Participants are exposed to an experimental trauma and evoked intrusive thoughts in daily life are monitored for three consecutive days. Functional magnetic resonance imaging is used to assess neural habituation to fearful faces and fear learning (conditioning and extinction) prior to trauma exposure. The results reveal a significant interaction between loneliness and sex such that loneliness is associated with more intrusions in men, but not in women. A similar pattern emerges at the neural level, with both reduced amygdala habituation to repeated fearful faces and amygdala hyperreactivity during the conditioning of fear signals in lonely men. The findings indicate that loneliness may confer vulnerability to intrusive memories after trauma exposure in healthy men and that this phenotype relates to altered limbic processing of fear signals.Loneliness exacerbates psychological distress and increases the risk of psychopathology after trauma exposure. However, it is still unclear whether a lack of social connectedness affects trauma‐related intrusions and the neural processing of fear signals. Moreover, it is uncertain, whether loneliness plays a different role in women and men. A prestratification strategy is used and  High‐lonely males exhibit amygdala hyperreactivity in fear conditioning and fear habituation processes during functional magnetic resonance imaging in contrast to high‐lonely women. The same pattern emerges in intrusive thought formation after experimental trauma exposure, highlighting loneliness as a risk factor for detrimental trauma coping in men related to altered limbic processing of fear signals. Loneliness can be considered as the social equivalent to hunger or pain to meet social needs and has been associated with increased mortality, resembling risk factors like obesity or smoking. Furthermore, loneliness is closely linked with various psychiatric disorders such as substance abuse, depression, and anxiety disorders. Importantly, loneliness also constitutes a risk factor for developing post‐traumatic stress disorder (PTSD) following a traumatic experience. In fact, loneliness predicts future PTSD and is predicted by past PTSD symptoms, indicating a bidirectional relationship between PTSD and social connectedness.Loneliness, defined as the discrepancy between desired and actual social connectedness,1 is a Intrusions are defined as involuntarily spontaneous memories of the distressing incident, mainly experienced as visual forms of mental imagery. The lifetime prevalence of PTSD varies substantially between sexes, with women being twice as likely to develop PTSD than men. Current neurocircuit models of PTSD highlight dysfunction of the amygdala–hippocampus complex as a core mechanism underlying the persistence of intrusive memories. Modern trauma‐focused psychotherapies for treating intrusions and other PTSD symptoms often include an exposure‐based intervention to reduce fear responses. Mechanistically, this decrease in fear responses can be achieved by both fear extinction and fear habituation. The former is characterized by a progressive decrement of a conditioned fear response (CR) when a conditioned stimulus (CS) is repeatedly presented in the absence of an aversive unconditioned stimulus (UCS) with which it has previously been paired, while the latter is based on repeated exposure to the (imagined) UCS. In fact, both fear extinction and habituation recruit overlapping forebrain structures including the amygdala.PTSD is a debilitating and frequently chronic condition characterized by intrusive thoughts about the traumatic experience as a key symptom.13 Int On a neural level, increased reactivity in the amygdala, hippocampus, insula, and anterior cingulate cortex during trauma exposure predicts increased intrusive thoughts. Interestingly, neural processing during fear extinction has also been linked to intrusion frequency in an experimental trauma paradigm and reduced extinction capacity predicts PTSD development. Furthermore, women reported more intrusive symptoms following the trauma paradigm than men, and this sex difference was related to peritraumatic responding and slowed extinction learning in women. Likewise, women showed a sustained amygdala response to negative evocative images relative to men.The experimental trauma paradigm is a widely used and reliable method to evoke intrusions by showing traumatizing film footage in a controlled laboratory setting.21 On  Given a crucial role of oxytocin in safety learning and a reduced oxytocin reactivity to positive social interactions in people experiencing loneliness, this raises the intriguing possibility that loneliness influences intrusions after trauma exposure by modulating self‐disclosure and amygdala‐related fear processing. Furthermore, a recent large‐scale study indicated a higher prevalence of loneliness in men than in women, and a growing number of studies reported sex‐specific effects of loneliness. For instance, loneliness was associated with more pronounced within‐network coupling of the default network in men than in women, and brain volume effects in the limbic system were linked to the frequency and intensity of social contact in a sex‐dependent manner. Surprisingly, however, the impact of loneliness on fear conditioning/extinction and fear habituation as well as the possible moderation by sex remain unclear. Therefore, this study aimed to examine loneliness‐associated neurobiological risk factors for intrusive thoughts in an experimental prospective study design.We previously found that strong trauma disclosure reduces intrusions and alters amygdala functional connectivity following trauma exposure only in individuals with heightened concentrations of the hypothalamic peptide oxytocin after intranasal administration.28 Giv Given previous findings about sex differences in the effects of loneliness and the formation of intrusive memories, we explored sex as a moderator variable.To this end, we recruited a prestratified sample of 82 healthy volunteers assigned to either a high‐lonely and low‐lonely group to test how loneliness interacts with sex to influence the neural processing of fear signals and the formation of intrusive thoughts. During functional magnetic resonance imaging (fMRI), subjects completed an emotional face‐matching task to assess neural responses to fearful faces and the habituation of these responses. In addition, we used a classical Pavlovian fear conditioning and extinction paradigm with two social and nonsocial stimuli one of each paired (CS+) and one without (CS−) an electric shock. To explore hormonal group differences, blood samples were taken before the fMRI session. Subsequently, we probed psychological , physiological , and hormonal (oxytocin) stress responses during an experimental trauma paradigm. The trauma paradigm consisted of a 24‐min‐long aversive video to mimic trauma exposure. Furthermore, evoked intrusions and communication behavior were monitored via online diaries during three consecutive days after trauma exposure. The total number of intrusions, trauma disclosure , intrusion stress ratings and the level of amygdala reactivity in neural fear processing served as primary study outcomes. We hypothesized that lonely individuals would exhibit more pronounced responses to the experimental trauma film and experience more intrusions. Furthermore, we expected to observe loneliness‐dependent hyperreactivity to fearful faces and fear‐conditioned stimuli in the amygdala, as well as changes in functional connectivity in a network responsible for fear processing.37 Giv22.1ps < 0.02; shown in Table ps < 0.03). In addition, across groups, women reported having more social support than men  = 5.12, p = 0.03, ηp2 = 0.06). There were no significant interactions between sex and loneliness in psychiatric symptoms and social network quality . Besides the expected sex differences, we found a significant sex*loneliness interaction in estradiol levels  = 7.60, p = 0.01, ηp2 = 0.11), showing that high‐lonely women exhibited higher estradiol levels than low‐lonely women at the fMRI session (t(16.55) = 2.62, pcor = 0.04, d = 0.87; shown in Table Table Psychiatric symptoms were measured via questionnaires during a screening interview. In addition, blood samples were taken before fMRI scanning. High‐lonely subjects reported more depressive symptoms, alexithymia, childhood maltreatment, social interaction anxiety, and subjective stress compared to low‐lonely participants  = 9.36, p < 0.01, d = 1.06) and reported high arousal (76.87 ± 23.53) induced by and low valence (9.35 ± 16.16) of the trauma film. Neither dissociative symptoms nor valence and arousal were affected by loneliness or sex . Subjects showed a decrease in positive affect  = 67.88, p < 0.01, ηp2 = 0.49; shown in FigureF = 139.58, p < 0.01, ηp2 = 0.66; shown in Figure F = 154.91, p < 0.01, ηp2 = 0.68; shown in Figure F = 4.44, p = 0.04, ηp2 = 0.06), such that lonely individuals displayed higher baseline state anxiety ratings (t(76) = 4.42, pcor < 0.01, d = 1.02) than low‐lonely individuals, but state anxiety significantly increased in both groups (high‐lonely: t(41) = 8.98, p < 0.01, d = 1.39; low‐lonely: t(33) = 7.99, p < 0.01, d = 1.37).An experimental trauma paradigm was conducted after the fMRI. Dissociative symptoms, positive and negative affect, state anxiety, and saliva oxytocin were measured before and after trauma exposure via questionnaires and saliva samples. Physiological stress markers  were measured during trauma exposure. After trauma exposure, subjects showed dissociative symptoms  = 13.57, p < 0.01, ηp2 = 0.18; shown in Figure F = 133.96, p < 0.01, ηp2 = 0.67; shown in Figure F(2130) = 3.39, p = 0.04, ηp2 = 0.05; post hoc t‐test: t(72) = 4.05, pcor < 0.01, d = 0.47; shown in Figure Physiologically, there was an increase in the skin conductance level  = 8.53, p = 0.01, ηp2 = 0.10). However, our results revealed a significant interaction between loneliness and sex  = 5.57, p = 0.02, ηp2 = 0.07), such that loneliness was associated with more intrusive memories in men but fewer intrusions in women  = 3.97, pcor < 0.01, d = 1.39), while there was no significant sex difference in high‐lonely individuals (t(44) = 0.39, p = 0.70, d = 0.12). Furthermore, analysis of the desire to talk about the trauma movie yielded a pattern consistent with intrusion effects  = 5.62, p = 0.02, ηp2 = 0.08; shown in Figure t‐tests revealed that low‐lonely women showed an increased desire to talk in contrast to low‐lonely men (t(32) = 2.66, pcor = 0.046, d = 0.91). In addition, high‐lonely subjects talked less about the movie  = 9.85, p < 0.01, ηp2 = 0.17; shown in Figure ps > 0.05).The primary study outcomes, intrusive thoughts, trauma disclosure , and intrusion stress ratings were measured via online questionnaires on three consecutive days after trauma exposure. Across loneliness groups, women experienced more intrusions than men  = 12.72, pFWE = 0.04; Fearful Block 1 > Fearful Block 3; shown in Figure r(76) = −0.22 p = 0.049; Fearful Block 1 > Fearful Block 3). In addition, a significant sex*loneliness interaction was observed for the left amygdala habituation to all faces which was reduced in high‐lonely women compared to high‐lonely men and the opposite pattern was evident in low‐lonely individuals  = 17.53, pFWE = 0.01; Faces Block 1 > Faces Block 3).fMRI scanning was conducted before trauma exposure and consisted of an emotional face‐matching task and a fear conditioning and extinction paradigm. In the emotional face‐matching task, participants had to match two simultaneously presented pictures at the bottom with a target picture presented at the top of the screen. In the fear conditioning and extinction paradigm, participants had to press a button before the UCS to indicate if they believed that they would receive an electric impulse. Responses were acquired with an fMRI compatible response grip system to measure reaction times and contingency ratings. Amygdala reactivity in both paradigms served as primary study outcome. There was no significant interaction effect of sex and loneliness on the neural response to fearful faces per se, but amygdala habituation was characterized by sex*loneliness interactions. Habituation to fearful faces in the right amygdala was reduced in high‐lonely men compared to high‐lonely women, while this pattern was reversed in low‐lonely individuals xyz: −34, −52, −58, k = 108, pFWE = 0.01; Fearful Block 1 > Fearful Block 3; shown in Figure Further habituation analyses revealed a sex*loneliness interaction in functional connectivity. High‐lonely men showed increased right amygdala coupling with the left superior parietal lobe in the habituation process to fearful faces  = 54.79, p < 0.01, ηp2 = 0.46). Likewise, a significant time*condition interaction  = 49.23, p < 0.01, ηp2 = 0.44) for the contingency ratings showed reduced shock expectations in the course of the EXT (extinction) task  = 5.41, p = 0.02, ηp2 = 0.08). There were no significant interactions with the factor sociality or sex * loneliness interactions in the EXT phase.Successful conditioning was evident in higher contingency ratings of the CS+ compared to the CS− in the second half of the COND (conditioning) task  and condition : 2.6 including the amygdala  = 12.62, pFWE = 0.046; COND CS+ > CS− > EXT CS+ > CS−; shown in Figure xyz: 30, 4, −20, F = 14.37, pFWE = 0.02; COND CS+ > CS−). High‐lonely men exhibited higher amygdala activation than high‐lonely women, while this effect was reversed in low‐lonely individuals.In the COND phase, the CS+ elicited activations in a fear conditioning network35 incxyz: −44, 28, −16, k = 98, pFWE = 0.02; COND CS+ > CS− > EXT CS+ > CS−; shown in Figure We also observed a loneliness*sex interaction in the functional connectivity of the amygdala during fear conditioning/extinction. High‐lonely men exhibited a stronger coupling between the left amygdala and orbitofrontal cortex , indicating that the loneliness‐related amygdala changes may be related to salience rather than arousal effects. The loneliness‐related amygdala activation changes during habituation and conditioning in men were evident across social and nonsocial stimuli, which is in line with previous studies suggesting that loneliness fosters hypervigilance for threat cues. Interestingly, high‐lonely women compared to high‐lonely men showed decreased left amygdala habituation to all faces, but we found decreased right amygdala habituation in response to social threat cues in high‐lonely men. Impaired right amygdala habituation has also been previously identified as a neural phenotype of patients with borderline personality disorder and trauma exposure. The absence of amygdala hypervigilance in high‐lonely women could be driven by hormonal factors with high‐lonely women showing increased estradiol levels compared to low‐lonely women in our sample. Estradiol administration improved extinction recall after fear extinction, and low levels of estradiol in women were linked to increased fear network responses to trauma films. However, the observed sex differences cannot be completely explained by hormonal factors either because women reported more intrusions across loneliness groups despite having higher estradiol levels than men. It is conceivable that the content of the trauma film was more distressing for women than men, but we did not detect significant sex differences in the acute stress responses, and a previous study found no evidence for an interaction between sex and intrusive memories induced by different trauma films. The unwillingness of men to admit loneliness and higher stigmatization of men who express feelings of loneliness might have contributed to the observed sex differences. Taken together, our data suggest that loneliness has a sex‐specific impact on the way threat cues are processed during fear conditioning and fear habituation.Furthermore, high‐lonely men exhibited heightened amygdala responses to the CS+ and functional connectivity with the orbitofrontal cortex during conditioning compared to low‐lonely men. Both amygdala and orbitofrontal cortex activity have been frequently linked to CS+/CS− differentiation during fear learning.60 NevThe present study had several limitations. First, our sample consisted of women with and without hormonal contraception. Although we did control for the use of hormonal contraception and measured hormonal blood levels to control for menstrual cycle‐related hormone changes, future studies are warranted to further delineate the hormonal basis of sex differences in the effects of loneliness. Second, the experimental trauma paradigm is widely used and well established to explore the neurobiological mechanisms underlying acute and prolonged trauma responses, but further clinical studies in a real‐life setting are required to gauge whether our findings can be extrapolated to patients with trauma exposure. Third, while we found sex‐specific associations between loneliness and amygdala reactivity consistently in two separate fMRI tasks, the results should be interpreted cautiously. The results were based on region of interest (ROI) analyses with lenient small‐volume corrections and the effects sizes were small. Replication studies are warranted to test the robustness of these effects. Fourth, we used a prestratification approach and we only included individuals without current psychological disorders. This way we were able to exclude possible confounding effects due to current psychotherapy or pharmacotherapy. Nevertheless, given that loneliness is closely linked to mental health, the absence of loneliness‐related effects in women may also reflect increased resilience in the group of high‐lonely women.Collectively, our results provide evidence that loneliness may confer vulnerability to increased intrusive thoughts in men following an experimental trauma. In addition, high‐lonely men were characterized by an increased desire to talk about the trauma film and reduced actual trauma disclosure. This phenotype relates to altered limbic processing driven by amygdala hyperreactivity during fear conditioning and habituation. Based on these findings, secondary prevention strategies should take sex differences in loneliness into account and focus on improving the social connectedness of high‐lonely men to mitigate the sequelae of traumatic experiences.4 High scorers (high‐lonely) were defined by a score above or equal to 50 , while low scorers (low‐lonely) were defined by a score of 25 or below . In total, 4515 participants completed the UCLA LS online questionnaire and a clinical screening interview was conducted with 97 subjects fulfilling the above‐mentioned loneliness criteria. The final sample consisted of 82 healthy subjects (mean age ± standard deviation (SD): 26.39 ± 5.83 years) assigned to either a high‐loneliness (n = 47 (20 women)) and a low‐loneliness control group (n = 35 (18 women)). In accordance with our preregistration, every subject included in the final sample was aged between 18 and 46 years and had no current physical or psychiatric disorder as assessed via self‐disclosure and the Mini‐International Neuropsychiatric Interview, no current psychotherapy, no current psychotropic medication, no illicit drug use in the previous four weeks, and was eligible for magnetic resonance imaging scanning . All participants gave written informed consent. The study was approved by the institutional review board of the medical faculty of the University of Bonn (number 248/16) and carried out in compliance with the latest revision of the Declaration of Helsinki.The present study used a quasi‐experimental design with a sample of prestratified healthy volunteers scoring high or low on the revised UCLA Loneliness Scale (UCLA LS).69 Hig and a well‐established emotional face‐matching paradigm. All magnetic resonance imaging (MRI) data were acquired using a 3T Siemens TRIO MRI system  with a Siemens 32‐channel head coil. Following fMRI acquisition, the participants completed an experimental trauma paradigm. To measure trauma disclosure and intrusive thoughts, subjects completed online diaries during the following three days after trauma exposure. Saliva samples were collected before the fMRI scan as baseline measure, and before and after the experimental trauma paradigm to measure oxytocin levels. In addition, blood samples were taken before the fMRI scan to measure the levels of gonadal steroids including estradiol and testosterone, as control variables. For a detailed list of the questionnaires and neuroendocrine parameters see the Supporting Information.In screening sessions, medical history and psychiatric symptoms were assessed  and houses as nonsocial control stimuli. Stimuli were presented in three blocks for every condition , with each block consisting of five trials. Participants had to match the face identity .The first fMRI paradigm consisted of an adapted version of a well‐established emotional face‐matching paradigm.75 Sub In the COND phase, subjects were shown four different pictures  and two houses ). One social and one nonsocial pictures were designated as fear‐associated CS (CS+) and the other picture of each category as safety signal (CS−). The choice of the picture that served as CS+ was counterbalanced within each group . Each stimulus was presented 16 times during the COND and EXT experiments. The trials were interleaved with an interstimulus interval (ISI) that was jittered between 5 and 7 s (mean: 6 s). In 75% of CS+ trials, subjects received an electric impulse (the UCS) 4 s after stimulus onset. The electric impulses were delivered by a Biopac System . To identify a stimulation intensity that was uncomfortable, but not painful, participants rated different intensities beforehand in an adaptive process (see the Supporting Information) while lying in the MRI on a scale from 0 to 100 . The stimulation intensity was set to reflect a rating of 60. In addition, the Biopac system measured electrodermal activity (EDA) and respiration during the experiment. After the COND phase, participants were informed that there would be another round of the same experiment. No electrical impulses were administered in the EXT phase. In both phases, participants had to press a button before the UCS to indicate if they believed that they would receive an electric impulse . For a detailed description of the data acquisition, preprocessing, and analyses of both tasks (see the Supporting Information).The COND/EXT paradigm was an adapted version of a Pavlovian fear conditioning paradigm described by Eckstein et al.73 In  valence , arousal  as well as state anxiety were measured prior and after the experimental trauma paradigm. The participants completed online intrusion diaries at home in the evening during three consecutive days following trauma exposure. For details about data collection and preprocessing, see the Supporting Information.Participants were seated in front of a Tobii TX300 binocular eye‐tracker  with a 23 in display to measure pupil sizes as the outcome indicating physical arousal during the movie alongside EDA. To evoke intrusive thoughts, participants were confronted with a 24‐min‐long movie clip derived from the movie "I spit on your grave" showing the multiple rape of a young woman by a group of men. EDA data were measured with a Biopac MP150 system. Positive and negative affect, dissociative symptoms ,76 valThe participants completed online intrusion diaries at home in the evening for three consecutive days after trauma exposure. In the intrusion diary, the participants stated the number of intrusions  and rated the distress caused by each of these intrusions on a visual analogue scale ranging from 0 (no distress) to 100 (extreme distress). Furthermore, participants were asked to state the time spent on talking about the trauma video (in minutes) and their desire (0 = no desire to 100 = strong desire) for trauma disclosure.pcor) post hoc t‐tests were calculated using SPSS 25  to examine changes in intrusive thoughts (sum of the three consecutive days following the trauma exposure), trauma disclosure , group differences in psychiatric symptoms and psychological as well as physiological and hormonal responses to the trauma exposure with the between‐subject factors of sex  and loneliness . Mixed‐design ANOVAs for contingency ratings included the additional within‐subject factors task  and condition . Additional mixed‐design ANOVAs for the COND/EXT paradigms included the between‐subject factors of sociality  and time . Partial eta‐squared and Cohen's d were calculated as measures of effect size.The primary outcomes included the number of intrusions and blood oxygen level‐dependent signal changes during fear learning and the processing of fearful faces. Fear habituation was assessed in an exploratory analysis. Other outcomes recorded were the psychological and physiological stress markers after trauma exposure and skin conductance response during fear conditioning. Mixed‐design analyses of variance (ANOVAs) and Bonferroni‐corrected (http://www.fil.ion.ucl.ac.uk/spm). On the first level, data were modeled using a fixed‐effects model. On the second level, the main contrasts of interest were compared between groups using a full factorial model with the two factors of loneliness and sex. Analyses were conducted using anatomically defined regions of interest (ROIs), including the amygdala, derived from the WFU PickAtlas . P values smaller than 0.05 after familywise error correction for multiple testing (pFWE) based on the size of the ROI  were considered significant. Whole‐brain analyses were calculated across groups for task‐validation . In addition, generalized psychophysiological interaction analysis was conducted to assess functional connectivity by using the CONN toolbox 18.a  with the same preprocessed data, ROIs, regressors, and contrasts that were used in the SPM analyses. Parameter estimates of significant contrasts were extracted using MarsBar  and further analyzed in SPSS 25 ). Pearson correlations between parameter estimates of significant ROI clusters and intrusive thoughts were calculated.To analyze the fMRI data, a two‐stage approach was used as implemented in the MATLAB toolbox  SPM12 (Wellcome Trust Center for Neuroimaging, London, UK; testing pFWE basedThe authors declare no conflict of interest.M.M. and D.S. designed the experiment. M.M., J.N., D.S., and J.D. conducted the experiments. M.M., B.S.‐W., and D.S. analyzed the data. M.M. and D.S. wrote the manuscript. All authors read and approved the manuscript in its current version.Supporting InformationClick here for additional data file.


Chemotherapy-induced peripheral neuropathy (CIPN) is a major issue for many cancer patients receiving chemotherapy agents such as platinums, taxanes, and vinca alkaloids, leading to dose reductions or early cessation of treatment  and inteThe putative mechanism for CIPN due to platinum chemotherapy is believed to be oxidation injury, secondary to mitochondrial damage . Other tWe commend Pfeiffer and colleagues  for 2 weP = .045).POLAR-A and POLAR-M were terminated early by the data safety monitoring board because of hypersensitivity reactions in the CaM-treated participants (12 of 336 CaM vs 2 of 246 placebo). This resulted in smaller than planned sample sizes with no participant completing either study according to the protocol and a lack of longer-term efficacy or safety data. To optimize the statistical power of the analyses, data from the 2 studies were pooled, and the authors used a modified intention-to-treat analysis that included 88% of the 434 participants randomly assigned, with data for the primary endpoint for 351 participants. In POLAR-A, participants were randomly assigned to intravenous CaM 5 μmol/kg (n = 120) or placebo (n = 119), and in POLAR-M, to CaM 2 μmol/kg (n = 54), CaM 5 μmol/kg (n = 55), or placebo (n = 57), but only 29%-38% received 12 cycles of study treatment. The trial findings did not indicate a beneficial effect of CaM on the primary endpoint of CIPN at 9 months after starting oxaliplatin, using the first 4 items of the patient-reported Functional Assessment of Cancer Therapy–Gynecologic Oncology Group–Neurotoxicity subscale questionnaire. Furthermore, moderate-to-severe CIPN at 9 months was greater in the participants receiving CaM 5 μmol/kg compared with placebo  and phase II trial in 173 participants with metastatic CRC randomly assigned to receive placebo (n = 60) or CaM at 2 (n = 57), 5 (n = 45), or 10 (n = 11) μmol/kg. (The 10 μmol/kg dose was reduced to 5 μmol because of greater non-neurotoxic grade 3 adverse events) , with iPfeiffer et al.  propose Overall, this was a negative study where people randomly assigned to the active treatment had more CIPN but also received more oxaliplatin. Clearly, CaM is not ready for “prime time” due to highlighted safety concerns combined with the perceived lack of benefit and potentially more neurotoxicity. There were challenges evaluating the efficacy of the therapy, meaning that a benefit from CaM, if it existed, might not have been identified. We concur with Pfeiffer et al.  that theNo funding was used for the writing of this editorial.Role of the funder: Not applicable.Disclosures: JV and CT have no disclosures to declare.Author contributions: JV: conceptualization, writing—original draft, writing—review & editing. CT: conceptualization, writing—original draft, writing—review & editing.


Red-green colour vision deficiency (CVD) affects ~ 4% of Caucasians. Notch filters exist to simulate CVD when worn by colour vision normal (CVN) observers (simulation tools), or to improve colour discrimination when worn by CVD observers (compensation tools). The current study assesses effects of simulation (Variantor) and compensation (EnChroma) filters on performance in a variety of tasks. Experiments were conducted on 20 CVN and 16 CVD participants under no-filter and filter conditions . Participants were tested on Ishihara and Farnsworth-Munsell 100 hue tests, CVA-UMinho colour discrimination and colour naming tasks and a board-game colour-sorting task. Repeated-measures ANOVAs found Variantor filters to significantly worsen CVN performance, mimicking protanopia. Mixed-model and repeated-measures ANOVAs demonstrate that EnChroma filters do not significantly enhance performance in CVD observers. Key EnChroma results were replicated in 8 CVD children (Ishihara test) and a sub-sample of 6 CVD adults (CVA-UMinho colour discrimination and colour naming tasks) for a smaller stimulus size. Pattern similarity exists across hue for discrimination thresholds and naming errors. Variantor filters are effective at mimicking congenital colour vision defects in CVN observers for all tasks, however EnChroma filters do not significantly compensate for CVD in any. The human retina has three types of cone cells responsible of colour vision: L, M and S cones . CVD observers suffer from abnormalities or absences in the cone photopigments. Anomalous trichromacy includes individuals with three photopigment types: S, M and M’ photopigments in protanomaly and S, L and L′ in deuteranomaly2, where M′ and L′ are variants of M and L photopigments. These variants differ in their peak spectral sensitivities, being 12 nm or less away from peaks for M or L photopigments3. In red-green dichromacy, one photopigment type is missing: L in protanopia; M in deuteranopia1. These abnormalities impact on CVD observers, reducing their colour discrimination performance. While colour vision normal (CVN) observers differentiate around 2 million colours7 this number is reduced theoretically to as few as 7%  in dichromatic CVD observers8, although for them, real world natural colours reveal higher degrees of discriminability9. A recent study simulating relative loss of colour discrimination in red-green CVD individuals highlights large differences between the impact of the CVD on perception in dichromacy versus in anomalous trichromacy, even when anomalous trichromacy is severe10.Red-green colour vision deficiency (CVD) is a genetic condition that affects 8% of males and 0.5% of females in the Caucasian population11, the existence of a third cone photopigment12, chromatic aberration13 and short-wave sensitive or “S” cone intrusion, spatial inhomogeneity of macular pigmentation, or differences in pigment optical density between central and parafoveal cones14. For foveal stimuli (sizes < 3°), compensation mechanisms in red-green dichromats might arise from gains and nonlinearities applied to receptor signals15. In anomalous trichromats (see Bosten3 for a review), optical density differences16, post-receptoral compensation19 and dynamic colour signals provided by changing illumination, or caused by eye movements as coloured light passes through different densities of macular pigment20, have been proposed. Other compensation mechanisms have been demonstrated in human blue-cone monochromats, such as variations of input to chromatic channels from rods21, or for cephalopods with a single photoreceptor, chromatic aberration can differentially affect intensity across wavelength22. In addition, observers with CVD could find benefit from a fifth photoreceptor type in peripheral vision: intrinsically photosensitive retinal ganglion cells (IPRGs)23.Several compensation mechanisms might explain improved discriminability in CVD individuals, above that calculated theoretically from spectral sensitivity curves. More acute use of luminance cues often associated with hue differences, is possible both for red-green dichromats and anomalous trichromats. Dichromat compensation is especially salient when foveal and peripheral vision act together (for stimuli over 3°). Large-field trichromacy could be due to rod intrusion26, education29, co-ordination of clothing25, recognition of natural colour or skin abnormality25, cooking25 and discrimination of rival teams when watching or playing sport25. A CVD diagnosis may prevent or impede access to some careers such as aviation, rail or marine transport, police forces, armed forces, fire services and electrical and electronic engineering30 and CVD observers have been found to more likely undertake technical studies, rather than traditional university degrees25. Past debate existed about the need to prevent protans from obtaining commercial driving licenses26. CVD observers might also face difficulties in carrying out some duties in professions such as medicine34, in which case those with CVD need to be made aware of potential difficulties and compensate for them.Despite availability of these many potential compensation mechanisms, the impact of CVD still reduces perception of a wide range of colours. The consequences of CVD impact on daily activities such as driving35 and (2) Can we improve colour discriminability in CVD observers to be like that in CVN observers? Simulation tools exist that use computerised algorithms (software) or notch filters (glasses) to allow CVN observers to discriminate colours in images like CVD observers might. Compensation tools exist in the form of notch filters (glasses), which aim to improve colour discriminability in CVD observers. Notch filters selectively alter light entering the eye from different parts of the spectrum. Depending on the wavelengths affected, a filter can potentially simulate a certain CVD type (simulation filters), or provide new luminance or chromatic cues to individuals with CVD, to enhance colour discrimination performance36 (compensation filters).Aiming to improve lives of CVD observers, two recurrent questions in colour vision research arise. They are: (1) What do the colour blind (or CVD) see?37 showed that Variantor filters mimic protanopia when colours (and their luminance properties) are compared along red-green pseudo-achromatic confusion axes . However this study did not test Variantor filter effects on discrimination of colours away from pseudo-achromatic confusion axes, or for other tasks, such as clinical colour vision tests, colour naming, or performance in real-world tasks.The Variantor filter  is the most known and widely available simulation filter. The logic underlying designs of simulation filters is to decrease colour contrast in normal trichromatic vision. To simulate protanopia and deuteranopia, the filter must block energy from parts of the spectrum that match the missing CVD cone type, so that the cone type simulated as missing in CVN observers, will have little or nothing to respond to. To simulate protanomaly and deuteranomaly, the filter must attenuate rather than block, certain parts of the spectrum. Degree and selectivity of attenuation would depend on severity of anomaly simulated. That is, if the spectral sensitivity curve of the abnormal cone is severely affected, attenuation will be high; if only slightly shifted, attenuation will be low. A previous study3. If the filter notch is positioned ideally, greater gain would be found for more severe anomalies. However, notch filters also modify luminance contrast across the spectral range, potentially giving new discrimination cues to both anomalous trichromats and dichromats. There have been several attempts to design compensation filters previously, but they have not demonstrated real compensation effects41. A decade ago, Moreland et al.41 assessed 43 commercially available filters without finding any evidence of a filter that improves discrimination along the L–M axis. A few studies44 have assessed EnChroma glasses , which incorporate more recently developed compensation filters. None of these studies have found statistically significant improvements in colour vision in results obtained for classical clinical colour vision tests22, or for digital versions of Ishihara and Farnsworth-Munsell 100 hue test (FM100)23, the Colour Assessment and Diagnosis test (CAD)44, or a 21 stimulus  colour naming task42. In one study45, it was predicted that EnChroma filters can result in gamut expansions near the protan and deutan confusion axes for spectrally broadband stimuli, while there might be no effect, or gamut contraction, for spectrally narrowband stimuli. Recent research seems to indicate that long term exposure to EnChroma filters might have positive impact on colour perception in CVD observers46. Specifically, Werner et al.46 found heightened chromatic responses after long-term exposure to EnChroma filters in observers with anomalous trichromacy. The authors suggest that “modifications of photoreceptor signals activate a plastic post-receptoral substrate that could potentially be exploited for visual rehabilitation”.Compensation filters, or CVD aids, are notch filters that aim to reshape M and/or L cone spectra to increase differences between peak spectral energies, so increasing colour contrast and providing more distinguishable responses arising from photoreceptors. This feature is particularly useful for anomalous trichromats, whose main characteristic is excessive overlap of spectral sensitivity profiles between M and M′ cones or L and L′ cones 43, we include intensive laboratory-based colour discrimination (CVA-UMinho) and colour appearance tasks (CVA-UMinho modified for colour naming), which use matching hue angles so that results can be compared, and a sorting task using real-world board-game pieces. This study aims to contribute to the debate about the value of using simulation/compensation filters for purposes of simulating CVD in CVN observers, and of enhancing colour perception in CVD observers. The impact of filters is assessed for coloured stimuli away from, as well as including pseudo-achromatic axes, and for computerised and non-computerised tasks of colour discrimination, colour appearance and real-world performance.The current study aims to measure and describe the effects of simulation (Variantor) filters in CVN observers and compensation (EnChroma) filters on colour perception in CVD and CVN observers. In addition to two clinical colour vision tests (Ishihara and FM100) also used in previous studies47 prior to experiments being conducted. CVD participants were 3 females and 13 males with an average age of 26.19 ± 5.92 years. Colour vision was screened using the Ishihara test and diagnosed using Rayleigh matches on the Oculus HMC anomaloscope. The CVD group consisted of 4 protanopic, 4 protanomalous, 4 deuteranopic and 4 deuteranomalous participants. Fifteen adult CVN (5 CVN from Variantor condition and 10 CVN from the EnChroma condition) and 9 adult CVD participants were recruited from the Cambridge campus of Anglia Ruskin University (ARU). Five adult CVN (from the EnChroma condition) and 7 adult CVD participants were recruited from the Braga campus of the University of Minho. In addition, data from 8 CVD children, who performed the Ishihara test only, with and without EnChroma filters are included to provide results from CVD observers less knowledgeable about both the Ishihara test and the impact of coloured filters. Anomaloscope testing found this group to consist of 2 protanomalous, 1 deuteranomalous and 5 deuteranopic participants . Child participants were recruited from a study about spatial vision involving colour conducted at a primary school in Cambridge (United Kingdom) and from the Anglia Vision Colour Investigations unit at the University Eye Clinic at ARU. Participants were excluded prior to experiments being conducted if visual acuity with best refractive correction was significantly worse than expected for age , or presence of any ocular abnormality or disease. One adult participant and one child participant were excluded due to the presence of systemic and ocular  disease. Only inherited binocular CVD conditions were eligible to take part. Participants performed all tasks binocularly.Research data for this study were collected in the United Kingdom, at Anglia Ruskin University (ARU) and in Portugal, at the University of Minho (UMinho). Twenty colour normal (CVN) and 16 red-green colour defective (CVD) adult participants performed experiments under two conditions: no-filter and filter (Variantor or EnChroma). The colour normal participants were 13 females and 7 males with an average age (± SD) of 23.35 ± 4.60 years. Five CVN participants  performed tasks with Variantor filters. Fifteen CVN participants  performed tasks with and without EnChroma filters. Normal colour vision was confirmed with the Ishihara testEthics approval was obtained from the Faculty of Science and Engineering Research Ethics Panel  at ARU, and the University of Minho Ethics Committee for Research in Life and Health Sciences  in line with the ethical principles of Helsinki declaration of 1975. All participants, as well as parents/guardians in the case of children, were provided with verbal and written explanations about the experiments. Written informed consent (and assent in the case of the child) was obtained before experiments were conducted.48 . The Variantor filter is said by their developers to “help you to understand what the world looks like to colour deficient people. You can easily see the confusing colour combinations just by wearing Variantor”49. The intended purpose of the Variantor filter is to provide its users with a colour impairment comparable to the one experienced by people with a red-green colour vision defect. The EnChroma filter is said by the developers to be “an optical assistive device for enhancement of colour discrimination in persons with colour blindness”48 that “gives those with colour blindness the ability to see more of the broad spectrum of bright colour most of us take for granted”50. The CX-65 lens was designed to enhance colour perception for indoor tasks under standard indoor lighting, including computer games and television. Variantor and EnChroma do not indicate the specific type of defect that they simulate or compensate for. We measured transmittance characteristics of these filters three times  and the compensation filter, EnChroma CX-65Whenever the simulation or compensation filters were needed to perform a colour discrimination task, they were worn over the top of the observer’s habitual refraction, which was checked to be optimal by an optometrist. As per EnChroma instructions, participants had a 10-min adaptation period prior to testing for both Variantor and EnChroma filters. Participants performed the tasks with and without filter (Variantor/EnChroma) in counterbalanced order across task and filter conditions. Colour discrimination tasks were carried out on the British adult participants with and without compensation/simulation filters twice, on different days, to gain an estimate of repeatability variance magnitude.The effect of filters was assessed for five colour vision tasks. Two were conventional clinical colour discrimination tests (Ishihara and FM100), two were computerised colour vision discrimination and naming tasks, and the final task was a sorting task with coloured board-game pieces. All tests are described below.The clinical colour vision tests and the sorting task, which required an external light source, were performed in a JUST Normlicht LED box  set to a metameric D65 at 100% brightness with uniform grey walls and an approximately uniform spatial distribution of the light source at Anglia Ruskin University. In Portugal a custom-built light box, with Munsell N5 paint  and a D65 simulator light source , was used. The dotted black line in Fig. 47 edition in Anglia Ruskin University and 199252 edition in University of Minho) was performed with the pseudoisochromatic test plates 1–25 placed inside the light box and following manufacturer instructions47. The participant was seated 75 cm from the plates and had 4 s to read each plate. The fail criterion was more than 3 errors excluding misreadings30. Protan and deutan deficiencies were further classified as “strong” or “mild” according to scoring instructions for responses to plates 22–25 of the test52.Standard versions of the Ishihara Test for Colour Deficiency  was used at both institutions. One of the four boxes of caps was placed under D65 lighting with caps in random order. Standard test instructions were given to the participant so that there was no time limit imposed on performing the task, and it was stated that accuracy was more important than speed in completing it. As indicated to participants in advance of taking the test, a 2-min reminder was provided so that they were aware of the length of time it was taking to complete each box. After each participant was content with the order of caps within a box, the task was repeated with the next box. Results were recorded and scored using Farnsworth–Munsell Test Scoring Software . The following parameters were scored for further analysis: (1) The total error score (TES), using a cut-off point of 100 for diagnosis of a CVD53, (2) the square root of TES (SQR), (3) the angle (previously known as mid-point), which identifies the primary axis of colour confusion, or orientation of the cap arrangement, (4) the confusion index (C-index), which describes the severity of the colour loss, a value of 1 corresponding to a perfect arrangement (> 1 indicating errors), (5) the similarity index (S-index), which identifies the specificity, polarity or randomness, a value of 1.28 corresponding to a perfect arrangement (> indicating stronger polarity) and (6) the time (s) it took to complete the entire test.A standard edition of the Farnsworth Munsell 100 Hue Test (FM100)54.The colour vision assessment task by the University of Minho  is a computer-based colour discrimination task, in which a coloured square target must be located against a grey background. The neutral grey background during stimulus presentation consisted of static luminance noise discs of variable diameter. The luminance of each disc was randomly selected from a predefined luminance range, to preclude colour discrimination judgements being based on luminance cues. A coloured square target appeared either to the left or the right side of the screen, the chromaticity of which was added to the grey background discs. The participant’s task was to identify the position of the square target, which had a different hue to the background. Target hue saturation commenced at 100% and followed a staircase paradigm until the chromaticity appeared to match that of the background for the participant. Below threshold level, the participant would be able to identify the position of the target only by chance. This task is based on similar principles to that of the Cambridge Colour Test (CCT) and The Colour Assessment and Diagnosis (CAD) test, however all parameters could be modified according to researcher needs. This task was also modified for the colour naming experiment. Further details of the CVA-UMinho colour vision discrimination task can be found elsewhere2), the same space-averaged luminance used for the background during stimulus presentation and in between stimulus trials. A subsample of 3 CVN and 6 CVD participants  also performed the CVA-UMinho discrimination task with a smaller stimulus square (1° size). The smaller size was tested as it was expected that the poorer resultant discrimination, more reliant on red-green mechanisms of foveal vision, might provide greater opportunity for compensation filters to demonstrate improvement.In these experiments, both at Anglia Ruskin University and the University of Minho, the CVA-UMinho stimuli were presented on a CRT colour computer screen  expressed on the CIE 1976 UCS chromaticity diagram. For each hue, a colour discrimination threshold was estimated by computing the Euclidean distance between the selected point and the neutral point. Thresholds were determined after 4 staircase reversals, or after presenting the stimulus for 25 trials, whichever occurred first. The last 15 trials contributed to threshold estimation where a minimum of 2 reversals had occurred.Sixteen hues were tested, 6 corresponding to hues on dichromatic  confusion lines. The other 10 hues were spaced approximately equally around the neutral point (the grey uniform background), which was CIE illuminant D65  or Portuguese58 . For clarity, in this paper, English equivalents are reported. Colour names were audio-recorded and mapped to the actual hue presented. “Hit scores” were computed using modal and loose criteria60. For the modal criterion, a response was deemed correct if it corresponded to the modal response of CVN participants not wearing any filter, for that stimulus condition. For the loose criterion, a response was deemed correct if at least one CVN not wearing any filter, gave the same name for that stimulus condition. Results reported in the paper are obtained using the modal criterion. Data for loose criterion are included in the supplementary material.The CVA-UMinho task was modified to present 5° chromatic squares on the neutral grey luminance noise background (and 1° squares for the participant subsample), for the same 16 hues , but for this experiment squares were drawn close to the centre of the screen to reduce positional uncertainty of the stimulus for naming. Saturation was fixed at 100%, 66% and 33%. Each hue-saturation combination was presented 3 times, which resulted in 144 trials (16 hues × 3 saturations × 3 presentations). Participants were instructed to name aloud each square presented, with one of the 11 basic colour terms Settlers of Catan 5th Edition61 and two sets (green and brown) from Catan Expansion—5–6 Player62 (Catan GmbH and Mayfair Games Inc) were presented  from the For almost all statistical analyses carried out in this project, participant data were grouped either as from CVN or CVD groups. This is because the literature produced about Variantor and EnChroma filters suggests that filters simulate, or compensate for, congenital red-green colour vision deficiency (CVD). This grouping does not affect overall outcomes see “”. For alBritish adult participants performed each task twice to gain information about the repeatability of our measures. Two types of analyses were performed. First for the Variantor filters in the CVN group, repeated measures ANOVAs were conducted for repeat  and filter  within-subject factors for different task measures. Second, for the EnChroma filters, three-way mixed-model ANOVAs were conducted with repeat  and filter  as within-subject factors, and participant group  as the between-group factor . For the sorting task, a separate three-way repeated measures ANOVA was conducted on the CVD group with within-subject factors of repeat , sorting task  and filter .Ps > 0.175). There were significant main effects of filter and participant group, as well as significant interactions with other factors in these analyses, however as these effects were also found in the main analyses to follow (using the average of the two runs), these are reported below. Means and standard errors across the two runs are provided in Table S1 in the supplementary information.Analyses of both Variantor and EnChroma data did not reveal any significant effect of repeat on Ishihara (error scores), FM100 (TES), CVA-UMinho discrimination thresholds, CVA-UMinho naming scores or sorting task results, nor any significant repeat interactions with other factors  = 256.32, P < 0.001, η2 = 0.99) was found. The mean error score without filter was 0.50 ± 0.35, whilst with Variantor filters it was 18.25 ± 0.78. All CVN participants were classified as such without any filter. When wearing Variantor filters, they were all diagnosed as “strong” protan, according to the Ishihara scoring instructions.A one-way repeated measures ANOVA with filter  as the within-subjects factor was conducted to test for the effect of Variantor filters on Ishihara error scores  as the within-subjects factor were conducted to test for the effect of Variantor filters on six FM100 indicators  = 87.94, P < 0.001, η2 = 0.96) and hue  = 11.23, P < 0.001, η2 = 0.74). The mean discrimination threshold without filter was 0.0028 ± 0.00032 whilst with Variantor it was 0.0088 ± 0.00088, so Variantor filters led to a significant worsening of discrimination thresholds in CVN participants. There was also a statistically significant interaction between filter and hue  = 14.96, P < 0.001, η2 = 0.79) so that the effect of Variantor filters depended on the hue. Tukey (HSD) post-hoc comparisons revealed that significant differences were found (filter vs no-filter) for hues near the protan axes of confusion and two neighbouring hues only  and hue (16 hues tested) was conducted to test the effect of Variantor filters on discrimination thresholds measured using the CVA-UMinho task with 5° stimuli Fig. a. This a2 and minimum saturation of 33%), some BCTs were never given as a response, . Variantor filters worsened modal hit scores for all 6 BCTs used by between 10% for brown, to 82% for blue , we calculated hit and error scores (in percent) per BCT using the modal criterion. Confusion matrices (11 × 11 BCTs) were created for CVN participants with and without Variantor filters and these are pictorially represented in Fig. F = 128.46, P < 0.001, η2 = 0.97; mean hit score without filter was 82.22 ± 3.39%, mean hit score with Variantor was 30.35 ± 5.21%—see red bars in Supplementary Fig. A one-way repeated measures ANOVA with filter  as the within-subjects factor was conducted on naming hit scores. This analysis revealed a significant main effect of Variantor filters , as three of the five participants had zero errors with and without Variantor filters. When proportional changes in time taken for sorting into groups of colour versus groups of shape for each participant  = 10.12, P = 0.033, η2 = 0.72; no-filter factor of 2.08 ± 0.24; Variantor, factor of 2.92 ± 0.28). Absolute time scores for the colour sorting task when Variantor filters were worn were significantly longer (268.80 ± 38.91 s) than when they were not  = 19.82, P = 0.011, η2 = 0.83).Mean sorting error scores with and without Variantor filters are shown in Fig. Ps > 0.134. Any significant effects of filter and participant group are described in the main analyses to follow, in which data from both laboratories are combined .Adults using EnChroma filters were tested in two different research laboratories . To check that participants in both laboratories produced equivalent results on our tasks, a mixed-model ANOVA was conducted with laboratory  and filter  as within-subject factors, and participant group  as the between-group factor. For the sorting task, a separate analysis was conducted on the CVD group only, as CVN adults did not perform this task with EnChroma filters. These analyses did not reveal any significant effect of laboratory, or any interaction between laboratory and measures for Ishihara (error scores), FM100 (TES), CVA-UMinho discrimination thresholds and CVA-UMinho naming (5°) hit scores, or sorting task (error scores): all F = 10.03, P = 0.004, η2 = 0.29). The mean error score without filter was 9.16 ± 0.47, whilst with EnChroma filters it was 8.40 ± 0.54. The overall effect of participant group was also significant  = 295.95, P < 0.001, η2 = 0.92). The mean error for CVN was 0.33 ± 0.65, whilst mean error for CVD was 17.23 ± 0.73. The interaction between filter and participant group was significant  = 18.26, P < 0.001, η2 = 0.42) and Tukey (HSD) post-hoc comparisons were conducted to test pairwise comparisons contributing to it. As expected, Ishihara error scores were significantly lower for CVN participants  than for CVD participants . For CVD adult participants, Ishihara error scores were significantly lower when using EnChroma filters (P < 0.001), however this reduction did not affect CVD clinical diagnosis in any individual case as error scores were much higher than the 3-error criterion line  as the within-subjects factor and adult participant group  as the between-subjects factor was conducted to test for the effect of EnChroma filters on Ishihara test error scores  = 0.172, P = 0.691, η2 = 0.97). The mean error score without filter was 14.75 ± 0.96, while with EnChroma filters it was 14.38 ± 1.03. When CVD child participants wore EnChroma filters, indicators of severity according to the Ishihara test did not change for six “strong” deutan children, or for one “mild” protan. One “mild” protan child was re-classified as a “strong” protan with EnChroma filters.For the CVD child sample, a one-way repeated measures ANOVA, with filter  as the within-subjects factor was conducted on Ishihara error scores , i.e., TES ; SQR ; Angle ; C-Index ; S-Index . Although there were no overall significant main effects of filter on any indicator , there were statistically significant interactions between filter and participant group for 4 key indicators: TES ; SQR , Angle  and C-Index , but not for 2 indicators . Tukey (HSD) post-hoc comparisons were used to investigate significant filter × group interactions. As expected, TES, SQR, Angle and C-Index scores were significantly lower for the CVN, than the CVD group for the no filter condition  and also the filter condition , but the effects of filter were not significant for either group alone .Six two-way mixed-model ANOVAs with filter  as the within-subjects factor and participant group  as the between-group factor, were conducted to test for the effects of the EnChroma filters on each of the six FM100 indicators listed in Table For all four indicators, EnChroma filters led to a trend of worse scores  compared to when no filter was worn in the CVN group, but for the CVD group, EnChroma filters led to improved scores , compared to when no filter was worn  were below (better than) the 100 score with and without the EnChroma filters, whilst the other CVD participants were above the 100 error score with and without them. If C-index were used as an indicator of colour discrimination loss , in our CVD group 5 participant scores worsened (mean ± 1 SE from 1.96 ± 0.21 to 2.14 ± 0.20), and 8 improved (from 2.18 ± 0.13 to 1.91 ± 0.12).The wearing of EnChroma filters did not influence clinical diagnoses with FM100 for any participant. According to the test scoring criteria in which a TES < 16 suggests superior discrimination, 16–100 is average discrimination and > 100 is low discriminationF = 11.06, P < 0.001, η2 = 0.28) and participant group  = 56.93, P < 0.001, η2 = 0.66) on discrimination threshold. The mean CVN discrimination threshold was 0.0029 ± 0.00061 and mean CVD discrimination threshold was 0.0093 ± 0.00059. As expected for red-green CVD versus CVN groups, there was a significant interaction between hue and participant group  = 13.49, P < 0.001, η2 = 0.32). However, our main interest here is to test the effects of EnChroma filters on hue discrimination. There was no overall effect of EnChroma filters on discrimination thresholds (P = 0.865). Nor were there significant interactions between filter and hue (P = 0.277), filter and participant group (P = 0.545), or between filter, hue and participant group (P = 0.639). An additional analysis on differences in thresholds  is provided in the Supplementary information.A three-way mixed-measures ANOVA with filter  and hue (16 hues tested) as within-subjects factors and participant group  as the between-group factor, was conducted to test effects of EnChroma filters on discrimination thresholds measured on the CVA-UMinho task with 5° stimuli  = 4.16, P = 0.023, η2 = 0.45). Tukey (HSD) post-hoc comparisons revealed that discrimination with the 1° size was significantly worse than the 5° size for hues at protan axes , a deutan axis , hue 18° , and 162° and 342° . However still, there were no significant effects of EnChroma filters on discrimination thresholds (P = 0.295) and there were no significant interactions between filter and hue (P = 0.413), filter and size (P = 0.111), or filter, size, and hue (P = 0.769).A subsample of 6 CVD participants also performed the CVA-UMinho discrimination task with a smaller stimulus size (1° versus 5° for main experiment). A three-way repeated-measures ANOVA with size , filter  and hue (16 hues tested) as within-subjects factors was conducted to test the effects of EnChroma filters  are very small, much smaller than the effect that Variantor filters had on discrimination thresholds for the CVN group. The magnitudes of effect of EnChroma filters in the CVD group are on average, too small to be of functional significance. Although participant data were usually grouped into CVN and CVD groups as previously mentioned, one question of interest could be, are EnChroma filters helpful for any specific CVD group? We plot differences in discrimination thresholds with and without filters in Fig. F = 23.47, P < 0.001, η2 = 0.78) and hue  = 57.75, P < 0.001, η2 = 0.69), but no overall filter effect (P = 0.921). There were significant interactions: hue × participant group  = 20.26, P < 0.001, η2 = 0.76), filter × hue  = 2.64, P = 0.035, η2 = 0.09) and hue × filter × participant group  = 2.52, P < 0.001, η2 = 0.28). Posthoc testing (using Tukey HSD at P < 0.05) revealed differences only between the no-filter and EnChroma condition for the group of protanopes  and for the group of deuteranopes . No significant differences in hue discriminability between the non-filter and EnChroma condition were found for groups of deuteranomalous or protanomalous participants for any hue axis tested.A three-way ANOVA with filter  and hue (16 hues tested) as within-subjects factors, and participant group  as the between-group factor on discrimination thresholds was conducted (thus excluding Variantor data). This analysis showed significant main effects of CVD group on discrimination thresholds  used for different hue angles, we calculated hit and error scores using the modal criterion. Confusion matrices (11 × 11 BCTs) were created for CVN and CVD participants with and without EnChroma filters and these are pictorially represented in Fig. On examining Fig. F = 13.48, P < 0.001, η2 = 0.32) and a significant effect of participant group on them  = 71.88, P < 0.001, η2 = 0.71). However, there was no significant overall effect of filter (P = 0.557) and no significant interaction between filter and participant group (P = 0.905). In summary, colour naming for the CVD group was significantly worse than for the CVN group across all hues but EnChroma filters had no significant impact on colour naming for CVN, or CVD participants , with and without EnChroma filters are provided for CVN and CVD groups in Fig. CVN participants showed improvements (reductions in naming errors by ≥ + 3%) with EnChroma for 98.5° (tritan axis) and 198°. CVN participants increased naming errors (by ≥ − 3%) at 167.3° and 347.3° (deutan axes) and 306°. CVD participants showed improvements with EnChroma (≥ + 3%) for 18°, 54°, 98.5 (tritan axis) and 306°, but increased naming errors (≥ − 3%) for 126°, 162°, 184.66° (protan axis), 198°, and 347.5° (deutan axis). Additional analyses on BCTs and on differences in hit-scores  are provided in the Supplementary information.For the subsample of 6 British CVD participants who performed the CVA-UMinho naming task with a small stimulus size (1°), EnChroma changed modal hit scores for all 6 BCTs see Fig. . For thiF = 14.21, P = 0.013, η2 = 0.74). The mean 1° size hit score was 37.41 ± 3.85% whereas the mean 5° hit score was 44.33 ± 4.35%). However, there was no significant effect of filter (P = 0.584) and no significant higher-order interactions between size, filter and hue . An additional analysis on the stimulus size effect on BCTs is provided in the Supplementary information.Specific CVD colour naming error scores across hue for 1° stimulus size, with and without EnChroma filters are provided in Fig. P = 0.349), but there was a significant increase in colour relative to shape sorting times for CVD participants with EnChroma  = 6.81, P = 0.020, η2 = 0.31). Without EnChroma filters, sorting by colour relative to shape in CVD participants was 2.52 ± 0.14 slower. With EnChroma filters it was 3.03 ± 0.24 times slower. Thus, although EnChroma filters did not result in a change in accuracy of sorting board-game pieces on the basis of colour, they did make CVD participants slower at completing the task.CVN observers using EnChroma filters did not perform this task. For CVD participants, sorting error scores  and their luminance properties were considered. The current study extends that result using multiple tasks and measurements involving the whole colour space. CVN observers using Variantor filters made significantly more errors in clinical colour vision tests (Ishihara and FM100) and in naming colours, than without. Naming errors are specific to the stimulus set used, but this study shows that CVN participants using Variantor filters made particular errors with colour categories that are problematic for protanopes, but not for deuteranopes . Variantor filters resulted in CVN participants being diagnosed as “strong” protans on the Ishihara test, and as having a CVD with the FM100. The FM100 colour loss estimate (C-Index) for CVN using Variantor (at 2.37–2.83) is similar to that scored by dichromats without filters (at 2.13–2.64), but greater on average than that scored by anomalous trichromats (at 1.25–2.41). CVN participants with Variantor filters had worse discrimination thresholds at protan axes (hues 4.66° and 184.66°) and neighbouring hues (18° and 198°), similar to our protanopes. This lack of specificity in hue angle may be due to the types of stimuli used as, except for monochromatic lights, paper and computerised stimuli are not spectrally pure. Spectrally broadband stimuli are not expected to separate deutan, from protan defects, or dichromacy from anomalous trichromacy. Actually, protan thresholds might be expected to cover not only the protan axes, but also neighbouring hue areas, as current results reflect. Our Variantor results demonstrate similarity in pattern across hue for both discrimination thresholds and naming error scores , as well as improved colour discrimination. EnChroma filters did not significantly affect performance accuracy in any colour discrimination task in the current study, although they did reduce the number of errors made on the Ishihara test by adult CVD participants and on the FM100 test. We also provide Ishihara test results for 8 CVD children. For them, Ishihara error scores were not statistically significantly different with and without EnChroma filters. Proportions of misreadings (misreadings/misreadings + errors) were higher in children without EnChroma, than in adults , in line with previous results65. There is a tendency for misreadings to proportionally increase with EnChroma filters  and this did reach statistical significance  = 5.05, P = 0.037, η2 = 0.22). Data collected on children are of interest here, as one could postulate that previously undiagnosed CVD children are less familiar with the Ishihara test and less likely to have knowledge about the effects of filters. For this group though, EnChroma filters had no impact on test results.A previous study46).If we look qualitatively at discrimination thresholds with the CVA-UMinho task, neglecting statistical significance, any threshold reduction seen in one part of the spectrum (such as at 162°) is compensated for by a threshold increase in another part of the spectrum, such as at (184.66° or 198°). Concurrent results appear in the colour naming task: improvements for certain BCTs  were accompanied by more errors in others . Patterns of quantitative improvement and degradation between discrimination thresholds and colour naming are also not clear, but the effects of wearing EnChroma filters are very small, even for small sized (1°) stimuli where their effects could be potentially greater, given the almost exclusive zone of red-green processing by the fovea and the poorer discrimination overall thresholds providing greater opportunity for improvement if it exists. Such small changes in discrimination with EnChroma filters are not likely to contribute to differences in colour perception .In dichromatic vision, notch filters (like EnChroma filters) cannot increase colour contrast as only 1 cone type exists in the range of the notch. However, the notch can result in changes in luminance contrast for dichromats. In addition, the ideal position of the notch is likely to vary, even within the same CVD classification . Apart from the variability in the sensitivity overlap of the L and M cones existent in anomalous trichromacyn = 16 CVD: Protanopia n = 4; Protanomaly n = 4; Deuteranopia n = 4; Deuteranomaly n = 4), although our results were also replicated in 8 children (Ishihara test) and for smaller sized stimuli (CVA-UMinho discrimination and naming tasks). Would we have found statistically significant differences if we increased our sample size? Fig. P = 0.002) and a significant enhancement for deuteranopes near a deuteranopic confusion line . We do not believe that the effects found for dichromats are due to increases in colour contrast but are more likely due to changes in luminance contrast. For the protanope, the luminance difference usually experienced between medium and long wavelengths was potentially reduced by the filter, leading to losses in hue discriminability. For deuteranopes, the filter potentially introduced new luminance differences not present without the filters, leading to improved hue discriminability. No significant differences were found for discrimination thresholds with and without the EnChroma filters for anomalous trichromats. The effects here were smaller than the variance (± 1 SD) of measures within the CVD group at specific hues, too small and variable to be of functional value. If a study were designed to seek statistical significance in anomalous trichromats for the greatest but very small enhancement effects , sample size estimates66 needed would be n = 39 and n = 62 for deuteranomalous and protanomalous CVD groups, respectively (using the larger of within and across group variance estimates for each hue and peak enhancement effects). It is also important to bear in mind that degradation effects on discrimination  are then also likely to be significant.Our results show no statistically significant differences on FM100 results, measures of hue discrimination and naming in laboratory tasks, or on performance on a real-world colour sorting task in CVD observers with versus without EnChroma filters, although sorting-by-colour took significantly longer in time for CVD participants when wearing them. Our sample size of CVD observers is modest  are greater for dichromats, than for anomalous trichromats, suggesting that altered luminance cues (rather than chromatic cues) were used in judgements. The luminance noise in the CVA-UMinho task is greater (Michelson contrast of 24 ± 4%) than in the Ishihara test (18 ± 1%), and changes randomly from trial to trial, increasing its power. Whereas a significant effect of EnChroma filter on error scores was found on the Ishihara test in adults with CVD see Fig. c. This sAny red-green CVD compensation filter can introduce different colour cues and specially luminance cues, which might result in improved clinical test scores (such as in the Ishihara test and the FM100) in CVD observers, and improved discriminability for some hues (but degraded discriminability for others). They might even help CVD observers in specific colour-combinations important in everyday tasks. However, the use of filters must be restricted to specific tasks, as our results also demonstrate that they may hinder other judgements and may lengthen times for completing some real-world tasks. To date, no study has demonstrated enhanced overall colour discrimination for persons with red-green colour vision deficiency when using compensation filters.Supplementary Information.


In response to 4‐7‐8 breathing control, low‐ and very‐low‐frequency powers significantly decreased (p < 0.05), whereas high‐frequency power significantly increased (p < 0.05) in the control group. Moreover, time domain, total power, and very‐low‐frequency power significantly decreased (p < 0.05) in the sleep deprivation group. Both groups had significantly decreased HR and systolic BP (p < 0.05). HRV, HR, and BP variables showed no significant differences between the groups. Healthy young adults with and without sleep deprivation may have similar HRV, BP, and FBG values. However, sleep deprivation may cause decreased endothelial function. Furthermore, 4‐7‐8 breathing control can help participants improve their HRV and BP, particularly in those without sleep deprivation.This study investigated the effects of sleep deprivation on heart rate variability (HRV), blood pressure (BP), fasting blood glucose (FBG), and endothelial function as well as the immediate effects of 4‐7‐8 breathing control on HRV and BP. In total, 43 healthy participants aged 19–25 years were classified into two groups: Twenty two in the with sleep deprivation group and 21 in the without sleep deprivation (control) group. Resting heart rate (HR), BP, HRV, FBG, and endothelial function were examined. Subsequently, participants practiced 4‐7‐8 breathing control for six cycles/set for three sets interspersed between each set by 1‐min normal breathing. Thereafter, the HR, BP, and HRV were immediately examined. The HRV, HR, and BP variables and FBG were not significantly different between the two groups. However, endothelial function was significantly lower in the sleep deprivation group than that in the control group ( Sleep deprivation may disturb heart rate variability, blood pressure, blood glucose, and endothelial function in healthy young adults. The 4‐7‐8 breathing control may improve these outcomes in those with and without sleep deprivation. Sleep duration for both the groups was categorized according to the NSF guidelines, which advise 7–9 h of sleep per night for the adults  and habitual sleep accumulation (how long that sleep duration has been sustained)—that is, for months or years—during the screening test. The Insomnia Severity Index Questionnaire was also used to assess the sleep quality. Participants were included in the study primarily based on their quantity of sleep. Regarding chronic sleep deprivation, the inclusion criteria, which were based on the American Academy of Sleep Medicine, required participants' sleep disturbances to last for at least 3 months. Additionally, the Participant's Health Screening Questionnaire was used to evaluate underlying diseases, treatment drugs, and illness history. The Thai General Health Questionnaire‐12 was used to evaluate mental health. Moreover, vital signs, including HR, BP, body temperature, and respiratory rate, were measured. To avoid confounding factors that could influence the main results and to ensure that the results mainly arose from the effect of sleep duration, we excluded participants who regularly smoked or drank; had diabetes mellitus, thyroid disease, cardiovascular disease, renal disease, or obesity; were currently on medication for depression or psychiatric disorders; or regularly practiced meditation or breathing control.The participants were informed of their role in the study both verbally and in writing before providing the consent form for participation. The consent form and the study protocols conformed to the ethical standards of the Human Ethics Committee of Burapha University , and of the 1964 Declaration of Helsinki and its later amendments.2.2Participants who passed the criteria and the screening test were asked to keep a consistent bed activity and wake time as usual and requested to come to the laboratory in the morning after 8‐h overnight fasting. Resting HR, BP, HRV, FBG, and endothelial function were then examined. Subsequently, a researcher taught and demonstrated to them the 4‐7‐8 breathing control until they were able to comprehend it well. They were then asked to practice the 4‐7‐8 breathing control. To practice this technique, the participants lay down in a supine position, with their eyes closed lightly and the arms and legs extended fully in a calm and quiet room. One cycle of the 4‐7‐8 breathing control included the following steps: (1) Let the lip part make a whooshing sound and exhale completely through the mouth; (2) close the lips, inhaling silently through the nose, count 1–4 in mind, and then hold breath for 7 seconds; and (3) make another whooshing, exhaling from the mouth, and count 1–8 in mind. They were asked to repeat the 4‐7‐8 breathing control for six cycles/set, for three sets interspersed between each set by 1‐min normal breathing. Finally, their posture was retained for 10 min further to measure their HR, BP, and HRV following the breathing control intervention.2.3After resting for 10 min and after the 4‐7‐8 breathing control intervention, HR and BP were assessed in the supine position using a digital automatic BP monitor (Microlife BP 3AQ1), which then provided the HR value. The BP cuff was thoroughly enveloped around the arm. The inferior edge of the cuff was nearly 1 inch above the bend of the elbow. We assessed the HR and BP twice with 1 min apart. The mean of the two readings served as the systolic and diastolic BP (SBP and DBP) and HR values. Pulse pressure (PP: SBP − DBP), mean arterial pressure (MAP: DBP + [PP/3]), and rate–pressure product (RPP: HR × SBP) were calculated based on the SBP, DBP, and HR values.2.4Following the HR and BP assessments, we assessed the HRV for 10 min. The HRV data were achieved by lead II electrocardiography  and analyzed by the HRV module . HRV variables included the time and frequency domains. The time‐domain consisted of the values of SDNN and the root mean square of successive R‐R interval differences (RMSSD). The frequency‐domain was analyzed using the values of total power, very‐low‐frequency power , low‐frequency power , high‐frequency power , and low‐frequency to high‐frequency ratio. Sympathetic and parasympathetic activities, as well as baroreceptor activity, are all described by HRV data. Low‐frequency power, for example, reflects sympathetic activity most of the time, whereas high‐frequency power reflects parasympathetic activity, and the low‐frequency to high‐frequency ratio reflects a balance of sympatho‐vagal activity , which consisted of lancets, test strips, and a glucometer. After lancing a participant's fingertip, we extracted a drop of capillary blood (approximately 0.6 μl), which was then collected by the test strip inserted into the glucometer. In a few seconds, the screen reported the blood glucose value in mg/dl unit.2.6Endothelial function was assessed using the endothelium‐dependent vasodilation technique. Forearm blood flow was measured using the Laser Doppler Flowmetry Module , whose probe was attached to the forearm skin in a perpendicular direction. To measure forearm blood flow during occlusion and recovery, a cuff from a standard mercury sphygmomanometer  was closely enclosed around the arm proximally to the probe. We inflated the cuff to gain suprasystolic pressure at approximately 200 mm Hg and to completely occlude the brachial artery. Subsequently, the cuff pressure was gradually released. The forearm blood flow was measured for three periods, namely, at rest, during occlusion, and after occlusion, which lasted for 5 min each, and reported in the perfusion unit. Thereafter, we analyzed the resting blood flow, blood flow during occlusion, peak blood flow after occlusion, peak blood flow after occlusion/resting blood flow ratio, and recovery time after occlusion. The recovery time after occlusion referred to the after occlusion duration when the blood flow was similar to the resting value.2.7Anthropometry and body composition were measured when the participants were standing and wearing only the minimum necessary clothing. A body composition analyzer  based on the principle of bioelectrical impedance analysis was used to determine body mass, body mass index, body fat percentage, fat mass, fat‐free mass, protein mass, mineral mass, and water mass. During inspiration, a stadiometer  was used to measure height. The mid‐point between the bottom rib and superior iliac spine was used to assess waist circumference at the end of a normal expiration. At maximum buttock extension, the hip circumference was determined. Before conducting any assessment, participants were asked to urinate. On the day before the assessments, they were also asked to refrain from eating and drinking, including alcohol and caffeine, smoking, and participating in strenuous exercise or exertion.2.8The quality of sleep was assessed using the Insomnia Severity Index Questionnaire , respectively. To assess differences within a group before and after intervention, we used the paired t‐test. All data are presented as mean ± standard deviation. A p value of less than 0.05 was considered significant.All statistical data were analyzed using the IBM SPSS Statistics (IBM). Data normality and equal variance were studied using the Shapiro–Wilk test and Levene's test, respectively. Differences between groups before and after the intervention were assessed using the independent 3One person in the control group, out of 44, was excluded from the analysis due to insufficient sleep on the night before. As a result, data from 43 participants were evaluated, including 21 participants in the control group (95.45%) and 22 participants in the sleep deprivation group (100%).3.1n = 36, 83.72%). Sex, height, body mass, body mass index, body fat percentage, fat mass, fat‐free mass, water mass, protein mass, mineral mass, visceral fat level, metabolic rate, waist and hip circumferences, and their ratios were not significantly different between the control and sleep deprivation groups. However, age was significantly higher in the control group (p = 0.002)  Table .3.2p < 0.05), and habitual sleep accumulation was not significantly different between these groups  and the recovery time after occlusion was significantly higher (p < 0.05) in the sleep deprivation group than in the control group  beat intervals, SDNN, RMSSD, total power, very‐low‐frequency power, low‐frequency power, low‐frequency power in normalized units, high‐frequency power, high‐frequency power in normalized units, and low‐frequency to high‐frequency ratio, were not significantly different between the control and sleep deprivation groups Figures .Similarly, baseline HR and BP measures, such as SBP, DBP, PP, MAP, and RPP, did not differ significantly between the sleep deprivation and control groups Figure .3.5p < 0.05), whereas, NN intervals and high‐frequency power in normalized units significantly increased in the control group (p < 0.05). In the sleep deprivation group, SDNN, RMSSD, total power, and very‐low‐frequency power were significantly reduced (p < 0.05) and NN intervals significantly increased (p < 0.05). However, the HRV variables showed no significant between‐group differences . Similarly, the sleep deprivation group had significantly decreased HR, SBP, PP, and RPP levels (p < 0.05). However, the HR and BP variables revealed no significant between‐group differences  in this study is smaller than that in Zhang et al.'s CONSORT study 2021) (sleep deprivation group: 3.78 ± 0.69 h versus control group: 7.63 ± 0.52 h). According to our study results, although participants with sleep deprivation may have impaired endothelial function, similarity in HRV, HR, and BP variables may indicate that their cardiovascular systems can adapt well to such physiological and psychological stresses , participants' respiratory rate was approximately 3 breaths/min. This breathing pattern is similar to deep and slow breathing, which has previously been investigated. Magnon et al.  conducteThe aforementioned data suggest that parasympathetic activity is augmented by breathing with a low inhale/exhale ratio. Moreover, additional holding of one's breath during inhalation can increase arterial oxygen saturation and subsequently decrease peripheral chemoreceptor stimulation, thereby enhancing the parasympathetic activity and lowering the BP (Turankar et al., In this study, very‐low‐frequency power significantly decreased in both the control and sleep deprivation groups. Although very‐low‐frequency power is not exactly known, it may be generated after exposure to mental stress (Usui and Nishida, Total power is the sum of the powers of the five frequencies, and the very‐low‐frequency and ultra‐low‐frequency powers account for most of the total power (Yılmaz et al., This study also observed that RMSSD significantly decreased in the sleep deprivation group. RMSSD, which is the principal time domain of HRV, reflects the parasympathetic activity and is associated with high‐frequency power (Shaffer and Ginsberg, We also found that the 4‐7‐8 breathing control decreased the HR and BP variables in almost all participants in both the control and sleep deprivation groups. The results of this study are consistent with previous findings (Mitsungnern et al., 4.3To the best of our knowledge, this is the first study evaluating the immediate effects of 4‐7‐8 breathing control on HRV and BP in people with sleep loss. However, there are several limitations to this research. The small sample size of males in this study appears to be insufficient to accurately reflect the general population's condition. As sex hormones can affect cardiac autonomic function, BP, and FBG, the results could have been influenced by the high female‐to‐male ratio. Furthermore, after the 4‐7‐8 breathing control, endothelial function was not evaluated. Hence, although the intervention's magnitude may not have been sufficiently high to affect endothelial function, this study may not provide complete data, and further research is needed. This study investigated the immediate effects of the 4‐7‐8 breathing control in people with sleep deprivation. Therefore, future research should investigate the long‐term effects of this breathing practice on cardiovascular and cardiac autonomic functioning over a period of months or years. It should also investigate the efficacy of 4‐7‐8 breathing control intervention in other populations, such as older people who may have sleep issues, patients with respiratory disorders who want to improve perfusion and gas exchange, and patients with mental illnesses who want to improve cognition, emotion, and behavior.5Healthy young adults with sleep deprivation and adequate sleep demonstrated similar HRV, BP, and FBG. Nevertheless, endothelial function may be lower in those with sleep deprivation. Furthermore, by increasing parasympathetic activity and decreasing sympathetic activity, the 4‐7‐8 breathing control may improve HRV and BP, especially in people without sleep deprivation. This intervention may also be beneficial for patients with cardiovascular disease or pulmonary disease in terms of reducing cardiac work and enhancing blood oxygenation.Piyapong Prasertsri designed the study. Jaruwan Vierra performed data collection. Piyapong Prasertsri performed statistical analysis and interpreted the data. Jaruwan Vierra, Orachorn Boonla, and Piyapong Prasertsri contributed to writing and proofing of the final version of the manuscript.No conflicts of interest are reported by the authors of this paper.