Cancer recurrence remains a great fear for many cancer survivors following their initial, apparently successful, therapy. Despite significant improvement in the overall survival of many types of cancer, metastasis accounts for ~90% of all cancer mortality. There is a growing understanding that future therapeutic practices must accommodate this unmet medical need in preventing metastatic recurrence. Accumulating evidence supports dormant disseminated tumor cells (DTCs) as a source of cancer recurrence and recognizes the need for novel strategies to target these tumor cells. This review presents strategies to target dormant quiescent DTCs that reside at secondary sites. These strategies aim to prevent recurrence by maintaining dormant DTCs at bay, or eradicating them. Various approaches are presented, including: reinforcing the niche where dormant DTCs reside in order to keep dormant DTCs at bay; promoting cell intrinsic mechanisms to induce dormancy; preventing the engagement of dormant DTCs with their supportive niche in order to prevent their reactivation; targeting cell-intrinsic mechanisms mediating long-term survival of dormant DTCs; sensitizing dormant DTCs to chemotherapy treatments; and, inhibiting the immune evasion of dormant DTCs, leading to their demise. Various therapeutic approaches, some of which utilize drugs that are already approved, or have been tested in clinical trials and may be considered for repurposing, will be discussed. In addition, clinical evidence for the presence of dormant DTCs will be reviewed, along with potential prognostic biomarkers to enable the identification and stratification of patients who are at high risk of recurrence, and who could benefit from novel dormant DTCs targeting therapies. Finally, we will address the shortcomings of current trial designs for determining activity against dormant DTCs and provide novel approaches. Recent years have seen great strides in the treatment of primary tumors, as well as in treating overt metastatic tumors. Nonetheless, the main cause of mortality of cancer patients remains metastasis and recurrence. Metastasis, the spread of tumor cells from the primary site to secondary organs accounts for 67-90% of all cancer mortality , 2. DespSome cancers are more notorious than others for delayed recurrence. These highly recurrent tumors include kidney cancer, acute myeloid leukemia (AML), non-small cell lung cancer (NSCLC), melanoma, prostate cancer, ovarian cancer, breast cancer and osteosarcoma. Kidney cancer and AML exhibit low single/double-digit recurrence rate –8, whileLung cancer is the leading cause of cancer-related death with NSCLC accounting for ~90% of new cases . Even whWhile metastasis and recurrence are the main cause of mortality in cancer patients, the mechanisms underlying metastatic recurrence years and decades after initial treatments are just beginning to unravel. Recurrence due to metastatic spread begins as a multi-step process that can take months or years until it becomes detectable. Recent studies have shown that although the probability of metastasis increases with the size of the primary tumor, cancer cells nonetheless leave primary tumors early –32 and sOnce cancer cells arrive at their new and foreign microenvironment (‘non-permissive soil’) they face several fates. The majority of them will undergo apoptosis and thus will meet their demise. Those that successfully launch adaptive and survival programs will enter a dormant state. Some dormant DTCs may reside as single solitary quiescent cells (cellular dormancy) and/or as small clusters of quiescent cells. Others may reside as small indolent micrometastases where cellular proliferation is balanced by apoptosis –36. ThesCell-intrinsic mechanisms governing DTC dormancy and escape from dormancy are influenced by signals arising at their foreign niche. Accumulating evidence in the literature attributes dormancy and survival of residing DTCs and their reactivation to the intricate cross- talk with their ‘non-permissive’ or their ‘permissive’ niche, respectively –43. HencThis review will focus on potential strategies, mechanisms and drugs to be considered for targeting quiescent dormant DTCs by manipulating their microenvironment and or their cell- intrinsic mechanisms. We will initially present clinical evidence for the presence of dormant DTCs. Potential biomarkers to enable the identification and stratification of patients who are at high risk of recurrence will also be discussed. Finally, we will address the shortcoming of current clinical trial designs for demonstrating activity against dormant DTCs, either quiescent or indolent, and provide novel trial designs.Demonstrating the presence of dormant DTCs and their subsequent progression in clinical settings has been challenging. However, advances in detection have provided new information and a growing body of evidence in the clinic to support the idea of early dissemination of tumor cells from the primary site, followed by subsequent dormancy and late recurrence.One clinical example that supports tumor dormancy is late-stage recurrence. Recurrence of a tumor after more than 5-years remission is in line with an initial dormancy period followed by reactivation and outgrowth. In support of this idea, it was found that breast cancer patients who do not have detectable disease can have circulating tumor cells (CTCs) found in their blood 20+ years after initial treatment . The inaAnother clinical observation in support of early metastasis and subsequent dormancy can be found in cases of unknown-primary carcinoma (UPC). UPC accounts for ~5% of metastatic cancer cases, and while some cases are later identified after more thorough evaluation, 30% of patients never have a primary site identified . These uOne of the best pieces of evidence for dormant DTCs that can recur after a long period of quiescence is seen in cases where patients develop tumors after organ transplantation. Accidental transmission of tumors from tissue transplants derived from seemingly cancer-free cadavers was first reported in the case of kidney transplants . In one Clinical data demonstrate the presence and persistence of dormant DTCs years and even decades after treatment. In some cases, these DTCs will remain dormant without relapsing. Hence, unraveling the mechanisms responsible for their long term ‘hibernation’ may set the premise to develop novel therapeutic strategies to prevent cancer from recurring by keeping them dormant indefinitely.Several restrictive signals have been described in the bone marrow (BM) and lung that maintain DTCs originating from breast, prostate, head and neck squamous carcinoma and multiple myeloma cells in their quiescent state Figure 1SPARC, also known as osteonectin, was shown to regulate tumor dormancy of prostate cancer cells by promoting the expression of BMP7 in BM stromal cells. SPARC was shown to be epigenetically silenced in aggressive cells by promoter methylation whereas treatment of prostate cancer cells with the DNA demethylating agent 5-azacytidine (5-AZA) or with the COX2 inhibitor NS398 could restore SPARC expression in malignant prostate cancer cells. This in turn promoted BMP7 expression in BM stromal cells leading to dormancy of prostate cancer cells . Hence, Another component that may be reinforced is thrombospondin-1 (TSP1), which is found in the suppressive BM and the lung niches. TSP1 secreted by stable microvasculature or by recruited BM-derived myeloid cells was shown to induce tumor dormancy of breast cancer cells at the perivascular niche (PVN) in the BM  and prevHence, the approach of inducing natural factors of the suppressive niche such as TSP1 and or SPARC is very appealing. However, this approach will require continuous treatment to ensure indefinite quiescence of the residing dormant DTCs and thus may lead to potential toxicities. Furthermore, TSP1 has been also reported to exert opposing effects which can be also attributed to multiple receptors that mediate TSP1 signaling . LikewisTherefore, it may be more practical to induce common cell-intrinsic mechanisms converging from the different microenvironmental cues comprising the dormant niches.high–ERKlow signaling axis) as hallmark of tumor dormancy in several types of tumors  induction vs. reduction in ERK -MAPK signaling  and in DTCs of prostate cancer patients . NR2F1 wprograms . Furtherprograms  and induprograms , 56. Henprograms Figure 1The microenvironment of the metastatic niche , 40, 71 via activation of focal adhesion kinase (FAK) by Src. This activation resulted in downstream activation of ERK, which in turn induced phosphorylation of myosin light chain (MLC) by myosin light chain kinase (MLCK), culminating in F-actin stress fiber organization and transition from quiescence to proliferation. Inhibition of MLCK activation  induced the outgrowth of dormant DTCs in the lungs by formation of neutrophil extracellular traps (NETs), which in turn lead to cleavage of laminin-111 by NET-derived elastase and MMP-9 and induction of the Intβ1/Src/FAK/MLCK axis   Figure 10536016) , 77.Therefore, if repurposing JSM6427 to treat cancer patients is being considered, the clinical regimen by which this drug will be administered must also be considered. In light of initial studies demonstrating Intβ1 plays a perquisite role in the reactivation of dormant DTCs [reviewed in ] while hIndeed, surgical trauma induces local and systemic inflammatory responses that can also contribute to the accelerated growth of residual and micrometastatic disease –91. Henc®), recently approved by the FDA for the treatment of myeloproliferative neoplasm-associated myelofibrosis  or lysyl oxidase like 2 (LOXL2), could prevent formation of the fibrotic milieu. Indeed, it was shown that LOX and/or LOXL2 inhibition significantly decreased pulmonary metastatic burden , 95. Furutgrowth , 97. Oveutgrowth Figure 1Interestingly, a recent report demonstrated how a systemic inflammatory response induced after surgery promotes the emergence of dormant immunogenic DTCs at distant anatomic sites while, preoperative anti-inflammatory treatment with meloxicam, a nonsteroidal anti-inflammatory drug (NSAID), prevented the outgrowth of DTCs in the lungs . NotablyOverall, the studies presented here emphasize the important role of inflammation and/or fibrosis in dormant DTCs outgrowth and also reinforce the notion that intervention in the perioperative stage and immediately after re-section may be critical to prevent local and/or distant recurrences.Once DTCs anchor in their new and “non-permissive niche” adaptive cell-intrinsic mechanisms ensure their long-term survival and escape from immune surveillance , 42, 100The mechanisms responsible for the long-term survival of DTCs are just beginning to emerge.in vivo model system to study tumor dormancy  on dormant breast DTCs was also found to ensure their survival once they colonize the lungs Figure 2Notably, senescence of tumor cells that have escaped cytotoxic therapy has been proposed by several groups as another form of cell-intrinsic mechanisms that confer tumor dormancy and survival of circulating tumor cells (CTCs) and/or DTCs residing at the perivascular niche [reviewed in ]. This eOverall, these promising studies highlight the importance in further investigating the cell- intrinsic mechanisms governing survival of dormant DTCs.via c-Src activation  has been reported to constitute aspects of the dormant niche and act as a survival factor. CXCL12 secreted by osteoblasts was shown to induce the survival of disseminated breast tumor cells expressing the CXCR4 receptor by upregulating the Akt pathway tivation . Furthertivation –122 and tivation . Plerixativation . This aptivation –127. Thutivation Figure 2F-box/WD repeat-containing protein 7 (FBXW7) was shown to promote quiescence by ubiquitylation and proteasomal degradation of cell cycle promoters. Ablation of FBXW7 in dormant breast cancer cells caused DTCs to exit their quiescent state, which sensitized the cells to paclitaxel treatments in mouse xenograft and allograft models .Recently, the TGFβ2/GAS6-Axl axis  was showAnother protein mediating the engagement of cancer cells to the BM niche is osteopontin. Osteopontin is an ECM protein secreted by osteoblasts in the BM niche and was shown to anchor acute lymphoblastic leukemia (ALL) cells to the BM culminating in their dormancy. Whereas, inhibition of osteopontin promoted ALL escape from tumor dormancy and sensitized them to cell-cycle–dependent Ara-C chemotherapy .Overall, these studies suggest that dissociating dormant DTCs from their ‘safe haven’ niche can promote the cells to exit their quiescent state and potentially sensitize them to chemotherapy treatments.Importantly, there are striking similarities between dormant DTCs and quiescent normal hematopoietic stem cells (HSCs), which reside in the BM. These similarities are exhibited by intrinsic mechanisms for survival, quiescence and their mobilization to the BM . TA recent report by Ghajar and his colleagues  demonstrHence, there exists a risk of DTCs mobilization from their dormant niche. Therefore, another strategy that we may contemplate to eradicate dormant DTCs is by preventing their immune evasion.A recent report by Malladie and colleagues demonstrated that cancer cells selected from lung and breast cancer cell lines for their competence to establish latent metastasis (dubbed LCC), acquired quiescence with stem-like characteristics by expressing the Wnt inhibitor DKK1. The authors demonstrated that autocrine DKK1 helps disseminated LCC cells enter quiescence. Furthermore, these quiescent cells downregulated natural killer (NK) cell ligands leading to evasion of immune surveillance. Specifically, quiescent LCCs downregulated cell surface UL16-binding protein (ULBP) activators of NK cell-mediated cytotoxicity, as well as receptors for cell death signals. Therefore, quiescent (dormant) LCCs escaped cytotoxic killing by NK cells whereas proliferating LCCs were eradicated by activated NK cells . Of noteOverall, these initial findings may open up in the future a novel therapeutic approach to selectively eradicate dormant DTCs by inhibiting their immune evasion. This will require re-expression of NK ligands in dormant DTCs. Interestingly, the inhibitor of histone deacetylases (HDACs) trichostatin A (TSA) was shown to induce ULBP expression in epithelial cancers . WhetherNotably, several studies demonstrated how autophagy in cancer cells can impair the susceptibility of the cancer cells to NK-mediated killing . Given tAnother mechanism by which DTCs were shown to evade the immune system is by downregulation of the expression of major histocompatibility complex class I protein (MHC-I), thus evading CD8+ T cell recognition . FurtherHence, upregulating MHC-I may be an attractive approach to reinstate DTCs vulnerability to immune surveillance Figure 2Overexpression of immune checkpoint proteins on dormant tumor cells was also shown to facilitate their immune escape. Dormant tumor cells in the DA1-3b/C3H mouse model of AML evade cytotoxic T-lymphocyte (CTL)-mediated killing because they overexpress PD-L1 (B7-H1) and CD80 (B7-1) . PD-L1 bMyeloid–derived suppressor cells (MDSCs) may also indirectly regulate the survival of dormant DTCs. MDSCs represent a population of special cells of the immune system, which consist of immature macrophages, immature granulocytes, and immature dendritic cells. MDSCs suppress activation of T and NK cells through the production of reactive oxygen species (ROS) and arginase 1 (Arg-1), along with recruitment of other immune suppressive cells such as regulatory T cells [reviewed in ]. AccumuHence, modulating MDSC immunosuppressive activity may in turn prevent immune evasion of dormant DTCs. Inhibitors of phosphodiesterase-5, sildenafil and tadalafil, were shown to inhibit the immunosuppressive activity of MDSCs in preclinical and clinical studies by the downregulation of inducible nitric oxide synthase (iNOS) and Arg-1 activities . PromisiOverall, several drugs that have already been tested in the clinical setting may be considered for inhibiting the immune evasion of dormant DTCs thus potentially leading to their demise Figure 2Current studies which are just beginning to unravel the intricate crosstalk between the residing dormant DTCs and their niche, highlight the complexity and the need to rethink the design of future therapeutic strategies to prevent dormant DTCs from ever emerging. If a common multi-faceted target that will both inhibit dormant DTCs and their niche can be identified, the cross-talk between them could be affected leading to their long-term hibernation or their demise.A potential multifaceted target that has been demonstrated to be a molecular hub in mediating tumor escape from immune surveillance  and reguHence, STAT3 may serve as an attractive clinical target given that inhibitors of STAT3 have already entered clinical trials along with newer inhibitors at the preclinical stage [reviewed in ]. Moreovvia the activation of leukemia inhibitory factor (LIF) receptor  to study combined 5-AZA and ATRA treatment on top of standard of care in recurrent prostate cancer patients based on rising prostate-specific antigen (PSA) only.Indeed, recent studies demonstrated that combining the epigenetic regulating drug 5-AZA with the differentiation agent trans-retinoic acid (ATRA), reinstated in part NR2F1 expression , while 5Overall, these studies may open up novel avenues to maintain DTCs dormancy.Once a therapy is developed to either target dormant DTCs for destruction or prevent their emergence from dormancy, the question arises as to which patients are at sufficient risk to warrant a therapy in the absence of overt disease. The decisions by clinicians to treat patients using such therapies will largely depend on the risk/benefit potential for such therapies, as well as healthcare-associated costs. Assuming that novel treatments to prevent recurrence will involve risks and substantial costs, the ability to stratify patients and determine for which patients the treatment is likely to provide benefit would be extremely valuable. This is true for clinical trials and post approval.via imaging. Fortunately, using innovative approaches, it seems like we are gaining ground. Various technologies that utilized liquid biopsies enable collection of surrogate markers. In addition, it may be possible to predict recurrence based on properties of cells in the primary tumor. Hence, identifying biomarkers in liquid biopsies and in primary tumors that will enable stratification of patients that are at high risk for recurrence, while remaining a great challenge, offers great hope.Patient prognosis and stratification in order to inform treatment decisions is far from being a simple endeavor. Dormancy and emergence from dormancy are presumably determined by properties of individual dormant DTCs and of the milieu in which they have lodged, as well as other factors such as the patient’s immune system. These properties may determine the risk/timing of relapse. Currently, detecting dormant DTCs through tissue biopsy is limited to BM. Furthermore, it is difficult to find biomarkers related to dormant DTCs, or provide prognosis predictions Current literature provides evidence for the identification of such markers specifically in breast cancer patients. A recent retrospective cohort study in breast cancer patients with a five-year follow-up after diagnosis found differential expression within the recurring tumors. The differential expression of the proteins was also related to breast cancer subtype. For instance, TNBC patients who recurred had significantly higher expression of Snail protein in their primary tumors compared to those without recurrence, whereas Twist expression was significantly higher in primary tumors of estrogen receptor and progesterone receptor positive breast cancer patients who recurred compared to those without recurrence . NotablyImportantly, a 21-gene recurrence-score assay  provides prognostic value. This gene-expression assay is used to assess risk of disease recurrence in hormone receptor-positive, HER2-negative breast cancer patients and to guide decisions regarding adjuvant chemotherapy . The assIndeed, a recent study demonstrated stratification of dormant DTCs in the BM of breast cancer patients based on their NR2F1 expression . ImportaImportantly, although some gene expression assays of the primary tumor can be used to determine both early and later recurrence and the efficacy of extended adjuvant endocrine therapy in ER+ breast cancer patients, the use of such assays is not recommended for guiding therapy beyond 5 years , 164. HeLiquid biopsies, which includes circulating tumor cells (CTCs), circulating cell-free tumor DNA (ctDNA) and extracellular vesicles (EVs), may hold great promise in clinical diagnosis. The presence of CTCs in the peripheral blood of patients after tumor resection denote the existence of minimal residual disease and may provide insight into the process of metastatic spread and enable real-time monitoring of disease progression and therapeutic response [reviewed in ]. SeveraA recent study by Sparano and colleagues demonstrated the prognostic value of CTCs in predicting late recurrence of breast cancer patients. The presence of CTCs in peripheral blood samples of hormone receptor–positive breast cancer patients obtained approximately 5 years after diagnosis provided independent prognostic information for late clinical recurrence . CTCs poThe prognostic value of CTCs was also illustrated in prostate cancer patients. A recent study demonstrated CTCs detection in patients who had undetectable PSA levels following radical prostatectomy. Furthermore, these patients with CTCs had an increased risk of biochemical recurrence (defined by an increase of PSA levels) . Hence, a priori to be present in the primary tumor based on previous genomic analysis of the primary tumor. Using this approach Garcia-Murillas and his colleagues performed mutation tracking in plasma DNA of early breast cancer patients receiving neoadjuvant chemotherapy. Detection of ctDNA and mutation tracking of several plasma samples after completion of apparently curative treatment predicted metastatic relapse with a median lead time of 7.9 months over clinical relapse , progression free survival (PFS) and overall survival (OS) identify only agents able to control dormant DTCs when this effect also controls other malignant cell growth of a measurable lesion. It is possible that drugs that are effective in current clinical trial designs also have an activity against outbreaking dormant DTCs. Therefore, a new approach that better aligns unmet therapeutic needs with drug development efforts is required.Practically, upfront trials, where patients who present with a primary tumor are treated with standard of care plus drug/s that can target dormant DTCs are more challenging than trials with patients that already have had metastatic recurrence. This is due to the fact that only a percentage of patient who present with a primary tumor will recur, and due to the fact that the time for a first recurrence can be anywhere from months to many years. As an example, if a trial design is based on EFS at two years, where about 40% of the patients are expected to have an event, the trials must be sufficiently powered to detect improved EFS in the subset of patients who entered the trial. These trials require larger number of patients and they require a longer time of treatment and observation compared to trials in patients who already had relapse. Furthermore, unless agents given to such patients have been specifically shown to eliminate DTCs, they will need to be continuously administered potentially for many years, which is often a challenge, especially when therapies given in conjunction with standard of care that includes chemotherapy. Therefore, it is also less likely that an open label study would be pursued; it is much more likely that a controlled study with a new agent on top of standard of care would be compared to standard of care alone. Implications for time to recruit patients and overall costs is very substantial compared to the option of an open label study. This is especially true given the challenges of identifying the dose that would be optimal for Phase II and III trials. Briefly, since a trial design that utilizes maximum tolerated dose to seek an effect on lesion size is not possible when targeting microscopic dormant DTCs, establishing a selected dose is quite challenging. This is described at more length in the study by Steward et al. .Alternatively, trials in patients that already had a relapse are expected in many cancers to require fewer patients, be shorter in duration of treatment and recruitment time, and importantly be suitable for open label study designs versus controlled studies. As an example, if patients with a certain cancer with well documented historical data have an EFS of 20% at 8 months post-surgical removal (resulting in no measurable disease), a study of a novel agent targeting dormant DTCs with an objective of doubling EFS to 40% at 8 months would require several dozen patients only. An alternate approach could be to target cancers that have a high prevalence of delayed metastasis but that occurs over a manageable time frame. Examples include osteosarcoma, TNBC and melanoma, although the latter also can recur a decade or more later.Therefore, a development plan in which efficacy of dormant DTCs targeting agents can be determined in smaller trials in patients that have already recurred followed by larger trials in upfront settings is more likely to have support by clinicians, advocates and industry. Drugs that demonstrate efficacy in patients that already recurred can then be tested in the upfront trial scenario, informed by safety, efficacy and PK/PD data from the previous trials.via imaging or biopsy), or indirectly  detect and follow such dormant DTCs and indolent micrometastases, we must rely on traditional clinical end points to assess the activity of drugs that may target and eliminate or control dormant DTCs. Unfortunately, there is no established trial design that completely accomplishes this task.As noted herein, dormant DTCs are undetectable at distant sites and reside as single/small cellular quiescent cells and or as small indolent micrometastases. Until it becomes possible to directly  provides an excellent disease model for this discussion. OS recurrence is observed in about 40% of patients presenting with localized disease who are treated with standard of care, most commonly to the lungs within a few years of completing therapy . About 8The hypothesis that we believe needs to be tested clinically involving dormant DTCs posits a limited number of dormant DTCs and/or dormant indolent tumors either residing alone or in the presence of undetectable micrometastases. Thus, in the presence of undetectable micrometastases even a 100% effective agent at either eliminating dormant DTCs or preventing their outgrowth may fail since the preformed micrometastases may progress clinically. Thus, in order to demonstrate the effectiveness of dormant DTCs targeting therapies in OS, novel trial designs are required. This study design would be based on the hypothesis that some undetectable tumors are too advanced to be affected by drugs that target dormant DTCs. In other words, if microscopic micrometastases resulting from outbreaking dormant DTCs are already present at the time of initial recurrence, they will emerge as additional lesions with or without DTCs treatment. Such a recurrence could be considered an “early failure” and its presence will not truly reflect the efficacy of an agent preventing the outgrowth of dormant DTCs or eliminating dormant DTCs Figure 4Metastatic recurrence is the major cause of mortality of cancer patients and poses a major unmet clinical therapeutic challenge. Unraveling the mechanisms underlying recurrence which arises years and decades following a protracted period of tumor dormancy may open up novel avenues to prevent disease from recurring. Being able to prevent delayed metastasis, either by specifically targeting dormant DTCs for destruction or maintaining them in the dormant state forever would represent a significant step forward and could save many lives. In this review, we introduced several potential strategies and drugs, some that can be repurposed and may prevent the outbreak of the hibernating DTCs based on growing studies in the field. These strategies aim to maintain the DTCs in indefinite dormancy and/or eradicate them. We also proposed to identify potential gatekeepers in regulation of dormant DTCs and their supportive microenvironment. These strategies could also potentially be complementary to each other. We have also provided various mechanisms and drugs, including some that may be repurposed with a shorter path to treating patients compared to novel compounds in preclinical stages. Notably, each strategy presented here has its complexities and must take into consideration the therapeutic window for treatment and patient stratification that would benefit from such treatment. Ideally, a preventive treatment should be started at the neoadjuvant/adjuvant setting to prevent the disease from ever emerging and/or prevent, delay, or minimize further progression in patients who present with a recurrence.In order to increase the ability to observe clinical impacts of novel dormant DTC targeting drugs, novel trial endpoints may be required. Since novel dormant DTC targeting drugs are designed to target such DTCs before they exit dormancy, non-dormant cells are likely to progress during treatment and therefore a clinical benefit may only be evident at a later time point . However, later outcomes that would be expected by controlling the dormant DTCs with novel drugs may be expected and could have meaningful impacts on disease progression and survival.Overall, these strategies could potentially open up novel avenues in the battle against cancer recurrence and may develop a strong foundation for developing drugs that would ensure that cancer will never recur.SS, MI, RH, and DW: research, writing, and editing. DR: conception and writing. DB: conception, research, writing, and editing. All authors contributed to the article and approved the submitted version.DB supported by a Research Career Development Award from the Israel Cancer Research Fund, Israel Cancer Association, United States-Israel Binational Science Foundation grant # 2017237 and Israel Science Foundation grant # 942/20. DR supported by the National Pediatric Cancer Foundation .SS, RH and DW were employed by Vuja De Sciences Inc. DB and MI have consulting agreements with Vuja De Sciences, Inc.The remaining author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Eukaryotic organisms share many common features in terms of endomembrane trafficking. This fact has helped plant scientists to propose testable hypotheses on how plant intracellular membrane trafficking is achieved and regulated based on knowledge from yeast and mammals. However, when a new compartment has been identified in a plant cell that has a vesicle tethering complex located at a position which is completely different to its counterpart in yeast and mammalian cells, caution is demanded when interpreting possible interactions with other trafficking elements. This is exemplified by the recently discovered EMAC (ER and microtubule-associated compartment). It has been postulated that this compartment is the recipient of vacuolar sorting receptors (VSRs) transported retrogradely via “retromer vesicles” from a post-Golgi location. Unfortunately, this suggestion was based entirely on our knowledge of retromer from yeast and mammalian cells, and did not take into account the available literature on the composition, localization, and function of the plant retromer. It also lacked reference to recent contradictory findings on VSR trafficking. In this short article, we have tried to rectify this situation, pointing out that plant retromer may not function as a pentameric complex of two subunits: the retromer core and the sorting nexins. Exchange of material between intracellular membrane compartments is mediated by vesicles and is essential for the function of eukaryotic organisms. Endomembrane vesicular transport is orchestrated by regulatory protein complexes, whose subunits are often evolutionary conserved between distantly related organisms (Bethune and Wieland trans-Golgi network (TGN) where it serves to tether incoming endosome-derived retrograde transport vesicles. Thus, it came as a surprise to discover that neither fluorescently tagged VPS51 nor VPS54 colcalized with standard TGN, Golgi, or multivesicular body (MVB) markers. Furthermore, whereas treatment with the Golgi-vesiculation inhibitor brefeldin A (BFA) typically leads to a redistribution of the Golgi membrane marker YFP-MEMB12 into the ER, the VPS51 and VPS54 signals remained unaffected. This was also the case after treatment with wortmannin which causes MVBs to enlarge via homotypic fusion  which are expressed in all eukaryotes. In yeast and mammalian cells, a second complex, usually containing two sorting nexins (SNX), is associated with the trimeric core retromer complex. Whereas Vps35p binds to endosomal membranes and was considered to interact with the cytosolic domain of sorting receptors for acid hydrolases , the SNXs, having a BAR-domain, were believed to be responsible for inducing membrane curvature and therefore driving tubule formation at the endosomal membrane   that bind to the recycling sorting motif WLM in the tail of the CI-MPR  and the site from which VSRs are recycled (the TGN). After the submission of this manuscript, we became aware of the recent opinions article of Scheuring and Kleine-Vehn  who also


Due to the COVID-19 pandemic, the Centers for Medicare and Medicaid Services (CMS) issued a memorandum for long term care facilities to restrict visitation by all nonessential personnel, including guardianship case managers. To enhance caseworker access to their guardianship clients, an eastern seaboard state agency distributed Amazon Echo Show devices to clients in long term care facilities. These touchscreen and voice-activated devices have both video and audio capabilities. This study reports the results of the first phase of a comprehensive evaluation, pilot testing the devices via a group of “superuser” case managers to understand the potential challenges and benefits of using these devices. Sixteen case managers participated in two virtual focus groups before and after the installation of an Echo device with one of their guardianship clients. Participants were asked to discuss experiences in accessing clients and client engagement before and after device installation. The focus groups were audio-recorded and transcribed verbatim and two researchers independently identified themes using open and axial coding. Major themes identified included: challenges to device installation and use, strategies to overcome challenges, benefits to using Echo devices, and ethical concerns. These findings suggest that touchscreen or voice-activated devices with video capability can assist case managers in protecting vulnerable clients and ensuring their well-being when in-person access is restricted. Additionally, the devices can be used to enable isolated residents to connect to the outside world, including family, friends, and case managers through technology. Strategies for replication of this innovative program will be discussed.


Autism spectrum disorder (ASD) is increasingly common with 1 in 59 children in the United States currently meeting the diagnostic criteria. Altered sensory processing is typical in ASD, with auditory sensitivities being especially common; in particular, people with ASD frequently show heightened sensitivity to environmental sounds and a poor ability to tolerate loud sounds. These sensitivities may contribute to impairments in language comprehension and to a worsened ability to distinguish relevant sounds from background noise. Event-related potential tests have found that individuals with ASD show altered cortical activity to both simple and speech-like sounds, which likely contribute to the observed processing impairments. Our goal in this review is to provide a description of ASD-related changes to the auditory system and how those changes contribute to the impairments seen in sound discrimination, sound-in-noise performance, and language processing. In particular, we emphasize how differences in the degree of cortical activation and in temporal processing may contribute to errors in sound discrimination. Autism spectrum disorder (ASD) is a developmental condition that is characterized by abnormalities in social communication, restricted behavior, and repetitive behavior. People with ASD also experience altered sensory processing and show both hyper- and hypo-reactivity to sensory input . For refThese ASD-related changes in auditory behavior and linguistic processing are reflected in dysfunction at the cortical level. Cortical function in ASD is hypothesized to be impacted by an increase in endogenous cortical “noise,” resulting from altered ratios of excitation to inhibition within neural circuits . This reA meta-analysis determined that 90% of individuals with ASD experience sensory abnormalities, with auditory hypersensitivity as the most common modality . One wayTo probe cortical responses to sound in ASD, studies generally present participants with various simple sound targets and then measure cortical response using electroencephalograms (EEG) or magnetoencephalograms (MEG). Both EEGs and MEGs represent cortical auditory processing as a series of waveforms (positivities and negativities) that occur in a stereotyped sequential manner as different portions of the cortex become active in response to sound. Generally, EEGs confer greater temporal resolution of responses, while MEGs have superior spatial resolution. Here, we will discuss how “sensory peaks”  respond to simple sounds in ASD, and in how changes in those sounds are represented by the mismatch negativity (MMN).P1 is an early cortical response to sound and is thought to reflect thalamocortical transmission along the ascending auditory pathway . In typiWhen processing sound, EEGs and MEGs show an event approximately 100 ms after stimulus presentation known as the N1 (EEG) or M100 (MEG). This response is thought to represent activity at the auditory cortex, superior temporal gyrus, and auditory association areas. There are also data suggesting parietal and frontal cortex involvement . In typiWork examining N1/M100 in people with ASD frequently reported abnormalities in lateralization. In typically developing people, the left temporal cortex response to sound is generally greater than that of the right temporal cortex . HoweverAs a general statement, people with ASD seem to show greater peak amplitudes and longer peak latencies for N1/M100. This trend is informative with regard to how auditory function may be fundamentally altered in ASD at early stages of cortical processing. Delays in peak latency correlate with stimulus complexity and the recruitment of neural resources. In which case, longer peak latencies and broader recruitment of neural resources could mean that individuals with ASD find simple stimuli to be more complex than control participants generally do . These fThe mismatch negativity is a waveform that reflects changes in stimuli; generally, it is considered to act as an automatic orienting reflex that marks changes in an environment. The MMN can be modulated by participants focusing their attention on a stimulus, but can still be elicited when attention is not directed at stimuli . The MMNResearch that tested the MMN amplitude in response to pure tone stimuli in ASD reported a range of findings that may reflect variation in participants’ sub-diagnosis, degree of intellectual impairment, tolerance of change, or age ; Table 3Similar to research describing MMN/MMF peak amplitude, data describing MMN/MMF latency vary by the study design and the participant composition. Consistent with the superior pitch discrimination observed in ASD, People with ASD demonstrate abnormal responses to phonemes and other speech-like sounds, which may stem from an altered perception of less complex stimuli or from impairments in auditory attention that are specific to linguistic components. Phonemes are the units of sound that comprise a language. Therefore, while phonemes have simple and complex tonal components, they have additional meaning in that they are used in an inherently social context. Aberrations in how “simple” stimuli are processed may hinder people with ASD in processing phonemes to some extent; however, there is also evidence that processing difficulties may be unique to linguistic elements.The MMN response to phonemes is somewhat varied, though some trends have emerged. Some studies reported that MMN/MMF peaks from participants with ASD were not different from those seen in typically developing children when presented with vowel sounds . HoweverMagnetic mismatch field latency was generally delayed in vowel discrimination oddball tasks for individuals with ASD . When teIn summary, work that studied the ASD MMN/MMF in response to speech stimuli varied with intellectual impairment, ASD subtype, and the nature of the auditory task. MMN did not seem to be especially sensitive to changes in the physical features of simple sounds (pitch and duration); however, it was markedly altered in response to vowel changes. This may imply (1) that there are stimulus features specific to language that slow auditory processing and (2) that impaired ability to rapidly detect changes in incoming speech stimuli may be fundamental to ASD language processing deficits. Consistent with this line of thought, The P3a is a response to unexpected or surprising stimuli and is thought to reflect activity in the frontal, temporal, and parietal lobes . As suchIn two different studies, reduced P300 amplitude was seen in response to phonemes but not to tonal stimulus. In the first, Research that studied how people with ASD process phonemes focused on how specific components of speech may impede processing in ASD. Those inquiries found that people with ASD had difficulty following phoneme changes, may become distracted by contextually irrelevant features of language, and may show attentional deficits with regard to linguistic stimuli. In addition to the challenges that seem to be intrinsic to speech processing in ASD, individuals with ASD experience particular difficulty processing sound when it is presented amid environmental noise. Background noise can impact simple sound detection, but can also impair linguistic processing to a greater or lesser degree depending on the features of the background noise stimuli. The presence of background stimuli could also contribute to the heightened cortical “noise” that is thought to interfere with auditory function in ASD .Even relatively simple background noise can increase the processing load in ASD to the point at which target identification is impaired. To investigate how the presence of background noise affected simple sound processing in ASD, In this vein, While background noise impairs speech detection in ASD generally, the features of the background noise in which those speech targets are presented can also impact how well participants are able to extract speech. For instance, people with ASD were found to perform poorly in speech discrimination tasks when the background noise contained temporal dips. Most often, tests aimed at determining how well individuals with ASD are able to extract speech sounds from background noise consist of having participants identify either a word or portions of a sentence while simultaneously presenting a background noise that competes with the target in some way. To test how spectral and temporal variations in background noise effected individuals with ASD’s ability to detect speech, As another possibility, individuals with ASD may find speech in noise conditions challenging as a result of altered voice perception. While individuals with ASD do seem to experience aberrations specific to processing speech and voices, given that speech is an inherently social activity, the desire to engage in social exchange must also be considered when testing speech-in-noise processing. When participants with ASD were presented with a conversation between two people that had competing “ecologically valid” background noise , typically developing and participants with ASD had comparable patterns of brain activity . HoweverDelayed or atypical development of sensory systems is a common feature of ASD and may underlie the peak latency and amplitude changes observed in EEG/MEG testing. In typically developing individuals, waveforms become more complex as children age. Importantly, auditory development in early childhood sees reductions in P1 latency and amplitude as N1 becomes more prominent . Early dThe majority of studies that characterized N1 also found increases in peak latency and amplitude that were almost exclusively seen over the right hemisphere, suggesting poor response lateralization in ASD. In typically developing people, auditory areas of the brain (including the STG) undergo synaptic pruning events that cause decreases in size relative to brain volume between childhood and young adulthood. During this developmental period, a left-right hemisphere asymmetry in auditory area volume is established. Symmetry in auditory area size in ASD seems to be rooted in a failure to loose volume in the right (or non-dominant) hemisphere of the brain . As suchWith regard to P1, peak amplitude was reduced in ASD in all studies reviewed, including studies with participants as young as 4 years old. This shows that even from a young age, sensory processing is abnormal in children with ASD . P1 alsoAt rest, individuals with ASD show differences in their states of arousal  relative to control groups . ParticiDifferences in how people with ASD direct their attention when undergoing auditory EEG/MEG testing could also account for some of the observed inconsistencies in the literature. Individuals with ASD consistently show anomalies in joint attention and orienting to speech, and frequently show co-morbidity with attention deficit/hyperactivity disorder (reviewed in Idiopathic ASD is the most common form of ASD; however, that diagnosis may describe a number of genetic conditions that potentially have slightly different presentations. Comparing non-syndromic ASD with known genetic conditions on the autism spectrum is useful in interpreting some of the variability found in ASD EEG/MEG data. For instance, Rett syndrome is a condition on the autism spectrum that presents with cognitive impairments. Consistent with ASD results, individuals with Rett syndrome show general increases in early sensory peak latencies , but divThe capacity to distinguish speech sounds and parse speech depends on the ability to follow rapid temporal cues on the order of milliseconds. As such, it is conceivable that even small changes in temporal processing may ultimately have a significant impact on an individuals’ ability to perceive and understand speech . Gap detIn review, some consistent themes emerged with regard to ASD EEG/MEG data, but still many EEG/MEG components reported a range of responses. This is at least somewhat expected given the variety of sub-diagnoses, intellectual ability, and developmental delays represented under to ASD umbrella. In order to better understand the driving factors in ASD-related sensory disability, future studies may incorporate the following five considerations. First, studies that required participants to direct their attention toward or away from a stimulus found consistent changes in EEG/MEG responses . Given tThe author researched and wrote the entire manuscript.The author declares 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 human prefrontal cortex is a structurally and functionally heterogenous brain region, including multiple subregions that have been linked to different large-scale brain networks. It contributes to a broad range of mental phenomena, from goal-directed thought and executive functions to mind-wandering and psychedelic experience. Here we review what is known about the functions of different prefrontal subregions and their affiliations with large-scale brain networks to examine how they may differentially contribute to the diversity of mental phenomena associated with prefrontal function. An important dimension that distinguishes across different kinds of conscious experience is the stability or variability of mental states across time. This dimension is a central feature of two recently introduced theoretical frameworks—the dynamic framework of thought (DFT) and the relaxed beliefs under psychedelics (REBUS) model—that treat neurocognitive dynamics as central to understanding and distinguishing between different mental phenomena. Here, we bring these two frameworks together to provide a synthesis of how prefrontal subregions may differentially contribute to the stability and variability of thought and conscious experience. We close by considering future directions for this work. The prefrontal cortex (PFC) is a broad swath of brain tissue encompassing numerous cytoarchitecturally and functionally heterogenous subregions. This heterogeneity has been explored throughout the last century of neuroscientific research, from cytoarchitectonic parcellations in the beginning of the last century , to funcThe DFT and the REBUS model distinguish among the diverse PFC-related mental phenomena by highlighting a central dimension of thought and conscious experiences: its stability vs. variability over time. The DFT describes human thought as a sequence of dynamically changing mental states that range from being highly variable to being highly stable over time: highly stable thought dynamics feature a narrow range of contents separated by predictable transitions, whereas highly variable thought dynamics feature a diverse range of contents interspersed by relatively unpredictable transitions. The DFT also describes how alterations in large-scale brain network interactions may underlie changes in the variability or stability of thought. The REBUS model, on the other hand, suggests that serotonergic psychedelics induce a heightened variability of conscious experience  throughPut together, these two frameworks can provide a conceptual basis for understanding how different prefrontal subregions may contribute to increased stability or variability in thought and conscious experience. Here, we combine insights afforded by the frameworks in an attempt to do that. We begin by reviewing how the two frameworks theoretically approach the stability and variability of thought and conscious experience. We then present a summary of how the primary concepts put forth by either framework are related to the functions of individual prefrontal subregions and the large-scale networks they form. We close by integrating perspectives from the two frameworks and considering future research directions.The DFT is a conceptual framework for understanding human thought  that disThe DFT proposes two general ways through which the dynamics of thought can be altered: deliberate and automatic constraints Fig. . These cDeliberate constraints are exemplified by cognitive control and executive processes that are supported by the brain’s control networks , leading to an overall increased variability of conscious experience. At the neural level this corresponds to decreased top-down signaling and increased informational entropy of underlying neural activity. The REBUS model reflects a growing body of functional neuroimaging findings that show the effects of psychedelics on major scale brain networks and their prefrontal nodes , 98–100.The DFT and REBUS model exhibit a variety of conceptual synergies. First, both frameworks treat cognitive control as a process that stabilizes thought and conscious experience. Within the DFT it is described as deliberate constraint upon thought, while the REBUS model describes it as deliberate influence on the psychedelic state that can be disruptive to positive therapeutic changes . Indeed, the standard protocol for psychedelic therapy encourages participants to “let go” or “surrender” to the experience , 102, whincreased variability of thought and conscious experience ., 32.5, 3Deliberate constraints are closely linked to recruitment of the brain’s frontoparietal and cingulo-opercular control networks Fig. . These nDeliberate constraints may primarily contribute to an increased stability of thought by constraining the contents and transitions between thoughts in line with current goals or task demands. This is associated with the recruitment of multiple prefrontal control network regions, including the dorsolateral PFC (DLPFC), rostrolateral PFC (RLPFC), and dorsal anterior cingulate cortex , while the RLPFC represents multiple rule-outcome relationships simultaneously to help guide rule selection. These rules may contribute to increased stability in thought and conscious experience by specifying a particular range of behaviors and cognitive process to occur over others.Thought suppression may be one example of this. During thought suppression unwanted thoughts are purged from one’s conscious experience . CertaiOther studies also show the DLPFC is closely associated with thought suppression , exhibitThe RLPFC may also affect thought dynamics, through its representation of multiple rule-outcome associations. Another study  that empThis difference in DLPFC vs. RLPFC connectivity with the default network core between these manipulations is consistent with the previously described difference in function between DLPFC and RLPFC according to level of abstraction in cognitive control: instructions to “think creatively” are more abstract and supraordinate than the rules that may have guided the rejection of primed, highly common verbs, and specific ideas. Such supraordinate-level information about how to “think creatively” may have included multiple rule-outcome associations connecting different thought dynamics to valued outcomes, perhaps including different executively demanding idea generation strategies that may be alternated . Their iIt is possible, however, that deliberate constraints supported by the frontoparietal control network may also contribute to increased variability in thought during creative thinking. For example, the frontoparietal control network has been found to “drive” brain dynamics toward unique network configurations that favor the generation of highly creative ideas . This maThe cingulo-opercular control network supports deliberate constraints in ways that may be distinct yet complimentary to that of the frontoparietal control network. While the frontoparietal control network primarily underlies deliberate constraints that seem to operate on shorter timescales, the cingulo-opercular control network appears to support deliberate constraints at relatively longer timescales. This is suggested by the functions associated with RLPFC and dorsal ACC , with one’s goals. This may contribute to an increased stability of conscious experience in contexts when the value of one action clearly outweighs the value of other possible actions, and when this difference in values remain relatively stable over time. Indeed, the dACC has been argued to underlie “tenacity”—the ability to maintain a consistent pattern of behavior over time . In indiAutomatic constraints are associated with the functions of multiple brain networks, including the default network with its three subcomponents, the salience network, and the ventral attentional network . Like deAround the turn of the 21st century, a set of brain regions became identified for being consistently deactivated during external task demands but being consistently activated when such external task demands were reduced . These rScientific understanding of the default network has evolved considerably in the last two decades. It is no longer thought of as a “task-negative network”, but is instead understood to support a variety of internal mental processes , 20, incCORE) is comprised of the anterior medial PFC, posterior cingulate cortex, and inferior parietal lobule, each of which are highly interconnected with many other brain regions [The core subcomponent of the default network  is the anterior-most node of the DNRE Figs.  and 2. I thought  that bia thought .Consistent with this, activation in aMPFC increases as a function of the self-relatedness of ongoing processing. This includes increased activation when judging whether an adjective applies to oneself , which aThese findings all involve the manipulation of relatively abstract self-related information: adjectives and personality traits are broad descriptors that generalize across specific information about a person . This maSchemas refer to complex and abstract knowledge structures formed by extracting and generalizing statistical regularities across previous individual experiences , 201. ThMTL has been observed during induced rumination in healthy controls [MTL were found in depressed individuals during a resting-state [MTL to fit with negative self-schemas. Such negative self-schemas may engender stereotypic sequences of thought, marked by highly predictable contents and manners of unfolding.For example, an excessive biasing towards abstract, personally significant thought contents may be a characteristic feature of ruminative thought –208. Rumcontrols , while lng-state . These fMTL) spans the hippocampus, parahippocampus, retrosplenial cortex, posterior inferior parietal lobule, and ventromedial PFC activating hippocampal-neocortical neural ensembles and enabling mental state transitions through associative cueing between mental contents [The medial temporal lobe subcomponent of the default network contents . This isMTL, the ventromedial PFC (vMPFC) may support increased automatic constraints on thought. Since the label “vMPFC” is somewhat inconsistently used in the literature [In contrast to the medial temporal lobe structures located in the DNterature , 201, heterature .MTL such as the hippocampus, parahippocampus, and retrosplenial cortex , that sis issue , 222–225is issue , 227 foris issue , 122, this issue  for selfis issue –195, andis issue  for infois issue –234. Andis issue , 235–239is issue , 240.Activation in mOFC positively correlates with the degree of rated familiarity and anticipated pleasantness for future episodic simulations , rated pThe mOFC’s role in automatic constraints may also be framed in terms of its association with schema-guided memory processes. The mOFC, as well as vMPFC more broadly, is closely linked to schema construction and maintenance . For insThought that is heavily schema-dependent may rely upon interactions between the mOFC and the hippocampus during initial construction, suggesting that the mOFC may serve to provide an inceptive schematic-template to help guide thought generation . Such a Combining its affiliations with value-based and schema-dependent thought, we suggest that the mOFC may contribute to an increased stability in thought and conscious experience by automatically constraining cognition and behavior around highly valued schemas. For example, if someone is planning to propose marriage to their significant other, they may be prone to engage in future-oriented thought about the proposal when even the slightest cue appears given the high value of the topic. This line of thought may be heavily dependent upon schemas for its elaboration since it is future-oriented and is likely to be structured by predictable cultural narratives (or schemas) about how the proposal ought to occur. As such, mOFC recruitment may underlie increased automatic constraints on thought by making highly valued thoughts more likely to be elaborated upon while also organizing their associated streams of thought around formularized schema. This is likely to result in a concomitant increase of stability in thought and conscious experience.SUB3), also known as the dorsomedial subcomponent, spans the temporopolar cortex, lateral temporal cortex, posterior inferior parietal lobule, inferior frontal gyrus, and dorsomedial PFC  node of the DNUB3 Fig. , which iSUB3), the dMPFC thus appears to underlie abstract, highly constructive processing more broadly, such that its close association with social processing may only reflect the high frequency with which social information is encoded and processed at an abstract level [single event, in contrast to the coarse-grained global features of multiple episodes referred to by the term schema [Similar to its parent network , the rACC may contribute to constraining conscious experience by continuously capturing the salience of the perceived pain and making it more likely to be continuously represented in thought and conscious experience over time. But in situations where viscero-sensory channels are more variable, such as during states of comorbid alexithymia and anxiety , the rACThe ventral attention network comprises the anterior insula, ventral extent of the temporoparietal junction, and inferior frontal gyrus  in the external environment. Therefore, the right IFG, through its affiliation with the ventral attention network, appears to support automatic constraints for salience detection for information in the external environment. So, while the rACC appears to detect salient stimuli that are interoceptive in nature, reflecting salient changes in the internal environment, the right IFG may perform a similar function for stimuli that are spatially localized in the external environment.The right IFG also may exerts automatic constraint on brain regions involved in motor control. Intracranial electrical recordings show the IFG to have downstream effects upon the primary motor cortex  and subtBecause the right IFG shows transient recruitment in response to salient changes in the external environment , it may The right IFG may also in some circumstances contribute to reducing the variability of conscious experience. This may occur, for example, when the external environment is stable and relatively invariable with respect to the relevance and salience of its constituents. In such circumstances—for example, driving on a quiet road along a familiar route—attention may become perceptually decoupled and turn toward the internal stream of thought, which may be relatively freely moving  and moreBeliefs (or predictions) serve to increase the brain’s processing efficiency by allowing for only the unpredicted portions of incoming information to be sent upwards along neurocognitive hierarchies . In thisCORE, salience network, and frontoparietal control network, as well as their connectivity to network nodes outside of the PFC, appear closely related to the process of ego-dissolution [Psychedelics disrupt the normative sense of self: one of their most consistent and profound effects is a subjectively experienced “ego-dissolution” , 329–332solution , 99, 100CORE significantly decrease following psilocybin and LSD administration [For instance, functional connectivity between the aMPFC and posterior cingulate cortex of the DNstration , 24, whistration . These rstration . These fHowever, the autobiographical sense of self is not the only aspect of self-experience that becomes weakened under psychedelics, and may not even be the first to undergo attenuation , 328. EaFurthermore, the early relaxation of bodily self-beliefs may play a causal role in the subsequent relaxation of mental self-beliefs. Given that the aMPFC occupies a relatively higher hierarchical position than the rACC along the principal gradient of functional brain organization , 30, 335In addition to relaxing mental and bodily beliefs about the self, psychedelics also reduce the capacity for goal-related beliefs to support an increased stability of thought and conscious experience , 30, 100Both control networks have been shown to exhibit a particularly high expression of serotonin 2A receptors . LSD andIn summary, psychedelics seem to reduce the constraining influence that mental self-beliefs, bodily self-beliefs, and goal-related beliefs have over lower levels of the neurocognitive hierarchy. By relaxing these beliefs, psychedelics reduce the extent to which incoming information is processed in terms of a mental self, bodily self, and goals, ultimately contributing to an increased variability of thought and conscious experience. This is due to an increase in the amount of bottom-up information flow that is no longer constrained according to hierarchically superior beliefs (or predictions).Recent theoretical advances through the DFT and REBUS frameworks have highlighted the importance understanding mental dynamics and the sources of variability and stability of mental states across time. Here we brought together these two frameworks and their predictions with what is known about the functions of different prefrontal subregions. The present discussion suggests a number of future directions for research that could give rise to an improved understanding of the dynamics of conscious experience and their clinical implications.First, an important conclusion from the preceding discussion is that constraints that may occur at the neural level do not necessarily result in increased stability of conscious experience over time. As noted, for example, in the discussion of regions such as the dACC, rACC, and inferior frontal gyrus, the neural constraints these regions contribute to could result in either increased or decreased variability of mental states over time, depending on the stability or variability of the internal and external environments. Therefore, it would be beneficial for future experimental and theoretical work to elucidate the complex and contextually dependent relationship between constraints at the neural level and constraints at the level of consciously experienced mental states.Second, regional differences in contributing to greater stability or variability in thought and conscious experience could be understood further by examining possible gradients of specialization and functional organization across adjacent cortical regions. For example, future work could examine the potential information processing gradient along the ventral-dorsal axis of the ACC. At the turn of the century, rostral portions of the ACC were proposed to underlie more emotional processes while dorsal areas were linked to more cognitive ones . Recent Third, to be able to make stronger regionally specific predictions regarding specific prefrontal subregions, neuroscientific work related to large-scale brain networks could benefit from more fine-grained and regionally specific analysis approaches. In particular, the field of psychedelics research could benefit from a more specific discrimination between different brain networks and regions. Neuroscientific work on psychedelics often makes use of whole-brain measurements , 99, 339Fourth, alongside functional gradients of cortical organization , 342, thFinally, existing knowledge and interventions for clinical conditions could be enhanced by paying greater attention to the stability or variability of thought and conscious experience that they feature. The DFT and REBUS model  have already led to new assessment tools and interventions for various clinical conditions. These include a behavioral assessment for detecting early-stage Alzheimer’s disease based off the prevalence of spontaneous thoughts , a new mOverall, the dynamics of thought and conscious experience hold promise to significantly advance our scientific understanding of brain function and organization, along with their clinically significant alterations. A more complete understanding of those dynamics, however, will be impossible without appreciating their significance and impact at multiple levels of brain organization. Although it can be challenging to combine different levels of explanation, such as large-scale network analyses and regionally specific functional localizations of brain function, we hope that our present undertaking hints at the benefits of doing so and motivates further work that attempts similar multi-level analyses of neural and mental functions.


Athletes and certain occupations  must navigate unique heat challenges as they perform physical tasks during prolonged heat stress, at times while wearing protective clothing that hinders heat dissipation. Such environments and activities elicit physiological adjustments that prioritize thermoregulatory skin perfusion at the expense of arterial blood pressure and may result in decreases in cerebral blood flow. High levels of skin blood flow combined with an upright body position augment venous pooling and transcapillary fluid shifts in the lower extremities. Combined with sweat-driven reductions in plasma volume, these cardiovascular alterations result in levels of cardiac output that do not meet requirements for brain blood flow, which can lead to orthostatic intolerance and occasionally syncope. Skin surface cooling countermeasures appear to be a promising means of improving orthostatic tolerance via autonomic mechanisms. Increases in transduction of sympathetic activity into vascular resistance, and an increased baroreflex set-point have been shown to be induced by surface cooling implemented after passive heating and other arterial pressure challenges. Considering the further contribution of exercise thermogenesis to orthostatic intolerance risk, our goal in this review is to provide an overview of post-exercise cooling strategies as they are capable of improving autonomic control of the circulation to optimize orthostatic tolerance. We aim to synthesize both basic and applied physiology knowledge available regarding real-world application of cooling strategies to reduce the likelihood of experiencing symptomatic orthostatic intolerance after exercise in the heat. Furthermore, autonomic regulation of blood flow, sweating and other responses during increases in internal temperature are coordinated with other essential processes to maintain normal physiological function even in environmental extremes. The goal of the present paper is to discuss the regulation of body temperature and blood pressure in a specific setting: post-exercise hyperthermia and its relationship with decreases in orthostatic tolerance (OI). We will then discuss the mechanisms by which post-exercise body cooling may be an effective countermeasure to both protect against heat illness and counteract any tendency for orthostatic intolerance that may occur in the post-exercise state.Occupations such as firefighting and the military often require work levels, clothing and/or ambient temperature exposures that are well-beyond thermoneutral “comfort” levels that most humans would consciously choose. Nonetheless, human physiological thermoregulation is remarkably capable of regulating elevations of core body temperature . This region contains temperature sensitive neurons that respond with changes in firing rate to their own  temperature as well as to inputs they receive from peripheral thermoreceptors Boulant, . ElevatiThe skin circulation works in concert with sweating to increase dissipation of heat from the body during increases in body temperature. Skin blood flow in humans is controlled by two branches of the sympathetic nervous system. Sympathetic noradrenergic vasoconstrictor nerves exhibit tonic activity at rest in thermoneutral environments, whereas the sympathetic active vasodilator system is only activated during increases in internal body temperature. During heat exposure, the initial thermoregulatory response in the skin is to withdraw the activity of the vasoconstrictor system. If body temperature continues to increase, the cutaneous vasodilator system is activated  and atrioventricular (AV) nodes) and across the myocardium to increase heart rate and contractility.In terms of human cardiovascular function, the parasympathetic nervous system is primarily limited to vagal control of heart rate. The vagus nerve releases acetylcholine at the SA and AV nodes, decreasing heart rate via a decrease in the slope of the pacemaker potential in these cells. With the onset of exercise and with assumption of upright posture, the first mechanism to increase heart rate is a withdrawal of parasympathetic activity, followed by an increase in sympathetic activity.The arterial baroreflex is the major autonomic reflex controlling blood pressure in humans. This reflex responds primarily to changes in blood pressure sensed by changes in activity of baroreceptors located in the carotid sinus and aortic arch :Cardiac Output (CO) × Total Peripheral Resistance (TPR)= [Heart Rate (HR) × Stroke Volume (SV)] × Total Peripheral Resistance (TPR)= We note that sympathetic and parasympathetic neural mechanisms are able to control all three of the major variables that contribute to the maintenance of a normal arterial pressure.Direct measurement of autonomic function in humans has proven challenging and thus has driven somewhat of a reliance on directly or indirectly measured cardiovascular components  to evaluate change in mean arterial pressure. In this section, we will briefly review some of the most common autonomic measurement techniques; the interested reader is referred to several comprehensive reviews on this topic . Under many  conditions, including rest, plasma norepinephrine is strongly correlated with directly measured activity of the sympathetic nervous system (see next). However, the concentration of norepinephrine in the plasma at a given time is the net result of release (spillover), reuptake and metabolism – so that changes in any of these could result in changes in the plasma [NE], without changes in actual sympathetic noradrenergic activity. Thus, while this approach is helpful for assessing sympathetic activity, plasma NE data should be interpreted in the context of its limitations.A method that has received increasing attention in recent decades is the approach of using frequency  analysis of cardiovascular variables  to give insight into the activity of sympathetic or parasympathetic nerves controlling those variables , contributed to an intravascular fluid shift, decreasing plasma volume by 205 mL. A shift in net filtration of plasma from the blood into the interstitium is postulated to result from an increase in capillary hydrostatic pressure as a result of increased cutaneous venomotor tone  exposure, with minimal influence on Tc and heart rate  has gained popularity as a post-exercise recovery technique due to its efficacy in recovering thermoregulatory variables including TOrthostatic tolerance is typically evaluated in a controlled laboratory setting using a head-up tilt test or lower body negative pressure  and was able to similarly reduce mean skin temperature during heated tilt to ~29.6°C (~Δ 8.5°C). Furthermore, a cool water perfused suit, applied during 5 min of active 70° head-up tilt, similarly decreased skin temperature to 28°C. Compared to post-tilt normothermia, cooling induced a tilt response marked by greater mean arterial pressure largely attributable to an exaggerated increase in total peripheral resistance . Durand et al.  began LBThe results of these studies clearly established the efficacy of whole-body skin surface cooling as a countermeasure for orthostatic intolerance. However, the feasibility of implementation of water perfused suits in real-world scenarios of orthostatic stress, which are often more reactive than preventative, is low. Johnson et al.  considerSkin surface cooling can clearly increase blood pressure; however, orthostatic stress resulting from exercise, heat, and/or a combination of both introduces additional circulatory stress, potentially complicating the effectiveness of cooling efforts. The majority of the aforementioned studies use skin surface cooling, in the absence of hyperthermia, to augment total peripheral resistance, enhance central venous return and thereby increase blood pressure. The degree to which these adjustments can be made when thermoregulatory demand remains high after the cessation of exercise is often overlooked. Logic may dictate that effectiveness in offsetting post-exercise blood pressure reduction may necessitate cooler water, greater body surface area exposure to cold, or alternative cooling media. A complicating factor in this context is the so-called “sympatholytic” effect of exercise and whole-body heat stress: vascular responses to sympathetic stimulation are blunted when compared with resting conditions . In contrast, when subjects are exposed to a neutral  or cool  post-exercise condition, both Tc and mean arterial pressure tend to return to baseline levels after 60 min. The likelihood of hypotension after exercise appears to be removed with cooler recovery conditions as a function of a quickened Tc recovery facilitated by a significant reduction in mean skin temperature. Furthermore, this study supports the notion that thermoregulatory mechanisms do play a significant role in the persistence of peripheral vasodilation post-exercise lending to the development of lowered blood pressure.Early work by Franklin et al.  suggestsAs mentioned previously, persistent vasodilation post-exercise combined with the loss of the skeletal muscle pump, leads to blood pooling in the extremities, decreasing venous return and consequently arterial pressure Rowell, . For exaOverall, elevated skeletal muscle temperature and skin perfusion following exercise in the heat contribute to a reduction in central venous pressure and a failure of TPR to increase appropriately with upright posture, leading to orthostatic intolerance. Cooling countermeasures appear to reduce both cutaneous and muscle blood flow to elicit a redistribution from the periphery to the thoracic vasculature at least when exercise is performed in thermoneutral conditions. Limited research has indicated that a reduction in large skeletal muscle microvascular perfusion following heated exercise is possible, although it appears to be smaller in magnitude than those changes seen following exercise performed in neutral ambient conditions. To improve our understanding of the influence of cooling countermeasures to prevent cardiovascular adjustments causing orthostatic intolerance, investigations examining the extent muscle and cutaneous vascular responsiveness may be blunted in response to varied cold stimuli following exercise performed in the heat are both warranted and necessary. Furthermore, it should be acknowledged that redistribution of cutaneous blood flow centrally could influence the degree of heat dissipation from the skin in a post-exercise setting. However, with a significantly widened thermal gradient elicited by skin surface cooling combined with a large preexisting degree of cutaneous vasodilation due to increased body temperatures, meaningful reductions in heat dissipation from the skin are likely minimal.Very few studies have specifically evaluated post-exercise cerebral blood flow modulation resultant from post-exercise cooling strategies. Post-exercise cooling may offset reductions in central venous pressure that would otherwise contribute to reductions in cerebral blood flow, reducing the risk of orthostatic intolerance. Skin surface cooling using a 15°C water-perfused suit immediately before head-up tilt induced orthostatic stress was successful in preventing the fall in cerebral blood flow velocity by increasing mean arterial pressure (Wilson et al., The ability of water immersion to increase central venous pressure via a shift of peripheral blood into the thoracic vasculature simultaneously stimulates high arterial pressure and low cardiopulmonary pressure baroreflexes (Pump et al., Successful orthostatic tolerance requires appropriate baroreflex responses to upright posture. During and after exercise in the heat, the ability of the baroreflex to cause vasoconstriction necessary to defend mean arterial pressure is limited by cutaneous vasodilation, elevated tissue temperature and peripheral venous pooling. Post-exercise cooling, especially cold water immersion, appears to augment both mean arterial pressure and cerebral vascular perfusion to minimize or prevent orthostatic intolerance after exercise in the heat . Still, All authors designed and outlined the work, performed literature reviews and interpreted findings, and drafted and revised the manuscript. All authors approved the final version of the manuscript and agree to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All persons designated as authors qualify for authorship, and all those who qualify for authorship are listed.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.