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Obtaining and maintaining cortical hand representation as evidenced from acquired and congenital handlessness

  1. Daan B Wesselink
  2. Fiona MZ van den Heiligenberg
  3. Naveed Ejaz
  4. Harriet Dempsey-Jones
  5. Lucilla Cardinali
  6. Aurelie Tarall-Jozwiak
  7. Jörn Diedrichsen
  8. Tamar R Makin  Is a corresponding author
  1. University College London, United Kingdom
  2. University of Oxford, United Kingdom
  3. University of Western Ontario, Canada
  4. Istituto Italiano di Tecnologia, Italy
  5. Queen Mary’s Hospital, United Kingdom
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Cite this article as: eLife 2019;8:e37227 doi: 10.7554/eLife.37227

Abstract

A key question in neuroscience is how cortical organisation relates to experience. Previously we showed that amputees experiencing highly vivid phantom sensations maintain cortical representation of their missing hand (Kikkert et al., 2016). Here, we examined the role of sensory hand experience on persistent hand representation by studying individuals with acquired and congenital hand loss. We used representational similarity analysis in primary somatosensory and motor cortex during missing and intact hand movements. We found that key aspects of acquired amputees’ missing hand representation persisted, despite varying vividness of phantom sensations. In contrast, missing hand representation of congenital one-handers, who do not experience phantom sensations, was significantly reduced. Across acquired amputees, individuals’ reported motor control over their phantom hand positively correlated with the extent to which their somatosensory hand representation was normally organised. We conclude that once cortical organisation is formed, it is remarkably persistent, despite long-term attenuation of peripheral signals.

https://doi.org/10.7554/eLife.37227.001

Introduction

A fundamental organising principle in the primary somatosensory cortex (SI) is somatotopic mapping, where adjacent body parts are represented more proximally on the cortical sheet than those further apart (Penfield and Rasmussen, 1950). In the cortical hand area, this topographic organising principle results in a detailed digit map (Kaas et al., 1979Penfield and Rasmussen, 1950), where neighbouring digits on the hand are represented closer together on the neocortex than non-neighbouring digits, as can be shown with functional MRI (fMRI) (Kolasinski et al., 2016). More generally, cortical activity patterns for neighbouring fingers overlap more – and are therefore more similar – than non-neighbouring fingers, independent of the exact spatial arrangement of these patterns (Ejaz et al., 2015). The representational structure (i.e. the relative dissimilarity of activity patterns for different movements) is thought to reflect the natural statistics of hand use over one’s life course (Graziano and Aflalo, 2007; Overduin et al., 2012): Concurrent inputs to neighbouring digits will increase representational similarity (Wang et al., 1995), while greater individuation of inputs will induce greater representational dissimilarity (Ejaz et al., 2015). Some have even suggested that alterations of these inter-digit representational boundaries after the hand map has been formed, for example due to digit overuse, can disrupt (musician’s dystonia [Elbert et al., 1998], but see Ejaz et al., 2016) or improve perception and action (tactile discrimination [Recanzone et al., 1992; Pleger et al., 2003]).

We (Kikkert et al., 2016) and others (Flesher et al., 2016; Bruurmijn et al., 2017) recently challenged the view that structured input from the periphery is required for preserving sensorimotor hand representation (Dempsey-Jones et al., 2016; see also Davis et al., 1998, Mercier et al., 2006, Garbarini et al., 2018 for related brain stimulation and behavioural findings). We took advantage of a well-documented phenomenon whereby some amputees report being able to volitionally move their phantom hand, resulting in kinaesthetic phantom sensations (Henderson and Smyth, 1948). Phantom limb movements have been shown to elicit both central and peripheral motor signals that are different from those found during movement imagery (Reilly et al., 2006; Raffin et al., 2012a; Raffin et al., 2012b; Makin et al., 2013b). Using 7T imaging we explored whether three amputees experiencing exceptionally vivid phantom sensations maintained the canonical hand representation, exemplified by somatotopically organised representation of individual digits. We found that although digit selectivity was reduced, digit order and the extent of the missing hand maps in SI were similar to what is observed in controls.

Our previous findings demonstrate the stability of SI hand organisation despite decades of amputation. It remains unknown, however, whether hand representation after amputation reflects phantom sensations, and as such only persists in individuals with highly vivid phantom sensations. Here, we asked whether persistent representation of a missing hand reflects an organisational principle in the sensorimotor cortex, and thus will even be observed in amputees with little phantom sensations. To address this question, we measured cortical hand representation in 18 acquired amputees with varying vividness of their phantom sensations (hereafter amputees). To test the idea that the development of a hand representation requires sensory experience, we also tested 13 individuals missing one hand from birth (due to congenital amelia; hereafter congenital one-handers). All participants underwent fMRI while performing a visually cued motor task involving individual digit movements (of both the missing hand and the intact hand). Activity patterns in the missing hand of SI and M1 were analysed using representational similarity analysis (Walther et al., 2016; Diedrichsen et al., 2016). We hypothesised that normal peripheral input is necessary to establish normal sensory hand representation, but not to maintain it.

Results

Phantom hand movements elicit typical hand representation in the missing hand area of acquired amputees

We first focused our analysis on the representation of the missing hand, as revealed by instructing individuals to move individual digits of their missing hand (or nondominant hand in controls). We interrogated fMRI activity in the SI hand area contralateral to the missing/nondominant hand (see Materials and methods for regions of interest (ROI) definition). We examined univariate task-related activity, as quantified by averaging the BOLD response across all the digit conditions within the missing hand ROIs (Figure 1A, see Figure 1—figure supplement 1 for M1 ROI results). Overall, all participants, including congenital one-handers, were able to engage the missing/nondominant hand area to some degree. Although activity was reduced in SI for the congenital one-handers’ missing hand compared to controls (t(23)=3.5, p=0.002), activity was significantly greater than baseline (t(12)=2.6, p=0.02).

Figure 1 with 1 supplement see all
Similar representation in primary somatosensory cortex (SI) for amputees’ missing hand and controls’ nondominant hand, but not for congenital one-handers’ missing hand.

(A) Activity (averaged digit movement versus rest) in SI for amputees (n = 18), two-handed controls (n = 12), and congenital one-handers (n = 13). (B–C) Mean dissimilarity and typicality of the representational structure of contralateral SI activity for the three groups. (D) Representational dissimilarity matrices for the three groups. D1-D5 correspond to the five digits (thumb-little finger). (E) Two-dimensional projection of the representational structure (D) (using multi-dimensional scaling; note that this is included for visualisation purposes only and was not used for statistical analysis). Dissimilarity is reflected by distance in the two dimensions; individual digits are reflected by different colours (see colour key, bottom right); and ellipses reflect the between-subject standard error after Procrustes alignment. Please note the different scale for one-handers compared to amputees and controls. Abbreviations: a.u.: arbitrary unit; *: significant difference, after accounting for multiple comparisons.

https://doi.org/10.7554/eLife.37227.002

To investigate digit discriminability in the hand area, we next estimated the dissimilarity between activity patterns for individual digit movements, measured using the cross-validated Mahalanobis distance (Nili et al., 2014). By comparing all possible pairs of digit-specific activity patterns, we obtained the representational structure (Figure 1D–E). The resulting inter-digit dissimilarity values were averaged across digit pairs and participants within each group (Figure 1B). Small inter-digit dissimilarity indicates that voxels in the hand area are similarly activated across individual digits; larger dissimilarity implies individuated digit representation. In amputees, mean dissimilarity was slightly, though inconclusively, reduced compared to controls (t(28)=1.13, p=0.27, BF = 0.772), but significantly greater than in congenital one-handers (t(29)=3.54, p=0.001). Congenital one-handers showed no differentiation between digits of their missing hand (mean dissimilarity not different from 0; t(12)=.73, p=0.48) and dissimilarity was significantly reduced compared to controls (t(23)=4.86, p<0.001).

While the extent of discriminability is greater in amputees than congenital one-handers, it is possible that the pattern of individuated digit activity is atypical in amputees. To determine whether the inter-digit organisation of a missing hand is normal, we next studied the representational structure’s typicality, that is the correlation of the representational dissimilarity matrix (RDM) with a dataset of hand RDMs in 2-handed controls drawn from a different study (Wesselink et al., 2018; see Figure 1C and Materials and methods). In amputees, on average, typicality was high (rho = 0.75) and was not significantly different from controls (t(28)=.991, p=0.33, BF = 0.128). Hence, the organisation of digit representation after hand-loss remained statistically unchanged, after an average of 18 years of handlessness. As expected, congenital one-handers’ missing hand representation did not correlate with a normal hand pattern, reflected in diminished representational typicality (mean rho = 0.29) compared to both controls (t(23)=5.86, p<0.001) and amputees (t(29)=6.09, p<0.001).

The results in M1 were generally in line with our findings in SI, but, as expected (Ejaz et al., 2015; Bruurmijn et al., 2017), digit individuation was weaker (see Figure 1—figure supplement 1). Univariate activity in congenital one-handers’ M1 was not significantly reduced compared to controls (t(23)=2.00, p=0.058) and greater than baseline (t(12)=4.60, p=0.001). Congenital one-handers showed significantly lower dissimilarity compared to controls (t(23)=4.23, p<0.001). In amputees, mean dissimilarity was slightly reduced compared to controls, but these differences were not significant (t(28)=.53, p=0.60, BF = 0.325). Typicality was also significantly lower in congenital one-handers than in either controls (t(23)=3.42, p=0.002) or amputees (t(29)=3.50, p=0.002), while the latter groups were not different from each other (t(28)=.11, p=0.91, BF = 0.253).

Although the inter-digit representational structure of congenital one-handers is atypical with respect to canonical hand representation, it is possible that it is still consistent within participants. To explore this idea, we split individual participants’ data to odd and even scans. For each participant, we calculated an RDM in the missing/nondominant hand area using the odd and even runs, and correlated the two RDMs. The correlation between odd and even RDMs was significantly lower in congenital one-handers (rho = -.02) compared to both amputees (rho = 0.41; p1H-AMP=.001) and controls (rho = 0.52; p1H-CTR=.001). We note that by splitting the data we are reducing the effectiveness of our analysis. Nevertheless, the relative reduction in split-half consistency indicates that there is no strongly consistent digit information in the missing hand area of congenital one-handers during this task.

Missing hand representation in acquired amputees is persistent even after phantom sensations have diminished

Next, we evaluated whether the consistency of hand representation in SI during missing hand movements correlates with amputees’ subjective reports of phantom sensations. We first carried out an exploratory forward stepwise regression with typicality as the dependent variable. The following factors were tested as independent variables: kinaesthesia of phantom sensations - the number of phantom digits perceived as independently moving during the phantom movement task; vividness of nonpainful phantom sensations as experienced both during the study and chronically; intensity of phantom limb pain, as experienced both acutely during the study and chronically; time since amputation; age at amputation, and; typicality of the intact hand (calculated from the intact hand SI area). The final model (F = 19.9, p<0.001, adjusted R2 = 0.645 included only kinaesthesia of phantom sensations (β = 0.07, t = 4.46, p<0.001) and the intercept (β = 0.52, t = 8.89, p<0.001). This regression was submitted to a bootstrapping analysis, allowing us to estimate the consistency of the final model (see Materials and methods). This bootstrapping analysis returned kinaesthesia as the final variable in 96.3% of the iterations (final model fit: median adjusted R2 = 0.645; 95% CI: 30-99%). The proportion of the other included factors in the final model was: typicality of the intact hand (7.0%); time since amputation (9.5%); age at amputation (9.6%); vividness of nonpainful phantom sensations (acute: 12.2%; chronic: 20.8%); phantom limb pain, acute: 22.7%; chronic: 10.1%).

Post-hoc analysis confirmed a significant correlation between typicality and kinaesthesia in amputees (Figure 2A; rho = 0.72, df = 16, p=0.001). No significant correlation was found with nonpainful phantom vividness (the chronicity of experiencing the missing hand as existing; Figure 2B; rho = 0.13, df = 16, p=0.61), or years since amputation (rho = 0.17, df = 16, p=0.49). The correlation between kinaesthesia and dissimilarity in SI approached significance (rho = 0.447, p=0.063).

Kinaesthetic sensations during individuated phantom hand movements in amputees correlate with typicality in the missing hand’s primary somatosensory cortex (SI).

Typicality is the correlation coefficient of the representational dissimilarity matrix (RDM) with an independent hand RDM in controls. Phantom kinaesthesia (A) shows the number of digits that produced a sensation of movement during volitional phantom digit movements, based on amputees’ self-reports. Grey and orange ranges show the mean and confidence intervals for typicality in one-handers and controls, respectively. The regression line is only presented for visualisation. Nonpainful phantom vividness (B) conveys the chronicity of the experience of the existence of a missing hand, where 0 indicates no sensations and 100 sensations identical to the intact hand.

https://doi.org/10.7554/eLife.37227.004

Further analysis confirmed that the correlation between kinaesthesia and typicality remained significant in SI after accounting for typicality in M1 (mean rho = 0.44) as a covariate (F = 17.73, p<0.001, Adjusted R2 = 0.51; βKinaesthesia =.74, p<0.001). This analysis suggests that although better recruitment of M1 is expected in individuals with clearer kinaesthetic sensations, the strong correlation between kinaesthesia and typicality in SI does not merely reflect information in M1.

Regardless of the positive relationship between kinaesthesia and typicality, amputees with little to no kinaesthetic sensations still showed missing hand representation. As stated above, the regression line between kinaesthesia and typicality had an intercept of βintercept=.52. This was also the case when phantom vividness was the (non-significant) dependent variable (F = 0.021, p=0.89, Adjusted R2 = -.061; βintercept=.75, p<0.001). These results predict that even amputees who do not experience any phantom sensations will retain some typical missing hand representation. To test this prediction directly, we examined the three amputees in our dataset showing weak to no chronic phantom vividness (below 10/100). Despite not being able to experience clearly their phantom hand when performing the phantom movements task, these individuals showed high typicality (average typicality (rho) = 0.83). Moreover, when comparing their typicality to that found in the congenital one-handers (who were arguably better matched to this sub-group in terms of task demands), the amputees with diminished phantom sensations showed significantly stronger correlations with the canonical hand structure (Mann-Whitney U = 38, p=0.007). Typicality was not different between these three amputees and controls (Mann-Whitney U = 29, p=0.52, BF = 0.089). Together, these additional analyses confirm that the representational structures’ typicality in SI of amputees is still present in those with little to no phantom or kinaesthetic sensations.

Diminished missing hand representation in congenital one-handers even when task performance is matched

While the task involving phantom hand movements was suitable to test the persistence of missing hand representation in individuals with phantom sensations, it was not designed to rule out the existence of missing hand representation in congenital one-handers. Indeed, it is possible that congenital one-handers have typical sensorimotor representation of their missing hand, but they did not access it due to unnatural task demands (see Striem-Amit et al., 2015 for analogous results regarding visual cortex organisation in congenitally blind individuals).

To probe digit structure in the missing hand cortex using an alternative task, we examined whether we could observe a representation of the ipsilateral (intact) hand in the missing hand cortex. In two-handed controls, finger movements lead to individuated digit representation in specific cortical patches in ipsilateral M1 and SI, which tightly correspond to the activity patches engaged in the movement of the mirror-symmetric contralateral finger (Diedrichsen et al., 2013). Importantly, this ipsilateral digit representation fully overlaps with the representation of the contralateral hand (Diedrichsen et al., 2018). Furthermore, ipsilateral representation disappears completely during asymmetric bimanual finger movements, during which activity in M1 and SI is fully determined by the contralateral hand (Diedrichsen et al., 2013). As such, the ipsilateral representation of one hand is likely elicited due to recruitment of the representation of the contralateral hand (Diedrichsen et al., 2018; Berlot et al., 2018). Ipsilateral representation of the intact hand can therefore provide an indirect assay into the representation of the missing hand, while controlling for task demands across groups. Importantly, all three groups were able to perform the individuated digit movement task equally well and contralateral representation of the intact hand was typical in all groups (see Materials and methods). We compared the intact/dominant inter-digit representational structure in the missing/nondominant hand area of one-handers/controls (respectively). We predicted that persistent missing hand representation in amputees should result in similar ipsilateral representation in their missing hand cortex as controls. If missing hand representation is diminished in congenital one-handers, then ipsilateral representation of their intact hand (in the missing hand area) should show reduced representational features compared to those found in amputees (see Discussion for an alternative mechanism, where the deprived cortex develops separate representations for both the contralateral (missing) and ipsilateral (intact) hands).

Mean (intact hand) ipsilateral digit dissimilarity and typicality were not significantly different between amputees and controls (Figure 3; dissimilarity: t(28)=1.42, p=0.166, BF = 0.209; typicality: t(28)=.69, p=0.498, BF = 0.244). In contrast, ipsilateral representation in congenital one-handers was significantly lower than in amputees (dissimilarity: t(29)=3.81, p<0.001; typicality: t(29)=3.05, p=0.005) and showed similar trends versus controls (dissimilarity: t(23)=2.20, p=0.038; typicality: t(23)=2.19, p=0.039). Together, these findings provide additional support for the reduced existence of inter-digit representational difference in the missing hand cortex of congenital one-handers versus amputees, independent of missing hand motor skill.

Similar ipsilateral hand representation in primary somatosensory cortex (SI) for amputees’ and controls’ intact hand.

(A–B) Mean dissimilarity and typicality of the representational structure of ipsilateral SI activity for the three groups. Both dissimilarity and typicality of ipsilateral hand representation indicate a difference between missing hand representation in congenital one-handers and amputees, independent of missing hand motor skill. The red error bars indicate the dissimilarity and typicality values (standard error of the mean) in a visual control area V5 for the same groups, designed to capture visuomotor representation that is not strictly somatosensory. While amputees and controls showed significantly greater digit representation in SI than V5 (both in terms of dissimilarity and typicality), congenital one-handers did not, further indicating reduced SI digit representation. Abbreviations: a.u.: arbitrary unit; *: significant difference; #: trending difference (.02 < p < 0.05).

https://doi.org/10.7554/eLife.37227.005

It is important to consider that we found some potential support for the existence of an ipsilateral digit representation in the missing hand cortex of congenital one-handers. The cross-validated dissimilarity measurement in this group was significantly larger than zero (t(12)=2.51, p=0.027), indicating that there were significant differences between the activity patterns associated with each finger. We therefore wished to determine whether the dissimilarity measures reflect meaningful (though reduced) sensorimotor digit information, or rather the increased sensitivity of RSA to other (not sensorimotor) inter-digit differences (e.g. visual task differences). For example, a recent study demonstrated that visual information about touch on the hand is sufficient to induce some residual digit-selective activity patterns in SI (Kuehn et al., 2018). We therefore compared representational measurements between SI and visual area V5, previously shown not to contain individual sensorimotor digit representation (Beauchamp et al., 2009). Although it is difficult to set a benchmark at a particular dissimilarity value, we suggest that representation crucial to somatosensation in SI should at least outperform V5. Congenital one-handers showed no significant differences in representation between SI and V5 (dissimilarity: t(12)=-1.03, p=0.322; typicality: t(12)=.78, p=0.448), whereas both amputees and controls showed significant differences (all p’s < 0.02), resulting in a significant group x area interaction (dissimilarity: F(2,85)=4.25, p=0.018; typicality: F(2,85)=4.42, p=0.015; Figure 3). These findings corroborate that the digit dissimilarity values in congenital one-handers are likely not specifically indicative of sensorimotor representation, but may rather reflect other task demands.

Discussion

Here, we demonstrated long-term stability of SI hand representation in a group of acquired amputees with diverse phantom sensations, including those experiencing limited phantom sensations. Using RSA, we find that amputees show individuated digit representation for their missing hand, as exemplified by significantly greater inter-digit dissimilarity values in amputees versus congenital one-handers. The inter-digit pattern comprising the missing hand representation was typical to SI hand representation in amputees and was not significantly different from controls (as supported by a Bayesian analysis). Importantly, by studying individuals with a varying range of phantom sensations, we were able to confirm stable hand representation even after phantom sensations have diminished. This result confirms the persistence of hand representation as a general principle in amputees, contrary to recent reports (Serino et al., 2017). Using the same task, we were unable to identify similar digit representation for the missing hand of congenital one-handers, demonstrated by significantly reduced pattern typicality compared to amputees (and even in comparison to those few amputees with little phantom sensations), as well as controls. This result confirms that the persistent hand representation observed in amputees does not reflect mere cognitive task demands (e.g. visual feedback, Kuehn et al., 2018; or attention, Puckett et al., 2017).

We also explored whether we could activate the representation of the missing hand indirectly by movements of the fingers of the intact hand. Previous studies in two-handers have demonstrated that contralateral and ipsilateral hand movements produce identical representational patterns (Diedrichsen et al., 2018). Since this ipsilateral representation is completely overwritten by the contralateral hand if the two hands are engaged in dissociated movements (Diedrichsen et al., 2013, Exp. 2), it has been proposed that the ipsilateral hand reactivates the cortical resources associated with the contralateral hand. Based on this evidence, the ipsilateral digit representation can serve as an indirect measure of the contralateral hand representation. This discovery provides us with a unique and novel opportunity to interrogate the information content underlying the ‘deprived’ hand cortex despite the physical absence of the hand. This approach provided converging evidence, but using different task demands, for similar (missing) hand representation for amputees and controls, but not for congenital one-handers. It might be worth considering whether congenital one-hander’s deprived cortex could have developed separate representations for both the contralateral (missing) and ipsilateral (intact) hand, which are uncorrelated due to anatomical or behavioural differences from two-handers. If this were possible, we would predict maintained, or even stronger, ipsilateral representation of the intact hand. Yet, our data does not reflect this hypothesis. Moreover, poor ipsilateral representation in the deprived cortex does not seem to stem from reduced inter-hemispheric connectivity, which appears to be functionally and structurally preserved in congenital one-handers, depending on lateralisation strategies in daily behaviour (Hahamy et al., 2015; Hahamy et al., 2017; Makin et al., 2013a). Finally, it is still possible that rudimentary missing hand representation, for example determined by genetic factors (Miyashita-Lin et al., 1999; Rubenstein et al., 1999), has originally formed in congenital one-handers but later diminished due to lack of consistent sensorimotor input. Bearing this caveat in mind, our findings suggest that early-life experience is potentially necessary to create typical functional sensory organisation, but not to maintain it.

It has previously been shown that restored peripheral input, for example via hand transplantation (Frey et al., 2008) or targeted reinnervation (Serino et al., 2017) can reinstate sensorimotor hand representation, indicating that the canonical hand representation is, to an extent, immutable to change. Moreover, we reported that the SI hand map can persist independently of the original peripheral input, as observed in a patient sustaining a brachial plexus avulsion injury, abolishing communication between the periphery and the central nervous system (Kikkert et al., 2016). Similarly, in the current study amputees showed, on average, persistent SI representation despite suffering diverse nerve injuries spanned varying degrees of amputation (Table 1). It is therefore necessary to consider alternative inputs that might contribute to the stability of the missing hand map in amputees. Considering that our task required active phantom movements, it is likely that the SI representation pattern is driven by motor efferent inputs. Indeed, while motor signals can no longer reach their final output muscle terminal, the motor cortex in amputees remains functional (Raffin et al., 2012a; see Kokotilo et al., 2009 for similar results in spinal cord injury patients). When a motor command is sent out (e.g. in the form of an attempted hand movement), efference signals are thought to reach SI and generate corollary discharge, suggested to resemble the expected sensory feedback activity pattern, resulting from the movement (London and Miller, 2013; Adams et al., 2013). Since congenital one-handers have never operated a hand, it is likely that this sensorimotor predictive coding architecture never formed in the first place, explaining the lack of inter-digit dissimilarity found in the present study.

It is important to consider how the finding of robust persistence of hand representation, despite the physical absence of a hand, conceptually aligns with other reports of brain reorganisation. Since the pioneering work of Hubel and Wiesel, demonstrating that input loss to visual cortex in early development leads to profound physiological changes (Wiesel and Hubel, 1965b; Hubel and Wiesel, 1965; Wiesel and Hubel, 1965a), it has long been established that sensory deprivation causes cortical reorganisation. Later seminal electrophysiological studies in monkeys further demonstrated that deprivation-driven reorganisation following peripheral input-loss also occurs in adults (Kaas et al., 1983). For example, following peripheral deafferentation of the hand and arm, the missing hand SI area becomes responsive to touch applied to the monkey’s lower face (Pons et al., 1991), likely due to subcortical re-routing of inputs (Kambi et al., 2014; Liao et al., 2016). Recent research in humans indicates extensive reorganisation of multiple body-part representations onto the deprived hand area of congenital one-handers (Hahamy et al., 2017; Striem-Amit et al., 2018; Stoeckel et al., 2009). In amputees sustaining input loss in adulthood, original reports emphasised facial remapping in SI, akin to the reorganisation observed in monkeys (Flor et al., 1995), as a driving mechanism for phantom limb pain (maladaptive plasticity; Flor et al., 2006). Later research challenged the notion that the deprived hand area gets taken over by facial inputs (Makin et al., 2013b; Makin et al., 2015; Raffin et al., 2016) and instead emphasised increased representation of the intact hand in the missing hand cortex as a potential neural correlate of adaptive plasticity (Makin et al., 2013a; Philip and Frey, 2014; see further discussion below). More recently, we have suggested that functional reorganisation is more limited than originally considered (Makin and Bensmaia, 2017). Regardless of the ongoing debate over the functional role of SI reorganisation in the adult (Andoh et al., 2018; Kuner and Flor, 2017) and developing brain (Hahamy et al., 2017; Striem-Amit et al., 2018), common to all these previous studies of reorganisation is that to activate the deprived cortex researchers studied representations of the spared body parts (e.g. the face or the intact hand). While this approach is suitable for documenting cortical remapping, it leaves unexplored the possibility that the original functional organisation of the now-deprived area may be preserved, though latent. We propose that reorganisation in the missing hand cortex does not necessarily abolish the original functional layout in sensory cortex. For example, persistent representation, in the form of efferent cortico-cortical input would engage a separate cortical layer (Felleman and Van Essen, 1991; Adams et al., 2013) than brainstem and thalamic facial inputs to the deprived cortex (Kambi et al., 2014). It still remains to be determined whether these two forms of persistent representation and reorganisation are functionally orthogonal, or interactive (Andoh et al., 2018).

As mentioned above, the missing hand cortex in amputees, but not in congenital one-handers, has been previously shown to respond to inputs from the intact hand (Bogdanov et al., 2012; Makin et al., 2013a; Hahamy et al., 2017; Philip and Frey, 2014), presumably through functional reorganisation. Here, we used RSA to dissect the information content underlying ipsilateral activity of the intact hand. We find that amputees, but not congenital one-handers, showed similar measures of dissimilarity and typicality as controls. However, the fact that ipsilateral dissimilarity was not significantly greater in amputees than in controls is inconsistent with the interpretation of increased intact hand activity as a neural correlate of adaptive reorganisation (Makin et al., 2013a). Regardless, the existence of ipsilateral digit-specific organisation in the missing hand cortex of amputees might provide an alterantive mechanism for the preservation of the missing hand digit maps. While we previously showed that the phantom hand map is activated by phantom hand movements independently of the intact hand (Kikkert et al., 2016; Philip and Frey, 2014), it is still possible that structured inputs from the intact hand (via ipsilateral pathways) sustains the missing hand map, despite the loss of the original peripheral inputs.

To conclude, here we show that once sensorimotor hand-representation is formed, it is generally immutable to change: We identified stable hand representation in amputees’ sensorimotor cortex using representational similarity analysis, despite years (and even decades) of amputation and irrespective of their phantom sensations vividness. In contrast, individuals born with a missing hand (congenital one-handers) did not show normal representation of their nonexisting hand. We therefore suggest that consistent sensory representation despite input loss may be a common organising principle (Striem-Amit et al., 2015; Collignon et al., 2013; Baseler et al., 2011). How can our findings of persistent representation, despite massive and long-lasting input change, be resolved with multiple observations of updated hand representation due to altered experience (e.g. due to nerve/digit deafferentation [Merzenich et al., 1983; Merzenich et al., 1984], increased usage [Jenkins et al., 1990], syndactyly [Allard et al., 1991; Wang et al., 1995], or mobile phone usage [Gindrat et al., 2015])? Here we show that amputees with greater phantom kinaesthetic sensations better retained their missing hand representation. In light of this, we suggest that daily life experience could shape the fine-grained aspects of hand representation, but the large-scale functional organisation of the hand area is fundamentally stable.

Materials and methods

Participants

We tested 18 acquired amputees with an average of 18 years since amputation (mean age: 50 ± 12; eight left-handed; four female), 13 congenital one-handers (mean age: 46 ± 10; four left-handers; six female), and 12 two-handed control participants (mean age: 45 ± 15; five left-handers; five female). All amputees reported experiencing phantom sensations after amputation, but vividness of these sensations varied across participants at the time of the study (mean chronic vividness score 58 ± 38 on a 0–100 scale, as assessed using questionnaires [Makin et al., 2013b; Makin et al., 2015Makin et al., 2013a; see Table 1 and Questionnaires section below for further details]). Three participants in the amputees group tested here also took part in our previous study (Kikkert et al., 2016). The congenital one-handers had never experienced any phantom sensations. In addition, we also recruited and excluded a further congenital one-hander (due to technical difficulties during data pre-processing) and two control participants (due to incomplete data collection and due to abnormal digit selectivity, i.e. more than three standard deviations from the mean).

Recruitment was carried out in accordance with the University of Oxford’s Medical Sciences inter-divisional research ethics committee (MS-IDREC-C2-2015-012). Informed consent and consent to publish was obtained in accordance with ethical standards set out by the Declaration of Helsinki. Control participants were recruited as to match the other two groups in term of age, gender and handedness (with respect to the intact hand). When possible, control participants were friends and family of the one-handed participants. All participants were compatible with local magnetic resonance imaging (MRI) safety guidelines.

Experimental procedures

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The experimental procedures described in this manuscript were run as part of a larger study (the full study protocol can be found on https://osf.io/gmvua/). Here we focus on procedures related to the representation of the missing hand in amputees and congenital one-handers.

Questionnaires

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To measure phantom sensations, as well as other demographic and clinical details of potential relevance to the missing hand representation, amputees and congenital one-handers completed a range of questionnaires (as summarised in Table 1). Amputees rated intensities of phantom sensations, using a 0–100 scale, as experienced during the last week (or in a typical week involving such sensations). Chronic phantom sensation was calculated by dividing intensity by sensations frequency (1- all the time; 2- daily; 3- weekly; 4- several times per month; and 5- once or less per month), as previously implemented (Makin et al., 2013b; Makin et al., 2015). Having used this measure in multiple studies with partly overlapping participant pools (Makin et al., 2013b; Kikkert et al., 2016; van den Heiligenberg et al., 2017) we can assess the consistency of this measure within participants and across studies (i.e. measure reliability). We found excellent inter-study consistency (intra-class correlation coefficient: 0.79, 95% CI: .48-.93, F(13,13)=8.46, p<0.001), when considering all amputees that participated in at least one other study (n = 14, earlier questionnaire taken 1–4 years before current study). In addition, participants reported the number of phantom digits that afford kinaesthetic sensations during volitional control of movements (kinaesthesia). This report was further validated by a demonstration of afforded phantom movements during the study’s main task with the intact hand, as detailed below.

MRI tasks

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All participants underwent one experimental session with four fMRI runs, using a block-design. The task involved individual digit-movement blocks for each of the five digits (12 s blocks) of either hand, as well as no movement (rest) blocks. Each condition was repeated three times in a semi-counterbalanced order within each run. Each run comprised a different block order.

To probe somatosensory digit representation, we used a visually cued active (motor) task. In an intact sensorimotor system, movement recruits a combination of peripheral receptors, encoding a range of somatosensory modalities (e.g. surface and deeper mechanoreceptors; proprioceptors), as well as efferent information from the motor system. Using an active task, we have previously shown high consistency of SI digit topography across multiple scanning sessions (Kolasinski et al., 2016, see also Ejaz et al., 2015 for validation using RSA). Participants were presented with five vertical bars, corresponding to the five digits, shown on a visual display projected into the scanner bore. To cue the participant which digit should be moved, the bar corresponding to this digit changed (i.e. by flashing in a different colour).

On ‘missing hand blocks’, participants were instructed to perform individual digit movements (1 Hz) with their nondominant (controls), phantom (amputees), or missing hand (congenital one-handers). Handless individuals were instructed to attempt performing actual movements with the digits of their missing hand, even when not being able to feel their digits, rather than using motor imagery. Controls moved their nondominant hand digits in mid-air. To ensure good understanding of these instructions, outside the scanner, the amputees were asked to demonstrate to the experimenter the extent of volitional movement they felt they were able to carry out in each of their phantom digits, by mirroring each movement onto their intact hand.

On ‘intact hand blocks’, all participants performed a comparable task with their intact/dominant hand by exerting force on a button box. Participants received real-time visual feedback of how much force each digit exerted by means of moving vertical bars on ‘intact hand blocks’, but not on ‘missing hand blocks’. The dominant hand of controls was paired up with the intact hand because, through intensive use, amputees’ and congenital one-handers’ intact hand becomes their de facto dominant hand (Philip and Frey, 2014). All groups were able to carry this task equally well, as verified in post-hoc analysis: each trial was assigned to the digit whose force output correlated most strongly with the instructed time course and the percentage of correctly performed trials, that is trials that were assigned to the instructed digit, was not different between congenital one-handers and amputees (74.2%, t(29)=1.13, p=0.266), or between congenital one-handers (81.1%) and controls (75.3%; t(23)=.93, p=0.362). This behaviour brought forth high representational typicality in intact SI for all three groups (controls: rho = 0.85; amputees: rho = 0.81; and congenital one-handers: rho = 0.89; group comparisons all p’s > 0.11).

MRI acquisition

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MRI images were acquired using a 3T MAGNETON Prisma MRI scanner (Siemens, Erlangen, Germany) with a 32-channel head coil. Functional images were collected using a multiband T2*-weighted pulse sequence with a between-slice acceleration factor of 4 and no in-slice acceleration. This provided the opportunity to acquire data with high spatial (2 mm isotropic) and temporal (TR: 1500 ms) resolution, covering the entire brain. The following acquisition parameters were used: TE: 32.40 ms; flip angle: 75°, 72 transversal slices. Field maps were acquired for field unwarping. A T1-weighted sequence was used to acquire an anatomical image (TR: 1900 ms, TE: 3.97 ms, flip angle: 8°, spatial resolution: 1 mm isotropic).

MRI analysis

MRI analysis was implemented using tools from FSL, SPM and Connectome Workbench software (Smith et al., 2004; Jenkinson et al., 2012, https://fsl.fmrib.ox.ac.uk/fsl/fslwiki/, https://www.fil.ion.ucl.ac.uk/spm/humanconnectome.org) in combination with other Matlab scripts (version R2016a), both developed in-house (Wesselink and Maimon-Mor, 2017) and as part of the RSA Toolbox (Nili et al., 2014). Cortical surface reconstructions were produced using FreeSurfer (Dale et al., 1999; Fischl et al., 2001, freesurfer.net).

fMRI pre-processing

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Functional data was first pre-processed in FSL 5.0. The following steps were included: Motion correction using MCFLIRT (Jenkinson et al., 2002), brain extraction using BET (Smith, 2002), and high pass temporal filtering with a cut-off of 100 s. Co-registration to each individual anatomical T1 scan was accomplished using FLIRT and, where needed, manual adjustments were performed to ensure precise co-registration around the hand knob of the central sulcus.

Anatomical T1 images were used to reconstruct the pial and white-grey matter surfaces using Freesurfer. Surface co-registration across hemispheres and participants was done using spherical alignment. Individual surfaces were nonlinearly fitted to a template surface, first in terms of the sulcal depth map, and then in terms of the local curvature, resulting in a nearly perfect overlap of the fundus of the central sulcus across participants (Fischl et al., 2008).

Regions of Interest (ROI) definition

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Since the focus of the study was on persistent sensory representation, our main analysis was restricted to the individualised hand-selective ROIs in SI. Further analysis was focused on the M1 hand areas. The ROIs were always in the hemisphere contralateral to the missing/nondominant hand. The anatomical ROIs were defined on the group surface using probabilistic cytotectonic maps aligned to the average surface (see Wiestler and Diedrichsen, 2013). These regions were then projected into the individual brains via the reconstructed individual anatomical surfaces. For the hand area of SI, we selected all surface nodes with the highest probability for any of BA3a, 3b, 1, and 2, surrounding the anatomical hand knob (Yousry et al., 1997). The hand area of M1 was selected similarly using BA 4. We note that given the probabilistic nature of these masks, the dissociation between SI and M1 is only an estimate. For one acquired amputee, surface alignment failed; for this subject, the ROIs were drawn manually within the surface ribbon using the above anatomical definitions. The ROIs were not significantly different in size across groups (one-way ANOVA on area volume: SI: F = 1.27, p=0.29; M1: F = 1.37, p=0.27). In addition, for control purposes, we also used an ROI of visual area V5 which we defined anatomically, based on the parameters previously published by Wiestler and Diedrichsen (2013). The ROI was constructed bilaterally and RSA outcome measures were averaged across both hemispheres.

fMRI analysis

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Voxel-wise General Linear Model (GLM) analysis was carried out, as implemented in SPM12. In brief, each of the experimental conditions was modelled for each run separately against rest. Regressors were created by convolving stimulus presentation (as a boxcar function) with a double-gamma hemodynamic response function (HRF). In the GLM estimation, the functional data was weighted using the robust Weighted Least Squares approach (Diedrichsen and Shadmehr, 2005), which estimates the heteroscedasticity of the time series and then ‘soft’-excludes noisy image volumes (e.g. due to movement). Task-related activity was quantified by averaging the BOLD response, averaged across all digits, versus baseline within each ROI. The voxel-wise parameter estimates (hereafter: activity patterns) and residuals from this analysis were also used to calculate the dissimilarity, as detailed below.

The dissimilarity between activity patterns within each ROI was measured for each digit pair using the cross-validated squared Mahalanobis distance, or ‘crossnobis’ distance (Nili et al., 2014). We calculated the distances using each possible pair of imaging runs and then averaged the resulting distances. Before estimating the dissimilarity for each digit pair, the activity patterns were pre-whitened using the residuals from the GLM. Due to cross-validation, the expected value of the distance is zero (but can go below 0) if two patterns are not statistically different from each other, and larger than zero if there is differentiation between the digits of the hand.

We extracted two measures from the resulting inter-digit representational dissimilarity matrix (RDM). As a measure of strength of the representation, we used the mean dissimilarity, the average dissimilarity between the ten unique digit pairs (excluding the diagonal). The typicality of the representational structure was assessed by calculating the Spearman’s rho correlation between the measured RDM and the average RDM of the dominant hand of two-handed controls (independently acquired; see below). Because the representational structure can be related to behavioural aspects of hand use and is highly invariant in controls (average correlation r = 0.9, Ejaz et al., 2015), this measure serves as a proxy for how ‘normal’ the hand representation is. Being able to study this measure was a main reason for using RSA in this study.

As an aid to visualise the RDMs, we also used classical multidimensional scaling (MDS). MDS projects the higher-dimensional RDM into a lower-dimensional space, while preserving the inter-digit dissimilarity values as well as possible (Borg and Groenen, 2005). MDS was performed on data from individual participants and averaged after Procrustes alignment to remove arbitrary rotation induced by MDS. Note that MDS is presented for intuitive visualisation purposes only, and was not used for statistical analysis.

As mentioned above, to determine typicality we correlated RDM from the current study with the average representational structure of the dominant hand of two-handed controls, in an independently acquired cohort of participants. The full details of the acquisition parameters are described in Wesselink and Maimon-Mor, 2017. In short, eight two-handed participants performed an active digit tapping task using a button box (four repetitions per digit of 8 s blocks of 1 Hz single-digit presses), without online visual feedback. The data was acquired at 7T (TR: 2000 ms, TE: 25 ms, voxel size: 1 × 1×1 mm). The ROI was defined similarly to the SI ROI used in the current study.

Statistical analysis

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Statistics were calculated using Matlab R2016a. Subsequent to normality validation (using the Shapiro-Wilk test), we used paired/independent-sample two-tailed Student’s t-tests to compare activity levels and distance measures within/between groups, and one-sample t-tests to compare group measures to zero. Correlations were calculated as Spearman’s rho. Partial correlation effects were calculated using linear regression. For the main analysis concerning RSA, within each ROI, group t-tests were adjusted to three comparisons, using the Bonferroni correction (α = 0.05/3), to account for the three inter-group comparisons. Post-hoc correlations between SI typicality and key clinical measurements (phantom sensations, phantom kinaesthesia and time since amputation) were also corrected for three comparisons (α = 0.05/3). One post-hoc comparison involving a small subset of amputees was done using a Mann-Whitney U test (see Results - section 2). For a control analysis, involving visual and somatosensory ROIs, we also used a mixed-design analysis of variance (ANOVA) to identify interactions across groups and ROIs.

In order to assess whether any aspect of the representational structure in the amputees was not different from that in controls, we used Bayesian statistics as implemented in Javascript (Dienes, 2014; Singh, 2018). Our alternative hypothesis is that amputees have no preserved hand representation. To construct our prior (i.e. to quantify the effect of having hand representation) we calculated the effect size of controls’ nondominant hand representation vs. congenital one-handers missing hand representation. We then compared the effect size of amputees’ missing hand representation (compared to controls) against that prior. More specifically, our alternative hypothesis assumes an effect size following a one-tailed t-distribution centred at 0 and a width of the difference between congenital one-handers and controls. The measured difference between amputees and controls (also modelled as a t-distribution) are tested against this hypothesis. Support for the null hypothesis was interpreted as supporting preserved hand representation. While it is generally agreed that it is difficult to establish a cut-off for what consists sufficient evidence, we used the threshold of BF<1/3 as positive evidence in support of the null, consistent with others in the field regarding this threshold as providing substantial evidence (Wetzels et al., 2011; Dienes, 2014). Note, however, that this threshold is not considered as providing strong evidence by all accounts (Kass and Raftery, 1995).

In order to gauge which aspects of phantom sensation and key demographics may relate to the amputees’ representational structure’s typicality, we performed an exploratory forward stepwise regression. The dependent variable was SI typicality in amputees. The following factors were used as independent variables (see also Table 1): typicality of the intact hand (rho; calculated from the intact hand SI area; time since amputation (in years); age at amputation (in years); vividness of nonpainful phantom sensations, as experienced during the study (on a 0–100 scale) and chronically (accounting for both intensity and frequency; Makin et al., 2013b); intensity of phantom limb pain, as experienced acutely during the study, and chronically (as detailed for nonpainful sensations). Only linear factors were considered, that is no interaction terms, and the criterion for inclusion was an increase in R2 >0.1. As a large number of predictor variables were included in the model and stepwise regression is generally only recommended for exploratory analysis, we aimed to establish internal replicability using bootstrap resampling (e.g. Thompson, 1995). In particular, we randomly sampled (with replacement) the full data matrix and repeated the stepwise regression 1000 times. We subsequently computed the proportion of bootstrap samples in which each factor was included in the final model, as well as confidence bounds on the model’s adjusted R2. We interpreted high proportion of inclusion (p>0.75) as evidence for internal replicability (Thompson, 1995).

Table 1
Summary demographic details and phantom sensations.

Data is shown for amputees (AMP), controls (CTR) and congenital one-handers (1H). Congenital one-handers did not feel any phantom limb sensations. All controls have full kinaesthetic sensations. F: female, M: male. Side: side of missing hand; L: left, R: right. Amputation level: 1: shoulder, 2: above elbow; 3: at elbow; 4: below elbow; 5: at wrist. Kin: Phantom limb kinaesthesia (number of independent controllable parts of the hand), Viv: Chronic phantom limb vividness (0: no sensation, 100: intact hand’s vividness), Pain: Chronic phantom limb pain (0: no pain, 100: worst pain imaginable), AViv/APain: Acute Viv/Pain (on the scanning day), Std: standard deviation, ND: nondominant.

https://doi.org/10.7554/eLife.37227.006
AMPAge
(years)
SexAmputationPhantom sensations
SideYears sinceAge at
(years)
LevelKin
(0-5)
Viv
(0-100)
Pain
(0-100)
AViv
(0-100)
APain
(0-100)
Mean50.417.632.9358466521
St. dev.12.110.411.8238373023
A0144MR15292510010010050
A0253ML322125501006070
A0340ML1129241005010020
A0451ML32192510001000
A0527FR720425040600
A0671MR16552120856020
A0746MR18282370907050
A0856ML263045640100
A0964ML3133241004010010
A1058ML25623900800
A1128ML820544040200
A1257MR29282180908040
A1350FL149400000
A1452MR272525100808050
A1568MR264241160800
A1639FR9303435405030
A1758ML12464520650
A1846FL14324380305030
CTRAge
(years)
SexND hand1HAge
(years)
SexMissing hand
SideSideLevel
Mean45.3Mean45.7
St. dev.14.9St. dev.10.4
C0129MRH0141ML4
C0224FLH0237MR4
C0347FLH0331FL4
C0439MLH0460ML4
C0532MRH0539FL4
C0653FRH0654FL4
C0738FRH0734ML4
C0867MRH0863ML4
C0942MRH0944FR4
C1041MRH1055FL4
C1169MLH1146MR4
C1263FLH1237MR4
H1353FL4

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Decision letter

  1. Eve Marder
    Senior and Reviewing Editor; Brandeis University, United States

In the interests of transparency, eLife includes the editorial decision letter and accompanying author responses. A lightly edited version of the letter sent to the authors after peer review is shown, indicating the most substantive concerns; minor comments are not usually included.

Thank you for submitting your article "Obtaining and maintaining cortical hand representation as evidenced from acquired and congenital handlessness" for consideration by eLife. Your article has been reviewed by Sabine Kastner as the Senior and Reviewing Editor, and two reviewers.

The reviewers have discussed the reviews with one another and the Reviewing Editor has drafted this decision to help you prepare a revised submission.

Summary:

This study uses fMRI to assess the somatotopic organisation of S1 (and M1) in healthy controls, in amputees and in congenitally one-handed participants. Two approaches are taken. The first is to instruct participants to "move" their (absent) digits. Then a representational similarity analysis is conducted to test for the representational structure across (digit-) conditions. BOLD activity, average representational dissimilarity and "typicality" (a comparison to controls) are similar between controls and amputees, but virtually absent in congenital one-handers. Second, as a control to ensure that this is not simply due to an inability of congenital one-handers in performing the task, a comparison is made using the ipsilateral representation of the other hand, confirming the initial findings.

The experiment builds upon and extends a previous paper by the group published in eLife. The difference is that the current paper includes congenital one-handed participants, which allows one to tell whether the representational structure is experience-dependent. The reviewers thought that this is a valuable addition to the previous study. However, there were a number of residual concerns that need to be addressed further.

Essential revisions:

1) For the first finding, the only support is a correlation between one imaging measure (representational typicality) and one questionnaire response (# of independently controllable parts of the hand). Given this, one would expect very large differences between those who can and cannot move their fingers independently – not only in representational typicality, but also in dissimilarity. Surprisingly, the correlation between kinaesthesia and dissimilarity is not reported in the manuscript. As stated in the Introduction, "greater individuation of inputs will induce greater representational dissimilarity." Therefore, if amputees vary on self-reported individuation, then there should be a correlation between these two variables. This needs to be reported. If there isn't a relationship between the two variables, then it makes it quite difficult to interpret the significant correlation between typicality and "kinaesthesia". Second, it is difficult to be confident about correlations with questionnaire data with a small sample size (e.g. the 95% CI for the reported correlation ranges from.33 to.89). Therefore, the relationship between typicality and kinaesthesia would be more convincing if there were other, related correlations that are also significant. (That is, it's hard for us to be confident about one correlation amongst a number of questionnaire and imaging variables, without some additional results that support the same narrative.) Further, the authors stated in the introduction that they would examine "whether persistent representation of a missing hand…will also be observed in amputees with little phantom sensations." Based on this, we were expecting an analysis of the specific individuals with little/no phantom sensations (i.e. some examination of these specific participants, showing that they do (or do not) have a phantom representation similar to intact controls). This should be included in a future submission.

2) The authors show that congenital amelics demonstrate reduced activity and digit discriminability in S1 for phantom movements compared to controls. We share some of the concerns of an earlier reviewer. Given that the authors suggest here and previously (Kikkert et al., 2016) that the S1 activity observed when moving the phantom is due to efference copy, it is unlikely that congenital amelics can generate efference copy for a limb that never existed. However, the lack of efference copy activation in S1 of congenital amelics is not the same as demonstrating that the hand representation has not developed without sensory experience – one of the goals of this paper (see Introduction). That is, there could be a hand representation that exists, but simply hasn't been accessed with the movement task. The manuscript, at times, seems to conflate a lack of activation in S1 for moving a phantom/non-existent hand as evidence for the lack of any hand representation. Although we are sympathetic to this view, we don't think the evidence is there given this single task – and so the claims should be softened.

3) The authors find that controls and amputees did not significantly differ across a number of variables. One example is that typicality did not differ between the two groups. The authors used Bayes factors, using a BF<1/3 as positive evidence in support of the null. Given the editor's concerns regarding exactly what "substantial" evidence means in the context of BFs, one suggestion would be to use frequentist tests designed specifically to examine whether two groups are equivalent (e.g. two one-sided tests for equivalence (TOSTER), Lakens, 2017). This provides a standard p-value that is more straightforward to interpret and is consistent with the (primarily) frequentist tests used throughout the manuscript.

4) The argument for using the ipsilateral hand representation to probe digit structure in the missing hand (Results section) was quite unclear. We understand that Diedrichsen showed a relationship between ipsilateral and contralateral digit representation. But this was in individuals with intact hands. Why would the fact that the "ipsilateral digit representation is a reliable predictor of contralateral hand representation" in two-handers mean that this would also be true for amputees or congenital one-handers? This logic should be laid out.

5) We suggest to briefly explain that the absence of information for the congenital one-handed participants means that the effect is not driven by the visual cue.

6) Is the MDS-analysis cross-validated?

7) To which degree is the typicality analysis influenced by SNR? Does the low dissimilarity of the congenital group (Figure 1) mean that the responses were reproducibly very similar across conditions (i.e. SNR presumably is not responsible for the absence of typicality)?

8) Is this a completely separate dataset than in the original paper? This should be clarified.

[Editors’ note: further revisions were requested before acceptance.]

Thank you for resubmitting your work entitled "Obtaining and maintaining cortical hand representation as evidenced from acquired and congenital handlessness" for further consideration at eLife. Your revised article has been favorably evaluated by Eve Marder (Senior Editor) and two reviewers.

The manuscript has been improved but there are some remaining issues that need to be addressed before acceptance, as outlined below: We assume that you will be able to address these issues by editorial revision of the text. Reviewer #1 is now satisfied, and Reviewer 2 agrees that modest textural revisions will be sufficient, and I see no reason why these would need to go back to the reviewers as long as you address the issues.

From Reviewer 2 "With regards to the comment starting with "Further, the authors stated in the Introduction that they would examine.…", my question was regarding whether those with little/no phantom sensations were SIMILAR TO INTACT CONTROLS. However, the author's comments seem to be related to whether these individuals were different from congenital one-handers. First, they say that the intercept for the regression line between phantom vividness and typicality was greater than zero. While this provides evidence for some representational similarity in those with low vividness, it does not mean that they are similar to intact controls. For their second point, they state that the 3 participants with little/no phantom vividness have higher typicality than congenital one-handers. While true, higher typicality than congenital one-handers is not the same as "similar to intact controls". This is not a major point, and doesn't need to be addressed, but could improve the manuscript.

More importantly, with regards to point #4 (and also point #2), we do understand the evidence showing a relationship between ipsilateral and contralateral digit representations in neurologically intact individuals, and the logic that the authors use here. However, no mechanistic argument is presented as to why such a relationship would occur in congenital one-handers, even if they had an intact "missing hand" representations. As it stands, the reader is left with the following implied argument: Since there is a precise correlation between the representational patterns for moving the contralateral hand and ipsilateral hand in the same is, then it must be that representational patterns for the ipsilateral hand assess the representation of the missing hand.

This argument seems to be based on the idea that a strong correlation = causal evidence. Just because this correlation was observed does not mean that moving the ipsilateral hand is necessarily a measure of the contralateral hand representation. For example, one possibility is that in individuals that have had hands at some point in their life, there is a strong relationship between the two representations such that the activity for moving the ipsilateral and contralateral hand in one hemisphere map perfectly. However, there is no mechanistic reason presented to think that this relationship would necessarily hold in those who have never had a hand….it could be that moving the ipsilateral hand does not result in any kind of organized activity in the "missing hand" hemisphere in this population.

Note that I think that this analysis is clever and creative, and I am sympathetic to it. However, as it stands, I don't see the clear mechanistic account for a) why activity for moving the ipsilateral hand is an index of the contralateral hand representation (apart from the correlational argument, which I fear is flawed) and b) why your argument would necessarily hold in congenital one-handers. I believe this could be improved with some mechanistic arguments to support the claim. There is some work on what ipsilateral activation indexes (see Diedrichsen, Wiestler & Krakauer, 2013; prior work on mirror movements)…maybe there is something in this literature that could strengthen the claims?"

https://doi.org/10.7554/eLife.37227.012

Author response

The experiment builds upon and extends a previous paper by the group published in eLife. The difference is that the current paper includes congenital one-handed participants, which allows one to tell whether the representational structure is experience-dependent. The reviewers thought that this is a valuable addition to the previous study. However, there were a number of residual concerns that need to be addressed further.

We agree with the reviewers’ summary but would like to add that the key innovation extends beyond the study of congenital one-handers. Primarily, whereas all previous studies (both from our own group and beyond) tested people with highly vivid phantom sensations, we have tested a large group of amputees with varying phantom sensations, allowing us to determine whether the experience of having a hand is necessary for the maintenance of the digit maps. Also, our M1 results further complement our original report and are of interest to many people in the field, particularly those working on brain-machine interfaces.

Essential revisions:

1) For the first finding, the only support is a correlation between one imaging measure (representational typicality) and one questionnaire response (# of independently controllable parts of the hand). Given this, one would expect very large differences between those who can and cannot move their fingers independently – not only in representational typicality, but also in dissimilarity. Surprisingly, the correlation between kinaesthesia and dissimilarity is not reported in the manuscript. As stated in the Introduction, "greater individuation of inputs will induce greater representational dissimilarity." Therefore, if amputees vary on self-reported individuation, then there should be a correlation between these two variables. This needs to be reported. If there isn't a relationship between the two variables, then it makes it quite difficult to interpret the significant correlation between typicality and "kinaesthesia".

We agree with the reviewers’ reasoning. Since our main prediction concerned typicality and not dissimilarity (based on our original report), we wished to limit the number of comparisons. Yet, based on the reviewers’ suggestion, we have now tested this correlation and added it to the manuscript. As stated below, we found a (trending) positive correlation between kinaesthesia and dissimilarity (rho=.447, p=.063).

Subsection “Missing hand representation in acquired amputees is persistent even after phantom sensations have diminished”: “The correlation between kinaesthesia and dissimilarity in SI approached significance (rho=.447, p=.063).”

Second, it is difficult to be confident about correlations with questionnaire data with a small sample size (e.g. the 95% CI for the reported correlation ranges from.33 to.89). Therefore, the relationship between typicality and kinaesthesia would be more convincing if there were other, related correlations that are also significant. (That is, it's hard for us to be confident about one correlation amongst a number of questionnaire and imaging variables, without some additional results that support the same narrative.)

We agree that, where possible, questionnaires should be validated with other empirical evidence. Since kinaesthesia was one of our two key predictors (along with chronic phantom sensations) we actually asked participants to show us the range of their missing hand movement by mimicking it with their intact hand (see the full study protocol on https://osf.io/gmvua/). This was recorded on video immediately after the scan. Unfortunately, due to a human error, we lost a substantial amount of this data and only have a sub-set of the first 8 acquired amputees tested. These existing videos strongly confirm the self-reports: rates estimated from the videos are strongly correlated with those estimated in the questionnaires (rho=.840). Considering the small sample size, we felt that this extended evidence does not substantially benefit the paper. However, should the editor or reviewers feel differently, we will be happy to include this supporting information.

Regardless, the reviewers make a strong point that we had multiple measures that could potentially be relevant to phantom finger representation. In our first submission, we attempted to deal with this by using a stepwise regression analysis as an exploratory tool. We had to be careful not to overfit the data, so we chose 7 measures that we felt were most appropriate for the missing hand’s typicality measure – time since amputation, age at amputation, kinaesthesia, non-painful phantom vividness (both acute and chronic), and phantom limb pain (both acute and chronic) – as well as typicality of the intact hand’s contralateral activity patterns. The analysis confirmed that kinaesthesia was most relevant to our fMRI measure, allowing us to focus on kinaesthesia in our Results section. We strongly agree that without our regression analysis this focus is not sufficiently justified. Yet, we were also attuned to the original reviewers’ concern that our regression model is not sufficiently reliable for its exploratory purpose.

In the revised manuscript we implemented a bootstrapping approach to provide a measure of confidence for the output of the regression analysis. The final model included an intercept and kinaesthesia as the only regressor (p<.001), with an adjusted R2 of.645 (95% confidence interval:.30-.99). Out of 1000 iterations, our measures were included in the final model 96% of the times. We believe that this analysis adds clarity to our Results section without reducing the rigidity of the overall findings. Nevertheless, as in our original finding, we suggest this analysis as an exploratory means only, and do not interpret it further. We have updated the manuscript as follows:

Subsection “Missing hand representation in acquired amputees is persistent even after phantom sensations have diminished”: “Next, we evaluated whether the consistency of hand representation in SI during missing hand movements correlates with amputees’ subjective reports of phantom sensations. We first carried out an exploratory forward stepwise regression with typicality as the dependent variable. The following factors were tested as independent variables: kinaesthesia of phantom sensations (the number of phantom digits perceived as independently moving during the phantom movement task; vividness of nonpainful phantom sensations as experienced both during the study and chronically; intensity of phantom limb pain, as experienced both acutely during the study and chronically; time since amputation; age at amputation and typicality of the intact hand (calculated from the intact hand SI area). The final model (F=19.9, p<.001, adjusted R2=.645 included only kinaesthesia of phantom sensations (β=.07, t=4.46, p<.001) and the intercept (β=.52, t=8.89, p<.001). This regression was submitted to a bootstrapping analysis, allowing us to estimate the consistency of the final model (see Materials and methods). This bootstrapping analysis returned kinaesthesia as the final variable in.963% of the iterations (final model fit: median adjusted R2=.645; 95% CI:.30-.99). The proportion of the other included factors in the final model was: typicality of the intact hand (7.0%); time since amputation (9.5%); age at amputation (9.6%); vividness of nonpainful phantom sensations (acute: 12.2%; chronic: 20.8%); phantom limb pain, acute: 22.7%; chronic: 10.1%).”

Subsection “Statistical Analysis”: “In order to gauge which aspects of phantom sensation and key demographics may relate to the amputees’ representational structure’s typicality, we performed an exploratory forward stepwise regression. The dependent variable was SI typicality in amputees. The following factors were used as independent variables (see also Table 1): typicality of the intact hand (rho; calculated from the intact hand SI area; time since amputation (in years); age at amputation (in years); vividness of nonpainful phantom sensations, as experienced during the study (on a 0-100 scale) and chronically (accounting for both intensity and frequency; Makin et al., 2013b); intensity of phantom limb pain, as experienced acutely during the study, and chronically (as detailed for nonpainful sensations). Only linear factors were considered, i.e. no interaction terms, and the criterion for inclusion was an increase in R2 > 0.1. As a large number of predictor variables were included in the model and stepwise regression is generally only recommended for exploratory analysis, we aimed to establish internal replicability using bootstrap resampling (e.g. Thompson, 1995). In particular, we randomly sampled (with replacement) the full data matrix and repeated the stepwise regression 1000 times. We subsequently computed the proportion of bootstrap samples in which each factor which included as well as confidence bounds on the model’s adjusted R2. We interpreted high proportion of inclusion (p >.75) as evidence for internal replicability.”

Further, the authors stated in the introduction that they would examine "whether persistent representation of a missing hand…will also be observed in amputees with little phantom sensations." Based on this, we were expecting an analysis of the specific individuals with little/no phantom sensations (i.e. some examination of these specific participants, showing that they do (or do not) have a phantom representation similar to intact controls). This should be included in a future submission.

Indeed, this question was of interest to us. We explored this in our previous submission in two different ways (see Results section of our previous submission). First, we showed that the intercept for the regression line between phantom vividness and typicality was significantly greater than 0 (.75). This means that, based on our population of n=18 amputees, individuals with no phantom sensation are still predicted to show some hand typicality. Secondly, we show that the 3 participants experiencing little to no phantom vividness (below 10/100) showed significantly higher typicality than congenital one-handers. In the revised manuscript, we have reordered the presentation of results and elaborated these results so that they are not easily missed by the readers.

Subsection “Missing hand representation in acquired amputees is persistent even after phantom sensations have diminished”: “Regardless of the positive relationship between kinaesthesia and typicality, amputees with little to no kinaesthetic sensations still showed missing hand representation. As stated above, the regression line between kinaesthesia and typicality included an intercept (rhointercept=.52). This was also the case when phantom vividness was the (non-significant) dependent variable (F=.021, p=.89, Adjusted R2 =-.061; rhointercept =.75, p<.001). These results predict that even amputees who do not experience any phantom sensations will retain some typical missing hand representation. To test this prediction directly, we examined the 3 amputees in our dataset showing weak to no chronic phantom vividness (below 10/100). Despite not being able to experience clearly their phantom hand when performing the phantom movements task, these individuals showed high typicality (average typicality (rho) =.83). Moreover, when comparing their typicality to that found in the congenital one-handers (who were arguably better matched to this sub-group in terms of task demands), the amputees with diminished phantom sensations showed significantly stronger correlations with the canonical hand structure (Mann-Whitney U = 38, p=.007). Together, these additional analyses confirm that the representational structures’ typicality in SI of amputees is still present in those with little to no phantom or kinaesthetic sensations.”

2) The authors show that congenital amelics demonstrate reduced activity and digit discriminability in S1 for phantom movements compared to controls. We share some of the concerns of an earlier reviewer. Given that the authors suggest here and previously (Kikkert et al., 2016) that the S1 activity observed when moving the phantom is due to efference copy, it is unlikely that congenital amelics can generate efference copy for a limb that never existed. However, the lack of efference copy activation in S of congenital amelics is not the same as demonstrating that the hand representation has not developed without sensory experience – one of the goals of this paper (see Introduction). That is, there could be a hand representation that exists, but simply hasn't been accessed with the movement task. The manuscript, at times, seems to conflate a lack of activation in SI for moving a phantom/non-existent hand as evidence for the lack of any hand representation. Although we are sympathetic to this view, we don't think the evidence is there given this single task – and so the claims should be softened.

We are grateful for this comment, as this is a point that we were very keen to clearly address in our previous submission, and we therefore appreciate the opportunity to improve our delivery. The task constraints imposed on congenital one-handers are similar to those encountered by the few acquired amputees that no longer experience phantom sensations. As highlighted above, these individuals were still able to show higher typicality then congenital one-handers (see above). Regardless, as we highlighted in the original manuscript (and emphasise further in the revised manuscript), we recognise that the task we used might have induced performance differences across the one-handed groups. For this reason, we designed two further analyses that are based on intact hand movements, which all participants could perform equally.

The representational structure of contralateral hand movement can be studied indirectly by examining activity in the same brain region induced by the ipsilateral hand (Diedrichsen et al., 2017). Just to reiterate this point, as an example – left hand movements would evoke strong activity patterns in the contralateral hand area (in the right hemisphere). When participants are moving their ipsilateral (right) hand, that same brain area would show activity patterns, that although smaller, crucially reflect the same representational inter-digit relationships (we elaborate on this key finding below; see comment 4). Because of this, the deprived hand area can be studied based on the ipsilateral intact hand, without imposing unnatural task demands on the congenital one-handers. If congenital one-handers truly have diminished hand representation for their missing hand, their ipsilateral movement should not induce typically structured activity patterns, whereas such activity patterns are expected for the acquired amputees and controls. To test this very specific prediction, we recorded individuated finger movements, which all groups were able to perform equally well. We find that ipsilateral activity following intact hand movement is structured similarly in acquired amputees and controls, but it is reduced in congenital one-handers. In fact, missing hand representation in congenital one-handers was found to be diminished to the point where it is not different from a control visual area.

As such, we are basing our interpretation on multiple converging pieces of evidence: diminished missing hand representation in congenital one-handers using our missing fingers movement task; persistent missing hand representation in acquired amputees with diminished-to-nonexisting phantom sensations; diminished ipsilateral intact hand representation in congneital one-handers; similar ipsilateral digit information in SI and V5 in congenital one-handers, but not amputees/controls.

Nevertheless, we have done our best to frame these results cautiously, e.g. in the abstract and elsewhere we refer to the congenital one-handers missing hand representation as “reduced” or “diminished” rather than non-existent. In the discussion we acknowledge the limitations of our results and call for further research to conclusively resolve this question. We also refrain from inferring that the missing hand representation never developed (as indicated by the reviewers), and instead suggest that the missing hand maps could have originally formed (e.g. based on a genetic blue-print) but later diminished due to insufficient relevant inputs.

In the revised manuscript we have re-written substantial portions of the text (including the abstract), and better structured the Results section (by grouping the findings into sub-sections) to communicate these ideas more clearly. Here are a few examples of revised text addressing this point:

Abstract:

“We used representational similarity analysis in primary somatosensory and motor cortex during missing and intact hand movements. We found that key aspects of acquired amputees’ missing hand representation persisted, despite varying vividness of phantom sensations. In contrast, missing hand representation of congenital one-handers, who do not experience phantom sensations, was significantly reduced. […] We conclude that once cortical organisation is formed, it is remarkably persistent, despite long-term attenuation of peripheral signals.”

Discussion section:

“Still, it is possible that rudimentary missing hand representation, e.g. determined by genetic factors (Miyashita-Lin et al., 1999, Rubenstein et al., 1999), has originally formed in congenital one-handers but later diminished due to lack of consistent sensorimotor input. Our results were insufficiently conclusive to address this important question. Bearing this caveat in mind, our findings suggest that early-life experience is necessary to create typical functional sensory organisation, but not to maintain it.”

3) The authors find that controls and amputees did not significantly differ across a number of variables. One example is that typicality did not differ between the two groups. The authors used Bayes factors, using a BF<1/3 as positive evidence in support of the null. Given the editor's concerns regarding exactly what "substantial" evidence means in the context of BFs, one suggestion would be to use frequentist tests designed specifically to examine whether two groups are equivalent (e.g. two one-sided tests for equivalence (TOSTER), Lakens, 2017). This provides a standard p-value that is more straightforward to interpret and is consistent with the (primarily) frequentist tests used throughout the manuscript.

We appreciate the suggestion to substitute our Bayesian analysis with the frequentist equivalence test TOSTER. After implementing this analysis, we conclude that while the resulting p value might be more intuitive to interpret, the effect size estimate required for conducting the test is conceptually quite tricky. The goal in the TOSTER approach is to specify lower and upper bounds, such that group differences falling within this range are deemed non-meaningful. But how do we choose the a priori equivalence bounds for the existence of a hand representation? One option was using the effect size of the group difference between the controls (who have a hand) and the congenital one-handers (who do not have a hand). Using this criterion, the typicality of the amputees was significantly equivalent to that of the controls (t=3.88, p<.000). But although appropriate for the Bayesian analysis, this boundary might be not suitable for TOSTER. This is because the lower bound should represent the smallest reasonable effect of having a hand, whereas we believe the congenitals do not have hand representation altogether. In other words, the congenital one-handers fall outside this lower boundary. Alternative measures of the lower hand representation boundary can come from the control participants: e.g. contralateral vs. ipsilateral univariate activity (t=6.68; p<.000) or the comparison between SI and V5 typicality (t=6.69; p<0.000). But the fact that these tests produce stronger evidence in favour of the null than our experimental results with the congenital one-handers lead us to doubt this boundary as meaningful as well.

While the TOSTER method clearly supports our main interpretation, in our view, there are two main reason why the Bayesian analysis is better suited to our paper. First, the inclusion of the congenital one-handers in our study was specifically designed to allow us to interpret the persistence of missing hand representation in the acquired amputees. Indeed, the Bayesian analysis allows us to compare the group difference ‘amputees vs controls’ as a measure of having a hand representation. Secondly, the TOSTER method is not quite standard in the field and much less widespread than the Bayesian statistics we have used. Therefore, we prefer implementing the current gold standard for interpreting null results (note that our BF for typicality is.128). Should the editor or reviewers feel that the TOSTER provides substantial further evidence, we will be happy to include it as well.

4) The argument for using the ipsilateral hand representation to probe digit structure in the missing hand (Results section) was quite unclear. We understand that Diedrichsen showed a relationship between ipsilateral and contralateral digit representation. But this was in individuals with intact hands. Why would the fact that the "ipsilateral digit representation is a reliable predictor of contralateral hand representation" in two-handers mean that this would also be true for amputees or congenital one-handers? This logic should be laid out.

We apologise for being unclear. We have adapted the text more explicitly address this crucial point.

Subsection “Diminished missing hand representation in congenital one-handers even when task performance is matched”: “To probe digit structure in the missing hand cortex using an alternative task, we examined ipsilateral hand representation of the intact hand (i.e. in the missing hand cortex). Ipsilateral digit representation in two-handed controls was previously shown to precisely predict contralateral hand representation (Diedrichsen et al., 2018). In other words, individuated digit movements with the ipsilateral hand in sensorimotor cortex produce an identical representational pattern to that of the contralateral hand (though this pattern can be masked by noise). As such, ipsilateral representation of the intact hand provides an indirect assay into the representation of the missing hand, while controlling for task demands across groups. Importantly, all three groups were able to perform the individuated digit movement task, with similar success (see Materials and methods). We compared the intact/dominant inter-digit representational structure in the missing/nondominant hand area of one-handers/controls (respectively). We predicted that persistent missing hand representation in amputees should result in similar ipsilateral representation in their missing hand cortex as controls. If missing hand representation is diminished in congenital one-handers, then ipsilateral representation of their intact hand (in the missing hand area) should show reduced representational features as those found in amputees.”

Discussion section:

“We further show that ipsilateral digit-representation in the missing hand cortex of the intact hand supports the existence of persistent missing hand representation in amputees and diminished representation in congenital one-handers. Previous studies demonstrate that contralateral and ipsilateral hand movements produce identical representational patterns (Diedrichsen et al., 2018). As such, the ipsilateral digit representational pattern is a reliable predictor of contralateral hand representation. This discovery provides us with a unique and novel opportunity to interrogate the information content underlying the “deprived” hand cortex despite the physical absence of the hand. This approach provided converging evidence, but using different task demands, for similar (missing) hand representation for amputees and controls, but not for congenital one-handers.”

5) We suggest to briefly explain that the absence of information for the congenital one-handed participants means that the effect is not driven by the visual cue.

That is a good point. We have added it to our revised manuscript.

Discussion section:

“Using the same task, we were unable to identify similar digit representation for the missing hand of congenital one-handers, demonstrated by significantly reduced pattern typicality compared to both amputees (and even in comparison to those few amputees with little phantom sensations), as well as controls. This result confirms that the persistent hand representation observed in amputees does not reflect mere cognitive task demands (e.g. visual feedback, Kuehn et al., 2018; attention, Puckett et al., 2017).”

6) Is the MDS-analysis cross-validated?

We believe that MDS does not easily lend itself to quantitative analysis, therefore in the manuscript we have relied on cross-validated Mahalanobis distances for all analyses and only used the MDS for visualisation purposes. This is now clarified in the Introduction, in both the Materials and methods section and the Figure legend.

Subsection “fMRI analysis”:

Note that MDS is presented for intuitive visualisation purposes only, and was not used for statistical analysis.

Figure 1:

E) Two-dimensional projection of the representational structure (D) (using multi-dimensional scaling; note that this is included for visualisation purposes only and was not used for statistical analysis).

7) To which degree is the typicality analysis influenced by SNR? Does the low dissimilarity of the congenital group (Figure 1) mean that the responses were reproducibly very similar across conditions (i.e. SNR presumably is not responsible for the absence of typicality)?

This is a good point that we didn’t address so far. When the SNR is low, dissimilarity will tend to 0. Typicality will therefore also be indirectly affected by SNR. In our view, this is not a design flaw, but, in the case of the congenital group, an indication that although the missing hand area of congenital one-handers was activated, it was not activated consistently across scans. To directly address the reviewers’ suggestion, we tested whether the representational patterns evoked by the congenital one-handers are reproducible, by calculating split-half consistency. For each participant, we calculated an RDM for the missing/non-dominant hand using the two odd runs, and one using the two even runs. The correlation between the odd and even RDMs was significantly lower in the congenital one-handers (rho=-.02) compared to both amputees (rho=.41; p1H-AMP=.001) and controls (rho=.52; p1H-CTR=.001). We note that by splitting the data we are reducing the effectiveness of our analysis (which was designed to rely on 4 scans for cross validation). Nevertheless, the relative difference in split-half consistency between the congenital one-handers and the other two groups indicates that there is no strongly consistent digit information in the missing hand area of the congenital one-handers during our task. We have added the following text to the manuscript to reflect this point:

Subsection “Phantom hand movements elicit typical hand representation in the missing hand area of acquired amputees”: “Although the inter-digit representational structure of congenital one-handers is atypical with respect to canonical hand representation, it is possible that it is still consistent within participants. To explore this idea, we split individual participants’ data to odd and even scans. For each participant, we calculated an RDM in the missing/nondominant hand area using the odd and even runs, and correlated the two RDMs. The correlation between odd and even RDMs was significantly lower in congenital one-handers (rho=-.02) compared to both amputees (rho=.41; p1H-AMP=.001) and controls (rho=.52; p1H-CTR=.001). We note that by splitting the data we are reducing the effectiveness of our analysis. Nevertheless, the relative reduction in split-half consistency indicates that there is no strongly consistent digit information in the missing hand area of congenital one-handers during this task.”

8) Is this a completely separate dataset than in the original paper? This should be clarified.

We apologise that we did not mention this in our original submission. All three participants from the original paper were scanned again in the current study, namely participants A4, A7, and A2 (respectively to their order in the original paper). After excluding these participants, our two main results still hold: typicality is not significantly reduced in the amputees compared to controls (t=-1.19, df=25, p=.247), and is significantly correlated with kinaesthesia (rho=.70, p=.004). We have added this information to the text.

Subsection “Participants”:

“3 participants in the amputees group tested here also took part in our previous study (Kikkert et al., 2016).”

[Editors’ note: further revisions were requested before acceptance.]

From reviewer 2 "With regards to the comment starting with "Further, the authors stated in the Introduction that they would examine…", my question was regarding whether those with little/no phantom sensations were SIMILAR TO INTACT CONTROLS. However, the author's comments seem to be related to whether these individuals were different from congenital one-handers. First, they say that the intercept for the regression line between phantom vividness and typicality was greater than zero. While this provides evidence for some representational similarity in those with low vividness, it does not mean that they are similar to intact controls. For their second point, they state that the 3 participants with little/no phantom vividness have higher typicality than congenital one-handers. While true, higher typicality than congenital one-handers is not the same as "similar to intact controls". This is not a major point, and doesn't need to be addressed, but could improve the manuscript.

We agree that adding the direct comparison between controls and amputees with little no phantom vividness improves the manuscript and apologise for not having included this initially. The difference between these groups is strongly non-significant (as supported by a Bayesian analysis). We’ve appended the following sentence to the manuscript:

Results section:

“These results predict that even amputees who do not experience any phantom sensations will retain some typical missing hand representation. To test this prediction directly, we examined the 3 amputees in our dataset showing weak to no chronic phantom vividness (below 10/100). Despite not being able to experience clearly their phantom hand when performing the phantom movements task, these individuals showed high typicality (average typicality (rho) =.83). Moreover, when comparing their typicality to that found in the congenital one-handers (who were arguably better matched to this sub-group in terms of task demands), the amputees with diminished phantom sensations showed significantly stronger correlations with the canonical hand structure (Mann-Whitney U = 38, p=.007). Typicality was not different between these 3 amputees and controls (Mann-Whitney U = 29, p=.52, BF=.089). Together, these additional analyses confirm that the representational structures’ typicality in SI of amputees is still present in those with little to no phantom or kinaesthetic sensations.”

More importantly, with regards to point #4 (and also point #2), we do understand the evidence showing a relationship between ipsilateral and contralateral digit representations in neurologically intact individuals, and the logic that the authors use here. However, no mechanistic argument is presented as to why such a relationship would occur in congenital one-handers, even if they had an intact "missing hand" representations. As it stands, the reader is left with the following implied argument: Since there is a precise correlation between the representational patterns for moving the contralateral hand and ipsilateral hand in the same is, then it must be that representational patterns for the ipsilateral hand assess the representation of the missing hand.

This argument seems to be based on the idea that a strong correlation = causal evidence. Just because this correlation was observed does not mean that moving the ipsilateral hand is necessarily a measure of the contralateral hand representation. For example, one possibility is that in individuals that have had hands at some point in their life, there is a strong relationship between the two representations such that the activity for moving the ipsilateral and contralateral hand in one hemisphere map perfectly. However, there is no mechanistic reason presented to think that this relationship would necessarily hold in those who have never had a hand….it could be that moving the ipsilateral hand does not result in any kind of organized activity in the "missing hand" hemisphere in this population.

Note that I think that this analysis is clever and creative, and I am sympathetic to it. However, as it stands, I don't see the clear mechanistic account for a) why activity for moving the ipsilateral hand is an index of the contralateral hand representation (apart from the correlational argument, which I fear is flawed) and b) why your argument would necessarily hold in congenital one-handers. I believe this could be improved with some mechanistic arguments to support the claim. There is some work on what ipsilateral activation indexes (see Diedrichsen, Wiestler and Krakauer, 2013; prior work on mirror movements)….maybe there is something in this literature that could strengthen the claims?"

We apologise for misunderstanding the concerns of reviewer 2 in our previous round of revisions. We agree that the reviewer raises an important issue that requires further consideration.

Recent evidence showing correspondence between ipsilateral and contralateral representations (across the two hands) sheds some light on the potential mechanism giving rise to ipsilateral hand representation in SI/M1. As we emphasised before, past studies have shown that the activity patterns underlying the ipsi- and contralateral representational structure in one hemisphere are spatially overlapping (Diedrichsen et al., 2013, Diedrichsen et al., 2018). This is generally interpreted as both hands’ representation being facilitated by the same architecture rather than two distinct representations with a correlating structure. In other words, the ipsilateral hand reactivates the patches of cortex associated with the contralateral hand. Strong evidence for this is indeed provided by Diedrichsen et al., (2013, Exp. 2): During bimanual asymmetric movement, SI activity patterns are dominated by contralateral actions only and ipsilateral representation disappears. Most recently, we also showed that ipsilateral representation is only present during active movement (as opposed to passive touch) and not related to uncrossed sensory inflow (Berlot et al., 2018). Taking this evidence together, we have good reason to suggest that in two-handers the motor command to move one hand activates (through the back door) the corresponding representations of the other hand and not a distinct (correlated) representation of the ipsilateral hand. We realise that some of the language used in our manuscript, e.g. emphasising that one representation predicts another, has not reflected this interpretation well and we have made some edits to elaborate on this suggested mechanism.

How does this shared architecture across hands emerge? As pointed out by the reviewers, the two key mechanisms are: Bilateral hand experience, emerging over life, and innate structural connections (e.g. relating to mirroring).

What does this mean for the congenital one-handers? As we see it, there are three possibilities: (1) The deprived cortex develops separate representations for both contralateral (missing) hand and ipsilateral (intact) hand. Here we assume that the overlap and functional interaction seen in controls require coordinated bimanual behaviour, so in one-handers both ipsi- and contralateral hand remain represented distinctly and independently within the deprived cortex. (2) The deprived cortex develops contralateral hand representation (of the missing hand), but intact hand movements do not recruit it, e.g. due to impaired connectivity across hemispheres. (3) The deprived cortex does not develop structured hand representation (neither contralateral nor ipsilateral).

If option 1 were true (though unlikely in light of the research in controls discussed above), then one would expect to find strong ipsilateral representation of the intact hand. Yet, our data does not reflect this hypothesis. Despite typical contralateral dissimilarity for the intact hand (in the intact cortex; not different from controls or acquired amputees’ intact hand; data included in revised manuscript), average ipsilateral dissimilarity (in the deprived cortex) is not higher than our control area, which does not represent hands (Figure 3A).

Option 2 cannot be disproven by our current data, but our previous findings suggest that callosal connections are functionally preserved in congenital one-handers, at least to some extent. We showed that the strength of functional connectivity across the two sensorimotor hand areas correlates with the extent to which congenital one-handers engage their stumps (residual arms) in performing “bi-manual” tasks (Hahamy et al., 2015). Moreover, congenital one-handers who strongly rely on stump usage show normal inter-hemispheric connectivity in comparison to two-handed controls. Since the stump has been shown to strongly activate the missing hand cortex in congenital one-handers (Hahamy et al., 2015; 2017, Makin et al., 2013), it appears that given the right input, the two hemispheres can engage. Please also note that in Makin et al. (2013) we explored structural connectivity changes (using DTI) and did not identify any striking callosal differences between congenital one-handers and controls.

This leaves us with option 3 (no hand representation in the deprived cortex). Taken together, it appears that sensorimotor experience is necessary for developing a hand area, including its connectivity to other areas in the sensorimotor network. We have clarified this both in the Results section and Discussion section, as follows:

Results section:

“To probe digit structure in the missing hand cortex using an alternative task, we examined whether we could observe a representation of the ipsilateral (intact) hand in the missing hand cortex. In two-handed controls, finger movements lead to activity in specific cortical patches in ipsilateral M1 and SI, which tightly correspond to the activity patches engaged in the movement of the mirror-symmetric contralateral finger (Diedrichsen et al., 2013). Indeed, this ipsilateral representation fully overlaps with the representation of the contralateral hand (Diedrichsen et al., 2018). Furthermore, ipsilateral representation disappears completely during asymmetric bimanual finger movements, during which activity in M1 and SI is fully determined by the contralateral hand (Diedrichsen et al., 2013). As such, the ipsilateral representation of one hand is likely elicited due to recruitment of the representation of the contralateral hand (Diedrichsen et al., 2018, Berlot et al., 2018). Ipsilateral representation of the intact hand can therefore provide an indirect assay into the representation of the missing hand, while controlling for task demands across groups.”

Results section:

“If missing hand representation is diminished in congenital one-handers, then ipsilateral representation of their intact hand (in the missing hand area) should show reduced representational features compared to those found in amputees [see Discussion for an alternative mechanism, where the deprived cortex develops separate representations for both the contralateral (missing) and ipsilateral (intact) hands].”

Discussion section:

“We also explored whether we could activate the representation of the missing hand indirectly by movements of the fingers of the intact hand. Previous studies in two-handers have demonstrated that contralateral and ipsilateral hand movements produce identical representational patterns (Diedrichsen et al., 2018). Since this ipsilateral representation is completely overwritten by the contralateral hand if the two hands are engaged in dissociated movements (Diedrichsen et al., 2013, Exp. 2), it has been proposed that the ipsilateral hand reactivates the cortical resources associated with the contralateral hand. Based on this evidence, the ipsilateral digit representation can serve as an indirect measure of the contralateral hand representation.”

Discussion section:

“It might be worth considering whether congenital one-hander’s deprived cortex could have developed separate representations for both the contralateral (missing) and ipsilateral (intact) hand, which are uncorrelated due to anatomical or behavioural differences from two-handers. If this were possible, we would predict maintained, or even stronger, ipsilateral representation of the intact hand. Yet, our data does not reflect this hypothesis. Moreover, poor ipsilateral representation in the deprived cortex does not seem to stem from reduced inter-hemispheric connectivity, which appears to be functionally and structurally preserved in congenital one-handers, depending on lateralisation strategies in daily behaviour (Hahamy et al., 2015, Hahamy et al., 2017, Makin et al., 2013a).”

https://doi.org/10.7554/eLife.37227.013

Article and author information

Author details

  1. Daan B Wesselink

    1. Institute of Cognitive Neuroscience, University College London, London, United Kingdom
    2. Wellcome Centre for Integrative Neuroimaging, University of Oxford, Oxford, United Kingdom
    Contribution
    Data curation, Software, Formal analysis, Validation, Investigation, Visualization, Methodology, Writing—original draft, Project administration, Writing—review and editing
    Contributed equally with
    Fiona MZ van den Heiligenberg
    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3229-0925
  2. Fiona MZ van den Heiligenberg

    1. Institute of Cognitive Neuroscience, University College London, London, United Kingdom
    2. Wellcome Centre for Integrative Neuroimaging, University of Oxford, Oxford, United Kingdom
    Contribution
    Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Project administration, Writing—review and editing
    Contributed equally with
    Daan B Wesselink
    Competing interests
    No competing interests declared
  3. Naveed Ejaz

    1. Brain and Mind Institute, University of Western Ontario, London, Canada
    2. Department of Computer Science, University of Western Ontario, London, Canada
    Contribution
    Conceptualization, Resources, Software, Formal analysis, Validation, Methodology, Writing—review and editing
    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8370-4588
  4. Harriet Dempsey-Jones

    1. Institute of Cognitive Neuroscience, University College London, London, United Kingdom
    2. Wellcome Centre for Integrative Neuroimaging, University of Oxford, Oxford, United Kingdom
    Contribution
    Investigation, Project administration, Writing—review and editing
    Competing interests
    No competing interests declared
  5. Lucilla Cardinali

    1. Brain and Mind Institute, University of Western Ontario, London, Canada
    2. Unit for Visually Impaired People, Istituto Italiano di Tecnologia, Genoa, Italy
    Contribution
    Investigation, Writing—review and editing
    Competing interests
    No competing interests declared
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0441-1806
  6. Aurelie Tarall-Jozwiak

    Queen Mary’s Hospital, London, United Kingdom
    Contribution
    Resources, Writing—review and editing
    Competing interests
    No competing interests declared
  7. Jörn Diedrichsen

    1. Brain and Mind Institute, University of Western Ontario, London, Canada
    2. Department of Computer Science, University of Western Ontario, London, Canada
    Contribution
    Conceptualization, Software, Supervision
    Competing interests
    Member of eLife BRE
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-0264-8532
  8. Tamar R Makin

    1. Institute of Cognitive Neuroscience, University College London, London, United Kingdom
    2. Wellcome Centre for Integrative Neuroimaging, University of Oxford, Oxford, United Kingdom
    3. Wellcome Centre for Human Neuroimaging, University College London, London, United Kingdom
    Contribution
    Conceptualization, Resources, Data curation, Software, Formal analysis, Supervision, Funding acquisition, Validation, Investigation, Visualization, Methodology, Writing—original draft, Writing—review and editing
    For correspondence
    t.makin@ucl.ac.uk
    Competing interests
    Member of eLife BRE
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-5816-8979

Funding

Wellcome (104128/Z/14/Z)

  • Tamar R Makin

Royal Society (104128/Z/14/Z)

  • Tamar R Makin

CREATE-IRTG

  • Lucilla Cardinali

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Acknowledgements

This work was supported by a Sir Henry Dale Fellowship jointly funded by the Wellcome Trust and the Royal Society (104128/Z/14/Z), awarded to TRM. LC was supported by a CREATE-IRTG grant. We thank our participants for taking part in the study and Opcare for invaluable help with participants’ recruitment. We thank Laurie Josephs, Paulina Kieliba, Gonzague de France and Liezel Wegner for their help with data collection. We thank Devin Terhune for providing us with a consistency analysis for the step-wise regression.

Ethics

Human subjects: Ethical approval was granted by Oxford University's Medical Sciences inter-divisional research ethics committee (MS-IDREC-C2-2015-012) and written informed consent was obtained from all participants prior to the study.

Senior and Reviewing Editor

  1. Eve Marder, Brandeis University, United States

Publication history

  1. Received: April 10, 2018
  2. Accepted: January 12, 2019
  3. Version of Record published: February 5, 2019 (version 1)

Copyright

© 2019, Wesselink et al.

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.

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