC group (Participants section). All interaction maps were corrected for multiple comparisons using the GRF correction with a threshold of voxel-wise P < 0.05, cluster-wise P < 0.025, corrected.75–77To assess group differences in ALFF between the EO compared with the EC condition, a 2 × 2 mixed effect model on standardized ALFF maps .74Since eyes open versus closed does not mean the same for the CB group as for the other groups and to identify whether the CC group shows different resting-state activity particularly in the EO condition (see hypotheses), the CC group and their matched SC group were separately compared in the EO and in the EC condition using voxel-wise t-tests . The anat-tests . In addi as implemented in the DPABI toolbox, was used to assign significant voxels to brain regions.Anonymized data and materials will be made available to the external scientists upon reasonable request to the corresponding author through data transfer agreements approved by the stakeholders, under stipulations of applicable law including but not limited to the General Data Protection Regulation (EU 2016/679).t-test was carried out . We found significantly higher ALFF in a cluster in left visual association areas , as well as in a cluster in the right visual association areas  and in a cluster in the left precuneus (BA7). In addition, ALFF was higher in the EO than in the EC condition in a cluster in the left frontal cortex . ALFF was significantly lower in the EO than in the EC condition bilaterally in sensorimotor and temporal (auditory) regions i.e. in clusters covering precentral  and postcentral gyrus (BA3), middle temporal gyrus, superior temporal gyrus and inferior temporal gyrus . In addition, ALFF was significantly lower in the EO than in the EC condition in a cluster in the left frontal regions  , lingual gyrus (BA18) and the middle occipital gyrus (BA19). Significantly lower ALFF in the EO compared with the EC condition was found in a cluster in the left cingulate gyrus (BA23) and in a cluster in the supramarginal (BA40) and postcentral gyrus   see .Z-scores) as the dependent variable. A significant interaction was found in a large cluster in right visual areas spanning the calcarine gyrus (BA17), lingual gyrus (BA18), middle occipital gyrus (BA19), and the fusiform gyrus (BA37) and in a cluster predominantly in the precuneus (BA7)  with the standardized ALFF maps , lingual gyrus (BA18) and the inferior and middle occipital gyrus (BA19). Significantly lower ALFF in the CC group than in the SC group was observed in a cluster in the right precuneus (BA7), and in clusters including parietal , temporal (BA22) and frontal regions  . Significantly lower ALFF in the EO compared with the EC condition was observed in a cluster in the right early visual areas i.e. in the calcarine gyrus (BA17) and lingual gyrus (BA18), and in a cluster in temporal regions (BA21). In addition, lower ALFF was found in a cluster in sensorimotor areas including the precentral (BA4) and postcentral gyrus   .Z-scores) as the dependent variable to test the interaction between group and condition. A significant interaction was found in a cluster in parietal cortex (BA7) (A 2 × 2 mixed effect model (DC group versus SC group x EO condition versus EC condition) was calculated with standardized ALFF maps (ex (BA7) ; Table 4We next compared ALFF between the DC and the SC group separately in the EO and in the EC condition with voxel-wise two-sample t-tests. In the EO condition, we observed significantly lower ALFF in the DC group than in the SC group in a cluster in parietal cortex, predominantly in the precuneus (BA7). We did not observe any region with significantly higher ALFF in the DC group compared with the SC group . In the In sum, ALFF patterns in the EO as compared with EC condition in the DC group resembled those of the SC group. Parietal cortex activity was lower in the DC than the SC group during EO.t-test was carried out. We found significantly higher ALFF in the EO compared with the EC condition in a cluster in frontal regions . Significantly lower ALFF in the EO compared with the EC condition was found in a cluster in parietal cortex (BA7)  .Z-scores) as the dependent variable. Significant interaction effects were observed in a cluster in parietal regions (BA7) and in a cluster in frontal regions   was carried out with standardized ALFF  ; Table 4t-tests. In the EO condition, we found significantly higher ALFF in the CB group than in the SC group in a cluster in the left visual areas spanning the middle occipital gyrus (BA19) and fusiform gyrus (BA37) and in a cluster in frontal cortex . We did not observe any regions with significantly lower ALFF for the CB group compared with the SC group . Significantly lower ALFF in the CC group than in the CB group was found in two clusters in parietal cortex   see . In the  40) see .In sum, compared with the CB group, CC individuals showed lower ALFF in a wide range of visual cortical regions in the EC condition.The goal of the present study was to identify whether the emergence of typical resting-state activity of the human brain, as the prerequisite of any task related processing, depends on experience during a sensitive period of early brain development. To this end, we tested to which degree BOLD resting-state activity in the low frequency range between 0.01 and 0.08 Hz during EO and EC recovered after a transient phase of congenital blindness due to congenital cataracts. Congenital cataract-reversal individuals (CC group) were compared with normally sighted controls (SC group) and to a group of congenitally blind humans (CB group). Developmental cataract-reversal individuals (DC group) served as additional control group. All groups were investigated in the same scanner, came from the same community and the groups were matched in age.73: ALFF was significantly higher in the EO than in the EC condition in visual association cortex and in parietal cortex. Moreover, ALFF was significantly lower in the EO than in the EC condition in sensorimotor and auditory cortices.First, we replicated the typical ALFF pattern in the SC groupImportantly, ALFF varied with EO versus EC in the CC group’s visual cortices as well: Similar to the SC group the amplitude of slow BOLD fluctuations was higher in the EO than in the EC condition. However, there were several group differences too: In the CC group’s early visual cortex, we did not observe a typical decrease of ALFF in the EO compared with the EC condition, instead ALFF was higher in this region in the EO than in the EC condition too. Moreover, in the EO condition visual cortex activity was overall higher in the CC than in the SC group. Furthermore, in the CC group an increase of ALFF in parietal cortex was not observed in the EO compared with the EC condition and thus was lower in the EO condition than in the SC group. Moreover, the typical decrease of ALFF for the EO compared with the EC condition was missing in auditory regions in the CC group and in sensorimotor regions the decrease was less extensive in the CC group than in the SC group. Finally, the CB group showed higher ALFF than SC individuals in both the EO and the EC condition, and higher ALFF than the CC group in the EC condition  individuals.Research in non-human primates showed that synaptic pruning in visual cortex is experience dependent and particularly affects the asymmetric, excitatory synapses, resulting in an experience dependent set-point for visual cortex excitability. Cortical thickness development runs parallel to the developmental trajectory of synaptogenesis.88 Non-human animal research has demonstrated that the elaboration of inhibitory neural networks is a hallmark of sensitive period plasticity.89 In fact, stabilization of inhibitory synapse and myelination ends the sensitive period.90Early visual cortex in sighted individuals is characterized by a high degree of inhibition, which results in a short time constant and thus the ability to process visual information at a fast rate.53–57 and in CB individuals higher gamma oscillatory activity.59 Alpha oscillatory activity has been considered to be an electrophysiological signature for the control of the excitatory–inhibitory (E/I) balance of neural circuits.91 In the present context, it is important to note that alpha oscillatory activity has been found to inversely correlate with ALFF49 whereas gamma band activity was found in monkeys to positively correlate with slow BOLD fluctuations.62 Thus, the higher posterior ALFF observed for the CC and the CB groups in the present study is consistent with reduced posterior alpha band activity in these groups and higher gamma band activity reported for CB individuals. All these findings converge to the hypothesis that overall excitation is enhanced in the visual cortex of CB and CC individuals. Moreover, the visually triggered BOLD signal seems to be correlated with changes in the glutamate level.92 In fact, there is evidence of higher glutamatergic93 and lower GABAergic activity in congenitally permanently blind individuals.94 Corresponding data in CC individuals are not available yet.Previous EEG studies in CB and CC individuals have repeatedly observed lower alpha oscillatory activityHowever, there was a crucial difference between the CC and the CB group. Higher ALFF was observed for the CC group than in the SC group only in the EO condition and in fact in the EC condition ALFF was lower in the CC than in the CB group. These group differences demonstrate that the visual cortical networks partially recovered in the CC group, that is, different resting-state activity levels were adopted as a function of whether or not light reaches the retina during rest.89 resulting in neural circuits95 which selectively respond only to certain input. Thus, we speculate that higher ALFF in the CC group in the EO condition indicates less selective processing and an impaired quick shutting down of visual-driven activation possibly due to a compromised intracortical inhibition. In fact, behavioural studies have shown longer lasting visual (motion) after-effects in CC individuals,22 which indirectly supports this speculation. By contrast, we hypothesize that higher ALFF in visual cortex in the CB group was predominantly due to higher spontaneous activity. Higher spontaneous activity in visual areas is an often reported finding in visually deprived non-human animals.97 In fact, a study in monkeys has observed a decrease of spontaneous activity in visual cortex after ending a phase of congenital lid suture.96 Thus, a decrease in spontaneous activity might explain the lower ALFF in visual areas in the CC than in the CB group during the EC condition and might explain as well indistinguishable ALFF of the CC and the SC groups in the EC condition. Lower spontaneous activity would be compatible with the idea of partial E/I balance recovery in visual cortex.Visual thalamo-cortical input excites pyramidal neurons in the granular layers of the cortex but in parallel entertains synapses to inhibitory interneurons, which allows a quick shutting down of excitation. From non-human animal research, it is known that these inhibitory circuits are shaped by experience and that they are stabilized by perineural networksIn sum, we suggest that higher slow BOLD fluctuations in CB and CC individuals might originate from a similar neural substrate, that is, not or less well attuned visual circuits. However, despite late availability of patterned visual input, the neural circuits in the CC group seem to had recovered to some degree too, such that spontaneous activity decreased. However, the fine-tuned neural (inhibitory) circuits which allow for a selective activation and quick shutting down of stimulus-driven activity might not have fully emerged, resulting in an enhanced and possibly longer-lasting excitation as a response to visual stimulation.8 Although sensorineural deafness might cause more extensive changes in E/I balance than visual deprivation due to cataracts which do not directly affect the sensorineural architecture, the same mechanism might contribute to cross-modal plasticity in blind humans. Here we argue that, visual entrainment of visual areas in the CC group might have enhanced the threshold for cross-modal activation and reduced spontaneous activity possibly via homeostatic plasticity mechanisms resulting in lower ALFF in the CC group compared with the CB group during EC.98Importantly, we interpret our results on slow BOLD fluctuations in the visual cortices of CC individuals as evidence for retracted cross-modal plasticity. ALFF in visual cortex was lower in the CC than in CB individuals and varied as a function of EO versus EC. In fact, in the context of cross-modal plasticity in deaf cats it has been argued that the higher level of excitation in their auditory cortex reflects a largely reduced threshold to allow for cross-modal activation.et al.99 reported a lower visual responsiveness in parietal area BA7 in monkeys who had been visually deprived for 7-11 months. In a follow-up study 1 year after the end of the deprivation period responsiveness to visual stimulation had further declined rather than increased, as would have been expected from restoring sight.100 Except one participant , all CC participants of the present study were scanned more than 1 year after cataract removal surgery. Here, we speculate that the observed lower resting-state activity in the CC individuals in parietal regions during EO might reflect a similar reduced regain of visual responsiveness in parietal cortex as observed by Hyvärinen et al.99 in non-human primates. Although parietal cortex is a multisensory region, many processes including multisensory spatial integration are visually dominated in sighted individuals. Thus, lower ALFF in parietal cortex in the CC compared with the SC group might indicate a lower visual influence on multisensory  processing. Recently, Smyre et al.101 reported an impaired multisensory detection and localization in dark reared cats. In fact, two behavioural studies have found a reduced visual impact on tactile spatial performance in congenital cataract-reversal individuals with a history of longer lasting visual deprivation.103In parietal cortex, we observed a lower ALFF in the CC than in the SC group in the EO condition. Hyvärinen 104 Crucially, such overall coupling between visual brain regions and both auditory and sensorimotor brain regions seems to be reduced in congenitally blind humans.67 Our new finding that resting-state activity in auditory regions is unaffected and in sensorimotor regions less affected by eyes opening in CC individuals might suggest, analogously to parietal cortex, a reduced impact of vision in multisensory processing. This idea is compatible with the previously reported lower lip reading specific activity in the superior temporal sulcus of CC individuals105 and the lack of audio–visual enhancements neither in this region18 nor in behaviour.106Altered multisensory processing is suggested by the third main result in the CC group too: Activity in auditory regions was, in contrast to the SC group (and the DC group) not lower during EO than during EC and in sensorimotor regions the decrease of ALFF in the EO as compared with EC was less robust in the CC group that is rather small compared with the SC group see . Functio107Finally, it has to be noted that similar group differences as found between the CC and the SC group were not observed for the DC group except the lower parietal cortex activity during EO. By contrast, the typical decrease in ALFF for resting-state activity with EO versus EC was highly robust in the DC group. Thus, a typical modulation of auditory and sensorimotor cortex activity by the visual system might crucially depend on connectivity elaborated in early brain development. By contrast, the lower impact of vision on parietal cortex activity condition might reflect a lower online weighting due to overall reduced reliability of visual input.Slow BOLD fluctuations indicating resting-state activity of neural circuits suggested a retraction of cross-modal plasticity after sight restoration in individuals with a history of blindness due to congenital cataracts. However, visual neural circuits seemed to be less tuned and activity thus was not as well-regulated as in normally sighted controls. This impairment might reflect remaining visual cortical circuit changes as a consequence of congenital blindness.A significant influence of the visual system on parietal areas, as well as the auditory system as typically found in sighted individuals had not recovered and in sensorimotor cortex not fully recovered in the congenital cataract-reversal individuals, suggesting a high influence of early experience on multisensory neural networks.Since resting-state brain activity builds the scaffold for task-related processing, we put forward the hypothesis that the incomplete recovery of typical resting-state activity patterns within the visual system and across sensory systems might contribute to the persisting visual and multisensory impairments after restoring sight in people with a congenital loss of pattern vision.fcac146_Supplementary_DataClick here for additional data file.


Median time from the arrival of the first-tier ambulance to successful airway management was 8 min for ETI performed by second tier and 4 min for SGA performed by the first or second tier (p = 0.001). No significant differences in the probability of good neurological outcome , overall survival , full neurological recovery  or sustained ROSC  were observed between ETI and SGA. Conclusions: Among the OHCA patients treated with trans-nasal evaporative intra-arrest cooling, the use of SGA was associated with a significantly shorter time to airway management and with similar outcomes compared to ETI. Background: Trans-nasal evaporative cooling is an effective method to induce intra-arrest therapeutic hypothermia in out-of-hospital cardiac arrest (OHCA). The use of supraglottic airway devices (SGA) instead of endotracheal intubation may enable shorter time intervals to induce cooling. We aimed to study the outcomes in OHCA patients receiving endotracheal intubation (ETI) or a SGA during intra-arrest trans-nasal evaporative cooling. Methods: This is a pre-specified sub-study of the PRINCESS trial (NCT01400373) that included witnessed OHCA patients randomized during resuscitation to trans-nasal intra-arrest cooling vs. standard care followed by temperature control at 33 °C for 24 h. For this study, patients randomized to intra-arrest cooling were stratified according to the use of ETI vs. SGA prior to the induction of cooling. SGA was placed by paramedics in the first-tier ambulance or by physicians or anesthetic nurses in the second tier while ETI was performed only after the arrival of the second tier. Propensity score matching was used to adjust for differences at the baseline between the two groups. The primary outcome was survival with good neurological outcome, defined as cerebral performance category (CPC) 1–2 at 90 days. Secondary outcomes included time to place airway, overall survival at 90 days, survival with complete neurologic recovery (CPC 1) at 90 days and sustained return of spontaneous circulation (ROSC). Results: Of the 343 patients randomized to the intervention arm , 328 received intra-arrest cooling and had data on the airway method ( Out-of-hospital cardiac arrest (OHCA) is a major public health concern, affecting approximately 275,000 individuals each year in Europe . The oveETI has been used by emergency medical services (EMS) since the 1970s . HoweverTargeted temperature management remains an important intervention that may influence survival with good neurological function among cardiac arrest patients . In partIn this sub-study of the PRINCESS trial, we aimed to compare the effect on neurologic outcome among OHCA patients that had received airway management with ETI versus SGA prior to trans-nasal evaporative intra-arrest cooling.We performed a post hoc sub-analysis of data from the PRINCESS trial, which is a multicenter randomized clinical trial that compared trans-nasal evaporative intra-arrest cooling to the standard ACLS in a bystander-witnessed OHCA  . Ethics We included bystander witnessed OHCA randomized to the intervention arm in the PRINCESS trial. The exclusion criteria were age ≥80 years; an etiology of cardiac arrest due to trauma, head trauma, severe bleeding, drug overdose, cerebrovascular accident, drowning, smoke inhalation, electrocution, or hanging; hypothermia at the time of evaluation; an anatomical barrier preventing the insertion of intra-nasal catheters; an existing do-not-attempt resuscitation order; known terminal illness; known or clinically apparent pregnancy; known coagulopathy ; need for supplemental oxygen; ROSC prior to randomization; and EMS response time  greater than 15 min. In this sub-study of the PRINCESS trial, we also excluded study participants who were randomized to the control group as well as study participants who were initially randomized to the intervention group but did not receive intra-arrest cooling. Patients were divided into two different treatment groups depending on the airway management technique used prior to trans-nasal evaporative cooling .2, but this was only recorded in a limited number of patients and not presented in this analysis. All study sites had two-tiered EMS systems where the first vehicle used bag mask ventilation only or SGA with bag-valve ventilation connected to the SGA prior to the arrival of the second tier. The second tier was manned by physicians or anesthetic nurses, trained in advance airway management including placing an SGA and endotracheal intubation. In addition, the intra-arrest cooling equipment was carried by the second tier. Thus, patients could have had the SGA placed by paramedics from the first vehicle and subsequently, cooling was started after randomization and application of trans-nasal evaporative cooling by the crew in the second tier. Among patients receiving bag mask ventilation by the paramedics, ETI was performed after the arrival of the second tier. The use of SGA could also be due to ETI being difficult to perform in the field. No data were collected on the number of intubation attempts or change in airway strategy. The confirmation of the tube was undertaken with end tidal COThe exposure of interest was defined as the type of airway management technique used prior to cooling. All of the study participants included in the PRINCESS trial were treated with advanced airway management prior to randomization using either ETI or an SGA ; patientThe RhinoChill™ device delivers a mixture of air or oxygen and a chemically inert cooling liquid (perfluorohexane) via nasal catheters directly into the nasal cavity, with the goal of primarily cooling the brain ,21,23. TNeurological outcome assessment was performed at 90 days via a structured telephone interview or during a follow-up appointment using the cerebral performance categories (CPC) scale . The priT-test, as appropriate. Group-wise differences in categorical variables were assessed using Pearson’s chi-squared test.Continuous variables were presented as means and standard deviations (SD) if normally distributed, or as medians and interquartile ranges (IQR) if not normally distributed. Categorical variables were reported as counts and percentages. We assessed the group differences in continuous variables using either the Mann–Whitney U-test or Student’s A range of factors may influence the decision of EMS personnel to use one advanced airway device or strategy over any other form of airway management. Therefore, we used propensity score matching to balance known confounding variables across the two treatment groups, in a manner reminiscent of that conducted in the work of Hasegawa et al. and McMullan et al. ,7. PropeWe used multiple imputation by chained equations (mice) to impute missing data, generating five imputed datasets . The anan = 63) with a laryngeal mask airway. Patients who were treated with SGA were older (p = 0.03) and had a higher BMI (p = 0.01) than patients who were treated with ETI , overall survival at 90 days , survival with complete neurologic recovery at 90 days  or hospital admission following sustained ROSC , as can be seen in p = 0.001 (p = 0.053 (The time until airway management was available for 276 out of 328 study participants (84%). The median time from EMS arrival at the scene until successful airway management was 8 (interquartile range 4–12 min) minutes in the ETI group and 4 min in the SGA group (interquartile range 2–7 min),  = 0.001 A. Howeve = 0.053 B. p = 0.045), overall survival , and survival with complete neurologic recovery at 90 days , as shown in the The time until the initiation of intra-arrest cooling therapy was associated with survival with good neurological outcome .The incidence of infections prior to hospital discharge was reported in conjunction with the PRINCESS trial by all participating centers . ApproxiIn this sub-analysis of the PRINCESS trial, we observed that SGA is feasible to use without any safety aspects reported prior to induction of trans-nasal evaporative cooling in OHCA and it significantly shortened the airway management time compared to ETI; SGA was not associated with any worsening in the gas exchange on arterial blood gas or with patient outcomes.Trans-nasal evaporative intra-arrest cooling is an emerging therapeutic option in the management of OHCA patients which has, as the only cooling method, been shown to be safe and effective in inducing intra-arrest cooling at the scene of the arrest. Although not currently part of routine medical practice, trans-nasal evaporative intra-arrest cooling therapy has already been implemented in some clinical settings for the treatment of OHCA. Furthermore, trans-nasal evaporative intra-arrest cooling will continue to be investigated in clinical trials. Therefore, it is important to establish the safety of advanced airway management in the setting of trans-nasal evaporative intra-arrest cooling. In this study, we observed a shorter time period to successful airway management in the SGA group compared to intubation, but no differences in terms of sustained ROSC, overall survival, neurological recovery, the prevalence of pneumonia, or arterial blood gas parameters.We observed that patients who were treated with SGA were on average older and had a higher BMI than patients who were treated with ETI. Although these imbalances were adjusted for in the subsequent analyses, we cannot dismiss the possibility of residual confounding due to one or several unmeasured parameters such as the lack of equipoise at the baseline including the expertise of centers. Therefore, we believe that there is a need for an external validation to confirm our results.Despite concerns regarding an increased aspiration risk following trans-nasal evaporative cooling, we did not find any significant differences between the ETI and SGA groups concerning the proportion of patients suffering from pneumonia or arterial blood gas parameters on admission to hospital. Despite a limited sample size, these findings further support our other results, which suggest that ETI and SGA have similar safety profiles in the setting of trans-nasal evaporative intra-arrest cooling.We observed a statistically significant difference between the ETI and SGA groups in the time elapsed until successful airway placement in the setting of trans-nasal evaporative intra-arrest cooling therapy. This result is in agreement with that of earlier studies on the topic of advanced airway management in OHCA ,11. HoweThis secondary analysis of the PRINCESS trial had several limitations. The study of two subgroups receiving different airway strategies within the intervention arm of the PRINCESS trial introduced a risk of selection bias. Although the treatment groups had similar baseline characteristics and propensity score matching was performed, the risk of residual confounding could not be eliminated. An additional limitation of this study is the fact that ETI and SGA could not be compared to bag-valve mask ventilation, as successful advanced airway management was specified as a requirement for inclusion in the PRINCESS trial . In addiIn this sub-study of the PRINCESS trial, a shorter time period to successful airway management was observed in the SGA group when compared to ETI. No differences in clinically relevant outcomes such as survival with good neurologic outcome and overall survival were observed between groups. This study might help to design future trials using trans-nasal evaporative cooling to minimize the time to induce cooling.


The primary aim of MULTI-MIBI is to assess feasibility of a multicentre study of 99mTc-sestamibi SPECT/CT against a reference standard of histopathology from surgical resection or biopsy. Secondary aims of the study include obtaining estimates of 99mTc-sestamibi SPECT/CT sensitivity and specificity and to inform the design and conduct of a future definitive trial.The incidence of renal tumours is increasing and anatomic imaging cannot reliably distinguish benign tumours from renal cell carcinoma. Up to 30% of renal tumours are benign, with oncocytomas the most common type. Biopsy has not been routinely adopted in many centres due to concerns surrounding non-diagnostic rate, bleeding and tumour seeding. As a result, benign masses are often unnecessarily surgically resected. 99mTc-sestamibi SPECT/CT (index test) compared with histopathology from biopsy or surgical resection (reference test). Interpretation of the index and reference tests will be blinded to the results of the other. Recruitment rate as well as estimates of sensitivity, specificity, positive and negative predictive value will be reported. Semistructured interviews with patients and clinicians will provide qualitative data to inform onward trial design and delivery. Training materials for 99mTc-sestamibi SPECT/CT interpretation will be developed, assessed and optimised. Early health economic modelling using a decision analytic approach for different diagnostic strategies will be performed to understand the potential cost-effectiveness of 99mTc-sestamibi SPECT/CT.A feasibility prospective multicentre study of participants with indeterminate, clinical T1 renal tumours to undergo Ethical approval has been granted (UK HRA REC 20/YH/0279) protocol V.5.0 dated 21/6/2022. Study outputs will be presented and published nationally and internationally.ISRCTN12572202. However, the literature on 99mTc-sestamibi SPECT/CT remains limited to single centres reporting relatively few tumours. We have recently reported on a pump-priming pilot study in the UK.99mTc-sestamibi SPECT/CT in the diagnostic pathway for renal tumours andGiven the excellent performance characteristics of Study methods are reported with reference to Standard Protocol Items: Recommendations for Interventional Trials Checklist (SPIRIT)2099mTC-sestamibi SPECT/CT in adults (n=50) with solid, enhancing clinical renal tumours (2–7 cm) on cross-sectional imaging. The study design is summarised in A prospective, multicentre study to assess the feasibility and diagnostic performance characteristics of 99mTc-sestamibi SPECT/CT can recruit successfully. Secondary aims are to assess patient and clinician acceptability, refine inclusion/exclusion criteria, sample size requirements and determine clinician training needs for 99mTc-sestamibi SPECT/CT interpretation.The primary aim of the study is to evaluate whether a multicentre diagnostic test evaluation study of The study objectives are to determine:99mTc-sestamibi SPECT/CT prior to surgery or biopsy, including those from under-represented and underserved groups?Will patients consent to have a What factors influence patient’s decisions to participate?99mTc-sestamibi SPECT/CT?What are the perceptions of clinicians and patients of 99mTc-sestamibi SPECT/CT?What barriers and facilitators are there for adoption of 99mTc-sestamibi SPECT/CT within the NHS?What is the potential cost-effectiveness of using 99mTc-sestamibi SPECT/CT?What are the minimally acceptable criteria (MAC) for the sensitivity and specificity of 99mTc-sestamibi SPECT/CT?Is it feasible to train nuclear medicine clinicians across the UK, including those serving under-represented and underserved communities, to interpret The study outcomes are as follows:Primary outcomeRecruitment rate.Secondary outcomesSensitivity and specificity of MIBI to detect benign lesions in this study.Define the MAC for MIBI-kidney to be adopted in clinical practice, to inform the design and parameters of the future definitive clinical trial.Interobserver variability and training requirements in the interpretation of MIBI-kidney .Patient and clinician perceptions of utility and experience of MIBI-kidney scans and training.The evidence requirements for a cost-effectiveness analysis.The study will be conducted in 3–6 NHS hospitals in England.99mTc-sestamibi and those unwilling or unable to undergo the study procedures.Consecutive patients discussed at specialist multidisciplinary team meetings will be screened for eligibility over a planned 15-month recruitment period. The inclusion criteria for entry to the study are adult patients (≥18 years) of any gender with a clinical T1 indeterminate solid renal tumour (2–7 cm) on cross-sectional imaging, willing and able to provide informed consent. Patients will be required to have surgery or renal tumour biopsy planned as part of their standard clinical care. Patients entering watchful waiting or active surveillance pathways without histologic diagnosis will be excluded. Other exclusion criteria will include cystic tumours, pregnant and breastfeeding patients, those with a known allergy to 99mTc-sestamibi SPECT/CT from international experts at the beginning of the recruitment period. The training will include a lecture on 99mTc-sestamibi SPECT/CT principles, ‘hands-on training’ supported by experienced faculty and a precourse and postcourse assessment.Nuclear medicine clinicians involved in the study will receive study-specific training on the interpretation of 99mTc-sestamibi will be injected intravenously in a single bolus, 75 min before SPECT/CT acquisition of the abdomen with the superior extent of the field-of-view set to the top of the liver dome. CT and SPECT image acquisition will follow manufacturer instructions and local experience. At minimum, we suggest that participating centres have SPECT/CT systems with the following specifications: at least two-slice helical diagnostic CT scanner, available low-energy all-purpose or low-energy high-resolution collimator, gamma camera or digital detector elements appropriate for 140-kEv photopeak acquisition and manufacturer-derived iterative reconstruction that includes scatter and attenuation correction.900 MBq of 99mTC-sestamibi SPECT/CT scans will be transferred for central review at the lead site  and discordant reports resolved by discussion and consensus. Local site clinicians will report a subset of studies a second time at the end of the recruitment period to allow assessment of intrarater reliability.The reporting clinician will document a qualitative assessment of the tumour as avid, non-avid or indeterminate on reconstructed SPECT/CT images, blinded to clinical information and the result of the histopathology reference test. A spherical region of interest will be drawn to measure maximum uptake in attenuation-corrected images within (a) the tumour and (b) the ipsilateral renal parenchyma. A ratio of maximum uptake between the tumour and normal renal parenchyma will be calculated. All Histopathology from the final surgical resection specimen is considered the ‘gold standard’ diagnostic test to determine renal tumour subtype. It is worth noting that although biopsy allows for histological diagnosis, questions remain about the accuracy of this technique for determining the precise histology of a renal tumour, mostly relating to an approximately 15% non-diagnostic rate of this procedure99mTc-sestamibi SPECT/CT result. Pathology slides/images will be exported for central review by a specialist uro-oncology pathologist and archiving at the lead site .Histopathological reporting of both biopsy and surgical samples will be performed by qualified pathologists at collaborating sites in accordance with the current WHO classification system for renal tumours,99mTC-sestamibi SPECT/CT in the diagnostic pathway for renal tumours. Data from the feasibility phase will be used to inform the design and sample size of the definitive trial. We will aim to recruit 50 patients from 3–6 centres. This sample size will allow us to assess if 80% (95% CI 70% to 90%) of approached patients agree to undergo the study scan. Additionally, this sample size will have sufficient power to detect if there is a significant difference in the estimates of sensitivity between our study population and those reported in the literature. A sample size of 40 patients would achieve 81% power to detect a sensitivity of 0.65 (representing an estimate outside the lower end of the 95% CI for sensitivity from the literature) using a two-sided binomial test at the 5% two-sided alpha level. A 20% inflation to 50 patients, will allow for possible dropouts and other methodological challenges.The aim of this study is to assess the feasibility of a multicentre study of Qualitative data obtained from semistructured interviews  with patients, carers and staff will be combined with documentary analysis  and will be used to inform within trial decision-making processes via a rapid feedback evaluation approach.99mTC-sestamibi SPECT/CT with other management scenarios. In addition, a survey will be conducted—informed by a rapid review of survey instruments reported in the published literature to capture the acceptability of interventions in clinical trials, to provide insights into the barriers or facilitators to patient decision-making and determine the degree of acceptability of 99mTC-sestamibi SPECT/CT.The findings from the interviews and documentary analysis will be used to develop a discrete choice experiment to gain an understanding of preferences for trial participation and how participants trade-off different attributes of 99mTc-sestamibi SPECT/CT will be estimated by generating 2×2 tables for both avid and non-avid qualitative assessment, and relative radiotracer uptake ratio >0.6 and ≤0.6 for external validation of a predefined threshold from the literature.99mTC-sestamibi SPECT/CT will be calculated in terms of sensitivity, specificity and predictive values along with their 95% CIs. The prevalence of renal oncocytoma and other histology subtypes will be calculated with a 95% CI.Diagnostic accuracy of 99mTC-sestamibi SPECT/CT results, for example, due to technical failure, will be reported. The proportion of valid but inconclusive results will also be reported, and their impact on estimates will be assessed by including them as either test positive or test negative in sensitivity analyses. This is to inform how 99mTc-sestamibi SPECT/CT might be used in the diagnostic pathway. If intended as a replacement test for histopathology, a valid but indeterminate 99mTc-sestamibi SPECT/CT would be considered non-avid to avoid misclassifying malignant tumours as benign. If, however, 99mTc-sestamibi SPECT/CT were to be used as a triage test, where avid tumours undergo confirmatory biopsy, then an indeterminate test could be considered avid to reduce the risk of surgery for benign pathology. The proportion of patients who do not complete the study schedule defined in the protocol will be calculated.Inconclusive test results will be reported.We will assess inter-rater and intrarater agreement using percentage agreement and Gwet’s first-order agreement coefficient.26We do not anticipate the need to adjust for diagnostic drift for the reference test, given the short study duration. However, if current pathologic guidelines for renal neoplasia are updated during the course of the study archived samples will be rereviewed and reported according to the latest guidelines.99mTC-sestamibi SPECT/CT in the evaluation of patients presenting with an indeterminate renal mass.Health economic modelling will be used to understand the potential cost-effectiveness of Patients have empiric surgery (current standard-of-care).Patients undergo tumour biopsy, those consistent with cancer have surgery and those with benign histology have active surveillance.99mTc-sestamibi SPECT/CT, those with a ‘cold’ scan (suggestive of cancer) have surgery and those with a ‘hot’ scan (suggestive of benign tumour) have active surveillance.Patients undergo 99mTc-sestamibi SPECT/CT, those with a ‘cold’ scan have surgery, and those with a ‘hot’ scan have a confirmatory biopsy (MIBI would be likened to a triage test to select patients for biopsy for tissue confirmation before embarking on active surveillance).Patients undergo The model will be populated with evidence from trial and published literature.99mTc-sestamibi SPECT/CT visit and thereafter by telephone or email according to participant preference, as shown in Case report forms (CRFs) in paper and electronic format will be trialled. The CRFs will not bear the participant’s name or other directly identifiable data. The participants’ study ID will be used for identification purposes. Study-related procedures will be carried out during the baseline routine clinical visit, 99mTc-sestamibi SPECT/CT scan. The decision to withdraw will be recorded in the CRF and medical notes. Participants withdrawing prior to 99mTc-sestamibi SPECT/CT will be replaced. If following 99mTc-sestamibi SPECT/CT, the participant states they do not wish to participate in scheduled follow-up (EQ-5D-5L completion), or deviate from the protocol, then data already collected will be kept and analysed. These patients will not be replaced.A participant may withdraw their consent to participate at any time prior to the Baseline data items will include the following:Baseline demographics .Baseline blood test results .Baseline imaging , contrast-enhanced CT or MRI of the abdomen).Renal tumour characteristics .Quality of life questionnaire (EQ-5D-5L).The following data on resource use will be collected at the time of the interventionDuration of visit to nuclear medicine department.Adverse events (AEs) during and immediately post-MIBI-kidney.The following data will be collected at postintervention follow-up by telephone or emailAEs following MIBI-kidney.Quality of life questionnaire (EQ-5D-5L).After participation in the trial participants will continue follow-up as per standard care.Patient and public involvement (PPI) has been central to the project concept and design. A prestudy PPI focus group informed the trial protocol and plain English summary. An online PPI survey received 231 responses and indicated 90% would be willing to participate in the proposed study. In addition to the qualitative workstream, PPI representatives from Kidney Cancer UK will form a study support group, meeting at regular intervals throughout the trial to provide advice and input on any trial challenges and developing/approving dissemination materials.99mTc-Sestamibi has been used for cardiac and parathyroid imaging globally for decades and is known to be a safe radiopharmaceutical. The radiation exposure from one MIBI-kidney scan is 14 mSv, equivalent to approximately 5 years of average UK background radiation.All AEs, whether related or unrelated to MIBI-kidney, will be documented in the patient’s notes, study CRF and the AE log. The AE log will be sent to the Sponsor  at least once per year. Incidental clinically significant abnormalities identified on MIBI-kidney will be recorded as AEs and communicated to the referring clinician and patient. All serious AEs will be recorded on an SAE form and reported to the Sponsor and relevant REC within 15 working days of the chief investigator becoming aware of the event.Investigators and sites will permit trial-related monitoring, audit, REC review and regulatory inspection(s) and provide access to required data and documents.Ethical approval for this study has been granted (UK HRA REC 20/YH/0279). Protocol amendments will be promptly disseminated to Sponsor, investigators and trial steering committee members. The study is recorded on the trial registration website. The trial involves the administration of unsealed radioactive substances. An Administration of Radioactive Substances Advisory Committee certificate has been granted (AA-3990).Study outputs will be presented at national and international conferences and published in peer-reviewed journals. Patient representatives will be involved in output dissemination to the public individual trial participants via study newsletter.


IMI2-PainCare-BioPain-RCT2 is one of four similarly designed clinical studies aiming at profiling a set of functional biomarkers of drug effects on specific compartments of the nociceptive system that could serve to accelerate the future development of analgesics. IMI2-PainCare-BioPain-RCT2 will focus on human spinal cord and brainstem activity using biomarkers derived from non-invasive neurophysiological measurements.This is a multisite, single-dose, double-blind, randomized, placebo-controlled, 4-period, 4-way crossover, pharmacodynamic (PD) and pharmacokinetic (PK) study in healthy subjects. Neurophysiological biomarkers of spinal and brainstem activity  and the R2 component of the blink reflex) will be recorded before and at three distinct time points after administration of three medications known to act on the nociceptive system , and placebo, given as a single oral dose in separate study periods. Medication effects on neurophysiological measures will be assessed in a clinically relevant hyperalgesic condition , and in a non-sensitized normal condition. Patient-reported outcome measures  will also be collected; and blood samples will be taken for pharmacokinetic modelling. A sequentially rejective multiple testing approach will be used with overall alpha error of the primary analysis split between the two primary endpoints, namely the percentage amplitude changes of the RIII area and N13 amplitude under tapentadol. Remaining treatment arm effects on RIII, N13 and R2 recovery cycle are key secondary confirmatory analyses. Complex statistical analyses and PK-PD modelling are exploratory.The RIII component of the flexion reflex is a pure nociceptive spinal reflex widely used for investigating pain processing at the spinal level. It is sensitive to different experimental pain models and to the antinociceptive activity of drugs. The N13 is mediated by large myelinated non-nociceptive fibers and reflects segmental postsynaptic response of wide dynamic range dorsal horn neurons at the level of cervical spinal cord, and it could be therefore sensitive to the action of drugs specifically targeting the dorsal horn. The R2 reflex is mediated by large myelinated non-nociceptive fibers, its circuit consists of a polysynaptic chain lying in the reticular formation of the pons and medulla. The recovery cycle of R2 is widely used for assessing brainstem excitability. For these reasons, IMI2-PainCare-BioPain-RCT2 hypothesizes that spinal and brainstem neurophysiological measures can serve as biomarkers of target engagement of analgesic drugs for future Phase 1 clinical trials. Phase 2 and 3 clinical trials could also benefit from these tools for patient stratification.2019-000755-14).This trial was registered on 02 February 2019 in EudraCT ( The order of the items has been modified to group similar items  has been extensively described previously  see  see  see  see  see  see [The plans to promote participant retention and complete follow-up are not the same as descibed in the protocol for our related trial [IMI2-PainCare-BioPain-RCT3] see [The data management is the same as described in the protocol for our related trial [IMI2-PainCare-BioPain-RCT3] see  see  see [This will be determined at the end of the project period of IMI-PainCare.The composition of the coordinating centre and trial steering committee are similar to the ones described in the protocol for our related trial [IMI2-PainCare-BioPain-RCT3] see  see [Any modifications to the protocol which may impact on the conduct of the study, potential benefit of the subject or may affect subject’s safety, including changes of study objectives, study design, subject population, sample sizes, study procedures, or significant administrative aspects will require a formal amendment to the protocol. Such amendment will be approved by the IEC and prior to implementation and notified to the health authorities in accordance with local regulations .A final report integrating trial results will be prepared. The Principal Investigator will provide the competent authority/ies and relevant IEC(s) with a summary of the trial results in accordance with applicable regulatory requirements.The results of this trial will be publicly disclosed (EudraCT). The results (or parts thereof) of this trial will be published according to the Grant Agreement and Consortium Agreement of IMI-PainCare (grant No 777 500).IMI2-PainCare-BioPain-RCT2 is one of four RCTs aiming at validating biomarkers of drug effects on nociceptive processing using the same trial design and IMPs. The design of IMI2-PainCare-BioPain-RCT3 on electrophysiological brain biomarkers has been published before [The RIII component of the flexor reflex is elicited by electrical stimulation of nociceptive A-delta fibers. It is sensitive to altered signal processing in ventral horn, dorsal horn, and their descending controls from the brainstem. The N13 component of the somatosensory evoked potential is elicited by electrical stimulation of tactile Aß fibers that converge with nociceptive fibers on dorsal horn wide-dynamic range nociceptive neurons. It is sensitive to altered dorsal horn signal processing and its modulation by descending controls. The R2 component of the blink reflex is also elicited by electrical stimulation of tactile Aß fibers, but the signal pathways that determine its recovery cycle include brainstem reticular formation that also processes nociceptive inputs.According to the sequentially rejective multiple testing design, two primary endpoints will be tested simultaneously at alpha/2; these are pairwise comparisons of one medication vs. placebo for one post-medication time point and one readout variable that are most likely to be significant, based on published literature . Three key secondary endpoints are assessed, given that alpha levels were passed successfully from one or both of the primary endpoints or from one or both of the other key secondary endpoints. All efficacy analyses will be done on data pooled across participating centers; efficiency of this multicenter study approach in human volunteers will be assessed by comparing across-center vs. within-center variability of the various readout variables.In addition to the analysis of the individual RCTs, biomarker results and pharmacokinetic and pharmacodynamics analyses will be compared across trials and to preclinical studies and will possibly try to establish a latent variable model of underlying mechanisms at peripheral, spinal, and brain levels. Pain ratings will be included as another readout variable, and questionnaires on psychological traits predictors  will also be analyzed as potential predictors of pain and analgesia. By identifying specific mechanisms within different compartments of the nociceptive system in healthy volunteers, these same quantitative neurophysiological biomarkers, if well validated against clinically meaningful outcomes in patients, may be used for patient stratification and enrichment in later clinical trials, as encouraged by the recent EMA/CHMP/970057/2011 Guideline. This will accelerate the development of novel analgesics in several ways: preclinical prediction will be improved by using translatable readouts across species; clinical Phase 1 trials will benefit from biomarkers of target engagement and from human surrogate models predictive of clinical efficacy; clinical Phase 2 and 3 studies will benefit from tools for patient stratification.This manuscript is based on protocol version 4.0 dated 12/06/2019. Recruitment started in Spring 2021 and is expected to be completed in March 2022.