Implications of structural connections within and between brain regions for their functional counterpart are timely points of discussion. White matter microstructural organization and functional activity can be assessed in unison. At first glance, however, the corresponding findings appear variable, both in the healthy brain and in numerous neuro-pathologies. To identify consistent associations between structural and functional connectivity and possible impacts for the clinic, we reviewed the literature of combined recordings of electro-encephalography (EEG) and diffusion-based magnetic resonance imaging (MRI). It appears that the strength of event-related EEG activity increases with increased integrity of structural connectivity, while latency drops. This agrees with a simple mechanistic perspective: the nature of microstructural white matter influences the transfer of activity. The EEG, however, is often assessed for its spectral content. Spectral power shows associations with structural connectivity that can be negative or positive often dependent on the frequencies under study. Functional connectivity shows even more variations, which are difficult to rank. This might be caused by the diversity of paradigms being investigated, from sleep and resting state to cognitive and motor tasks, from healthy participants to patients. More challenging, though, is the potential dependency of findings on the kind of analysis applied. While this does not diminish the principal capacity of EEG and diffusion-based MRI co-registration, it highlights the urgency to standardize especially EEG analysis. With the latter, typically based on functional MRI (fMRI) and blood-oxygen-level dependent (BOLD) contrasts, one assesses (correlated) changes in the metabolic demand, providing indirect insight into this neural functioning.The human brain is characterized by structural and functional connectivity within and between regions. Structural connectivity refers to the anatomical organization of the brain by means of fiber tracts. Recent advances in magnetic resonance imaging (MRI) and image processing provide various means to quantify structural connectivity in a non-invasive way using short-range local measures and/or long-range tract tracing procedures, called diffusion tractography. Functional connectivity refers to statistical dependence between time series of electro-physiological activity and (de)oxygenated blood levels in distinct regions of the brain. The first can be assessed non-invasively using encephalographic recordings. Electro- and magnetoencephalography  reflect changes in averaged post-synaptic potential and dendritic currents of neural populations and, hence, provide Structural connectivity is known to shape functional connectivity  but the Electro-encephalography signals are recorded from the scalp using surface electrodes at positions typically based on the 10–20 system ; see FigDiffusion MRI encodes diffusion-driven displacements of water molecules over time into MRI signals. It allows to assess tissue characteristics in vivo , three forms of diffusivity, and kurtosis anisotropy (KA). FA quantifies the amount of diffusion along the principal direction on a normalized scale, i.e., a value of 0 indicates undirected or isotropic diffusion, whereas a value of 1 indicates full directedness or anisotropy. FA is thought to reflect various tissue properties such as axonal myelination, axonal diameter, axonal density, and fiber organization . By contmechanistic perspective one may expect that the nature of WM microstructural organization influences the transfer of activity between different neuronal populations. That is, if EEG measures serve to quantify such transfer of activity directly, one may expect them to be strongly associated with the afore-listed DWI metrics. By hypothesis, a higher degree of microstructural organization is, hence, accompanied by stronger and faster neural transmission. As will be shown, this is indeed the case for some of the EEG outcomes but not for all of them.From a We conducted a keyword-based search in PubMed and ISI Web-of-Science. Search terms [“DTI” AND “EEG”], [“DWI” AND “EEG”], and [“DKI” AND “EEG”], which provided 270, 220, and 3 hits, respectively. Animal and computational studies, technical/methods studies as well as clinical studies that employed the modalities for their isolated diagnostic value were excluded. We also excluded review papers and non-English publications. This selection yielded 40 combined dMRI/EEG studies including 17 experimental ones in healthy participants and 23 investigating pathology. The latter included neurodegenerative disease like epilepsy and Alzheimer, trauma-induced cases like stroke and traumatic brain injury (TBI), and psychiatric conditions ; an overview of some of them can be found as i) selecting a set of ROIs, either a priori or based on the statistical significance of FA group differences or signal power as a function of location; (ii) defining appropriate  methods to determine the structural and functional characteristics of interest; and (iii) selecting the proper statistical means to identify possible associations between them. Several studies (N = 13) refrained from conducting step (iii) – for example because they referred to case studies – and only reported the individual EEG- and dMRI-based results. Yet, these studies revealed novel insight into the structure/function relationship and were included in this review.Estimating the relationship between structural and functional brain characteristics requires three important steps:  relation between functional EEG and structural dMRI measures. Given the long tradition of using EEG both in fundamental research and in the clinic, its experimental designs and analyses form the backbone of our review, from assessments of event-related potentials, different spectral analyses and functional connectivity approaches to more recent studies involving source-localization techniques. In what follows we will particularly highlight the reported association between these functional activities and co-registered dMRI measures before discussing the potential benefits and shortcomings of such combined recordings.The most used approach in experimental EEG studies comprises of evoking activity patterns using repeated stimuli that may differ between studies. This design yields so-called event-related potentials (ERPs), highlights activity that is time-locked to an event  and eliminates non-time-locked background activity via averaging to event-aligned epochs. According to the aforementioned mechanistic view (see Section “A Mechanistic View”), the possibility of time-locked activity to emerge depends on the myelination of connecting axons and dendrites . An incrtask-related studies, where participants were asked to perform a cognitive, sensory, or motor task during EEG recording. We grouped them under the headings error-related, early attention-related and later cognitive-related responses. Error-related responses are ERPs evoked by commission errors when participants are asked to respond as accurately and quickly as possible to a cognitive task  activity entails both resting state and task-related assessments, a classification we adopt to group the here-included ones. The so-called resting state condition refers to EEG recordings during which participants are asked to relax and ‘do nothing’ for some time period . Recently, resting state studies have become quite popular but more traditionally the EEG’s spectral content has been associated with task-related functional state like attention or motor performance. Given the diversity of EEG studies supporting the latter, spectral power in distinct frequency bands and its association with structural measures has in fact been the primary target in the search for functional expression of WM microstructural changes . They can be observed, e.g., during the deepest level of relaxation or sleep but also mark anxiety, poor emotional awareness, stress, depression, inattentiveness and dementia .Given the close relation between sleep, cognitive performance and memory consolidation, Alpha oscillations (∼8–14 Hz) are spectral components that were already observed by Hans Berger about a century ago . For manMuch in line with this, As already discussed above, Despite the strong link of beta oscillations to motor performance, all studies collected here linked dMRI outcomes with beta power during rest.As mentioned earlier, Coming back to the study by The combination of dMRI and EEG power at different frequencies can reveal how neural structure and function are coupled. Our review of the literature suggests that when associating with dMRI, the EEG spectral power in the lower frequency bands  can be considered a functional expression of structural abnormalities, especially when applied to the resting state condition or during sleep. Analyzing dMRI together with beta power showed the sensitivity of beta power  to changes in structural characteristics of cognitive and motor networks, as observed in patients with Alzheimer’s or stroke; cf. The brain’s structural connectivity contributes to shaping neurophysiological activity, and thereby influences functional connectivity among neuronal populations  and braiChanges in phase synchronization due to mild traumatic brain injury (mTBI) and its association with structural abnormality of WM axons were estimated from eyes-closed resting state EEG . The we allowed The interaction between structure and function during performance of a cognitive control task in mTBI patients was investigated by With the aim to identify long-range structural networks that support error processing function, The combination of dMRI and coherence/phase synchronization allows for studying the link between functional and structural connectivity and to identify structural networks underlying task-related or resting state functional networks at different frequency bands . All theSections “Event-Related Potentials,” “Spectral Analysis – Local Functional Connectivity,” and “Synchronization Analysis – Distant Functional Connectivity”.An important aim of functional neuroimaging is to localize active brain regions. By now, there is a plenitude of methods available to estimate source activity potentially generating experimentally observed EEG. Inverse calculations yield virtual sensors or sources in the brain that may undergo analysis identical to what has been outlined in In patients with epilepsy during eyes-closed resting state, Similar to the More recently, Likewise recently, To investigate hand motor outcome in chronic stroke patients, The EEG source localization yields  brain activity at voxel level. When combined with dMRI one may estimate structural connectivity between the functionally active brain regions. This may reveal the underlying structural connectivity of the functional networks. For example, We reviewed a variety of studies that combined EEG with dMRI. Before discussing them, we would like to note that despite the considerable insights into the relationship between GM/WM microstructural organization and neural functioning, the combination of EEG and dMRI comes with many challenges, often involving defining details of the measurement procedure and the analysis. While there are several technical hurdles to take, one must realize that the major bottleneck in this field is that most studies are low-powered, sometimes covering only a handful of patients with a very specific phenotype. We highlighted this within the current review but realized that the generalizability of many findings remains limited and calls for more research. Yet, in the following we briefly recapitulate some of the insights gained from the here-reviewed studies and put them in a broader context of relating functional and structural properties of the brain.With respect to studies using event-related designs, most of the findings can be explained by accounting for the conceptual differences in EEG outcome measures – latency vis-à-vis amplitude changes – and their relation to neural processing. In line with a mechanistic view (see Section “A Mechanistic View”), changes in the WM microstructural organization affect the rate of information transfer  between different brain regions. ERP latencies do address this information transfer directly. Hence, one may expect a close relationship between conduction delays and the structural properties of WM measured via dMRI . ApparenChanges in WM microstructural organization also affect amplitude of ERP components. Higher ERP amplitudes appear to be related to WM microstructure, as, e.g., shown by the positive correlation between N2 amplitudes and FA in the superior longitudinal fascicle linking the parietal and frontal cortices , or by aIn contrast to the studies on event-related potentials, the findings on spectral EEG content are more diverse, as we found both positive and negative correlations, albeit not always significant. Slow wave power changes were often investigated in the context of neuropathology to identify associations between abnormal functional activity  and structural abnormalities underlying a specific disorder. All studies claimed a relationship between neurodegeneration and spectral power, indicating that in combination with dMRI the spectral power may serve to identify structural disorder. For instance, the observed reduction of power across cortical areas which coincided with elevated MD in the thalamus and the cingulate cortex in a fatal familial insomnia patient  supportsThe more recent studies on functional connectivity largely confirm the findings on local power changes. They demonstrate a close relationship between structural and functional connectivity both in resting state and during task execution. In mild cognitive impairment patients, frontal alpha coherence and FA values were positively correlated in frontal fiber tracts , while iWhile in many studies the EEG source localization merely served to select seed regions for DTI tractography , it has Compared to fMRI, EEG has several advantages which may help for better understanding the relationship between structural and functional properties. First, EEG records electrophysiological activity at a high temporal resolution that can be crucial for monitoring rapid, functional changes. Second, EEG does not necessarily have to be recorded in a lab-setting. In fact, some current EEG systems are portable, which allows for recording functional brain activity during daily tasks . Yet, itb-values) with large numbers of diffusion directions can elicit differential tissue responses that are useful to model in greater detail WM microstructural organization. Interestingly, the impact of in-scanner motion on multi-shell measures of microstructure appears to be reduced, at least on a global level  can improve source localization by increasing the signal-to-noise ratio to detect functional networks , potential level .In addition, more advanced methods of analysis can greatly improve the accuracy of outcome measures. For example, even though the DTI model has clinical relevance, it is not suitable for complex fiber configurations, such as crossing and ‘kissing’ fibers . Multi-cIn very recent years, an increasing number of studies extended the combination of encephalographic assessments and diffusion-based imaging with electrically or magnetically stimulating. This includes transcranial magnetic stimulation, direct current stimulation as well as deep brain stimulation. This field is rapidly growing and certainly worthwhile monitoring. For the sake of legibility, we provide a glance on this literature as a A review entitled “Brain structural and functional connectivity” cannot ignore the developments on network assessments using graph theoretical approaches. The field is vast, and we consider a proper overview to be beyond the scope of our review. We summarized two recent studies employing graph theory , which h1, Brain Imaging Data Structure (BIDS)2 or John Richard’s neurodevelopmental MRI database3. This list could readily be extended, but again we consider digging deeper into these exciting developments beyond the scope of our current review. When it comes to analysis tools, the list is even longer, ranging from EEGLAB . One step toward standardization is to provide open access to both data and analysis. Given the very nature of the topic under research, privacy-related issues may limit availability of data. Yet, there are quite recent initiatives to provide openly accessible data sets, like the Healthy Brain Network (HBN) , OpenNeum EEGLAB , Fieldtrm EEGLAB , Statistm EEGLAB , Brainstm EEGLAB , MNE-pytm EEGLAB , and NUTm EEGLAB  to Explom EEGLAB , FMRIB Sm EEGLAB , and MRtm EEGLAB . We cleam EEGLAB .We reviewed various studies that combined EEG and dMRI assessments to relate functional activity with structural connectivity. The variety of techniques, methods and outcomes renders comparison a challenge, especially when it comes to the additive value of combining EEG with dMRI. Yet, the different ERP analyses revealed clear associations with dMRI measures. Apparently, the speed and strength of broadcasting between distant brain regions quantified by, e.g., ERP latencies and amplitudes can largely express the WM microstructural organization. This can be expected when adopting a simple mechanistic standpoint, i.e., that an increased WM microstructural organization and/or structural connectivity strength ‘simply’ facilitates neuronal information transfer. The EEG’s spectral power and synchronization appeared less sensitive unless used to assess the effects of neurodegenerative diseases. Nonetheless, when combined with dMRI, properties of the WM microstructural organization may eventually yield proper biomarkers for functional decline accompanying structural changes. Yet, the diversity of findings requires a very careful interpretation of the results and highlights the need to standardize EEG analysis. This is certainly true when searching for a more general relationship between functional and structural characteristics of the human brain, let alone when aiming for employing EEG-dMRI co-registration in the clinic.PB and AD designed the study and reviewed all the included manuscript. LR-D reviewed in particular the EEG studies. JG and SS focussed on the dMRI studies. All authors wrote the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Biological rhythms pervade physiology and pathophysiology across multiple timescales. Because of the limited sensing and algorithm capabilities of neuromodulation device technology to-date, insight into the influence of these rhythms on the efficacy of bioelectronic medicine has been infeasible. As the development of new devices begins to mitigate previous technology limitations, we propose that future devices should integrate chronobiological considerations in their control structures to maximize the benefits of neuromodulation therapy. We motivate this proposition with preliminary longitudinal data recorded from patients with Parkinson's disease and epilepsy during deep brain stimulation therapy, where periodic symptom biomarkers are synchronized to sub-daily, daily, and longer timescale rhythms. We suggest a physiological control structure for future bioelectronic devices that incorporates time-based adaptation of stimulation control, locked to patient-specific biological rhythms, as an adjunct to classical control methods and illustrate the concept with initial results from three of our recent case studies using chronotherapy-enabled prototypes.  Biological sciences; Neuroscience; Biotechnology; Bioelectronics Throughout physiology, homeostasis is observed at multiple spatial and temporal scales. Physiological processes such as body temperature, blood pressure, and blood hormone concentrations are carefully regulated during healthy conditions, whereas dysregulation can lead to the initiation, progression, and expression of diseases . From a In contrast to the definition of adaptive control above, the term ‘adaptive’ is used throughout bioelectronic medicine to describe neuromodulation devices capable of instantaneous parameter adjustments in response to monitored physiomarker signals. These ‘adaptive’ devices in truth rely solely on static implementations of either feedforward, such as activity-based cardiac pacemakers  and motiAlthough living organisms maintain constant homeostasis, their underlying physiological processes are not time invariant. Physiological processes display rhythmic variations which recur periodically at ultradian (less than 24 h), circadian (24 h), and infradian  and infradian (1 month) time scales. Circadian rhythms are generated endogenously with a spontaneous period close to but not exactly 24 h. They are synchronized to zeitgebers, or cues from the external environment, the most notable of which is the light-dark cycle . The ligDue to limited sensing capabilities of previous generations of bioelectronic devices, the influence of biological rhythms on neuromodulation efficacy has been largely underexplored. Traditionally, therapy optimization for bioelectronic devices has been limited to daytime scheduled clinical visits, a single phase of the day-night cycle. As a result, tonic stimulation parameters or control goals for ‘adaptive’ bioelectronic devices may provide effective therapeutic benefits for the specific phase or the day-night cycle that they are optimized for, but may perform better or worse, and in worst cases induce side-effects during other phases of the circadian cycle . For thiParkinson's disease (PD) is one of the fastest growing neurological conditions with enormous human and economic costs. It is estimated that PD will affect 1.5 million people in the United States and would cost more than $79 billion by 2037 . MedicatOver time, the management of PD becomes significantly oriented around the optimization of therapy toward symptom fluctuations that occur according to biological and exogenously driven rhythms. Although PD symptoms are described to be improved on waking — the “sleep benefit”  — overalOverall, it has been found that open-loop DBS improves sleep structure as a fortuitous byproduct . HoweverEpilepsy is a neurological disease in which brain activity becomes abnormal, resulting in seizures or periods of unusual behavior, sensations, and sometimes loss of awareness. Epilepsy affects close to 50 million people worldwide and has been estimated to cost the United States economy $9.6 billion annually . AlthougHowever, the optimization of stimulation is difficult and long-term seizure freedom is rare . The relDisruption of the sleep-wake cycle is also common in epilepsy; however, stimulation at present is not optimized for nighttime or within sleep stage operation . ANT stiThe above case studies illustrate two examples of neurological disorders which demonstrate strong rhythmic variations across several timescales. Many other biological processes and neurological disorders are reported to display similar variations. For example, brain temperature, cortical excitability, and brain responses to a performance task are all modulated by circadian and sleep-wake cycles ; the cirWe have undertaken preliminary work to integrate patient-specific rhythms in DBS therapy. In the following sections, we briefly highlight this work in two investigational DBS devices for the treatment of PD and epilepsy.In Sleep comorbidities are commonly experienced by patients with epilepsy. To improve sleep quality and seizure mitigation for patients with epilepsy, we presented the Mayo Epilepsy Personal Assistant Device (EPAD) in . The EPAIn The above examples highlight potential technological infrastructures required for integrating chronotherapy in bioelectronic devices. In practice, future devices will require hybrid frameworks to leverage the benefits of both embedded and distributed device functionalities. In this manner flexible device platforms will provide clinicians with improved insight into disease pathophysiology across a breadth of neurological disorders, while facilitating long-term optimization of neuromodulation chronotherapy on a disease-specific and patient-specific basis. Furthermore, flexible platforms such as this would facilitate exploration of alternative algorithms for regulating stimulation parameters in accordance with the relevant biological rhythms. On-off and proportional-integral-derivative strategies from classical control theory may be readily suitable for embedded implementations on current device hardware. In contrast, implementations of modern control or machine learning algorithms which are computationally expensive, such as Bayesian optimization and reinforcement learning , and mayHowever, limitations may also be encountered in the development of these future chronotherapeutic-bioelectronic devices. The ability to trial new computationally expensive algorithms comes with an increased algorithm training complexity. Moreover, the ability to adapt the control strategy or stimulation parameters implemented in response to specific phases of biological rhythms may also result in increased clinical burden and programming time requirements. However, as highlighted in this article, the state-of-the-art for many neuromodulation therapies at present is still oriented around the delivery of open-loop stimulation optimized for maximizing daytime therapeutic benefits. Therefore, we propose that first generation chronoadaptive-bioelectronic devices may readily improve patient therapeutic outcomes by simply implementing time-based adjustments to open-loop stimulation parameters aligned to patient-specific chronotypes and medication schedules. Following this, if further therapy refinement is necessary, additional feedforward-based or feedback-based strategies whose control goals are appropriately synchronized to specific phases of these patient rhythms can be gradually layered in to provide additional stimulation refinement in a time-localized manner, only when required. This framework should help to constrain the multiple degrees of freedom encountered when developing new strategies for automatic therapy optimization in a tractable manner. At a minimum, deployment of chronoadaptation in this manner should immediately enhance the therapeutic benefits of open-loop stimulation at nighttime during sleep, while maintaining the performance of daytime optimized therapy. Trialing of future feedforward-based and feedback-based control strategies may then focus on optimization of therapy goals during specific phases of the day-night cycle.With the continued development of bioelectronic device technology, previous technical limitations that have limited access to longitudinal data recordings from patients with neurological disorders in the past are gradually being eased. As a result, clinical neuroscience researchers are beginning to gain insight into rhythmic variations in patient physiology over a variety of timescales and explore the influence of these biological rhythms on the therapeutic outcomes of neuromodulation therapies. With this in mind, it is becoming increasingly clear that there is a need to incorporate considerations for these biological rhythms in future bioelectronic devices. To optimize patient-specific therapeutic outcomes, we emphasize that future devices should incorporate feedforward, feedback, and adaptive control strategies together to maximize therapeutic benefits for patients and provide more natural regulation of patient pathophysiology to restore healthy physiological homeostasis.


Mutations in the gene encoding TNAP leading to severe hypophosphatasia result in strongly reduced mineralization and perinatal death. Fortunately, the relatively recent development of a recombinant TNAP with a bone anchor has allowed to correct the bone defects and prolong the life of affected babies and children. Researches on TNAP must however not be slowed down, because accumulating evidence indicates that TNAP activation in individuals with metabolic syndrome (MetS) is associated with enhanced cardiovascular mortality, presumably in relation with cardiovascular calcification. On the other hand, TNAP appears to be necessary to prevent the development of steatohepatitis in mice, suggesting that TNAP plays protective roles. The aim of the present review is to highlight the known or suspected functions of TNAP in energy metabolism that may be associated with the development of MetS. The location of TNAP in liver and its function in bile excretion, lipopolysaccharide (LPS) detoxification and fatty acid transport will be presented. The expression and function of TNAP in adipocyte differentiation and thermogenesis will also be discussed. Given that TNAP is a tissue- and substrate-nonspecific phosphatase, we believe that it exerts several crucial pathophysiological functions that are just beginning to be discovered. Tissue-nonspecific alkaline phosphatase (TNAP) is mainly known for its necessary role in skeletal and dental mineralization, which relies on the hydrolysis of the mineralization inhibitor inorganic pyrophosphate (PP With ffication . The hypfication  and is nfication ,13, we wAlpl+/− mice develop liver steatosis when fed a high fat diet (HFD) [The group of Olga Martinez-Augustin, in this journal issue, reported that et (HFD) . This suet (HFD) ,16. Thiset (HFD) . This reThe liver localization of TNAP suggests two different functions . First, An important function of hepatocytes is to produce the bile, which consists of bile acids, the end-products of cholesterol metabolism, cholesterol itself, phosphatidylcholine and toxic metabolites . It is ha of approximately 4 and 1–2 respectively, and therefore, at these high concentrations, a significant amount of bile acids might be protonated and toxic [− secretion through cystic fibrosis transmembrane regulator (CFTR). The rise of extracellular Cl− then stimulates the Cl−/HCO3− exchanger AE2 (anion exchanger 2) to allow carbonate secretion and increase the pH [Alpl+/− mice fed a high fat diet is unknown but deserves consideration [The most likely function of TNAP in bile excretion is to participate in the regulation of bile pH and of bile excretion during the postprandial period . nd toxic , hepatoce the pH . The hore the pH . They moe the pH . Interese the pH . In summderation .The function of liver TNAP may not be restricted to bile excretion but might extend to intestinal digestion and absorption of lipids and/or bacterial compounds. A fraction of liver TNAP is indeed excreted in the intestinal lumen within the bile. TNAP is released with fragments of hepatocytes and/or cholangiocytes upon the action of bile acids . In addiTlr4-deficient mice [4 exposure, oral IAP administration attenuated the compromised intestinal barrier permeability and liver fibrosis, in association with reduced LPS absorption [IAP at least exerts two suspected important functions. First, IAP dephosphorylates bacterial lipopolysaccharide (LPS) in the intestinal track ,35, to lent mice . In addisorption . Furthersorption . A seconsorption ,45. CD36sorption ). CD36 hsorption ).Thr92 is a consensus site for protein kinase C (PKC) and Ser237 for PKA . In platAkp3, one of the two genes encoding IAP, have higher circulating TNAP levels than wildtype mice during aging [i hydrolysis as compared to LPS, while the reverse is true for IAP [Whether TNAP that is eliminated in the duodenum within the bile exerts the same functions as IAP is unknown and remains purely speculative. On one hand, since mice deficient in ng aging , and sin for IAP . This su for IAP , where A for IAP . In the While the location of liver TNAP at the hepatocyte and cholangiocyte membranes facing bile canaliculi and ducts suggests a specific role in bile excretion and function, the presence of TNAP in endothelial cells of hepatic arteries in the portal triad suggests additional functions, likely associated with the dephosphorylation of circulating molecules . Among tLike in the intestinal lumen, another possible TNAP substrate in the liver is CD36. In HepG2 hepatocytes cultured in presence of high glucose or FA levels, TNAP expression increased in parallel with that of CD36 ,72, and Alpl+/− mice develop an exacerbated steatosis as compared to Alpl+/+ mice when the steatosis is induced by a HFD, but not when the steatosis is induced by a choline deficient-diet [Finally, the fact that ent-diet  suggestsent-diet ,78). AboN-methyltransferase (PEMT) [Pemt−/− mice fed a HFD develop non-alcoholic fatty liver disease (NAFLD) resulting from lack of sufficient phosphatidylcholine to export TG in VLDL [Phosphatidylcholine from the diet or from circulating lipoproteins is converted to phosphocholine by several enzymes that are still incompletely identified ,79. Phose (PEMT) . Pemt−/− in VLDL ,86, and  in VLDL . How ext in VLDL  strongly in VLDL . It shou in VLDL ). If nevIt seems now accepted that adipose tissue expandability relies on both the hyperplasia of existing adipocytes and the formation of new adipocytes from mesenchymal progenitors (reviewed in ). These Alpl−/− or Alpl+/− mice or from patients with hypophosphatasia generate more adipocytes than bone marrow stromal cells from corresponding controls [Alpl+/− MSCs appear to have higher levels of extracellular ATP than Alpl+/+ MSCs, suggesting that TNAP might inhibit adipocyte differentiation through ATP dephosphorylation and decreased activation of purinergic receptors [While historically, TNAP has been associated with the differentiation of mesenchymal progenitors into mineralizing osteoblasts and chondrocytes, relatively recent data suggest that it is already functional in mesenchymal stem cells (MSCs), and also involved in MSC commitment into other lineages, and in particular adipocytes (reviewed in ). Publiscontrols ,93,94, scontrols . Similarcontrols ,96,97. Teceptors , TNAP waeceptors . When ceeceptors . Despiteeceptors  merits ti instead of an ATP [Alpl deletion in adipocytes in vivo induced obesity when mice were fed a HFD [Recently, TNAP in adipocytes was shown to participate in thermogenesis . The mosf an ATP  (Figure f an ATP , and thef an ATP . Spiegeled a HFD . This suSeveral important points remain to be explored after these pioneer studies on TNAP in adipocyte mitochondria. The first one is how TNAP localization to the cell membrane or to the mitochondria is controlled. To our knowledge, TNAP lacks classical mitochondrial addressing signals. Interestingly, an article published in this issue reports that TNAP is also likely associated with mitochondria in bone and muscle progenitor cells . In thesCirculating TNAP levels are associated with both the development of MetS and with the cardiovascular risk ,6,7,8,9.


The discovery of induced pluripotent stem cells (iPSCs) has made an invaluable contribution to the field of regenerative medicine, paving way for identifying the true potential of human embryonic stem cells (ESCs). Since the controversy around ethicality of ESCs continue to be debated, iPSCs have been used to circumvent the process around destruction of the human embryo. The use of iPSCs have transformed biological research, wherein increasing number of studies are documenting nuclear reprogramming strategies to make them beneficial models for drug screening as well as disease modelling. The flexibility around the use of iPSCs include compatibility to non-invasive harvesting, and ability to source from patients with rare diseases. iPSCs have been widely used in cardiac disease modelling, studying inherited arrhythmias, neural disorders including Alzheimer’s disease, liver disease, and spinal cord injury. Extensive research around identifying factors that are involved in maintaining the identity of ESCs during induction of pluripotency in somatic cells is undertaken. The focus of the current review is to detail all the clinical translation research around iPSCs and the strength of its ever-growing potential in the clinical space. The science around terminal inactivation and deletion of genetic codes of heredity in somatic cells was postulated by the Weismann barrier theory . The somc-Myc has been replaced with miR-291-3p, miR-294, and miR-295 to generate homogenous colonies of human iPSCs  out and angiotensin-II. This indicates mutated CM from DCM patients to express blunted inotropic response [2+ levels, that becomes causative for HCM-specific electrophysiological abnormalities [In case of DCM, characterized by ventricular chamber enlargement, and dilation as well as systolic dysfunction, human derived iPSCs have been used to investigate the excitation-contraction-coupling machinery, response to positive inotropic interventions, and study the proteome profile. This study utilized DCM patient specific-iPSC derived from skin fibroblasts and identified defects in assembly and maintenance of sarcomeric structure in the mutated iPSC-CM, as well as lower response to β-adrenergic stimulation with isoproterenol, and increased [Caresponse . In casemalities . Recent malities .The iPSC models around FPAH have identified modification of BMPR2 signaling causing reduced endothelial cell adhesion, migration, survival, as well as angiogenesis. The autosomal dominant BMPR2 disease causing mutation has been found to be only 20% penetrant and the use of iPSC identified increased BIRC3 to be related to improved survival, indicating the potential to use protective modifiers of FPAH for developing treatment strategies in the future . The iPSThe iPSC technology is also largely viewed to promote pre-clinical drug trials and screening over animal models to overcome differences in electrophysiological properties between human and animal cardiomyocytes. Studies have shown patient-derived iPSCs to exhibit higher sensitivity towards cardiotoxic drugs that could be the cause for change in action potential and arrhythmia . StudiesThus, iPSC has been successfully used to model and understand pathogenesis of different cardiac diseases, providing insights on pathways around progression as well as for assessment of drug toxicity. These highlight the potential to use iPSC-based models for precision medicine in clinical use.DISC1) and used gene editing to generate isogenic iPS cell lines. This study found depletion of DISC1 protein among the mutation carriers, along with dysregulation of genes associated with synapses and psychiatric disorders in the forebrain. This mutation causes deficit of synaptic vesicles among the iPS-cell derived forebrain neurons. This identification of transcriptional dysregulation in human neurons, highlights a new facet involving synaptic dysregulation in mental disorders [Theoretically iPSC has the potential to be programmed to form any cell in the human body, and coupled with improvements in reprogramming techniques, this technology has advanced our knowledge on disease pathology, developing precise therapeutics, as well as fuel advances in regenerative medicine . In caseisorders . The tecisorders .SOD1) gene that is associated with a slowly progressing form of ALS. This study found successful reprogramming to be possible with only four factors; KLF4, SOX2, OCT4, and c-MYC. Further, the severe disability state of the patients used for harvesting in this case did not seem to block the transformation process or efficiency [OCT4, SOX2, KLF4, c-MYC. A batch of genetically corrected somatic cells using lentiviral vectors encoding FANCA or FANCD2 was also used for reprogramming to overcome the predisposition to apoptosis found in FA cells. The FANCA involved fibroblasts also underwent successful transformation to generate iPSCs. This study also found restoration of the FA pathway as a necessity to generate iPS from somatic cells of FA patients. The persistent FANCA expression in the FA-iPS cells indicated successful generation of genetically corrected FA-iPSCs with functional FA pathway, and disease-free status [Degenerative disease generally progresses through multiple differentiation stages, and using iPSC models, these pathways of transition can be easily identified to assess cause as well as etiopathology better. Amyotrophic lateral sclerosis (ALS) involves loss of neurons from the spinal cord and motor cortex causing paralysis and death. The research around advancement of therapeutics, requires supply of human motor neurons positive for the causative genetic mutations that will also aid in understanding the root cause of motor neuron death. One study documented the production of iPS from ALS patient specific-skin fibroblasts from two sisters. Both were identified to be positive for the L144F (Leu144 → Phe) mutation of the superoxide dismutase (ficiency . Fanconie status .+)/CD29(low) sorting to result in enriching ventral midbrain dopaminergic neurons from the pluripotent stem cell-derived neural cell populations. Further, these neurons also exhibited increased expression of FOXA2, LMX1A, TH, GIRK2, PITX3, EN1, and NURR1 mRNA. These neurons were also found to bear the potential to restore motor function among the 6-hydroxydopamine lesioned rats, 16 weeks after transplantation. Further, the primate iPSC-derived neural cell was found to have survived without any immunosuppression after one year of autologous transplant, highlighting the proof-of-concept around feasibility and safety of iPSC-derived transplantation for PD [Parkinson’s disease (PD) is a common chronic progressive disorder due to loss of nigrostriatal dopaminergic neurons. The pathophysiology of the disease is complex and research till date lacks complete understanding. Further, sporadic cases are not linked to any genetic variation. Development of patient-specific invitro iPSC models have been attempted to understand disease etiology better. Studies have worked upon generating iPSCs from sporadic cases of PD, which have been successfully reprogrammed to form dopaminergic neurons free of the reprogramming factors. This study utilized doxycycline-inducible lentiviral vectors that were excised with Cre-/lox-recombinase, resulting in generation of iPSC free of programming factors, and which retained all the pluripotent characteristics after removal of transgenes. This removal of promoter and transgene sequences from the vector reduced risk of oncogenic transformation and re-expression of the transduced transcription factors. This study highlighted the possibility of generating stable iPS-cell line in PD for better disease modelling . Anothern for PD .Type 1 diabetes is an autoimmune condition involving destruction of the β-cells of the pancreas wherein transplantation with β-cells as islet tissues or the entire pancreas is suggested as an alternative over the traditional exogenous insulin supplementation. However, these come with risk of rejection, need of immunosuppression, apart from difficulty in the physiological control on blood glucose levels. To circumvent this block, generation of β-cells or islet tissues from human pluripotent stem cells like iPSCs has been attempted. Many studies have generated pancreatic β-like cells which secrete insulin in response to stimuli like potassium chloride . HoweverThese studies highlight the development around iPSCs and transplantation technology for treatment of degenerative diseases as well as use them as disease models. The ability to generate patient-specific iPSC from skin biopsies, increases safety of autologous transplants without risk of immunorejection.The treatment for blood disorders involves need for mature red blood cells/erythrocytes from the bone marrow or umbilical cord blood, for blood transfusion, and is limited due to incompatibility in blood group and Rh antigens, and risk of infections . ErythroBCR-ABL gene fusion is the major disease driver, and treatment involves use of tyrosine kinase inhibitor (TKI), causing remission in the vast majority of the cases. Studies have shown the CML-iPSCs to not be affected by TKI even in presence of BCR-ABL expression, indicating absence of dependency in this state of differentiation. The CML-iPSCs factors essential for maintenance of BCR-ABL positive and iPSCs including phosphorylation of AKT, JNK, ERK1/2 remained unchanged while the expression of STAT5 and CRKL was decreased. Further, the hematopoietic cells derived from CML-iPSC regained TKI sensitivity thus facilitating understanding on the disease pathogenesis better [SRSF2 P95L, each of these connected to a specific phenotype and drug response [Studies have also worked on developing iPSC models for blood malignancies including myelodysplastic syndromes (MDS), acute myeloid leukemia (AML), and myeloproliferative neoplasms (MPN). A study worked on generating iPSC clones from bone marrow and blood of patients by integrating mutational analysis with cell programming to generate different iPSC clones which represent different disease stage as well as spectrum of the diseases including predisposition, low- and high-risk conditions. Additionally, the researchers also utilized the CRISPR/Cas9 system to introduce as well as correct mutations in the iPSCs. This study found iPSC from AML patients upon differentiation exhibited the leukemic phenotype, and the derived hematopoietic stem cells contained two immunophenotypically distinct cell populations; an adherent and non-adherent fraction, wherein the adherent fraction cells continuously renewed and generated the non-adherent cells. The AML-iPSC thus generated was found to exhibit characteristics of the leukemia stem cell model thus becoming an efficient model for molecular analysis and studying key functional aspects to be utilized for developing better therapeutics . In cases better ,72. In cs better . To highresponse .+ hematopoietic progenitor cells in beta-thalassemia major adults. Further, the CD34+ were transduced with globin lentiviral vector, wherein the vector-encoded beta-chain was found to be expressed at normal hemizygous protein output levels in NSG mice. This trial thus validated an effective protocol for beta-globin gene transfer among thalassemia major CD34+ hematopoietic progenitor cells [CYBB gene to exhibit antimicrobial activity through generation of ROS and phagocytosis [Human iPSC preclinical models also exist for monogenic blood disorders including thalassemia, and hemoglobinopathies for gene and cell therapy. Pilot trial investigations have explored the safety and effectiveness of mobilizing CD34or cells . The risor cells . Studiesor cells . Human iocytosis .Thus, the potential of iPSCs to study etiology of complex diseases which manifest late in life, as well as to identify markers for precision therapeutics, is worth exploring in the arena of clinical biomedical research. Human iPSC-based models are a true success in our understanding of disease pathogenesis away from the animal models.Organ donations are a key clinical need to treat end-stage organ failure conditions, and in often cases, patients are left to fight the acute shortage for the same. This apart, from identifying HLA-matched donors, handling risk of infections and rejection, as well as life-long immunosuppression, to a great extent damages quality of life for the affected as well as leads to loss of crucial time. Human iPSCs are being evaluated as a potential source for generating organs that can overcome roadblocks of shortage as well as risk of rejection. Studies have explored the possibility of generating a three-dimensional vascularized and functional liver organ from human iPSCs ,80,81. GHepatocytes represent 80% of the liver mass and are the specialized epithelial cells crucial for maintaining homeostasis. The hepatic differentiation involves induction of endoderm differentiation by activin A, fibroblast growth factor 2 (FGF2), and bone morphogenetic protein 4 (BMP4), and such generated hepatocytes have been found to retain features of human liver including lipid and glycogen storage, urea synthesis, etc. Cholangiocytes in the inner space of the bile duct tree have also been generated from the common progenitor hepatoblast, through downregulation of signaling factors including epidermal growth factor (EGF), interleukin 6 (IL-6), Jagged 1, sodium taurocholate, and the generated cholangiocytes have been detected to express mature markers including SOX9 (SRY-Box Transcription Factor 9), OPN (Osteopontin), CK7 (Cytokeratin 7), CK19 (Cytokeratin 19), etc. The kupffer cells are the largest population of resident macrophages in the human body and also facilitate liver regeneration after an ischemic injury. Studies have demonstrated generation of iPSC-derived kupffer cells from macrophage precursors by adding a hepatic stimulus ,84.Another study evaluated lung regeneration by endogenous and exogenous stem cell mediated therapeutic approaches. Physiologically the tissue turnover rate in lung is slow and any insult to the regeneration process can lead to development of chronic obstructive pulmonary disease (COPD) as well as idiopathic pulmonary fibrosis. Bone marrow stem cells, embryonic stem cells, as well as iPSCs have shown excellent regenerative capacity to repair injured lung by generating whole lung in the lab using de-cellularized tissue scaffold and stem cells . Lung orStudies have also utilized iPSC-derived organ models to study pathogenesis of the coronavirus disease-2019 (COVID-19). One study established a screening strategy to identify drugs that reduce angiotensin converting enzyme 2 (ACE2) using human ESCs-derived cardiac cells and lung organoids, as the infection occurs due to binding of the virus to ACE2 on the cell membrane. Target analysis revealed treatment with antiandrogenic drugs to reduce ACE2 expression, thus protecting the lung organoids from the SARS-CoV-2 infection. Clinical studies on COVID-19 identified patients with prostate disease, with elevated levels of circulating androgen to pose increased risk for high disease severity . AnotherDevelopment of organ-specific progenitor cells which progress into the complete three-dimensional organ in a lab highlights the potential of iPSCs in regenerative medicine. Further, the impact of organ-system models to study infection pathology, highlights the wide clinical arena in which iPSC-technology can be used.p53 in development of osteosarcoma. LFS being a heterogenous cancer condition, osteosarcoma is one of the types wherein relevance of germline p53 mutations have been highly reported. The pre-existing murine LFS models have been insufficient in charting the entire tumor landscape and patient-derived iPSCs in this regard have demonstrated the feasibility to effectively study human cancer syndromes. Studies have found the LFS-derived mesenchymal stem cells to exhibit low expression of targets of p53 including p21 and MDM2; highlighting their ability to retain the defective p53 function from the parental fibroblasts. Further, p53 knockdown was found to cause upregulation of osteogenic markers in LFS osteoblasts, and the possibility to attain osteosarcoma-related phenotypes in LFS iPSC-derived osteoblasts was found. Further, gene expression analysis in LFS-derived osteoblasts was found to correlate with poor patient survival, and decreased time for recurrence. The impaired H19 restoration was also found to repress tumorigenic potential [PTPN11 mutations, studies which have derived hiPSCs from hematopoietic cells and which harbor the PTPN11 mutations were found to successfully recapitulate features of NS. The iPSC-derived NS myeloid cells were found to exhibit increased STAT5 signaling and enhanced expression of micro-RNAs viz. miR-223 and miR-15a. Further, reducing miR-223 function was found to normalize myelogenesis, highlighting the role of micro-RNA dysregulation in early oncogenesis [BRCA1-deleted tumor niche to be the cause for disease progression [The iPSCs have been generated for modelling pathogenesis of many diseases, and one of the most notable additions to the same is cancer, including models for familial cancer syndromes. One such study reports on the successful establishment of Li-Fraumeni Syndrome (LFS) patient-derived iPSC to study role of otential . Anotherotential . In caseogenesis . Human igression .The iPSC models around cancer aid in overcoming the hurdles posed by traditional cancer cell line systems, which may lose the characteristics of the original tumor with time, and further harnessing primary cancer cells at different stages of carcinogenesis is not feasible. The established iPSC reprogramming strategies can aid in differentiation of cancer cells to target cell lineages which can aid in studying each of the different stages in cancer progression . The iPSThe iPSCs plug the very fundamental requirement for advancing scientific and clinical research by aiding insight into the fundamental growth and regenerative processes that happen early-on in life. They have emerged as an efficient alternative to ESC in clinical settings especially for screening new pharmaceutical compounds. The ability to produce patient-specific iPSCs has made studying disease pathogenesis and experiments around treating and managing inherited disease conditions, a close reality. They provide an unlimited repertoire of highly differentiated cells from focus area, fueling multiple investigations without roadblock on availability. The fact that iPSCs have also demonstrated success in treating few conditions including substantial correction of SCA and FA, highlights their therapeutic potential . With orScientific and technological advancements have also aided in designing high throughput drug screening systems based on hiPSC stem cell-derived atrial myocytes for detecting cardiac toxicity in atrial arrhythmias. The platform established by human iPS-derived cardiomyocytes was found to be useful in detecting propensity for drug-induced tachyarrhythmias . The potAlthough the impressive prospects of iPSC in clinical settings have been proven, the challenges that need to be realized and worked upon include tumorigenicity, heterogeneity, and immunogenicity. Teratoma formation continues to be serious challenge and a limiting factor in hiPSC transplantation. Though there are multiple elimination methods including cell sorting systems, antibody conjugates, the risk of tumor arising from proliferating progeny still exists. These apart, the reprogramming factors, and occurrence of genetic alterations during in vitro culturing also add to risk of tumorigenicity. Immunogenicity characterized by rejection especially in allograft transplantation deems the need to use immunosuppressants. Heterogeneity has also been reported as a limitation in iPSC which to a great extent is an outcome of genetics and differences in gene expression . Apart fInduced pluripotent stem cells are also now being investigated in clinical trials for diseases and one such involves the Hirschsprung disease initiated in the year 2020. This case-control involves use of patient-derived iPSCs to develop models of the disease to understand genetic factors contributing to disease pathogenesis . The gloHuman iPSCs have become a powerful tool in basic as well as translational and clinical research because of their ability to be maintained indefinitely whilst preserving the genetic makeup of the host. Further, technological advancements which enable genetic manipulation of iPSCs, ensure they become a rich repertoire for cell replacement therapy. Identification of GSH regions further adds to the feasibility to reduce risk of transplantation to a great extent. Their ability to provide cell-specific information, makes studying interaction between genetics, epigenetics, as well as extracellular environment a possibility. Complex relationships involving multiple cell types becomes dissectible with iPSC-models, and their ability to harbor large chromosomal alterations as well as deletions, makes them a favorable model for cancer studies. Though highly researched upon, the technological advancements to decipher the impact of genetic and epigenetic alterations as well as variability in cell clones or colonies is still naïve. Moreover, development of engraftable cell lines need to be devised considering the safety and efficacy aspect.


Drug resistance represents an ultimate challenge for cancer treatment. The initial rationale behind using chemo-treatment was the preferred elimination of fast-growing cancer cells, and the drugs' killing power became the priority. Enormous efforts have been made to develop new types of increasingly potent drugs, improve drug delivery, and test various combinations of therapeutic methods , extra cautions were taken to explain their biological meanings. First, despite some differences comparing to in vivo models, culture cells are truly biological systems, which follow the laws of somatic evolution. Second, the above observations have been confirmed by various in vitro and in vivo models of different types of cancers and patient samples , regardless of their mechanisms , can all eliminate many cancer cells. At the same time, these can also induce genome chaos, a rapid and massive process for re-organizing the genomes, resulting in the emergence of resistant clones with altered karyotypes following a few weeks in culture.The common feature for the survivor cells was the altered karyotype and transcriptome profiles. Initially, the treatment-induced chaotic genomes are highly dynamic, coupled with massive cell death. Later on, many survivor cells are relatively stable, reflected by clonal karyotypes. Comparing the products of multiple runs of parallel experiments resulted in survivor cells that shared similar phenotypes but displayed different karyotypes.The highly diverse types of chaotic genomes detected immediately following drug treatment indicate that different mechanisms can contribute to genome re-organization including structural and numerical genome chaos .Despite that drug treatment-induced karyotype chaos mediated rapid resistance was previously reported  of the chaotic genome, but rather the consequence of this stress-induced process to create new genomes with new genomic information. For example, these drug-induced giant cells are unstable, which can generate drug-resistant cell populations with re-organized genomes. The general model that integrates “high stress in crisis,” “chaotic genome like giant cell,” “new karyotypes,” and “drug-resistance” can be described as a three-stage-process based on two-phased cancer evolution: (1) Under high stresses with cell-killing power, a conservative developmental-related process is active leading to polyploidization. (2) The self-organization process generates large numbers of smaller cells with new genomes selected by genome chaos mediated macroevolution. (3) The survivor cells can further grow into dominant populations via microevolution, often with help of diverse cancer genes. In other words, high stress is the trigger factor, giant cells are the transitional phenotype, genome chaos is the process for rapid and massive genome re-organization, and the newly formed karyotypes of the survivor cell populations are the genotype of drug resistance. Alternatively, some cells can directly re-organize their genome without going through the polyploidization process. All roads lead to new information creation. This explanation fits well with Genome Architecture Theory where karyotype codes system inheritance, and re-organization of the genome represents a powerful way to generate new genomic information essential for cancer macroevolution. The formed cancer systems could grow into dominant cell populations with the help of gene mutation/epigenetic alteration-mediated microevolution Heng, . A recenThe above understanding has diagnosed the conceptual limitations of current treatment strategies, which are mainly focused on killing cancer cells based on the stepwise microevolution cancer model. Knowing that drug treatment can paradoxically harm patients by promoting cancer macroevolution through the creation of new karyotypes, urgent actions are needed for the following priorities: First, validation that drug-induced genome chaos-mediated drug resistance is common in the clinical setting. It is important to know to what extent can this phenomenon be avoided by modifying treatment strategies. Second, we hypothesize that reduced genome chaos might be the reason behind the success of adaptive therapy (Gatenby et al., JY and HH drafted the manuscript. All authors participated in the discussion, literature search, and editing of the manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The everchanging policies and inability to utilize university students due to COVID-19 impacted both residents living in long-term care as well as the next generation of students pursuing careers in the field. University Wisconsin-La Crosse (UWL) faculty strategized solutions as restrictions threatened to impact hands-on opportunities for students. Was there a safe and effective solution to offer residents evidence-based programming while also providing students with vital field experience? Simply stated, the answer was yes. Thus, the UWL Happiness Project was born. This session will outline the UWL Happiness Project, a ten-week, telehealth program implemented between a skilled nursing facility in rural Wisconsin and the UWL Therapeutic Recreation Program, an AGHE Program of Merit for Health Professions designated program. The evidence-based curriculum was developed by an emerging UWL graduate student scholar with faculty mentorship. The innovative curriculum focuses on increasing feelings of happiness using PERMA, a theoretical model grounded in positive psychology. During virtual sessions, older adult residents (ages 65-85) and students built connection while working through weekly focus areas . An overview of AGHE competencies addressed within the project, online course demonstration, and assignment development will be discussed along with information about how these connections fostered an opportunity for students to see aging from a different perspective. This is the first time we are presenting results from the newly developed program. In this, we look forward to sharing student measurements and outcomes, as well as lessons learned during this meaningful, stimulating, and insightful educational session.


Premature ventricular beats (PVBs) in athletes are not rare. The risk of PVBs depends on the presence of an underlying pathological myocardial substrate predisposing the subject to sudden cardiac death. The standard diagnostic work-up of athletes with PVBs includes an examination of family and personal history, resting electrocardiogram (ECG), 24 h ambulatory ECG (possibly with a 12-lead configuration and including a training session), maximal exercise testing and echocardiography. Despite its fundamental role in the diagnostic assessment of athletes with PVBs, echocardiography has very limited sensitivity in detecting the presence of non-ischemic left ventricular scars, which can be revealed only through more in-depth studies, particularly with the use of contrast-enhanced cardiac magnetic resonance (CMR) imaging. The morphology, complexity and exercise inducibility of PVBs can help estimate the probability of an underlying heart disease. Based on these features, CMR imaging may be indicated even when echocardiography is normal. This review focuses on interpreting PVBs, and on the indication and role of CMR imaging in the diagnostic evaluation of athletes, with a special focus on non-ischemic left ventricular scars that are an emerging substrate of cardiac arrest during sport. Adolescents and young adults engaged in competitive sports are at increased risk of sudden cardiac death (SCD) compared with their sedentary counterparts. Sport activity can act as a trigger for life-threatening ventricular arrhythmias (VAs) in athletes with an underlying cardiovascular disease, predisposing them to adrenergic-induced ventricular fibrillation ,2,3,4. SDocumentation of premature ventricular beats (PVBs) on 12-lead electrocardiography (ECG), ambulatory ECG monitoring or ECG stress tests should raise the suspicion of an underlying cardiomyopathy. PVBs with common morphologies such as infundibular or fascicular ectopic beats are often not associated with structural cardiac abnormalities, whereas other PVBs may represent a red flag for the presence of a pathological substrate and must be properly investigated ,9. StandThis review focuses on interpreting PVBs, and on the indication and role of CMR imaging in the diagnostic evaluation of athletes, with a special focus on non-ischemic left ventricular scars that are an emerging substrate of cardiac arrest during sport.Malignant arrhythmic events may occur in athletes with a structurally normal heart as a result of genetic channelopathies , some ofIn master athletes (age >35 years), the most important arrhythmic substrate is ischemic heart disease. Male master endurance athletes, even with a low atherosclerotic risk profile, are more likely to have a higher CAC score compared with sedentary males with a similar risk profile ; the meaBeyond channelopathies and ischemic heart disease, different structural cardiomyopathies can be associated with PVBs, including genetic and acquired heart muscle diseases. Hypertrophic cardiomyopathy (HCM) has been reported to cause more than one third of SCDs in the USA ,15, whilNLVSs have been associated with cardiac arrest occurring during effort ,18. In 2Non-ischemic myocardial fibrosis, in contrast to the ischemic one, involves the mid-mural and subepicardial myocardial layers but spares the endocardium. Since the latter contributes most to myocardial contractility, echocardiographic wall motion evaluation has limited sensitivity in detecting the presence of a NLVS . ECG abnCardiac magnetic resonance is the gold standard for the assessment of biventricular volume, mass and global and regional function ,28; LV fRecently, myocardial strain analysis on feature-tracking CMR, following the footsteps of speckle-tracking echocardiography, has emerged as a surrogate marker of LV systolic function, analyzing the deformation of a fixed myocardial point through the cardiac cycle; it is of prognostic value in different cardiomyopathies such as HCM, dilated cardiomyopathy, ischemic cardiomyopathy or myocarditis ,36,37,38Beyond the role of functional indices, CMR offers the unique ability to identify myocardial tissue abnormalities that may have been missed by other imaging techniques. Tissue abnormalities typical of cardiomyopathies reflect chronic injury, inflammation, infiltration or abnormal storage that can be disclosed by specific CMR sequences. Standard CMR techniques for tissue characterization include T2-weighted CMR images, with an increased signal intensity reflecting inflammation-related myocardial edema ,40; T1-wOver the last two decades, it has been demonstrated that a higher scar volume quantified with late gadolinium enhancement (LGE) predicts the risk of life-threatening VA recurrence in patients with aborted SCD , and theMyocardial mapping, a technique that has been recently introduced in clinical practice, allows measuring the different magnetic properties of each myocardial volume as expressed by magnetic relaxation times  and the extracellular volume (ECV) derived from post-contrast T1. Changes in magnetic properties are typical of diseases characterized by intracellular cardiomyocyte disturbances , extracellular disturbances in the myocardial interstitial tissue  or both  ,51. ConvFinally, arrhythmogenic substrates characterized by inflammation, such as cardiac sarcoidosis, may be better disclosed by integrating CMR with nuclear imaging, particularly positron emission tomography (PET) ,57. CardAccording to the current criteria for ECG interpretation in athletes, abnormalities are labeled as either “common” or “uncommon” ,60. The The modern classification of PVBs follows a similar approach: “common” PVBs are usually idiopathic , whereas “uncommon” PVBs carry a higher probability of an underlying myocardial disease .PVB morphology is the most relevant feature. Athletes with frequent PVBs in the absence of cardiac disease most often show a morphology suggesting a right or left ventricular (LV) outflow tract origin -like with a vertical axis pattern) or from the posterior fascicle of the left bundle branch  with a right bundle branch block (RBBB)-like pattern and superior axis). These PVBs usually decrease during effort ,62.PVBs with an LBBB-like morphology and intermediate or superior axis (denoting a right ventricular (RV) or septal origin), and those with an RBBB-like morphology, wide QRS and superior axis , are classified as “uncommon” because they carry a higher risk of being associated with an underlying disease. The suspect is reinforced when arrhythmias increase in number or complexity during effort ,67,68,69The traditional concept that arrhythmic risk is predicted by “PVB burden”  has been disproved by recent evidence. Indeed, it has been demonstrated that benign extrasystolic foci (more often located in the outflow tract) can give rise to a very high number of PVBs on 24 h Holter ECG in the absence of a pathological substrate ,62. SeveThe evaluation of athletes with PVBs should take into consideration other clinical data: in particular, a positive family history of SCD or cardiomyopathy, history of unexplained syncope or the presence of other major ECG abnormalities (such as T wave inversion) on resting ECG.In the last year, CMR imaging has been increasingly prescribed to investigate athletes with PVBs even when other clinical investigations (particularly echocardiography) are normal. This examination has led to the discovery of isolated NLVSs as the origin of PVBs in some cases that would otherwise have been considered idiopathic.The first systematic study on athletes with VAs in association with an isolated NLVS was reported by Zorzi et al., in 2016 , althougSeveral studies have demonstrated that PVB morphology and behavior during exercise can predict the presence of an underlying NLVS in athletes with apparently idiopathic VAs. Cipriani et al. reported CMR findings in a series of athletes with PVBs who were referred to a tertiary center . FindingAnother investigation aimed to evaluate the PVB burden in apparently healthy young athletes who volunteered to undergo a 12-lead 24 h Holter ECG, including a training session. Seventeen subjects were subsequently referred to CMR for frequent (>500/day), repetitive or training-induced PVBs . In threIn a multicenter study reporting on 251 competitive athletes with a negative family history and unremarkable ECG and normal echocardiography findings who underwent CMR for evaluation of VAs, LV LGE was found in 28 (11%). The scar often showed a subepicardial/midmyocardial distribution (non-ischemic) and was more likely when PVBs depicted an RBBB-like pattern or multiple morphologies, or when non-sustained VT was recorded during exercise testing. Conversely, a high number of PVBs (>3300/day) predicted a normal CMR  23]..23].The systematic evaluation of PVBs occurring during exercise testing increases the diagnostic sensitivity of preparticipation screening compared to traditional first-line investigations . In a population of 10,985 young athletes, adding exercise testing for VAs led to a 75% increase in the diagnosis of pathological cardiac substrates, mainly NLVSs . The morFinally, the correlation between the characteristics of PVBs and the probability of an isolated NLVS as well as the prognostic significance of this CMR finding has also been documented in the general population. In a study by Muser et al. on 518 subjects with frequent PVBs, LGE was diagnosed by CMR in 16%. Polymorphic PVBs and PVBs with a non-infundibular morphology were predictors of pathological myocardial substrates. Nearly one third of the subjects with LGE suffered major arrhythmic events during a median follow-up of 67 months (three died suddenly during effort), underscoring, once again, the precipitating role of adrenergic stimulation as an arrhythmic trigger in patients with a NLVS . The samIn summary, NLVSs are a relevant substrate for malignant VAs, mostly for those occurring during sport activity. The two main PVB characteristics that should trigger CMR prescription to rule out an underlying scar in athletes with otherwise normal clinical findings are the morphology  and the increase in number and complexity during exercise.In athletes with PVBs, a careful clinical examination is required to rule out an underlying myocardial disease that could potentially cause SCD. Although more recent recommendations suggest performing additional testing only when two or more PVBs are recorded on the resting ECG, we believe that even a single “uncommon” PVB should be considered as a red flag .First-line investigations in athletes with PVBs should include history, physical examination, exercise testing, 24 h ambulatory ECG including a training session  and echocardiography . The preExercise testing is a key tool for unmasking electrocardiographic abnormalities or arrhythmias absent at baseline. The effort should be maximal, and the test should be carried out until muscular exhaustion, not just limited to 85% of the predicted maximal heart rate. This is important not only for the documentation of ischemic ST-T changes, but also for eliciting adrenergic-dependent PVBs that may occur only at the peak of effort. Echocardiography is essential to rule out valvular defects, cardiomyopathies, congenital disorders and the anomalous origin of coronary arteries ,83.In case one of the above investigations is abnormal, additional tests may be needed based on clinical suspicion . If first-line investigations are normal, indications for CMR depend on the characteristics of the PVBs  that have been discussed above. When analyzing CMR imaging, attention must be paid to some pitfalls. It is known that CMR shows systematically larger chamber dimensions and volumes, but a smaller wall thickness and mass, compared with echocardiography ; thus, cNot all non-ischemic LGE patterns should be considered markers of disease. Isolated right ventricular insertion point  LGE, i.e., LGE confined to the insertion points of the right ventricle to the anterior and/or posterior ventricular septum, has been described in many pathological conditions such as pulmonary hypertension  or hyperIn both the European Society of Cardiology and the American Heart Association guidelines on competitive athletes, NLVSs are addressed in the myocarditis chapter ,93. The Systematic investigations of athletes with PVBs through CMR imaging in recent years have served to clarify that substrates of SCD, particularly NLVSs, may be missed in routine clinical tests. In the presence of at-risk PVB characteristics, based on their morphology, complexity and relation to effort, CMR should be performed even when echocardiography and baseline ECG are normal. Besides standard CMR scans and post-contrast images, novel techniques such as mapping and feature tracking are also emerging in the subset of athletes’ evaluation.Beyond NLVS, ischemic heart disease represents an important substrate of VAs, especially in master athletes. In the evaluation of older athletes with PVBs, stress CMR in addition to the standard CMR protocol might be useful for ruling out coronary artery stenosis, although further studies are needed to evaluate its diagnostic value in this peculiar clinical setting.


This article describes an emerging non-invasive neuromodulatory technology, called low intensity focused ultrasound (LIFU). This technology is potentially paradigm shifting as it can deliver non-invasive and reversible deep brain neuromodulation through acoustic sonication, at millimeter precision. Low intensity focused ultrasound's spatial precision, yet non-invasive nature sets it apart from current technologies, such as transcranial magnetic or electrical stimulation and deep brain stimulation. Additionally, its reversible effects allow for the causal study of deep brain regions implicated in psychiatric illness. Studies to date have demonstrated that LIFU can safely modulate human brain activity at cortical and subcortical levels. Due to its novelty, most researchers and clinicians are not aware of the potential applications and promise of this technique, underscoring the need for foundational papers to introduce the community to LIFU. This mini-review and synthesis of recent advances examines several key papers on LIFU administered to humans, describes the population under study, parameters used, and relevant findings that may guide future research. We conclude with a concise overview of some of the more pressing questions to date, considerations when interpreting new data from an emerging field, and highlight the opportunities and challenges in this exciting new area of study. Non-invasive brain stimulation is a rapidly growing area of psychiatric research and clinical practice. As our understanding of the neurocircuitry underlying psychiatric disorders has grown, so too has the development of technologies to modulate disease-relevant brain targets. Though promising, most non-invasive stimulation approaches lack the spatial precision of other invasive techniques, limiting their therapeutic utility.The most widely used non-invasive brain stimulation modalities are transcranial magnetic simulation (TMS) and transcranial electrical stimulation. Transcranial magnetic simulation has garnered considerable attention following its success in treating pharmacoresistant depression , 2 and oTranscranial electrical stimulation also has promise in psychiatric research and treatment –13. ThisLow intensity focused ultrasound (LIFU) holds great promise as a novel approach to brain stimulation . Unlike Ultrasound is defined as a sound or acoustic wave higher than 20 kHz . In a typical neuromodulation experiment, a pulse generator emits an electrical waveform, which is amplified and transferred to a transducer housing a piezoelectric element. This amplified electrical signal excites the element, causing the active face of the transducer to oscillate and produce ultrasound waves. The transducer is affixed to the head, often using a head strap, and with important technical developments  can be uLike many other areas of brain stimulation, and more so because of its novelty, the exact mechanism underlying LIFU neuromodulation remains, yet, unknown , 22, 23.SPTA) measures the average intensity during an entire sonication and the spatial-peak pulse average (ISPPA) measures the average intensity over a single pulse. The current US Food and Drug Administration safety guidelines for diagnostic ultrasound imaging devices suggest a maximum derated ISPTA of 720 mW/cm2 and a maximum derated ISPPA of 190 W/cm2 (SPTA.3) seeks to account for attenuation that occurs as the LIFU passes through tissue.Each transducer is composed of a piezoelectric element(s) that transforms electrical signal into ultrasound waves; the construction of this element also provides focality with a specific associated focal length (measured in mm) that defines a three-dimensional zone of maximum applied ultrasound, called “sonication.” A sonication protocol is defined by five parameters: (1) fundamental frequency, (2) pulse repetition frequency (PRF), (3) duty cycle, (4) sonication duration, and (5) intensity . The fun90 W/cm2  to avoidIn recent years, researchers have begun to use LIFU in humans, studying how different parameters can induce reversible physiological effects on the nervous system. Several foundational studies examined the effect of LIFU on neural activity using electroencephalography (EEG). The first study demonstrated that LIFU focally applied to the somatosensory cortex  attenuatIn addition to SEP modulation, recent studies have examined LIFU on the motor cortex. Using a simultaneous ultrasound and TMS paradigm, LIFU to the motor cortex  reduced Badran et al. demonstrated that LIFU may be able to modulate pain sensitivity via thalamic sonication . ResearcPresently there are very few reports of LIFU in patients with neurological or psychiatric disorders . The firRecently, investigators have investigated whether ultrasound may modulate mood and worry. Several studies have used transcranial ultrasound, a less focal ultrasound technique, to modulate activity in regions associated with mood and anxiety symptoms. In one double-blind pilot study, researchers applied continuous ultrasound over the posterior frontal cortex in 31 healthy participants and observed significant mood improvements at 10- and 40-min post-sonication . AnotherSimilarly, in a double-blind pilot study, researchers investigated the effect of right fronto-temporal cortex non-focal transcranial ultrasound on mood and worry in depressed patients . In thisTaken together, the above work represents initial evidence of the feasibility of ultrasound neuromodulation in humans. Non-focal transcranial ultrasound methods point to this technique's ability to alter mood states and set the stage for more focal LIFU experiments. This work also demonstrates that LIFU can target both cortical and deeper brain structures with lasting neuromodulatory effects and high spatial precision.2 ISPTA.3  and the impact of altering parameters awaits systematic examination. Effects of repeated LIFU administration are also in need of evaluation. Furthermore, because LIFU administers low amounts of energy, it is likely that the brain state or context during sonication  may affect the dose–response. The ideal way to combine sonication and manipulation of the neurocognitive state remains unclear  and may The effect of variable skull anatomy across individuals is another important issue in LIFU research. Skull density and composition, particularly presence of trabecular (porous) bone between the cortical bone inner and outer tables, affect ultrasound conduction, as this porous nature leads to scattering. There are clear examples of this when using high intensity focused ultrasound , 48, howA potential confound with respect to LIFU's ability to modulate deep brain structures is that it may activate intermediate regions like auditory cortex , 51. SevWhether LIFU settings are more likely to excite, inhibit, or disrupt neural activity remains an important area of inquiry. To this effect, studies have tested whether PRF  or duty https://itrusst.github.io/).Reflecting the very early nature of the field, there remains no quantifiable metric of LIFU-related physiological effects, with prior work in clinically relevant studies either relying upon behavioral output , 32 or iLow intensity focused ultrasound holds appeal for its unique combination of non-invasiveness, likely safety, precision, and broad range of possible targets. Recent studies provide initial evidence of the safety and feasibility. These advances set the stage for pioneering studies to test causal hypotheses in neuropsychiatric research, utilizing focal, and reversible suppression of core brain regions involved in psychiatric illnesses. A virtually infinite parameter space and brain targets remain to be studied, and further prospective studies are needed to determine dose–response effects on neural activity. Yet, for those parametric studies to be successful, research is urgently needed (and currently ongoing) on safety, particularly in patient populations, as well as inquiry into targeting, acoustic confounds, and mechanisms of action. While the field's understanding of neural mechanisms underlying LIFU remain at a very early stage of study, it appears that the clinically relevant LIFU studies are consistently suppressive. Of note, we explicitly do not use the word inhibitory, as this conveys a degree of mechanistic characterization that is not yet supported by the extant literature.Caveats aside, this technology has tremendous potential for clinical applications. Focal, non-invasive, deep-brain stimulation is currently impossible with available technologies. If the promise of LIFU-induced neuromodulation is realized, it may become not only a powerful tool to test causal relationships between brain activity and function, and transform clinical therapeutics.AA wrote the original draft. All authors reviewed, edited, and approved the final version of the manuscript.This work was supported in part by the National Institutes of Health grants U01MH123427 and P20 GM130452, and the US Department of Veterans Affairs RR&D Center for Neurorestoration and Neurotechnology (I50 RX002864) and a CSR&D Career Development Award (IK2 CX001824). The contents do not represent the views of the National Institutes of Health, U.S. Department of Veterans Affairs, or the United States Government.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The use of alcohol and prescription opioids is common among people in pain and poses significant public health burdens. This review identifies factors associated with motivation to use alcohol and prescription opioids in the context of pain. Pain-relevant, cognitive-affective, transdiagnostic vulnerability factors—expectancies/motives, pain catastrophizing, pain-related anxiety, distress intolerance, anxiety sensitivity, and perceived interrelations—were selected from theoretical conceptualizations of pain and substance use. Searches conducted in PubMed, PsycINFO, and Embase returned 25 studies that examined associations between identified variables of interest and the use of alcohol and prescription opioids in the context of pain. Consistent with a larger literature on pain and substance use, the studies included in this review demonstrated that people with chronic pain are motivated to use alcohol and opioids in response to negative affect and hold expectancies/motives for coping with pain. Vulnerabilities that engender difficulty managing aversive internal states (distress intolerance and anxiety sensitivity) and maladaptive responses to pain (pain-related anxiety and pain catastrophizing) also were implicated in motivation for alcohol and opioid use. Although one study found that pain-related anxiety was associated with co-use of alcohol and opioids, no studies examined simultaneous use. Future research directions that can explicate causal associations, identify patterns of alcohol and opioid co-use, clarify the role of pain in cessation processes, and inform treatment development are discussed. Pain is a complex, near-universal phenomenon, which can be conceptualized as a motivational state that engenders goal-directed action.9The sections that follow begin with a brief overview of alcohol and opioid use, acute and chronic pain, and guiding theoretical frameworks. The results of studies that examined associations between pain, selected transdiagnostic cognitive-affective factors , and alcohol/prescription opioid use patterns/trajectories are then reviewed. Finally, the relevant extant literature is discussed with an emphasis on explicating clinical implications and generating recommendations to help guide future research in this emerging domain.Approximately 50% of American adults consume alcohol each month,21Although definitions vary in the literature, in the context of alcohol and opioids, co-use may be characterized as concurrent use  or simultaneous use .7Although a handful of documented cases indicate a rare congenital inability to perceive pain,42The extant literature suggests that pain co-occurs at high rates with both alcohol and opioid use, potentially in a dose-dependent fashion.Pain is an inherently aversive experience that reliably elicits negative affect and motivates escape/avoidance behaviors.5In drawing upon motivation theory, negative reinforcement frameworks, and the four-stage model of pain processing, a leading reciprocal model posits that pain and substance use interrelate in the manner of a positive feedback loop, resulting in more severe pain, greater functional impairment, and the maintenance of addiction.5Given the high degree of co-occurrence and significant individual/societal costs associated with alcohol and prescription opioid use, the goal of this review is to explicate potentially modifiable factors that are associated with motivation to use and co-use alcohol and prescription opioids in the context of pain.64The constructs of interest in this review were derived from theoretical frameworks of motivation and pain–substance use relations, with a focus on cognitive-affective constructs that are hypothesized mechanisms of the pain–to–substance use pathway and have been implicated as vulnerabilities to multiple psychiatric disorders.For the current review, expectancies, motives, and perceived interrelations broadly refer to the extent to which people perceive associations between their pain and substance use.69Pain-related anxiety is the tendency to respond to actual or anticipated pain with anxiety or fear, which may motivate avoidance behaviors.75Anxiety sensitivity 82Distress intolerance also may contribute to pain and substance use reciprocity. Research has consistently demonstrated positive associations between distress intolerance ,89Literature searches were conducted in PubMed, PsycINFO, and Embase using the terms alcohol drinking OR alcohol-related disorders OR analgesics, opioid OR opioid-related disorders; pain; and expectancies OR motives OR perceived interrelations OR negative affect OR pain-related anxiety OR catastrophizing OR anxiety sensitivity OR distress intolerance. All search criteria were limited to human species and peer-reviewed journals published in English before December 2020. Searches yielded 124 unique records after duplicates were removed. Given this review’s focus on the pain–to–substance use pathway, the authors sought to identify studies that examined alcohol/opioid criterion variables in relation to at least one of the selected constructs. They included studies that utilized pain-related predictor variables  or were conducted among relevant pain populations . Studies conducted among healthy, non–treatment-seeking samples were included only if (a) the sample was necessary to answer a pain-related research question , and (b) the study included at least one other variable of interest. Primary reasons for exclusion were (a) it was not a behavioral study of the pain–to–substance use pathway (81 studies) or not a relevant population (32 studies).Twenty-five studies were identified for inclusion see .N = 128), high-intensity pain ratings (vs. no or low-intensity pain) were associated with a greater number of self-reported drinks per day and higher alcohol craving, and participants with high-intensity pain were more likely to report normalizing motives for drinking .92Initial qualitative and cross-sectional evidence suggest that people with chronic pain hold unique cognitions about how pain and alcohol use are related. First, a qualitative study of 10 people living with HIV who had chronic pain and reported heavy drinking  provided evidence that alcohol may be seen by people with chronic pain as a primary means of coping with both pain and pain-related distress.N = 2,125), pain severity at the end of treatment predicted frequency and quantity of alcohol use at long-term (9- to 12-month) follow-up. Moreover, pain predicted increased negative affect, which mediated the effect of pain on drinking outcomes.Consistent with the larger substance use literature and theoretical conceptualizations of negative affect as a primary motivator of substance use behavior,Although several studies have implicated pain-related anxiety in associations between pain and other substances ,Like pain-related anxiety, pain catastrophizing has been widely studied in relation to tobacco smoking and cannabis use.N = 45). Over the 90-day assessment period, greater levels of pain and negative affect were individually associated with use of opioids at higher doses during the same day, although negative affect was not statistically tested as a mediator.96Initial evidence suggests that negative affect mediates proximal associations between pain and prescription opioid use, similar to alcohol use. Although the search did not return any experiments that tested causal association between pain and opioid use in humans, prospective studies that utilized repeated assessments provide evidence for the effects of pain on prescription opioid use via negative affect. First, real-time ecological momentary assessment among 34 patients on long-term opioid therapy for chronic pain indicated that, over the 2-week assessment period , patients were more likely to report opioid use during occasions of increased pain, and they consumed higher doses when pain was accompanied by increased negative affect.N = 256) who endorsed moderate to severe past-month pain, greater pain-related anxiety was associated with increased likelihood of self-reported addiction to opioids, history of family concern about opioid use, past use of opioid detoxification, and more opioid-related problems.100Results from several cross-sectional studies suggest that pain-related anxiety is associated with multiple indices of prescription opioid misuse among people with chronic pain. First, in an online survey of young adults  and 3% met cut-offs for both hazardous drinking and opioid misuse in the past month .Prior research has demonstrated that pain motivates substance use and may contribute to the maintenance of addiction.90Motivational models of both pain and substance use highlight the role of negative affect as a primary determinant of escape/avoidance behaviors, and negative reinforcement is likely a key mechanism by which pain motivates and ultimately maintains substance use.Pain-related anxiety and pain catastrophizing are both thought to motivate maladaptive attempts to avoid or alleviate pain.64Although both alcohol and prescription opioids present health risks when used individually, co-use is associated with increased morbidity and mortality and presents a significant public health threat.Studies included in the current review consistently yielded evidence suggesting that negative affect and other maladaptive cognitive-affective responses to pain and distress may cause alcohol and prescription opioids to take on greater salience in the context of pain. As shown in Future research also is needed to better understand motivation to use alcohol and prescription opioids in the context of other comorbidities. First, co-use of alcohol and opioids with other substances —particularly those that increase risk for medical consequences and public health burden—should be examined. For example, 22% of opioid-related overdose deaths involve co-use of alcohol, opioids, and benzodiazepines.Given the central role of motives and expectancies in motivational models of substance use,Although only a few included studies examined sex/gender differences, this review did observe initial evidence that pain and pain-related constructs may hold greater motivational salience for alcohol and prescription opioid use among men, relative to women. Future research should investigate reasons why men may be more motivated to use substances in the context of pain. For example, men and women likely experience varying pharmacologic effects of alcohol and opioids as a function of biological sex,Finally, this review identified studies that predominantly focused on motivational processes in the context of ongoing substance use. Additional research is needed to better understand whether cognitive-affective and transdiagnostic vulnerability factors are also associated with motivational processes in the context of substance-related cessation, reduction, and abstinence. For example, several constructs  previously were associated with relapse to tobacco smoking among people with chronic pain.Although based on a relatively nascent empirical literature, the current review highlights the importance of assessing pain when treating alcohol and prescription opioid use. Integrated treatments may be especially useful for addressing co-occurring pain and substance use because, relative to traditional approaches , they can be more efficient and cost-effective and can focus on both conditions as treatment targets.Motivational enhancement interventions that target treatment engagement  and motivation to reduce or abstain from alcohol and opioid use also are needed. Although interventions to increase awareness of opioid overdose risk behaviors have been developed,160Finally, personalized feedback interventions (PFIs) are a subset of motivational interventions that show greater promise for treating co-occurring pain and substance use, and can be delivered via scalable treatment modalities .Chronic pain and use of alcohol and prescription opioids co-occur frequently, and pain is a potent motivator of alcohol and opioid use. People with chronic pain may be motivated to use alcohol and opioids in response to negative affect or in response to expectancies/motives for pain coping. Transdiagnostic vulnerabilities for maladaptive responses to pain  and difficulty managing or tolerating aversive states and negative affect  also may motivate alcohol or opioid use in the context of pain. Future research should examine the role of transdiagnostic factors in motivating patterns of alcohol or opioid use or co-use over time . Integrated interventions for alcohol and prescription opioid use that address pain-relevant, cognitive-affective processes  also should be developed and tested.